Publications

[No abstract available]

JTD Keywords: Alpha synuclein, Animal cell, Article, Astrocyte, Brain blood flow, Capillary endothelial cell, Cardiovascular system, Cell interaction, Coculture, Degenerative disease, Differential expression analysis, Endothelium cell, Entactin, Extracellular matrix, Fibronectin, Gene expression, Human, Human cell, Huntington chorea, Hydroxyapatite, In vitro study, Induced pluripotent stem cell, Laminin, Macrophage, Maturity, Microglia, Nervous system, Nervous system inflammation, Neuroprotection, Neurotoxicity, Nonhuman, Parkinson disease, Pericyte, Perivascular space, Personalized medicine, Shear stress, Smooth muscle cell, Three dimensional printing

The application of 3D printing to calcium phosphates has opened unprecedented possibilities for the fabrication of personalized bone grafts. However, their biocompatibility and bioactivity are counterbalanced by their high brittleness. In this work we aim at overcoming this problem by developing a self-hardening ink containing reactive ceramic particles in a polycaprolactone solution instead of the traditional approach that use hydrogels as binders. The presence of polycaprolactone preserved the printability of the ink and was compatible with the hydrolysis-based hardening process, despite the absence of water in the ink and its hydrophobicity. The microstructure evolved from a continuous polymeric phase with loose ceramic particles to a continuous network of hydroxyapatite nanocrystals intertwined with the polymer, in a configuration radically different from the polymer/ceramic composites obtained by fused deposition modelling. This resulted in the evolution from a ductile behavior, dominated by the polymer, to a stiffer behavior as the ceramic phase reacted. The polycaprolactone binder provides two highly relevant benefits compared to hydrogel-based inks. First, the handleability and elasticity of the as-printed scaffolds, together with the proven possibility of eliminating the solvent, opens the door to implanting the scaffolds freshly printed once lyophilized, while in a ductile state, and the hardening process to take place inside the body, as in the case of calcium phosphate cements. Second, even with a hydroxyapatite content of more than 92 wt.%, the flexural strength and toughness of the scaffolds after hardening are twice and five times those of the all-ceramic scaffolds obtained with the hydrogel-based inks, respectively. Statement of significance: Overcoming the brittleness of ceramic scaffolds would extend the applicability of synthetic bone grafts to high load-bearing situations. In this work we developed a 3D printing ink by replacing the conventional hydrogel binder with a water-free polycaprolactone solution. The presence of polycaprolactone not only enhanced significantly the strength and toughness of the scaffolds while keeping the proportion of bioactive ceramic phase larger than 90 wt.%, but it also conferred flexibility and manipulability to the as-printed scaffolds. Since they are able to harden upon contact with water under physiological conditions, this opens up the possibility of implanting them immediately after printing, while they are still in a ductile state, with clear advantages for fixation and press-fit in the bone defect. © 2024 The Authors

JTD Keywords: 3-d printing, 3d printing, 3d-printing, Binders, Biocompatibility, Biomimetic hydroxyapatites, Biomimetics, Bone cement, Bone scaffolds, Brittleness, Calcium phosphate, Ceramic phase, Ceramic scaffolds, Ceramics particles, Fracture mechanics, Hardening, Hardening process, Hydrogels, Hydroxyapatite, Mechanical properties, Plasticity, Polycaprolactone, Scaffolds, Scaffolds (biology), Self hardening, Strength and toughness

Polypropylene (PP), thermoplastic polyurethane (TPU), polyethylene terephthalate glycol (PETG) and polylactic acid (PLA) 3D printed specimens, which are intrinsically non-electroresponsive materials, have been converted into electroresponsive electrodes applying a low-pressure oxygen plasma treatment. After complete chemical, morphological and electrochemical characterization, plasma treated samples have been applied as integrated electrochemical sensors for detecting dopamine and serotonin by cyclic voltammetry and chronoamperometry. Results show differences in the sensing behavior, which have been explained on the basis of the chemical structure of the pristine materials. While plasma treated PLA exhibits the highest performance as electrochemical sensor in terms of sensitivity (lowest limits of detection and quantification) and selectivity (against uric acid and ascorbic acid as interfering substances), plasma treated PP displays the poorest behavior due to its low polarity compared to PLA 3D-printed electrodes. Instead, plasma treated TPU and PETG shows a very good response, much closer to PLA, as sensitive electrodes towards neurotransmitter molecules (dopamine and serotonin). Overall, results open a new door for the fabrication of electrochemical conductive sensors using intrinsically insulating materials, without the need of chemical functionalization processes.

JTD Keywords: 3d printing, Amines, Ascorbic acid, Chemical characterization, Cyclic voltammetry, Dopamine, Electrochemical characterizations, Electrochemical sensor s, Electrochemical sensors, Electrode materials, Electroresponsive materials, Low-pressure oxygen-plasma treatments, Morphological characterization, Multiwalled carbon nanotubes (mwcn), Neurophysiology, Oxygen, Oxygen plasmas, Plastic bottles, Polyethylene terephthalate glycol, Polyethylene terephthalate glycols, Polyethylene terephthalates, Polylact i c acid, Polylactic acid, Polylactic acid pla, Polyols, Polypropylene, Polypropylene oxides, Polypropylenes, Polyurethanes, Reinforced plastics, Supercapacitors, Thermoplast i c polyurethane, Thermoplastic polyurethane, Thermoplastic polyurethanes

Background: Early detection of postoperative complications after colorectal cancer (CRC) surgery is associated with improved outcomes. The aim was to investigate early metabolomics signatures capable to detect patients at risk for severe postoperative complications after CRC surgery. Materials and methods: Prospective cohort study of patients undergoing CRC surgery from 2015 to 2018. Plasma samples were collected before and after surgery, and analyzed by mass spectrometry obtaining 188 metabolites and 21 ratios. Postoperative complications were registered with Clavien-Dindo Classification and Comprehensive Complication Index. Results: One hundred forty-six patients were included. Surgery substantially modified metabolome and metabolic changes after surgery were quantitatively associated with the severity of postoperative complications. The strongest positive relationship with both Clavien-Dindo and Comprehensive Complication Index (beta=4.09 and 63.05, P<0.001) corresponded to kynurenine/tryptophan, against an inverse relationship with lysophosphatidylcholines (LPCs) and phosphatidylcholines (PCs). Patients with LPC18:2/PCa36:2 below the cut-off 0.084 mu M/mu M resulted in a sevenfold higher risk of major complications (OR=7.38, 95% CI: 2.82-21.25, P<0.001), while kynurenine/tryptophan above 0.067 mu M/mu M a ninefold (OR=9.35, 95% CI: 3.03-32.66, P<0.001). Hexadecanoylcarnitine below 0.093 mu M displayed a 12-fold higher risk of anastomotic leakage-related complications (OR=11.99, 95% CI: 2.62-80.79, P=0.004). Conclusion: Surgery-induced phospholipids and amino acid dysregulation is associated with the severity of postoperative complications after CRC surgery, including anastomotic leakage-related outcomes. The authors provide quantitative insight on metabolic markers, measuring vulnerability to postoperative morbidity that might help guide early decision-making and improve surgical outcomes.

JTD Keywords: Acids, Anastomotic leakage, Bypass, Clinical-practice guidelines, Colon, Colorectal cancer, Metabolomics, Postoperative complications, Predict, Sepsis, Trauma, Tryptophan degradation

Malaria, caused by different species of protists of the genus Plasmodium, remains among the most common causes of death due to parasitic diseases worldwide, mainly for children aged under 5. One of the main obstacles to malaria eradication is the speed with which the pathogen evolves resistance to the drug schemes developed against it. For this reason, it remains urgent to find innovative therapeutic strategies offering sufficient specificity against the parasite to minimize resistance evolution and drug side effects. In this context, nanotechnology-based approaches are now being explored for their use as antimalarial drug delivery platforms due to the wide range of advantages and tuneable properties that they offer. However, major challenges remain to be addressed to provide a cost-efficient and targeted therapeutic strategy contributing to malaria eradication. The present work contains a systematic review of nanotechnology-based antimalarial drug delivery systems generated during the last 10 years. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Infectious Disease

JTD Keywords: Adjuvant system, Antimalarial activities, Antimalarial agent, Antimalarial drug, Antimalarial drugs, Antimalarials, Artemisinin resistance, Causes of death, Child, Controlled drug delivery, Diseases, Drug delivery system, Drug delivery systems, Drug interactions, Drug side-effects, Drug-delivery, Experimental modelling, Heparan-sulfate, Human, Humans, In-vitro, Malaria, Malaria vaccine, Mannosylated liposomes, Medical nanotechnology, Models, theoretical, Nanocarriers, Nanomedicine, Nanotechnology, Parasite-, Parasitics, Plasmodium, Plasmodium-falciparum malaria, Red-blood-cells, Targeted delivery, Targeted drug delivery, Theoretical model, Therapeutic strategy

Microbial biofilms are complex three-dimensional structures where sessile microbes are embedded in a polymeric extracellular matrix. Their resistance toward the host immune system as well as to a diverse range of antimicrobial treatments poses a serious health and development threat, being in the top 10 global public health threats declared by the World Health Organization. In an effort to combat biofilm-related microbial infections, several strategies have been developed to independently eliminate biofilms or to complement conventional antibiotic therapies. However, their limitations leave room for other treatment alternatives, where the application of nanotechnology to biofilm eradication has gained significant relevance in recent years. Their small size, penetration efficiency, and the design flexibility that they present makes them a promising alternative for biofilm infection treatment, although they also present set-backs. This review aims to describe the main possibilities and limitations of nanomedicine against biofilms, while covering the main aspects of biofilm formation and study, and the current therapies for biofilm treatment. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Infectious Disease Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicine.

JTD Keywords: Anti-bacterial agents, Anti-infective agents, Antiinfective agent, Antimicrobial, Antimicrobials, Antimicrobials,bacteria,biofilm,infectious diseases,microorganism, Bacteria, Biofilm, Biofilm infections, Biofilms, Complex three dimensional structures, Diseases, Diverse range, Drug-delivery systems,in-vitro,cellular toxicity,nanoparticles,penetration,model,biocompatibility,perspectives,hyperthermia,diagnosi, Extracellular matrices, Global public health, Health risks, Infectious disease, Infectious diseases, Medical nanotechnology, Microbial biofilm, Microorganisms, Nanomedicine, Polymer, Polymers, Regulatory issues, Roadmap

The performance of single-chain polymeric nanoparticles (SCPNs) in biomedical applications highly depends on their conformational stability in cellular environments. Until now, such stability studies are limited to 2D cell culture models, which do not recapitulate the 3D tumor microenvironment well. Here, a microfluidic tumor-on-a-chip model is introduced that recreates the tumor milieu and allows in-depth insights into the diffusion, cellular uptake, and stability of SCPNs. The chip contains Matrigel/collagen-hyaluronic acid as extracellular matrix (ECM) models and is seeded with cancer cell MCF7 spheroids. With this 3D platform, it is assessed how the polymer's microstructure affects the SCPN's behavior when crossing the ECM, and evaluates SCPN internalization in 3D cancer cells. A library of SCPNs varying in microstructure is prepared. All SCPNs show efficient ECM penetration but their cellular uptake/stability behavior depends on the microstructure. Glucose-based nanoparticles display the highest spheroid uptake, followed by charged nanoparticles. Charged nanoparticles possess an open conformation while nanoparticles stabilized by internal hydrogen bonding retain a folded structure inside the tumor spheroids. The 3D microfluidic tumor-on-a-chip platform is an efficient tool to elucidate the interplay between polymer microstructure and SCPN's stability, a key factor for the rational design of nanoparticles for targeted biological applications.© 2024 The Authors. Small Methods published by Wiley-VCH GmbH.

JTD Keywords: 3d cancer cell uptake, Cancer cells, Cell culture, Cell uptake, Cellular uptake, Diseases, Ecm penetration, Extracellular matrices, Extracellular matrix penetration, Functional polymers, Hydrogen bonds, Medical applications, Microfluidics, Microstructure, Nanoparticles, Polymeric nanoparticles, Scpns, Single chains, Single-chain polymeric nanoparticle, Stability, Tumor-on-a-chip, Tumors

Interactions between living cells and nanoparticles are extensively studied to enhance the delivery of therapeutics. Nanoparticles size, shape, stiffness, and surface charge are regarded as the main features able to control the fate of cell-nanoparticle interactions. However, the clinical translation of nanotherapies has so far been limited, and there is a need to better understand the biology of cell-nanoparticle interactions. This study investigates the role of cellular mechanosensitive components in cell-nanoparticle interactions. It is demonstrated that the genetic and pharmacologic inhibition of yes-associated protein (YAP), a key component of cancer cell mechanosensing apparatus and Hippo pathway effector, improves nanoparticle internalization in triple-negative breast cancer cells regardless of nanoparticle properties or substrate characteristics. This process occurs through YAP-dependent regulation of endocytic pathways, cell mechanics, and membrane organization. Hence, the study proposes targeting YAP may sensitize triple-negative breast cancer cells to chemotherapy and increase the selectivity of nanotherapy.© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.

JTD Keywords: cancer treatment, cells, differentiation, hippo pathway, mechanics, mechanobiology, mechanotransduction, nanoparticles, progression, protein, resistance, yap-signaling, yap/taz, Adaptor proteins, signal transducing, Bio-nano interaction, Bio-nano interactions, Breast cancer cells, Cancer cells, Cancer treatment, Cells, Cellular therapeutics, Cellular uptake, Chemotherapy, Cytology, Diseases, Extracellular-matrix, Human, Humans, Mechano-biology, Mechanobiology, Metabolism, Nanoparticle, Nanoparticle interaction, Nanoparticles, Physiology, Protein serine threonine kinase, Protein serine-threonine kinases, Protein signaling, Signal transducing adaptor protein, Signal transduction, Therapeutic effects, Triple negative breast cancer, Triple negative breast neoplasms, Triple-negative breast cancers, Yap-signaling, Yes-associated protein-signaling

The accurate spatial segregation into distinct phases within cell membranes coordinates vital biochemical processes and functionalities in living organisms. One of nature's strategies to localize reactivity is the formation of dynamic raft domains. Most raft models rely on liquid-ordered L-0 phases in a liquid-disordered L-d phase lacking correlation and remaining static, often necessitating external agents for phase separation. Here, we introduce a synthetic system of bicomponent glycodendrimersomes coassembled from Janus dendrimers and Janus glycodendrimers (JGDs), where lactose-lactose interactions exclusively drive lateral organization. This mechanism results in modulated phases across two length scales, yielding raft-like microdomains featuring nanoarrays at the nanoscale. By varying the density of lactose and molecular architecture of JGDs, the nanoarray type and size, shape, and spacing of the domains were controlled. Our findings offer insight into the potential primordial origins of rudimentary raft domains and highlight the crucial role of glycans within the glycocalyx.

JTD Keywords: Article, Artificial cells, Atomic force microscopy, Bicomponents, Bilayer, Bilayer membrane, Biochemical functionality, Biochemical process, Biological-membranes, Cell component, Cell membrane, Cellular parameters, Chemical interaction, Chemical structure, Chemistry, Cytology, Defined janus glycodendrimers, Dehydration, Dendrimer, Dendrimers, Dilution, Dimer, External agents, Fourier transform, Giant vesicles, Glycan, Glycans, Glycocalyx, Glycodendrimers, Janus dendrimer, Janus glycodendrimer, Lactose, Lateral organization, Lectin, Lipid rafts, Living organisms, Membrane damage, Membrane microdomain, Membrane microdomains, Membrane structure, Metabolism, Modulated phases, Molecule, Monomer, Nanoarrays, Oligosaccharide, Organization, Periodicity, Phase separation, Phase-separation, Phospholipids, Polysaccharide, Polysaccharides, Raft like domain, Relative humidity, Spatial segregation, Structure analysis, Sugars, Synthetic systems, Tetramer, Unclassified drug, Unilamellar vesicles, Water

Although we have learned much about how the brain fuels its functions over the last decades, there remains much still to discover in an organ that is so complex. This article lays out major gaps in our knowledge of interrelationships between brain metabolism and brain function, including biochemical, cellular, and subcellular aspects of functional metabolism and its imaging in adult brain, as well as during development, aging, and disease. The focus is on unknowns in metabolism of major brain substrates and associated transporters, the roles of insulin and of lipid droplets, the emerging role of metabolism in microglia, mysteries about the major brain cofactor and signaling molecule NAD+, as well as unsolved problems underlying brain metabolism in pathologies such as traumatic brain injury, epilepsy, and metabolic downregulation during hibernation. It describes our current level of understanding of these facets of brain energy metabolism as well as a roadmap for future research. This article details current knowledge and major unknowns in brain energy metabolism and lays out a roadmap for future research.image

JTD Keywords: Acetate, Acetyl-coa, Aerobic glycolysis, Atp-citrate lyase, Extracellular glutamate concentration, Fatty-acid transport, Glucose-metabolism, Glut4, In-vivo, Insulin, Lipid droplet accumulation, Nicotinamide adenine-dinucleotide, Noradrenaline, Obese zucker rats, Rat cerebral-cortex

Surface electromyography (sEMG) can be used to measure the electrical activity of the respiratory muscles. The possible applications of sEMG span from patients suffering from acute respiratory failure to patients receiving chronic home mechanical ventilation, to evaluate muscle function, titrate ventilatory support and guide treatment. However, sEMG is mainly used as a monitoring tool for research and its use in clinical practice is still limited-in part due to a lack of standardization and transparent reporting. During this round table meeting, recommendations on data acquisition, processing, interpretation, and potential clinical applications of respiratory sEMG were discussed. This paper informs the clinical researcher interested in respiratory muscle monitoring about the current state of the art on sEMG, knowledge gaps and potential future applications for patients with respiratory failure.

JTD Keywords: Acute respiratory failure, Artificial ventilation, Asthmatic-children, Breathing muscle, Clinical monitoring, Clinical practice, Clinical research, Consensus development, Data interpretation, Disease exacerbation, Drive, Electrode positioning, Electrode removal, Electromyography, Force, Home care, Human, Human diaphragm, Humans, Information processing, Inspiratory muscle training, Inspiratory muscles, Intensive care unit, Knowledge gap, Long term care, Mechanical ventilation, Medical procedures, Muscle contraction, Muscle fatigue, Muscle function, Muscle training, Muscle, skeletal, Muscle-activity, Noninvasive ventilation, Patient monitoring, Patient-ventilator asynchrony, Physiology, Prognosis, Quality of life, Reporting and data system, Respiratory failure, Respiratory muscles, Review, Severe exacerbations, Signal processing, Skeletal muscle, Standardization, Surface electromyography, Time factor

Degradable polymeric micelles are promising drug delivery systems due to their hydrophobic core and responsive design. When applying micellar nanocarriers for tumor delivery, one of the bottlenecks encountered in vivo is the tumor tissue barrier: crossing the dense mesh of cells and the extracellular matrix (ECM). Sometimes overlooked, the extracellular matrix can trap nanoformulations based on charge, size, and hydrophobicity. Here, we used a simple design of a microfluidic chip with two types of ECM and MCF7 spheroids to allow high-throughput screening of the interactions between biological interfaces and polymeric micelles. To demonstrate the applicability of the chip, a small library of fluorescently labeled polymeric micelles varying in their hydrophilic shell and hydrophobic core forming blocks was studied. Three widely used hydrophilic shells were tested and compared, namely, poly(ethylene glycol), poly(2-ethyl-2-oxazoline), and poly(acrylic acid), along with two enzymatically degradable dendritic hydrophobic cores (based on hexyl or nonyl end groups). Using ratiometric imaging of unimer:micelle fluorescence and FRAP inside the chip model, we obtained the local assembly state and dynamics inside the chip. Notably, we observed different micelle behaviors in the basal lamina ECM, from avoidance of the ECM structure to binding of the poly(acrylic acid) formulations. Binding to the basal lamina correlated with higher uptake into MCF7 spheroids. Overall, we proposed a simple microfluidic chip containing dual ECM and spheroids for the assessment of the interactions of polymeric nanocarriers with biological interfaces and evaluating nanoformulations' capacity to cross the tumor tissue barrier.

JTD Keywords: Extracellular matrix, Microfluidics, Nanoparticle mobility, Polymeric micelles, Tumor-on-a-chip

The immobilization of vesicles has been conceptualized as a method to functionalize biointerfaces. However, the preservation of their integrity post immobilization remains a considerable challenge. Interfacial interactions can cause vesicle rupture upon close surface contact and non-specific protein adsorption impairing surface functions. To date, immobilization of vesicles has relied solely on either entrapment or prior modification of vesicles, both of which require laborious preparation and limit their applications. This work develops a bioinspired strategy to pin vesicles without prior modification while preserving their intact shape. This work introduces antifouling diblock copolymers and ultrathin surface-attached hydrogels containing a brush-like interface consisting of a bottle brush copolymer of N-(2-hydroxypropyl) methacrylamide (HPMA) and N-(3-methacrylamidopropyl)-N,N-dimethyldodecan-1-aminiumiodide (C12+). The presence of positive charges generates an attractive force that pulls vesicles toward the surface. At the surface, the amphiphilic properties of the combs facilitate their insertion into the membrane, mimicking the harpooning mechanism observed in antimicrobial peptides. Importantly, the antifouling poly(HPMA) backdrop serves to safeguard the vesicles by preventing deformation and breakage. Using a combination of thermodynamic analysis, surface plasmon resonance, and confocal laser scanning microscopy, this work demonstrates the efficiency of this biomimetic system to capture vesicles while maintaining an antifouling interface necessary for bioapplications. This work presents a novel supramolecular approach that combines three key elements: long-range attraction, vesicle pinning, and short-range repulsion to attract and harpoon vesicles, while protecting them at the surface. This work envisions these coatings as universal and biocompatible platforms that can be used not only to study vesicle interactions, but also as tools for biomedical applications.image

JTD Keywords: Antifouling coatings, Coatings, Delivery, Extracellular vesicles, Fabrication, Hydrogel, Janus dendrimers, Lipid vesicles, Liposomes, Membrane insertion, Polymer brushes, Proteins, Surface-energy components, Ultrathin surface-attached hydrogels, Vesicle pinning

Using nanoparticles (NPs) in drug delivery has exhibited promising therapeutic potential in various cancer types. Nevertheless, several challenges must be addressed, including the formation of the protein corona, reduced targeting efficiency and specificity, potential immune responses, and issues related to NP penetration and distribution within 3-dimensional tissues. To tackle these challenges, we have successfully integrated iron oxide nanoparticles into neuroblastoma-derived extracellular vesicles (EVs) using the parental labeling method. We first developed a tissue-engineered (TE) neuroblastoma model, confirming the viability and proliferation of neuroblastoma cells for at least 12 days, supporting its utility for EV isolation. Importantly, EVs from long-term cultures exhibited no differences compared to short-term cultures. Concurrently, we designed Rhodamine (Rh) and Polyacrylic acid (PAA)-functionalized magnetite nanoparticles (Fe3O4@PAA-Rh) with high crystallinity, purity, and superparamagnetic properties (average size: 9.2 +/- 2.5 nm). We then investigated the internalization of Fe3O4@PAA-Rh nanoparticles within neuroblastoma cells within the TE model. Maximum accumulation was observed overnight while ensuring robust cell viability. However, nanoparticle internalization was low. Taking advantage of the enhanced glucose metabolism exhibited by cancer cells, glucose (Glc)-functionalized nanoparticles (Fe3O4@PAA-Rh-Glc) were synthesized, showing superior cell uptake within the 3D model without inducing toxicity. These glucose-modified nanoparticles were selected for parental labeling of the TE models, showing effective NP encapsulation into EVs. Our research introduces innovative approaches to advance NP delivery, by partially addressing the challenges associated with 3D systems, optimizing internalization, and enhancing NP stability and specificity through EV-based carriers. Also, our findings hold the promise of more precise and effective cancer therapies while minimizing potential side effects.

JTD Keywords: Biomimetic models, Extracellular vesicles, Iron oxide nanoparticles, Neuroblastoma, Precision medicine

Recently, organoids have emerged as revolutionizing tools with the unprecedented potential to recreate organ-specific microanatomy in vitro. Upon their derivation from human pluripotent stem cells (hPSCs), organoids reveal the blueprints of human organogenesis, further allowing the faithful recapitulation of their physiology. Nevertheless, along with the evolution of this field, advanced research exposed the organoids' shortcomings, particularly regarding poor reproducibility rates and overall immatureness. To resolve these challenges, many studies have started to underscore the relevance of mechanical cues as a relevant source to induce and externally control hPSCs differentiation. Indeed, established organoid generation protocols from hPSCs have mainly relyed on the biochemical induction of fundamental signalling pathways present during kidney formation in mammals, whereas mechanical cues have largely been unexplored. This review aims to discuss the pertinence of (bio) physical cues within hPSCs-derived organoid cultures, while deciphering their effect on morphogenesis. Moreover, we will explore state-of-the-art mechanobiology techniques as revolutionizing means for understanding the underlying role of mechanical forces in biological processes in organoid model systems.

JTD Keywords: development, hpscs, mechanobiology, nephrogenesis, Activated ion-channel, Development, Extracellular-matrix viscoelasticity, Forces, Hpscs, Ips cells, Mechanical regulation, Mechanobiology, Nephrogenesis, Nephron progenitors, Organoids, Pluripotent stem-cells, Self-renewal, Substrate mechanics, Tissue

Many neurodegenerative diseases are identified but their causes and cure are far from being well-known. The problem resides in the complexity of the neural tissue and its location which hinders its easy evaluation. Although necessary in the drug discovery process, in vivo animal models need to be reduced and show relevant differences with the human tissues that guide scientists to inquire about other possible options which lead to in vitro models being explored. From organoids to organ-on-a-chips, 3D models are considered the cutting-edge technology in cell culture. Cell choice is a big parameter to take into consideration when planning an in vitro model and cells capable of mimicking both healthy and diseased tissue, such as induced pluripotent stem cells (iPSC), are recognized as good candidates. Hence, we present a critical review of the latest models used to study neurodegenerative disease, how these models have evolved introducing microfluidics platforms, 3D cell cultures, and the use of induced pluripotent cells to better mimic the neural tissue environment in pathological conditions.

JTD Keywords: 3d in vitro models, bioprinting, ipsc cell culture, microfluidic device, 3d in vitro models, Bioprinting, Blood-brain-barrier, Cerebral organoids, Culture model, Endothelial-cells, Expression profile, Extracellular-matrix, Ipsc cell culture, Microfluidic device, Neurodegenerative diseases, On-a-chip, Pluripotent stem-cells, Shear-stress, Substrate stiffness

Decellularized extracellular matrix (dECM) hydrogels have emerged as promising materials in tissue engineering. The steps to produce dECM hydrogels containing the bioactive epitopes found in the native matrix are often laborious, including the initial harvesting and decellularization of the animal organ. Furthermore, resulting hydrogels often exhibit weak mechanical properties that require the use of additional crosslinkers such as genipin to truly simulate the mechanical properties of the desired study tissue. In this work, we have developed a protocol to readily obtain tens of thin dECM hydrogel cryosections attached to a glass slide as support, to serve as scaffolds for two-dimensional (2D) or three-dimensional (3D) cell culture. Following extensive atomic force microscopy (AFM)-based mechanical characterization of dECM hydrogels crosslinked with increasing genipin concentrations (5 mM, 10 mM, and 20 mM), we provide detailed protocol recommendations for achieving dECM hydrogels of any biologically relevant stiffness. Given that our protocol requires hydrogel freezing, we also confirm that the approach taken can be further used to increase the mechanical properties of the scaffold in a controlled manner exhibiting twice the stiffness in highly crosslinked arrays. Finally, we explored the effect of ethanol-based short- and long-term sterilization on dECM hydrogels, showing that in some situations it may give rise to significant changes in hydrogel mechanical properties that need to be taken into account in experimental design. The hydrogel cryosections produced were shown to be biocompatible and support cell attachment and spreading for at least 72 h in culture. In brief, our proposed method may provide several advantages for tissue engineering: (1) easy availability and reduction in preparation time, (2) increase in the total hydrogel volume eventually used for experiments being able to obtain 15-22 slides from a 250 µL hydrogel) with a (3) reduction in scaffold variability (only a 17.5 ± 9.5% intraslide variability provided by the method), and (4) compatibility with live-cell imaging techniques or further cell characterization of cells.

JTD Keywords: atomic force microscopy, cryogel, cryosectioning, decellularization, extracellular matrix, genipin, sterilization, stiffness, young's modulus, Atomic force microscopy, Cryogel, Cryosectioning, Decellularization, Extracellular matrix, Genipin, Hydrogel, Sterilization, Stiffness, Young’s modulus

The intestine is a complex tissue with a characteristic three-dimensional (3D) crypt-villus architecture, which plays a key role in the intestinal function. This function is also regulated by the intestinal stroma that actively supports the intestinal epithelium, maintaining the homeostasis of the tissue. Efforts to account for the 3D complex structure of the intestinal tissue have been focused mainly in mimicking the epithelial barrier, while solutions to include the stromal compartment are scarce and unpractical to be used in routine experiments. Here we demonstrate that by employing an optimized bioink formulation and the suitable printing parameters it is possible to produce fibroblast-laden crypt-villus structures by means of digital light projection stereolithography (DLP-SLA). This process provides excellent cell viability, accurate spatial resolution, and high printing throughput, resulting in a robust biofabrication approach that yields functional gut mucosa tissues compatible with conventional testing techniques.Copyright © 2023 Elsevier B.V. All rights reserved.

JTD Keywords: 3d microstructure, barrier, cells, epithelial-stromal interactions, gelma-pegda soft hydrogels, growth, hydrogel, intestinal mucosa model, methacrylamide, microfabrication, proliferation, scaffold, stereolithography, 3d bioprinting, 3d microstructure, Epithelial-stromal interactions, Fibroblasts, Gelma-pegda soft hydrogels, Intestinal mucosa model

The small intestine is a complex organ with a characteristic architecture and a major site for drug and nutrient absorption. The three-dimensional (3D) topography organized in finger-like protrusions called villi increases surface area remarkably, granting a more efficient absorption process. The intestinal mucosa, where this process occurs, is a multilayered and multicell-type tissue barrier. In vitro intestinal models are routinely used to study different physiological and pathological processes in the gut, including compound absorption. Still, standard models are typically two-dimensional (2D) and represent only the epithelial barrier, lacking the cues offered by the 3D architecture and the stromal components present in vivo, often leading to inaccurate results. In this work, we studied the impact of the 3D architecture of the gut on drug transport using a bioprinted 3D model of the intestinal mucosa containing both the epithelial and the stromal compartments. Human intestinal fibroblasts were embedded in a previously optimized hydrogel bioink, and enterocytes and goblet cells were seeded on top to mimic the intestinal mucosa. The embedded fibroblasts thrived inside the hydrogel, remodeling the surrounding extracellular matrix. The epithelial cells fully covered the hydrogel scaffolds and formed a uniform cell layer with barrier properties close to in vivo. In particular, the villus-like model revealed overall increased permeability compared to a flat counterpart composed by the same hydrogel and cells. In addition, the efflux activity of the P-glycoprotein (P-gp) transporter was significantly reduced in the villus-like scaffold compared to a flat model, and the genetic expression of other drugs transporters was, in general, more relevant in the villus-like model. Globally, this study corroborates that the presence of the 3D architecture promotes a more physiological differentiation of the epithelial barrier, providing more accurate data on drug absorbance measurements.Copyright © 2023. Published by Elsevier B.V.

JTD Keywords: 3d architecture, alkaline-phosphatase, caco-2 cells, culture, drug development, efflux proteins, gene-expression, human-colon, intestinal absorption, intestinal models, microenvironment, paracellular transport, permeability, photopolymerization, villi, 3d architecture, 3d bioprinting, Drug development, In-vitro, Intestinal absorption, Intestinal models, Photopolymerization, Villi

As cells migrate and experience forces from their surroundings, they constantly undergo mechanical deformations which reshape their plasma membrane (PM). To maintain homeostasis, cells need to detect and restore such changes, not only in terms of overall PM area and tension as previously described, but also in terms of local, nanoscale topography. Here, we describe a novel phenomenon, by which cells sense and restore mechanically induced PM nanoscale deformations. We show that cell stretch and subsequent compression reshape the PM in a way that generates local membrane evaginations in the 100 nm scale. These evaginations are recognized by I-BAR proteins, which triggers a burst of actin polymerization mediated by Rac1 and Arp2/3. The actin polymerization burst subsequently re-flattens the evagination, completing the mechanochemical feedback loop. Our results demonstrate a new mechanosensing mechanism for PM shape homeostasis, with potential applicability in different physiological scenarios.© 2023, Quiroga et al.

JTD Keywords: arp2/3 complex, bar, bar proteins, cdc42, cells, domain, human, irsp53, membrane biophysics, mouse, proteins, rac, tension, Actin polymerization, Bar proteins, Cell biology, Human, Mechanobiology, Membrane biophysics, Mouse, Physics of living systems

Since the emergence of 3D bioprinting technology, both synthetic and natural materials have been used to develop bioinks for producing cell-laden cardiac grafts. To this end, extracellular-matrix (ECM)-derived hydrogels can be used to develop scaffolds that closely mimic the complex 3D environments for cell culture. This study presents a novel cardiac bioink based on hydrogels exclusively derived from decellularized porcine myocardium loaded with human-bone-marrow-derived mesenchymal stromal cells. Hence, the hydrogel can be used to develop cell-laden cardiac patches without the need to add other biomaterials or use additional crosslinkers. The scaffold ultrastructure and mechanical properties of the bioink were characterized to optimize its production, specifically focusing on the matrix enzymatic digestion time. The cells were cultured in 3D within the developed hydrogels to assess their response. The results indicate that the hydrogels fostered inter-cell and cell-matrix crosstalk after 1 week of culture. In conclusion, the bioink developed and presented in this study holds great potential for developing cell-laden customized patches for cardiac repair.

JTD Keywords: biology, biomaterials, collagen, decellularized cardiac tissue, extracellular matrix, hydrogels, mesenchymal stromal cells, 3d bioprinting, Biomaterials, Decellularized cardiac tissue, Extracellular matrix, Hydrogels, Mesenchymal stem-cells, Mesenchymal stromal cells

There is a growing interest in developing natural hydrogel-based scaffolds to culture cells in a three-dimensional (3D) millieu that better mimics the in vivo cells' microenvironment. A promising approach is to use hydrogels from animal tissues, such as decellularized extracellular matrices; however, they usually exhibit suboptimal mechanical properties compared to native tissue and their composition with hundreds of different protein complicates to elucidate which stimulus triggers cell's responses. As simpler scaffolds, type I collagen hydrogels are used to study cell behavior in mechanobiology even though they are also softer than native tissues. In this work, type I collagen is mixed with bacterial nanocellulose fibers (BCf) to develop reinforced scaffolds with mechanical properties suitable for 3D cell culture. BCf were produced from blended pellicles biosynthesized from Komagataeibacter xylinus. Then, BCf were mixed with concentrated collagen from rat-tail tendons to form composite hydrogels. Confocal laser scanning microscopy and scanning electron microscopy images confirmed the homogeneous macro- and microdistribution of both natural polymers. Porosity analysis confirmed that BCf do not disrupt the scaffold structure. Tensile strength and rheology measurements demonstrated the reinforcement action of BCf (43% increased stiffness) compared to the collagen hydrogel while maintaining the same viscoelastic response. Additionally, this reinforcement of collagen hydrogels with BCf offers the possibility to mix cells before gelation and then proceed to the culture of the 3D cell scaffolds. We obtained scaffolds with human bone marrow-derived mesenchymal stromal cells or human fibroblasts within the composite hydrogels, allowing a homogeneous 3D viable culture for at least 7 days. A smaller surface shrinkage in the reinforced hydrogels compared to type I collagen hydrogels confirmed the strengthening of the composite hydrogels. These collagen hydrogels reinforced with BCf might emerge as a promising platform for 3D in vitro organ modeling, tissue-engineering applications, and suitable to conduct fundamental mechanobiology studies.

JTD Keywords: 3d cell culture, bacterial cellulose, collagen, composite hydrogels, 3d cell culture, Bacterial cellulose, Cellulose/collagen composite, Collagen, Composite hydrogels, Contraction, Cross-linking, Cytocompatibility, Fibroblasts, Matrix, Mechanical-properties, Reinforcement, Stiffness, Tissue engineering

Cancer progression is caused by genetic changes and associated with various alterations in cell properties, which also affect a tumor's mechanical state. While an increased stiffness has been well known for long for solid tumors, it has limited prognostic power. It is hypothesized that cancer progression is accompanied by tissue fluidization, where portions of the tissue can change position across different length scales. Supported by tabletop magnetic resonance elastography (MRE) on stroma mimicking collagen gels and microscopic analysis of live cells inside patient derived tumor explants, an overview is provided of how cancer associated mechanisms, including cellular unjamming, proliferation, microenvironment composition, and remodeling can alter a tissue's fluidity and stiffness. In vivo, state-of-the-art multifrequency MRE can distinguish tumors from their surrounding host tissue by their rheological fingerprints. Most importantly, a meta-analysis on the currently available clinical studies is conducted and universal trends are identified. The results and conclusions are condensed into a gedankenexperiment about how a tumor can grow and eventually metastasize into its environment from a physics perspective to deduce corresponding mechanical properties. Based on stiffness, fluidity, spatial heterogeneity, and texture of the tumor front a roadmap for a prognosis of a tumor's aggressiveness and metastatic potential is presented.© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.

JTD Keywords: brain, cancer, cells, collective migration, elastic energy, elastography, in vivo magnetic resonance elastography, invasion, medical imaging, solid stress, tissue fluidity, tumor mechanics, viscoelastic properties, Cancer, Extracellular-matrix, In vivo magnetic resonance elastography, Medical imaging, Tissue fluidity, Tumor mechanics

Ratiometric fluorescent nanothermometers with near-infrared emission play an important role in in vivo sensing since they can be used as intracellular thermal sensing probes with high spatial resolution and high sensitivity, to investigate cellular functions of interest in diagnosis and therapy, where current approaches are not effective. Herein, the temperature-dependent fluorescence of organic nanoparticles is designed, synthesized, and studied based on the dual emission, generated by monomer and excimer species, of the tris(2,4,6-trichlorophenyl)methyl radical (TTM) doping organic nanoparticles (TTMd-ONPs), made of optically neutral tris(2,4,6-trichlorophenyl)methane (TTM-αH), acting as a matrix. The excimer emission intensity of TTMd-ONPs decreases with increasing temperatures whereas the monomer emission is almost independent and can be used as an internal reference. TTMd-ONPs show a great temperature sensitivity (3.4% K-1 at 328 K) and a wide temperature response at ambient conditions with excellent reversibility and high colloidal stability. In addition, TTMd-ONPs are not cytotoxic and their ratiometric outputs are unaffected by changes in the environment. Individual TTMd-ONPs are able to sense temperature changes at the nano-microscale. In vivo thermometry experiments in Caenorhabditis elegans (C. elegans) worms show that TTMd-ONPs can locally monitor internal body temperature changes with spatio-temporal resolution and high sensitivity, offering multiple applications in the biological nanothermometry field.© 2023 The Authors. Small published by Wiley-VCH GmbH.

JTD Keywords: dual emission, elegans, excimer emission, fluorescence, in vivo sensing, luminescence, nanoparticles, organic radical nanoparticles, ratiometric nanothermometers, sensors, thermometry, trityl radicals, Caenorhabditis elegans, Excimer emission, In vivo sensing, Intracellular ph, Luminescence, Organic radical nanoparticles, Ratiometric nanothermometers, Trityl radicals

JTD Keywords: extracellular vesicles, microfluidic platforms, micrornas, obesity, plasma, rnas, Biomarkers, Mirnas

The formation of a biological seal around the neck of titanium (Ti) implants is critical for ensuring integration at the gingival site and for preventing bacterial colonization that may lead to periimplantitis. This process is guided by activated fibroblasts, named myofibroblasts, which secrete extracellular matrix (ECM) proteins and ECM-degrading enzymes resolving the wound. However, in some cases, Ti is not able to attract and activate fibroblasts to a sufficient extent, which may compromise the success of the implant. Fibronectin (FN) is an ECM component found in wounds that is able to guide soft tissue healing through the adhesion of cells and attraction of growth factors (GFs). However, clinical use of FN functionalized Ti implants is problematic because FN is difficult to obtain, and is sensitive to degradation. Herein, functionalizing Ti with a modified recombinant heparin binding II (HBII) domain of FN, mutated to include an Arg-Gly-Asp (RGD) sequence for promoting both fibroblast adhesion and GF attraction, is aimed at. The HBII-RGD domain is able to stimulate fibroblast adhesion, spreading, proliferation, migration, and activation to a greater extent than the native HBII, reaching values closer to those of full-length FN suggesting that it might induce the formation of a biological sealing.© 2023 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.

JTD Keywords: alpha-4-beta-1, beta, cell-binding, collagen, extracellular-matrix, fibroblast activation, fibronectin, growth factors, integrins, metalloproteinases, myofibroblasts, proliferation, soft-tissue integration, titanium, Biological-activities, Fibroblast activation, Titanium

The Bin/amphiphysin/Rvs (BAR) superfamily proteins have a crescent binding domain and bend biomembranes along the domain axis. However, their anisotropic bending rigidities and spontaneous curvatures have not been experimentally determined. Here, we estimated these values from the bound protein densities on tethered vesicles using a mean-field theory of anisotropic bending energy and orientation-dependent excluded volume. The dependence curves of the protein density on the membrane curvature are fitted to the experimental data for the I-BAR and N-BAR domains reported by C. Prevost et al. Nat. Commun., 2015, 6, 8529 and F.-C. Tsai et al. Soft Matter, 2021, 17, 4254-4265, respectively. For the I-BAR domain, all three density curves of different chemical potentials exhibit excellent fits with a single parameter set of anisotropic bending energy. When the classical isotropic bending energy is used instead, one of the curves can be fitted well, but the others exhibit large deviations. In contrast, for the N-BAR domain, two curves are not well fitted simultaneously the anisotropic model, although it is significantly improved compared to the isotropic model. This deviation likely suggests a cluster formation of the N-BAR domains.

JTD Keywords: Membrane-mediated interactions,elastic properties,bar,shape,mechanisms,inclusions,generation,polymers,driven,bod

Fibrillar collagens are the most abundant extracellular matrix components in non-small cell lung cancer (NSCLC). However, the potential of collagen fiber descriptors as a source of clinically relevant biomarkers in NSCLC is largely unknown. Similarly, our understanding of the aberrant collagen organization and associated tumor-promoting effects is very scarce. To address these limitations, we identified a digital pathology approach that can be easily implemented in pa-thology units based on CT-FIRE software (version 2; https://loci.wisc.edu/software/ctfire) analysis of Picrosirius red (PSR) stains of fibrillar collagens imaged with polarized light (PL). CT-FIRE set-tings were pre-optimized to assess a panel of collagen fiber descriptors in PSR-PL images of tissue microarrays from surgical NSCLC patients (106 adenocarcinomas [ADC] and 89 squamous cell carcinomas [SCC]). Using this approach, we identified straightness as the single high-accuracy diagnostic collagen fiber descriptor (average area under the curve 1/4 0.92) and fiber density as the single descriptor consistently associated with a poor prognosis in both ADC and SCC inde-pendently of the gold standard based on the TNM staging (hazard ratio, 2.69; 95% CI, 1.55-4.66; P < .001). Moreover, we found that collagen fibers were markedly straighter, longer, and more aligned in tumor samples compared to paired samples from uninvolved pulmonary tissue, particularly in ADC, which is indicative of increased tumor stiffening. Consistently, we observed an increase in a panel of stiffness-associated processes in the high collagen fiber density patient group selectively in ADC, including venous/lymphatic invasion, fibroblast activation (a-smooth muscle actin), and immune evasion (programmed death-ligand 1). Similarly, a transcriptional correlation analysis supported the potential involvement of the major YAP/TAZ pathway in ADC. Our results provide a proof-of-principle to use CT-FIRE analysis of PSR-PL images to assess new collagen fiber-based diagnostic and prognostic biomarkers in pathology units, which may improve the clinical management of patients with surgical NSCLC. Our findings also unveil an aberrant stiff micro -environment in lung ADC that may foster immune evasion and dissemination, encouraging future work to identify therapeutic opportunities. (c) 2023 THE AUTHORS. Published by Elsevier Inc. on behalf of the United States & Canadian Academy of Pathology. This is an open access article under the CC BY-NC-ND license (http://creativecommo ns.org/licenses/by-nc-nd/4.0/).

JTD Keywords: biomarkers, collagen, ct-fire, lung cancer, mechanobiology, Adenocarcinoma, Association, Biomarkers, Collagen, Ct-fire, Differentiation, Expression, Extracellular-matrix, I collagen, Invasion, Lung cancer, Mechanobiology, Microenvironment, Signature, Survival, Tumor microenvironment

JTD Keywords: actin cytoskeleton, extracellular vesicles, vesicular transport, virulence factors, Actin cytoskeleton, Extracellular vesicles, Host-parasite interactions, Vesicular transport, Virulence factors

Sprouting angiogenesis is a core biological process critical to vascular development. Its accurate simulation, relevant to multiple facets of human health, is of broad, interdisciplinary appeal. This study presents an in-silico model replicating a microfluidic assay where endothelial cells sprout into a biomimetic extracellular matrix, specifically, a large-pore, low-concentration fibrin-based porous hydrogel, influenced by chemotactic factors. We introduce a novel approach by incorporating the extracellular matrix and chemotactic factor effects into a unified term using a single parameter, primarily focusing on modelling sprouting dynamics and morphology. This continuous model naturally describes chemotactic-induced sprouting with no need for additional rules. In addition, we extended our base model to account for matrix sensing and degradation, crucial aspects of angiogenesis. We validate our model via a hybrid in-silico experimental method, comparing the model predictions with experimental results derived from the microfluidic setup. Our results underscore the intricate relationship between the extracellular matrix structure and angiogenic sprouting, proposing a promising method for predicting the influence of the extracellular matrix on angiogenesis.Copyright © 2023 Ferre-Torres, Noguera-Monteagudo, Lopez-Canosa, Romero-Arias, Barrio, Castaño and Hernandez-Machado.

JTD Keywords: angiogenesis, biomimmetic, chemotaxis, endothelial cells, filopodia, growth, in silico model, mathematical models, mechanisms, metalloproteinase, migration, morphogenesis, phase field, pore-size, simulation, Angiogenesis, Biomimmetic, Chemotaxis, Endothelial cells, Extracellular matrix, In silico model, Mathematical models, Phase field, Tip cells

The bacterium Pseudomonas aeruginosa is involved in chronic infections of cystic fibrosis lungs and chronic wounds. In these infections the bacteria are present as aggregates suspended in host secretions. During the course of the infections there is a selection for mutants that overproduce exopolysaccharides, suggesting that the exopolysaccharides play a role in the persistence and antibiotic tolerance of the aggregated bacteria. Here, we investigated the role of individual P. aeruginosa exopolysaccharides in aggregate-associated antibiotic tolerance. We employed an aggregate-based antibiotic tolerance assay on a set of P. aeruginosa strains that were genetically engineered to over-produce a single, none, or all of the three exopolysaccharides Pel, Psl, and alginate. The antibiotic tolerance assays were conducted with the clinically relevant antibiotics tobramycin, ciprofloxacin and meropenem. Our study suggests that alginate plays a role in the tolerance of P. aeruginosa aggregates toward tobramycin and meropenem, but not ciprofloxacin. However, contrary to previous studies we did not observe a role for Psl or Pel in the tolerance of P. aeruginosa aggregates toward tobramycin, ciprofloxacin, and meropenem.Copyright © 2023 Liang, Nilsson, Kragh, Hedal, Alcàcer-Almansa, Kiilerich, Andersen and Tolker-Nielsen.

JTD Keywords: aggregates, antibiotic tolerance, biofilm formation, extracellular matrix, genome, growth, lungs, molecular-mechanisms, mutations, polysaccharide, pseudomonas aeruginosa, psl, system, Aggregates, Antibiotic tolerance, Biofilm, Extracellular matrix, Pseudomonas aeruginosa, Small-colony variants

Tumor secreted extracellular vesicles (EVs) are potent intercellular signaling platforms. They are responsible for the accommodation of the premetastatic niche (PMN) to support cancer cell engraftment and metastatic growth. However, complex cancer cell composition within the tumor increases also the heterogeneity among cancer secreted EVs subsets, a functional diversity that has been poorly explored. This phenomenon is particularly relevant in highly plastic and heterogenous triple-negative breast cancer (TNBC), in which a significant representation of malignant cancer stem cells (CSCs) is displayed. Herein, we selectively isolated and characterized EVs from CSC or differentiated cancer cells (DCC; EVsCSC and EVsDCC , respectively) from the MDA-MB-231 TNBC cell line. Our results showed that EVsCSC and EVsDCC contain distinct bioactive cargos and therefore elicit a differential effect on stromal cells in the TME. Specifically, EVsDCC activated secretory cancer associated fibroblasts (CAFs), triggering IL-6/IL-8 signaling and sustaining CSC phenotype maintenance. Complementarily, EVsCSC promoted the activation of α-SMA+ myofibroblastic CAFs subpopulations and increased the endothelial remodeling, enhancing the invasive potential of TNBC cells in vitro and in vivo. In addition, solely the EVsCSC mediated signaling prompted the transformation of healthy lungs into receptive niches able to support metastatic growth of breast cancer cells.© 2023 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.

JTD Keywords: chemoresistance, dormancy, drives, extracellular vesicles, invasion, plasticity, premetastatic niche, triple-negative breast cancer, tumor microenvironment, Cancer cell plasticity, Extracellular vesicles, Fibroblasts, Premetastatic niche, Triple-negative breast cancer, Tumor microenvironment

Here we revisit tau protein aggregation at primary, secondary, tertiary and quaternary structures. In addition, the presence of non-aggregated tau protein, which has been recently discovered, is also commented on.Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.

JTD Keywords: alpha-helix, alzheimer-disease, antigenic determinants, binding, isomerase pin1, microtubule-binding repeats, neurofibrillary tangles, paired helical filaments, repeat domain, structural-characterization, tau conformations, w-tau isoform, Microtubule-associated protein, Microtubule-binding repeats, Tau, Tau conformations, W-tau isoform

The term 'mechanosensation' describes the capacity of cells to translate mechanical stimuli into the coordinated regulation of intracellular signals, cellular function, gene expression and epigenetic programming. This capacity is related not only to the sensitivity of the cells to tissue motion, but also to the decryption of tissue geometric arrangement and mechanical properties. The cardiac stroma, composed of fibroblasts, has been historically considered a mechanically passive component of the heart. However, the latest research suggests that the mechanical functions of these cells are an active and necessary component of the developmental biology programme of the heart that is involved in myocardial growth and homeostasis, and a crucial determinant of cardiac repair and disease. In this Review, we discuss the general concept of cell mechanosensation and force generation as potent regulators in heart development and pathology, and describe the integration of mechanical and biohumoral pathways predisposing the heart to fibrosis and failure. Next, we address the use of 3D culture systems to integrate tissue mechanics to mimic cardiac remodelling. Finally, we highlight the potential of mechanotherapeutic strategies, including pharmacological treatment and device-mediated left ventricular unloading, to reverse remodelling in the failing heart.© 2022. Springer Nature Limited.

JTD Keywords: cardiomyocyte proliferation, cross-linking, extracellular-matrix, focal adhesions, gene-expression, mechanical regulation, myocardial-infarction, substrate stiffness affects, t-cells, Ventricular assist device

JTD Keywords: current-voltage characteristics, electrolyte gated organic field effect transistors, Analytical model, Thin-film transistors

JTD Keywords: deep generative models, eatris european infrastructure for translational medicine, integrative omics, machine learning, multi-omics, personalized medicine, Deep generative models, Eatris european infrastructure for translational medicine, Integrative omics, Machine learning, Multi-omics, Personalized medicine, Protein protein interaction (ppi) network

Mechanical changes in tumors have long been linked to increased malignancy and therapy resistance and attributed to mechanical changes in the tumor extracellular matrix (ECM). However, to the best of our knowledge, there have been no mechanical studies on decellularized tumors. Here, we studied the biochemical and mechanical progression of the tumor ECM in two models of lung metastases: lung carcinoma (CAR) and melanoma (MEL). We decellularized the metastatic lung sections, measured the micromechanics of the tumor ECM, and stained the sections for ECM proteins, proliferation, and cell death markers. The same methodology was applied to MEL mice treated with the clinically approved anti-fibrotic drug nintedanib. When compared to healthy ECM (~0.40 kPa), CAR and MEL lung macrometastases produced a highly dense and stiff ECM (1.79 ± 1.32 kPa, CAR and 6.39 ± 3.37 kPa, MEL). Fibronectin was overexpressed from the early stages (~118%) to developed macrometastases (~260%) in both models. Surprisingly, nintedanib caused a 4-fold increase in ECM-occupied tumor area (5.1 ± 1.6% to 18.6 ± 8.9%) and a 2-fold in-crease in ECM stiffness (6.39 ± 3.37 kPa to 12.35 ± 5.74 kPa). This increase in stiffness strongly correlated with an increase in necrosis, which reveals a potential link between tumor hypoxia and ECM deposition and stiffness. Our findings highlight fibronectin and tumor ECM mechanics as attractive targets in cancer therapy and support the need to identify new anti-fibrotic drugs to abrogate aberrant ECM mechanics in metastases.

JTD Keywords: atomic force microscopy, basement membrane, breast-cancer, decellularization, expression, extracellular matrix, extracellular-matrix, fibronectin, intermittent hypoxia, lung carcinoma, lung metastases, melanoma, metastatic niche formation, micromechanical properties, nintedanib, signature, stiffness, tumor-growth, Colorectal-cancer progression, Lung metastases, Stiffness

Surface curvature both emerges from, and influences the behavior of, living objects at length scales ranging from cell membranes to single cells to tissues and organs. The relevance of surface curvature in biology is supported by numerous experimental and theoretical investigations in recent years. In this review, first, a brief introduction to the key ideas of surface curvature in the context of biological systems is given and the challenges that arise when measuring surface curvature are discussed. Giving an overview of the emergence of curvature in biological systems, its significance at different length scales becomes apparent. On the other hand, summarizing current findings also shows that both single cells and entire cell sheets, tissues or organisms respond to curvature by modulating their shape and their migration behavior. Finally, the interplay between the distribution of morphogens or micro-organisms and the emergence of curvature across length scales is addressed with examples demonstrating these key mechanistic principles of morphogenesis. Overall, this review highlights that curved interfaces are not merely a passive by-product of the chemical, biological, and mechanical processes but that curvature acts also as a signal that co-determines these processes.© 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.

JTD Keywords: biological systems, butterfly wing scales, cubic membranes, extracellular-matrix, geometry, mechanotransduction, membrane curvature, morphogenesis, neotissue growth, pattern-formation, soft materials, surface curvature, tissue-growth, Biological systems, Collective cell-migration, Surface curvature

Introduction: Three-dimensional (3D) bioprinting is a promising technique for the development of neuronal in vitro models because it controls the deposition of materials and cells. Finding a biomaterial that supports neural differentiation in vitro while ensuring compatibility with the technique of 3D bioprinting of a self-standing construct is a challenge.Methods: In this study, gelatin methacryloyl (GelMA), methacrylated alginate (AlgMA), and hyaluronic acid (HA) were examined by exploiting their biocompatibility and tunable mechanical properties to resemble the extracellular matrix (ECM) and to create a suitable material for printing neural progenitor cells (NPCs), supporting their long-term differentiation. NPCs were printed and differentiated for up to 15 days, and cell viability and neuronal differentiation markers were assessed throughout the culture.Results and Discussion: This composite biomaterial presented the desired physical properties to mimic the ECM of the brain with high water intake, low stiffness, and slow degradation while allowing the printing of defined structures. The viability rates were maintained at approximately 80% at all time points. However, the levels of beta-III tubulin marker increased over time, demonstrating the compatibility of this biomaterial with neuronal cell culture and differentiation. Furthermore, these cells showed increased maturation with corresponding functional properties, which was also demonstrated by the formation of a neuronal network that was observed by recording spontaneous activity via Ca2+ imaging.

JTD Keywords: biomaterials, bioprinting, differentiation, in vitro models, neural progenitor cells, 2d, Biomaterials, Bioprinting, C17.2, Differentiation, Extracellular-matrix, Hydrogels, In vitro models, In-vitro, Neural progenitor cells, Neuronal models, Proliferation, Scaffolds, Stem-cells, Substrate stiffness

Lately, there has been an increasing demand for materials that could improve tissue regenerative therapies and provide antimicrobial effects. Similarly, there is a growing need to develop or modify biomaterials for the diagnosis and treatment of different pathologies. In this scenario, hydroxyapatite (HAp) appears as a bioceramic with extended functionalities. Nevertheless, there are certain disadvantages related to the mechanical properties and lack of antimicrobial capacity. To circumvent them, the doping of HAp with a variety of cationic ions is emerging as a good alterative due to the different biological roles of each ion. Among many elements, lanthanides are understudied despite their great potential in the biomedical field. For this reason, the present review focuses on the biological benefits of lanthanides and how their incorporation into HAp can alter its morphology and physical properties. A comprehensive section of the applications of lanthanides-substituted HAp nanoparticles (HAp NPs) is presented to unveil the potential biomedical uses of these systems. Finally, the need to study the tolerable and non-toxic percentages of substitution with these elements is highlighted.

JTD Keywords: biolabeling, biomedicine, biosensors, bone regeneration, calcium, cancer treatment, cationic ions, cell imaging, cerium, doped hap, hydroxyapatite, implants, in-vitro bioactivity, lanthanides-substitutions, lanthanidessubstitutions, nanoparticles, radiation synovectomy, sm-153 particulate hydroxyapatite, structural-characterization, theragnostics, theranostic nanoplatforms, Europium-doped hydroxyapatite, Hydroxyapatite, Theragnostics

Human induced pluripotent stem cell (hiPSC) technologies offer a unique resource for modeling neurological diseases. However, iPSC models are fraught with technical limitations including abnormal aggregation and inefficient maturation of differentiated neurons. These problems are in part due to the absence of synergistic cues of the native extracellular matrix (ECM). We report on the use of three artificial ECMs based on peptide amphiphile (PA) supramolecular nanofibers. All nanofibers display the laminin-derived IKVAV signal on their surface but differ in the nature of their non-bioactive domains. We find that nanofibers with greater intensity of internal supramolecular motion have enhanced bioactivity toward hiPSC-derived motor and cortical neurons. Proteomic, biochemical, and functional assays reveal that highly mobile PA scaffolds caused enhanced β1-integrin pathway activation, reduced aggregation, increased arborization, and matured electrophysiological activity of neurons. Our work highlights the importance of designing biomimetic ECMs to study the development, function, and dysfunction of human neurons.Copyright © 2022 Elsevier Inc. All rights reserved.

JTD Keywords: differentiation, force-field, laminin, migration, nanostructures, peptide amphiphiles, spinal-cord, statistical-model, supramolecular materials, Coarse-grained model, Dynamics, Extracellular matrix, Ikvav, Ipsc-derived neurons, Laminin, Neuronal maturation, Peptide amphiphiles, Supramolecular motion, Supramolecular nanofibers

In response to different types and intensities of mechanical force, cells modulate their physical properties and adapt their plasma membrane (PM). Caveolae are PM nano-invaginations that contribute to mechanoadaptation, buffering tension changes. However, whether core caveolar proteins contribute to PM tension accommodation independently from the caveolar assembly is unknown. Here we provide experimental and computational evidence supporting that caveolin-1 confers deformability and mechanoprotection independently from caveolae, through modulation of PM curvature. Freeze-fracture electron microscopy reveals that caveolin-1 stabilizes non-caveolar invaginations-dolines-capable of responding to low-medium mechanical forces, impacting downstream mechanotransduction and conferring mechanoprotection to cells devoid of caveolae. Upon cavin-1/PTRF binding, doline size is restricted and membrane buffering is limited to relatively high forces, capable of flattening caveolae. Thus, caveolae and dolines constitute two distinct albeit complementary components of a buffering system that allows cells to adapt efficiently to a broad range of mechanical stimuli.© 2022. The Author(s).

JTD Keywords: cavin, cell-migration, cholesterol, extracellular-matrix, nanoscale organization, particle-size, polarization, size distribution, tension, Plasma-membrane

Various locomotion strategies employed by microorganisms are observed in complex biological environments. Spermatozoa assemble into bundles to improve their swimming efficiency compared to individual cells. However, the dynamic mechanisms for the formation of sperm bundles have not been fully characterized. In this study, we numerically and experimentally investigate the locomotion of spermatozoa during the transition from individual cells to bundles of two cells. Three consecutive dynamic behaviors are found across the course of the transition: hydrodynamic attraction/repulsion, alignment, and synchronization. The hydrodynamic attraction/repulsion depends on the relative orientation and distance between spermatozoa as well as their flagellar wave patterns and phase shift. Once the heads are attached, we find a stable equilibrium of the rotational hydrodynamics resulting in the alignment of the heads. The synchronization results from the combined influence of hydrodynamic and mechanical cell-to-cell interactions. Additionally, we find that the flagellar beat is regulated by the interactions during the bundle formation, whereby spermatozoa can synchronize their beats to enhance their swimming velocity.

JTD Keywords: behavior, cilia, collective locomotion, collective motion, competition, flagellar propulsion, hydrodynamics, motility, propulsion, sperm cooperation, tracking, Collective locomotion, Flagellar propulsion, Flagellar synchronization, Spermatozoa bundle

The extracellular matrix (ECM) of the lung is a filamentous network composed mainly of collagens, elastin, and proteoglycans that provides structural and physical support to its populating cells. Proliferation, migration and overall behaviour of those cells is greatly determined by micromechanical queues provided by the ECM. Lung fibrosis displays an aberrant increased deposition of ECM which likely changes filament organization and stiffens the ECM, thus upregulating the profibrotic profile of pulmonary cells. We have previously used AFM to assess changes in the Young’s Modulus (E) of the ECM in the lung. Here, we perform further ECM topographical, mechanical and viscoelastic analysis at the micro- and nano-scale throughout fibrosis development. Furthermore, we provide nanoscale correlations between topographical and elastic properties of the ECM fibres. Firstly, we identify a softening of the ECM after rats are instilled with media associated with recovery of mechanical homeostasis, which is hindered in bleomycin-instilled lungs. Moreover, we find opposite correlations between fibre stiffness and roughness in PBS- vs bleomycin-treated lung. Our findings suggest that changes in ECM nanoscale organization take place at different stages of fibrosis, with the potential to help identify pharmacological targets to hinder its progression.

JTD Keywords: atomic force microscopy, cells, deposition, extracellular matrix, idiopathic pulmonary fibrosis, mechanisms, mechanosensing, membranes, micromechanical properties, pathogenesis, stiffness, tissues, viscoelasticity, Extracellular matrix, Induced lung fibrosis, Mechanosensing

Prion-like domains (PrLDs) are intrinsically disordered regions (IDRs) of low sequence complexity with a similar composition to yeast prion domains. PrLDs-containing proteins have been involved in different organisms' regulatory processes. Regions of moderate amyloid propensity within IDRs have been shown to assemble autonomously into amyloid fibrils. These sequences tend to be rich in polar amino acids and often escape from the detection of classical bioinformatics screenings that look for highly aggregation-prone hydrophobic sequence stretches. We defined them as cryptic amyloidogenic regions (CARs) and recently developed an integrated database that collects thousands of predicted CARs in IDRs. CARs seem to be evolutionary conserved among disordered regions because of their potential to stablish functional contacts with other biomolecules. Here we have focused on identifying and characterizing CARs in prion-like proteins (pCARs) from plants, a lineage that has been poorly studied in comparison with other prionomes. We confirmed the intrinsic amyloid potential for a selected pCAR from Arabidopsis thaliana and explored functional enrichments and compositional bias of pCARs in plant prion-like proteins.Copyright © 2023 Pintado-Grima, Santos, Iglesias, Manglano-Artuñedo, Pallarès and Ventura.

JTD Keywords: aggregation, aromatic residues, bioinformatics, domains, functional interactions, identify proteins, plants, prediction, prion-like domains, q/n-rich, regulator, sup35, yeast, Bioinformatics, Cryptic amyloidogenic regions, Functional interactions, Plants, Prion-like domains, Rna-binding proteins

Sperm cells undergo complex interactions with external environments, such as a solid-boundary, fluid flow, as well as other cells before arriving at the fertilization site. The interaction with the oviductal epithelium, as a site of sperm storage, is one type of cell-to-cell interaction that serves as a selection mechanism. Abnormal sperm cells with poor swimming performance, the major cause of male infertility, are filtered out by this selection mechanism. In this study, collinear bundles, consisting of two sperm cells, generate propulsive thrusts along opposite directions and allow to observe the influence of cell-to-cell interaction on flagellar wave-patterns. The developed elasto-hydrodynamic model demonstrates that steric and adhesive forces lead to highly symmetrical wave-pattern and reduce the bending amplitude of the propagating wave. It is measured that the free cells exhibit a mean flagellar curvature of 6.4 +/- 3.5 rad mm(-1) and a bending amplitude of 13.8 +/- 2.8 rad mm(-1). After forming the collinear bundle, the mean flagellar curvature and bending amplitude are decreased to 1.8 +/- 1.1 and 9.6 +/- 1.4 rad mm(-1), respectively. This study presents consistent theoretical and experimental results important for understanding the adaptive behavior of sperm cells to the external time-periodic force encountered during sperm-egg interaction.

JTD Keywords: bovine sperm cells, cell-to-cell interaction, flagellar propulsion, Bovine sperm cells, Cell-to-cell interaction, Cilia, Filaments, Flagellar propulsion, Hydrodynamic models, Mechanism, Micro-video, Model, Motility, Thermotaxis, Transformations, Transition

Mechanical force modulates the conformation and function of individual proteins, and this underpins many mechanically driven cellular processes. This Review addresses single-molecule force spectroscopy experiments conducted on proteins with a known role in mechanosensing and mechanotransduction in eukaryotic cells.; In addition to biochemical signals and genetic considerations, mechanical forces are rapidly emerging as a master regulator of human physiology. However, the molecular mechanisms that regulate force-induced functionalities across a wide range of scales, encompassing the cell, tissue or organ levels, are not well understood in comparison. With the advent, development and refining of single-molecule nanomechanical techniques that enable the conformational dynamics of individual proteins under the effect of a calibrated force to be probed, we have begun to acquire a comprehensive knowledge of the diverse physicochemical principles that regulate the elasticity of single proteins. Here, we review the major advances underpinning our current understanding of how the elasticity of single proteins regulates mechanosensing and mechanotransduction. We discuss the present limitations and future challenges of this prolific and burgeoning field.

JTD Keywords: Cadherin adhesion, Energy landscape, Extracellular-matrix protein, Focal adhesion kinase, Mechanical stability, Molecule force spectroscopy, Muscle protein, N2b element, Stranded-dna, Structural basis

The involvement of the extracellular matrix (ECM) in tumor progression has motivated the development of biomaterials mimicking the tumor ECM to develop more predictive cancer models. Particularly, polypeptides based on elastin could be an interesting approach to mimic the ECM due to their tunable properties. Here, we demonstrated that elastin-like recombinamer (ELR) hydrogels can be suitable biomaterials to develop breast cancer models. This hydrogel was formed by two ELR polypeptides, one containing sequences biodegradable by matrix metalloproteinase and cyclooctyne and the other carrying arginylglycylaspartic acid and azide groups to allow cell adhesion, biodegradability, and suitable stiffness through "click-chemistry" cross-linking. Our findings show that breast cancer or nontumorigenic breast cells showed high viability and cell proliferation for up to 7 days. MCF7 and MCF10A formed spheroids whereas MDA-MB-231 formed cell networks, with the expression of ECM and high drug resistance in all cases, evidencing that ELR hydrogels are a promising biomaterial for breast cancer modeling.

JTD Keywords: clinical-trials, collagen i, discovery, mcf-7 cells, phenotype, progression, spheroids, translation, tumor microenvironment, Extracellular-matrix

Abstract Oxidative stress, which occurs when an organism is exposed to an adverse stimulus that results in a misbalance of antioxidant and pro-oxidants species, is the common denominator of diseases considered as a risk factor for SARS-CoV-2 lethality. Indeed, reactive oxygen species caused by oxidative stress have been related to many virus pathogenicity. In this work, simulations have been performed on the receptor binding domain of SARS-CoV-2 spike glycoprotein to study what residues are more susceptible to be attacked by ·OH, which is one of the most reactive radicals associated to oxidative stress. The results indicate that isoleucine (ILE) probably plays a crucial role in modification processes driven by radicals. Accordingly, QM/MM-MD simulations have been conducted to study both the ·OH-mediated hydrogen abstraction of ILE residues and the induced modification of the resulting ILE radical through hydroxylation or nitrosylation reactions. All in all, in silico studies show the importance of the chemical environment triggered by oxidative stress on the modifications of the virus, which is expected to help for foreseeing the identification or development of antioxidants as therapeutic drugs. Graphic abstract

JTD Keywords: atom abstraction, damage, density functionals, hydrogen abstraction, isoleucine, molecular dynamics, pathogenesis, protein, reactive oxygen species, receptor binding domain, residues, spike protein, Amino-acids, Hydrogen abstraction, Isoleucine, Molecular dynamics, Reactive oxygen species, Receptor binding domain, Spike protein

The study and use of decellularized extracellular matrix (dECM) in tissue engineering, regenerative medicine, and pathophysiology have become more prevalent in recent years. To obtain dECM, numerous decellularization procedures have been developed for the entire organ or tissue blocks, employing either perfusion of decellularizing agents through the tissue's vessels or submersion of large sections in decellularizing solutions. However, none of these protocols are suitable for thin tissue slices (less than 100 µm) or allow side-by-side analysis of native and dECM consecutive tissue slices. Here, we present a detailed protocol to decellularize tissue sections while maintaining the sample attached to a glass slide. This protocol consists of consecutive washes and incubations of simple decellularizing agents: ultrapure water, sodium deoxycholate (SD) 2%, and deoxyribonuclease I solution 0.3 mg/mL (DNase I). This novel method has been optimized for a faster decellularization time (2-3 h) and a better correlation between dECM properties and native tissue-specific biomarkers, and has been tested in different types of tissues and species, obtaining similar results. Furthermore, this method can be used for scarce and valuable samples such as clinical biopsies. This protocol was validated in: Front Bioeng Biotechnol (2022), DOI: 10.3389/fbioe.2022.832178.Copyright © 2022 The Authors; exclusive licensee Bio-protocol LLC.

JTD Keywords: decellularization, extracellular matrix, glass slide, mechanobiology, sodium deoxycholate, tissue slices, Decellularization, Extracellular-matrix, Tissue slices

Around 30-40% of patients with colorectal cancer (CRC) undergoing curative resection of the primary tumour will develop metastases in the subsequent years1. Therapies to prevent disease relapse remain an unmet medical need. Here we uncover the identity and features of the residual tumour cells responsible for CRC relapse. An analysis of single-cell transcriptomes of samples from patients with CRC revealed that the majority of genes associated with a poor prognosis are expressed by a unique tumour cell population that we named high-relapse cells (HRCs). We established a human-like mouse model of microsatellite-stable CRC that undergoes metastatic relapse after surgical resection of the primary tumour. Residual HRCs occult in mouse livers after primary CRC surgery gave rise to multiple cell types over time, including LGR5+ stem-like tumour cells2-4, and caused overt metastatic disease. Using Emp1 (encoding epithelial membrane protein 1) as a marker gene for HRCs, we tracked and selectively eliminated this cell population. Genetic ablation of EMP1high cells prevented metastatic recurrence and mice remained disease-free after surgery. We also found that HRC-rich micrometastases were infiltrated with T cells, yet became progressively immune-excluded during outgrowth. Treatment with neoadjuvant immunotherapy eliminated residual metastatic cells and prevented mice from relapsing after surgery. Together, our findings reveal the cell-state dynamics of residual disease in CRC and anticipate that therapies targeting HRCs may help to avoid metastatic relapse.© 2022. The Author(s), under exclusive licence to Springer Nature Limited.

JTD Keywords: colonization, defines, human colon, mutations, plasticity, retrieval, stem-cells, subtypes, underlie, Animal, Animal cell, Animal experiment, Animal model, Animal tissue, Animals, Article, Cancer, Cancer growth, Cancer immunotherapy, Cancer inhibition, Cancer recurrence, Cancer staging, Cell, Cell adhesion, Cell migration, Cell population, Cell surface receptor, Cohort analysis, Colorectal cancer, Colorectal neoplasms, Colorectal tumor, Comprehensive molecular characterization, Controlled study, Crispr-cas9 system, Cytoskeleton, Disease exacerbation, Disease progression, Dynamics, Emp1 gene, Epithelial membrane protein-1, Extracellular matrix, Flow cytometry, Fluorescence intensity, Gene expression, Genetics, Human, Human cell, Humans, Immune response, Immunofluorescence, In situ hybridization, Marker gene, Metastasis potential, Mice, Minimal residual disease, Mouse, Neoplasm proteins, Neoplasm recurrence, local, Neoplasm, residual, Nonhuman, Pathology, Phenotype, Prevention and control, Protein, Receptors, cell surface, Single cell rna seq, Tumor, Tumor protein, Tumor recurrence

The use of physiomimetic decellularized extracellular matrix-derived hydrogels is attracting interest since they can modulate the therapeutic capacity of numerous cell types, including mesenchymal stromal cells (MSCs). Remarkably, extracellular vesicles (EVs) derived from MSCs display similar functions as their parental cells, mitigating tissue damage in lung diseases. However, recent data have shown that ECM-derived hydrogels could release other resident vesicles similar to EVs. Here, we aim to better understand the contribution of EVs and ECM-vesicles released from MSCs and/or lung-derived hydrogel (L-HG) in lung repair by using an in vitro lung injury model. L-HG derived-vesicles and MSCs EVs cultured either in L-HG or conventional plates were isolated and characterized. The therapeutic capacity of vesicles obtained from each experimental condition was tested by using an alveolar epithelial wound-healing assay. The number of ECM-vesicles released from acellular L-HG was 10-fold greater than EVs from conventional MSCs cell culture revealing that L-HG is an important source of bioactive vesicles. MSCs-derived EVs and L-HG vesicles have similar therapeutic capacity in lung repair. However, when wound closure rate was normalized by total proteins, the MSCs-derived EVs shows higher therapeutic potential to those released by L-HG. The EVs released from L-HG must be considered when HG is used as substrate for cell culture and EVs isolation.

JTD Keywords: cell, extracellular vesicles, hydrogel, lung epithelial cells, lung repair, mesenchymal stem cells, Extracellular matrix, Extracellular vesicles, Hydrogel, Lung epithelial cells, Lung repair, Mesenchymal stem cells, Respiratory-distress-syndrome

JTD Keywords: anti-fxiia antibody, artificial surfaces, blood compatibility, complement activation, factor xii, fibrinolytic system, hemocompatible coatings, interactive hemocompatibility, poly(2-methacryloyloxyethyl phosphorylcholine), polyethylene oxide, polymer brushes, radical polymerization, sequential coimmobilization, Antifouling polymer brushes, Protein adsorption

Several factors present in the extracellular environment regulate epithelial cell adhesion and dynamics. Among them, growth factors such as EGF, upon binding to their receptors at the cell surface, get internalized and directly activate the acto-myosin machinery. In this study we present the effects of EGF on the contractility of epithelial cancer cell colonies in confined geometry of different sizes. We show that the extent to which EGF triggers contractility scales with the cluster size and thus the number of cells. Moreover, the collective contractility results in a radial distribution of traction forces, which are dependent on integrin β1 peripheral adhesions and transmitted to neighboring cells through adherens junctions. Taken together, EGF-induced contractility acts on the mechanical crosstalk and linkage between the cell-cell and cell-matrix compartments, regulating collective responses.Copyright © 2022 The Authors. Published by Elsevier GmbH.. All rights reserved.

JTD Keywords: actin, activation, actomyosin, adherens junctions, adhesion, e-cadherin, egf, maturation, mechanical regulation, micropatterning, migration, traction forces, transduction, transmission, Actomyosin, Adherens junctions, Collective contractility, Egf, Epidermal-growth-factor, Micropatterning, Traction forces

Hydroxyapatite nanoparticles are popular tools in bone regeneration, but they have also been used for gene delivery and as anticancer drugs. Understanding their mechanism of action, particularly for the latter application, is crucial to predict their toxicity. To this end, we aimed to elucidate the importance of nanoparticle membrane interactions in the cytotoxicity of MG-63 cells using two different types of nanoparticles. In addition, conventional techniques for studying nanoparticle internalisation were evaluated and compared with newer and less exploited approaches. Hydroxyapatite and magnesium-doped hydroxyapatite nanoparticles were used as suspensions or compacted as specular discs. Comparison between cells seeded on the discs and those supplemented with the nanoparticles allowed direct interaction of the cell membrane with the material to be ruled out as the main mechanism of toxicity. In addition, standard techniques such as flow cytometry were inconclusive when used to assess nanoparticles toxicity. Interestingly, the use of intracellular calcium fluorescent probes revealed the presence of a high number of calcium-rich vesicles after nanoparticle supplementation in cell culture. These structures could not be detected by transmission electron microscopy due to their liquid content. However, by using cryo-soft X-ray imaging, which was used to visualise the cellular ultrastructure without further treatment other than vitrification and to quantify the linear absorption coefficient of each organelle, it was possible to identify them as multivesicular bodies, potentially acting as calcium stores. In the study, an advanced state of degradation of the hydroxyapatite and magnesium-doped hydroxyapatite nanoparticles within MG-63 cells was observed. Overall, we demonstrate that the combination of fluorescent calcium probes together with cryo-SXT is an excellent approach to investigate intracellular calcium, especially when found in its soluble form.Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.

JTD Keywords: adsorption, cryo-soft x-ray tomography, cytotoxicity, expression, flow cytometry, internalisation, intracellular calcium, magnesium, nano, nanomaterials, nanoparticles, proliferation, protein corona, ultrastructure, Calcium-phosphate nanoparticles, Cryo-soft x-ray tomography, Flow cytometry, Hydroxyapatite, Internalisation, Intracellular calcium, Nanoparticles

Single-molecule localization microscopy (SMLM) is a powerful super-resolution technique for elucidating structure and dynamics in the life- and material sciences. Simultaneously acquiring spectral information (spectrally resolved SMLM, sSMLM) has been hampered by several challenges: an increased complexity of the optical detection pathway, lower accessible emitter densities, and compromised spatio-spectral resolution. Here we present a single-component, low-cost implementation of sSMLM that addresses these challenges. Using a low-dispersion transmission grating positioned close to the image plane, the +1stdiffraction order is minimally elongated and is analyzed using existing single-molecule localization algorithms. The distance between the 0th and 1st order provides accurate information on the spectral properties of individual emitters. This method enables a 5-fold higher emitter density while discriminating between fluorophores whose peak emissions are less than 15 nm apart. Our approach can find widespread use in single-molecule applications that rely on distinguishing spectrally different fluorophores under low photon conditions.

JTD Keywords: cells, multicolor imaging, nanoscopy, particle tracking, point accumulation for imaging in nanoscale topography (paint), precision, single-molecule fo?rster resonance energy transfer (smfret), stochastic optical reconstruction microscopy (storm), Diffraction-limit, Multicolor imaging, Point accumulation for imaging in nanoscale topography (paint), Single-molecule förster resonance energy transfer (smfret), Single-molecule spectroscopy, Stochastic optical reconstruction microscopy (storm)

The synthesis and study of the tripeptide Arg-Gly-Asp (RGD), the binding site of different extracellular matrix proteins, e.g., fibronectin and vitronectin, has allowed the production of a wide range of cell adhesive surfaces. Although the surface density and spacing of the RGD peptide at the nanoscale have already shown a significant influence on cell adhesion, the impact of its hierarchical nanostructure is still rather unexplored. Accordingly, a versatile colloidal system named quatsomes, based on fluid nanovesicles formed by the self-assembling of cholesterol and surfactant molecules, has been devised as a novel template to achieve hierarchical nanostructures of the RGD peptide. To this end, RGD was anchored on the vesicle's fluid membrane of quatsomes, and the RGD-functionalized nanovesicles were covalently anchored to planar gold surfaces, forming a state of quasi-suspension, through a long poly(ethylene glycol) (PEG) chain with a thiol termination. An underlying self-assembled monolayer (SAM) of a shorter PEG was introduced for vesicle stabilization and to avoid unspecific cell adhesion. In comparison with substrates featuring a homogeneous distribution of RGD peptides, the resulting hierarchical nanoarchitectonic dramatically enhanced cell adhesion, despite lower overall RGD molecules on the surface. The new versatile platform was thoroughly characterized using a multitechnique approach, proving its enhanced performance. These findings open new methods for the hierarchical immobilization of biomolecules on surfaces using quatsomes as a robust and novel tissue engineering strategy.

JTD Keywords: activation, arg-gly-asp (rgd), cell adhesion, extracellular-matrix, growth, integrins, ligands, nanopatterns, quatsomes, scaffolds, self-assembled monolayers, surface engineering, tissue engineering, Arg-gly-asp (rgd), Cell adhesion, Integrins, Nano-structured surfaces, Nanovesicles, Quatsomes, Self-assembled monolayers, Surface engineering, Tissue engineering

Cooperative behaviour of sperm is one of the mechanisms that plays a role in sperm competition. It has been observed in several species that spermatozoa interact with each other to form agglomerates or bundles. In this study, we investigate the effect of physical and biochemical factors that will most likely promote bundle formation in bull sperm. These factors include fluid viscosity, swim-up process, post-thaw incubation time and media additives which promote capacitation. While viscosity does not seem to influence the degree of sperm bundling, swim-up, post-thaw migration time and suppressed capacitation increase the occurrence of sperm bundles. This leads to the conclusion that sperm bundling is a result of hydrodynamic and adhesive interactions between the cells which occurs frequently during prolonged incubation times.Copyright © 2022 Morcillo i Soler, Hidalgo, Fekete, Zalanyi, Khalil, Yeste and Magdanz.

JTD Keywords: acrosome reaction, adhesion, bundling, capacitation, cell-cell interaction, cooperation, cooperative behaviour, fertilization, mammals, membrane, motility, progesterone, sperm competition, sperm migration, sperm selection, Bovine spermatozoa, Bundling, Cell-cell interaction, Cooperative behaviour, Sperm competition, Sperm migration, Sperm selection, Spermatozoa

Upon the initiation of collective cell migration, the cells at the free edge are specified as leader cells; however, the mechanism underlying the leader cell specification remains elusive. Here, we show that lamellipodial extension after the release from mechanical confinement causes sustained extracellular signal-regulated kinase (ERK) activation and underlies the leader cell specification. Live-imaging of Madin-Darby canine kidney (MDCK) cells and mouse epidermis through the use of Förster resonance energy transfer (FRET)-based biosensors showed that leader cells exhibit sustained ERK activation in a hepatocyte growth factor (HGF)-dependent manner. Meanwhile, follower cells exhibit oscillatory ERK activation waves in an epidermal growth factor (EGF) signaling-dependent manner. Lamellipodial extension at the free edge increases the cellular sensitivity to HGF. The HGF-dependent ERK activation, in turn, promotes lamellipodial extension, thereby forming a positive feedback loop between cell extension and ERK activation and specifying the cells at the free edge as the leader cells. Our findings show that the integration of physical and biochemical cues underlies the leader cell specification during collective cell migration.Copyright © 2022 Elsevier Inc. All rights reserved.

JTD Keywords: activation, c-met, contact inhibition, focal adhesions, heparan-sulfate, mechanical forces, morphogenesis, rho, stress fibers, Collective cell migration, Erk, Feedback regulation, Fret, Growth-factor receptor, Hgf, Lamellipodia, Leader cell specification, Signal transduction, Traction force, Wound healing

Most of the conventional in vitro models to test biomaterial-driven vascularization are too simplistic to recapitulate the complex interactions taking place in the actual cell microenvironment, which results in a poor prediction of the in vivo performance of the material. However, during the last decade, cell culture models based on microfluidic technology have allowed attaining unprecedented levels of tissue biomimicry. In this work, we propose a microfluidic-based 3D model to evaluate the effect of bioactive biomaterials capable of releasing signalling cues (such as ions or proteins) in the recruitment of endogenous endothelial progenitor cells, a key step in the vascularization process. The usability of the platform is demonstrated using experimentally-validated finite element models and migration and proliferation studies with rat endothelial progenitor cells (rEPCs) and bone marrow-derived rat mesenchymal stromal cells (BM-rMSCs). As a proof of concept of biomaterial evaluation, the response of rEPCs to an electrospun composite made of polylactic acid with calcium phosphates nanoparticles (PLA+CaP) was compared in a co-culture microenvironment with BM-rMSC to a regular PLA control. Our results show a significantly higher rEPCs migration and the upregulation of several pro-inflammatory and proangiogenic proteins in the case of the PLA+CaP. The effects of osteopontin (OPN) on the rEPCs migratory response were also studied using this platform, suggesting its important role in mediating their recruitment to a calcium-rich microenvironment. This new tool could be applied to screen the capacity of a variety of bioactive scaffolds to induce vascularization and accelerate the preclinical testing of biomaterials. STATEMENT OF SIGNIFICANCE: : For many years researchers have used neovascularization models to evaluate bioactive biomaterials both in vitro, with low predictive results due to their poor biomimicry and minimal control over cell cues such as spatiotemporal biomolecule signaling, and in vivo models, presenting drawbacks such as being highly costly, time-consuming, poor human extrapolation, and ethically controversial. We describe a compact microphysiological platform designed for the evaluation of proangiogenesis in biomaterials through the quantification of the level of sprouting in a mimicked endothelium able to react to gradients of biomaterial-released signals in a fibrin-based extracellular matrix. This model is a useful tool to perform preclinical trustworthy studies in tissue regeneration and to better understand the different elements involved in the complex process of vascularization.Copyright © 2022. Published by Elsevier Ltd.

JTD Keywords: angiogenesis, bioactive materials, bone regeneration, bone-formation, calcium-phosphate, extracellular calcium, in-vitro, interstitial flow, ion release, microfluidic model, signalling gradient, substitutes, tissue engineering, vascularization, vegf, Ion release, Mesenchymal stem-cells, Tissue engineering, Vascularization

Tauopathies are a group of neurodegenerative diseases characterized by the hyperphosphorylation and deposition of tau proteins in the brain. In Alzheimer's disease, and other related tauopathies, the pattern of tau deposition follows a stereotypical progression between anatomically connected brain regions. Increasing evidence suggests that tau behaves in a "prion-like" manner, and that seeding and spreading of pathological tau drive progressive neurodegeneration. Although several advances have been made in recent years, the exact cellular and molecular mechanisms involved remain largely unknown. Since there are no effective therapies for any tauopathy, there is a growing need for reliable experimental models that would provide us with better knowledge and understanding of their etiology and identify novel molecular targets. In this review, we will summarize the development of cellular models for modeling tau pathology. We will discuss their different applications and contributions to our current understanding of the "prion-like" nature of pathological tau.

JTD Keywords: neurodegeneration, seeding, spreading, Culture model, Efficient generation, Extracellular tau, Familial alzheimers-disease, Microtubule-associated protein, Mouse model, Neurodegeneration, Neurofibrillary tangles, Paired helical filaments, Pathogenic tau, Pluripotent stem-cells, Seeding, Spreading, Tauopathies

This work aimed at the antimicrobial functionalization of 3D-printed polymer-infiltrated biomimetic ceramic networks (PICN). The anti-microbial properties of the polymer-ceramic composites were achieved by coating them with human-and environmentally safe silver nanoparticles trapped in a phenolated lignin matrix (Ag@PL NPs). Lignin was enzymatically phenolated and used as a biobased reducing agent to obtain stable Ag@PL NPs, which were then formulated in a silane (gamma-MPS) solution and deposited to the PICN surface. The presence of the NPs and their proper attachment to the surface were analyzed with spectroscopic methods (FTIR and Raman) and X-ray photoelectron spectroscopy (XPS). Homogeneous distribution of 13.4 +/- 3.2 nm NPs was observed in the transmission electron microscopy (TEM) images. The functionalized samples were tested against Gram-positive (Staphylococcus aureus) and Gram-negative (Pseudomonas aeruginosa) bacteria, validating their antimicrobial efficiency in 24 h. The bacterial reduction of S. aureus was 90% in comparison with the pristine surface of PICN. To confirm that the Ag-functionalized PICN scaffold is a safe material to be used in the biomedical field, its biocompatibility was demonstrated with human fibroblast (BJ-5ta) and keratinocyte (HaCaT) cells, which was higher than 80% in both cell lines.

JTD Keywords: accuracy, antibacterial activity, disease, facile, laccase enzyme, lignin, polyacrylates, polymer-infiltrated ceramic network, silver nanoparticles, zirconia, Mechanical-properties, Mechanical-properties,zirconia,lignin,accuracy,disease,facil, Polymer-infiltrated ceramic network, Polymer-infiltrated ceramic network,polyacrylates,lignin,laccase enzyme,silver nanoparticles,antibacterial activit, Silver nanoparticles

Cells are constantly exposed to various chemical and physical stimuli. While much has been learned about the biochemical factors that regulate secretory trafficking from the endoplasmic reticulum (ER), much less is known about whether and how this trafficking is subject to regulation by mechanical signals. Here, we show that subjecting cells to mechanical strain both induces the formation of ER exit sites (ERES) and accelerates ER-to-Golgi trafficking. We found that cells with impaired ERES function were less capable of expanding their surface area when placed under mechanical stress and were more prone to develop plasma membrane defects when subjected to stretching. Thus, coupling of ERES function to mechanotransduction appears to confer resistance of cells to mechanical stress. Furthermore, we show that the coupling of mechanotransduction to ERES formation was mediated via a previously unappreciated ER-localized pool of the small GTPase Rac1. Mechanistically, we show that Rac1 interacts with the small GTPase Sar1 to drive budding of COPII carriers and stimulates ER-to-Golgi transport. This interaction therefore represents an unprecedented link between mechanical strain and export from the ER.© 2022 The Authors. Published under the terms of the CC BY 4.0 license.

JTD Keywords: cells, copii, docking, endoplasmic reticulum, endoplasmic-reticulum, er, gtpase, mechanobiology, proliferation, protein, reticulum exit sites, web server, Copii, Fast interaction refinement, Mechanobiology

Photosynthesis is a fundamental process that converts photons into chemical energy, driven by large protein complexes at the thylakoid membranes of plants, cyanobacteria, and algae. In plants, water-soluble plastocyanin (Pc) is responsible for shuttling electrons between cytochrome b6f complex and the photosystem I (PSI) complex in the photosynthetic electron transport chain (PETC). For an efficient turnover, a transient complex must form between PSI and Pc in the PETC, which implies a balance between specificity and binding strength. Here, we studied the binding frequency and the unbinding force between suitably oriented plant PSI and Pc under redox control using single molecule force spectroscopy (SMFS). The binding frequency (observation of binding-unbinding events) between PSI and Pc depends on their respective redox states. The interaction between PSI and Pc is independent of the redox state of PSI when Pc is reduced, and it is disfavored in the dark (reduced P700) when Pc is oxidized. The frequency of interaction between PSI and Pc is higher when at least one of the partners is in a redox state ready for electron transfer (ET), and the post-ET situation (PSIRed-PcOx) leads to lower binding. In addition, we show that the binding of ET-ready PcRed to PSI can be regulated externally by Mg2+ ions in solution.

JTD Keywords: architecture, binding-site, complexes, ferredoxin, force spectroscopy, induced structural-changes, interprotein electron transfer, light-dependent interaction, mg2+ concentration, photosystem i, plastocyanin, probe, recognition, reduction, Force spectroscopy, Interprotein electron transfer, Light-dependent interaction, Photosynthetic reaction-center, Photosystem i, Plastocyanin, Single molecule measurements

Acute Respiratory Distress Syndrome is one of the more common fatal complications in COVID-19, characterized by a highly aberrant inflammatory response. Pre-clinical models to study the effect of cell therapy and anti-inflammatory treatments have not comprehensively reproduced the disease due to its high complexity. This work presents a novel physiomimetic in vitro model for Acute Respiratory Distress Syndrome using lung extracellular matrix-derived hydrogels and organ-on-a-chip devices. Monolayres of primary alveolar epithelial cells were cultured on top of decellullarized lung hydrogels containing primary lung mesenchymal stromal cells. Then, cyclic stretch was applied to mimic breathing, and an inflammatory response was induced by using a bacteriotoxin hit. Having simulated the inflamed breathing lung environment, we assessed the effect of an anti-inflammatory drug (i.e., dexamethasone) by studying the secretion of the most relevant inflammatory cytokines. To better identify key players in our model, the impact of the individual factors (cyclic stretch, decellularized lung hydrogel scaffold, and the presence of mesenchymal stromal cells) was studied separately. Results showed that developed model presented a more reduced inflammatory response than traditional models, which is in line with what is expected from the response commonly observed in patients. Further, from the individual analysis of the different stimuli, it was observed that the use of extracellular matrix hydrogels obtained from decellularized lungs had the most significant impact on the change of the inflammatory response. The developed model then opens the door for further in vitro studies with a better-adjusted response to the inflammatory hit and more robust results in the test of different drugs or cell therapy.

JTD Keywords: alveolar epithelial cells, ards, extracellular matrix, hydrogels, inflammation, lung-on-a-chip, Acute lung injury, Alveolar epithelial cells, Ards, Dexamethasone, Epithelial-mesenchymal transition, Extracellular matrix, Extracellular-matrix, Hydrogels, Inflammation, Lung-on-a-chip, Mesenchymal stromal cells, Oxygen, Stem-cells

Chemically responsive polymers are macromolecules that respond to local variations of the chemical composition of the solution by changing their conformation, with notable examples including polyelectrolytes, proteins, and DNA. The polymer conformation changes can occur in response to changes in the pH, the ionic strength, or the concentration of a generic solute that interacts with the polymer. These chemical stimuli can lead to drastic variations of the polymer flexibility and even trigger a transition from a coil to a globule polymer conformation. In many situations, the spatial distribution of the chemical stimuli can be highly inhomogeneous, which can lead to large spatial variations of polymer conformation and of the rheological properties of the mixture. In this paper, we develop a theory for the flow of a mixture of solute and chemically responsive polymers. The approach is valid for generic flows and inhomogeneous distributions of polymers and solutes. To model the polymer conformation changes introduced by the interactions with the solute, we consider the polymers as linear elastic dumbbells whose spring stiffness depends on the solute concentration. We use Onsager's variational formalism to derive the equations governing the evolution of the variables, which unveils novel couplings between the distribution of dumbbells and that of the solute. Finally, we use a linear stability analysis to show that the governing equations predict an equilibrium phase separation and a distinct shear-induced phase separation whereby a homogeneous distribution of solute and dumbbells spontaneously demix. Similar phase transitions have been observed in previous experiments using stimuli-responsive polymers and may play an important role in living systems. (C) 2022 The Society of Rheology.

JTD Keywords: Coil-globule transition, Constitutive equation, Dilute-solutions, Dumbbell model, Dynamics, Macromolecules, Nonequilibrium thermodynamics, Polyelectrolytes, Polymer migration, Polymer phase separation, Polymers, Predictions, Rheology, Shear-induced phase separation, Solute-polymer interactions, Stress, Viscoelasticity

In recent years, 3D in vitro modeling of human skeletal muscle has emerged as a subject of increasing interest, due to its applicability in basic studies or screening platforms. These models strive to recapitulate key features of muscle architecture and function, such as cell alignment, maturation, and contractility in response to different stimuli. To this end, it is required to culture cells in biomimetic hydrogels suspended between two anchors. Currently available protocols are often complex to produce, have a high rate of breakage, or are not adapted to imaging and stimulation. Therefore, we sought to develop a simplified and reliable protocol, which still enabled versatility in the study of muscle function. In our method, we have used human immortalized myoblasts cultured in a hydrogel composed of MatrigelTM and fibrinogen, to create muscle strips suspended between two VELCROTM anchors. The resulting muscle constructs show a differentiated phenotype and contractile activity in response to electrical, chemical and optical stimulation. This activity is analyzed by two alternative methods, namely contraction analysis and calcium analysis with Fluo-4 AM. In all, our protocol provides an optimized version of previously published methods, enabling individual imaging of muscle bundles and straightforward analysis of muscle response with standard image analysis software. This system provides a start-to-finish guide on how to produce, validate, stimulate, and analyze bioengineered muscle. This ensures that the system can be quickly established by researchers with varying degrees of expertise, while maintaining reliability and similarity to native muscle.

JTD Keywords: cells, contraction, models, Differentiation

Peptide derivatives and, most specifically, their self-assembled supramolecular structures are being considered in the design of novel biofunctional materials. Although the self-assembly of triphenylalanine homopeptides has been found to be more versatile than that of homopeptides containing an even number of residues (i.e. diphe-nylalanine and tetraphenylalanine), only uncapped triphenylalanine (FFF) and a highly aromatic analog blocked at both the N-and C-termini with fluorenyl-containing groups (Fmoc-FFF-OFm), have been deeply studied before. In this work, we have examined the self-assembly of a triphenylalanine derivative bearing 9-fluorenylme-thyloxycarbonyl and benzyl ester end-capping groups at the N-and C-termini, respectively (Fmoc-FFF-OBzl). The antiparallel arrangement clearly dominates in beta-sheets formed by Fmoc-FFF-OBzl, whereas the parallel and antiparallel dispositions are almost isoenergetic in Fmoc-FFF-OFm beta-sheets and the parallel one is slightly favored for FFF. The effects of both the peptide concentration and the mediu m on the self-assembly process have been examined considering Fmoc-FFF-OBzl solutions in a wide variety of solvent:co-solvent mixtures. In addi-tion, Fmoc-FFF-OBzl supramolecular structures have been compared to those obtained for FFF and Fmoc-FFF-OFm under identical experimental conditions. The strength of pi-pi stacking interactions involving the end-capping groups plays a crucial role in the nucleation and growth of supramolecular structures, which de-termines the resulting morphology. Finally, the influence of a non-invasive external stimulus, ultrasounds, on the nucleation and growth of supramolecular structures has been examined. Overall, FFF-based peptides provide a wide range of supramolecular structures that can be of interest in the biotechnological field.

JTD Keywords: aromatic interactions, beta-sheet, hierarchical structures, phenylalanine homopeptides, supramolecular structures, Amino-acids, Aromatic interactions, Beta-sheet, Fmoc, Hierarchical struc tures, Hydrogels, Phenylalanine homopeptides, Solvent, Spectroscopy, Supramolecular structures, Triphenylalanine

The intestinal mucus lines the luminal surface of the intestinal epithelium. This mucus is a dynamic semipermeable barrier and one of the first-line defense mechanisms against the outside environment, protecting the body against chemical, mechanical, or biological external insults. At the same time, the intestinal mucus accommodates the resident microbiota, providing nutrients and attachment sites, and therefore playing an essential role in the host–pathogen interactions and gut homeostasis. Underneath this mucus layer, the intestinal epithelium is organized into finger-like protrusions called villi and invaginations called crypts. This characteristic 3D architecture is known to influence the epithelial cell differentiation and function. However, when modelling in vitro the intestinal host–pathogen interactions, these two essential features, the intestinal mucus and the 3D topography are often not represented, thus limiting the relevance of the models. Here we present an in vitro model that mimics the small intestinal mucosa and its interactions with intestinal pathogens in a relevant manner, containing the secreted mucus layer and the epithelial barrier in a 3D villus-like hydrogel scaffold. This 3D architecture significantly enhanced the secretion of mucus. In infection with the pathogenic adherent invasive E. coli strain LF82, characteristic of Crohn’s disease, we observed that this secreted mucus promoted the adhesion of the pathogen and at the same time had a protective effect upon its invasion. This pathogenic strain was able to survive inside the epithelial cells and trigger an inflammatory response that was milder when a thick mucus layer was present. Thus, we demonstrated that our model faithfully mimics the key features of the intestinal mucosa necessary to study the interactions with intestinal pathogens.

JTD Keywords: 3d in vitro models, barrier function, bile-salts, cells, drug-delivery, host-pathogen interaction, host–pathogen interaction, hydrogels, ileal mucosa, infection, intestinal models, intestinal mucus, microbiome, patient, responses, 3d in vitro models, Intestinal mucus, Invasive escherichia-coli

According to circular economy, wine-making by-products represent a fascinating biomass, which can be used for the sustainable exploitation of polyphenols and the development of new nanotechnological health-promoting products. In this study, polyphenols contained in the grape pomace were extracted by maceration with ethanol in an easy and low dissipative way. The obtained extract, rich in malvidin-3-glucoside, quercetin, pro-cyanidin B2 and gallic acid, was incorporated into phospholipid vesicles tailored for intestinal delivery. To improve their performances, vesicles were enriched with gelatine or a maltodextrin (Nutriose (R)), or their com-bination (gelatine-liposomes, nutriosomes and gelatine-nutriosomes). The small (-147 nm) and negatively charged (--50mV) vesicles were stable at different pH values mimicking saliva (6.75), gastric (1.20) and intestinal (7.00) environments. Vesicles effectively protected intestinal cells (Caco-2) from the oxidative stress and promoted the biofilm formation by probiotic bacteria. A preliminary evaluation of the vesicle feasibility at industrial levels was also performed, analysing the economic and energetic costs needed for their production.

JTD Keywords: Adhesion, Antioxidant activity, Caco-2 cells, Dextrin, Grape pomace extract, Lactobacillus-reuteri, Manufacturing costs, Oxidative stress, Ph, Phospholipid vesicles, Polyphenols, Probiotic bacteria, Protein

3DNA holds promise as a carrier for drugs that can be intercalated into its core or linked to surface arms. Coupling 3DNA to an antibody targeting intercellular adhesion molecule 1 (ICAM-1) results in high lung-specific biodistributions in vivo. While the role of individual parameters on ICAM-1 targeting has been studied for other nanocarriers, it has never been examined for 3DNA or in a manner capable of revealing the hierarchic interplay among said parameters. In this study, we used 2-layer vs. 4-layer anti-ICAM 3DNA and radiotracing to examine biodistribution in mice. We found that, below saturating conditions and within the ranges tested, the density of targeting antibodies on 3DNA is the most relevant parameter driving lung targeting over liver clearance, compared to the number of antibodies per carrier, total antibody dose, 3DNA dose, 3DNA size, or the administered concentration, which influenced the dose in organs but not the lung specific-over-liver clearance ratio. Data predicts that lung-specific delivery of intercalating (core loaded) drugs can be tuned using this biodistribution pattern, while that of arm-linked (surface loaded) drugs requires a careful parametric balance because increasing anti-ICAM density reduces the number of 3DNA arms available for drug loading.

JTD Keywords: 3dna nanocarrier, acid sphingomyelinase, antibody, carrier design parameters, carriers, dna nanostructures, doxorubicin, drug type, icam-1, inflammation, lung targeting, multiparametric hierarchy, nanoparticles, size, 3dna nanocarrier, Intracellular delivery, Multiparametric hierarchy

JTD Keywords: coordination, embodied cognition, human-robot interaction, sensorimotor coupling, Coordination, Embodied cognition, Human-robot interaction, Sensorimotor coupling, Social cognition

The extracellular matrix protects biofilm cells by reducing diffusion of antimicrobials. Tobramycin is an antibiotic used extensively to treat P. aeruginosa biofilms, but it is sequestered in the biofilm periphery by the extracellular negative charge matrix and loses its efficacy significantly. Dispersal of the biofilm extracellular matrix with enzymes such as DNase I is another promising therapy that enhances antibiotic diffusion into the biofilm. Here, we combine the charge neutralization of tobramycin provided by dextran-based single-chain polymer nanoparticles (SCPNs) together with DNase I to break the biofilm matrix. Our study demonstrates that the SCPNs improve the activity of tobramycin and DNase I by neutralizing the ionic interactions that keep this antibiotic in the biofilm periphery. Moreover, the detailed effects and interactions of nanoformulations with extracellular matrix components were revealed through time-lapse imaging of the P. aeruginosa biofilms by laser scanning confocal microscopy with specific labeling of the different biofilm components.

JTD Keywords: Cystic-fibrosis sputum, Delivery, Extracellular dna, Infections, Pseudomonas-aeruginosa, Transport

Effective inhibition of sperm motility using a spermicide can be a promising approach in developing non-invasive male contraceptive agents. Copper is known to have contraceptive properties and has been used clinically for decades as intrauterine contraceptive devices (IUDs) for contraception in females. Beyond that, the spermicidal use of copper is not explored much further, even though its use can also subdue the harmful effects caused by the hormonal female contraceptive agents on the environment. Herein, the size, concentration, and time-dependent in vitro inhibition of bovine spermatozoa by copper microparticles are studied. The effectivity in inhibiting sperm motility is correlated with the amount of Cu2+ ions released by the particles during incubation. The copper particles cause direct suppression of sperm motility and viability upon incubation and thereby show potential as sperm-inhibiting, hormone-free candidate for male contraception. In addition, biocompatibility tests using a cervical cell line help optimizing the size and concentration of the copper particles for the best spermicidal action while avoiding toxicity to the surrounding tissue.

JTD Keywords: Bovine spermatozoa, Clinical-trial, Copper, Human-spermatozoa, Ions, Male contraception, Metallic copper, Microparticles, Progestins, Sperm motility, Sperm viability, Spermicide, Viability

Pulmonary fibrosis (PF) is characterized by aberrant extracellular matrix (ECM) deposition, activation of fibroblasts to myofibroblasts and parenchymal disorganization, which have an impact on the biomechanical traits of the lung. In this context, the balance between matrix metalloproteinases (MMPs) and their tissue inhibitors of metalloproteinases (TIMPs) is lost. Interestingly, several MMPs are overexpressed during PF and exhibit a clear profibrotic role (MMP-2, -3, -8, -11, -12 and -28), but a few are antifibrotic (MMP-19), have both profibrotic and antifibrotic capacity (MMP7), or execute an unclear (MMP-1, -9, -10, -13, -14) or unknown function. TIMPs are also overexpressed in PF; hence, the modulation and function of MMPs and TIMP are more complex than expected. EMMPRIN/CD147 (also known as basigin) is a transmembrane glycoprotein from the immunoglobulin superfamily (IgSF) that was first described to induce MMP activity in fibroblasts. It also interacts with other molecules to execute non-related MMP aactions well-described in cancer progression, migration, and invasion. Emerging evidence strongly suggests that CD147 plays a key role in PF not only by MMP induction but also by stimulating fibroblast myofibroblast transition. In this review, we study the structure and function of MMPs, TIMPs and CD147 in PF and their complex crosstalk between them.

JTD Keywords: basigin, cd147, emmprin, mmps, timps, Basigin, Cd147, Cell-surface, Emmprin, Extracellular-matrix, Gelatinase-b, Gene-expression profiles, Growth-factor-beta, Immunoglobulin superfamily, Induced lung injury, Inducer emmprin, Mmps, Pulmonary fibrosis, Timps, Tissue inhibitor, Transforming growth-factor-beta-1

The tumor extracellular matrix (ECM) plays a vital role in tumor progression and drug resistance. Previous studies have shown that breast tissue-derived matrices could be an important biomaterial to recreate the complexity of the tumor ECM. We have developed a method for decellularizing and delipidating a porcine breast tissue (TDM) compatible with hydrogel formation. The addition of gelatin methacrylamide and alginate allows this TDM to be bioprinted by itself with good printability, shape fidelity, and cytocompatibility. Furthermore, this bioink has been tuned to more closely recreate the breast tumor by incorporating collagen type I (Col1). Breast cancer cells (BCCs) proliferate in both TDM bioinks forming cell clusters and spheroids. The addition of Col1 improves the printability of the bioink as well as increases BCC proliferation and reduces doxorubicin sensitivity due to a downregulation of HSP90. TDM bioinks also allow a precise three-dimensional printing of scaffolds containing BCCs and stromal cells and could be used to fabricate artificial tumors. Taken together, we have proven that these novel bioinks are good candidates for biofabricating breast cancer models.

JTD Keywords: 3d in vitro cancer model, bioprinting, breast tissue, 3d in vitro cancer model, Bioink, Bioprinting, Breast tissue, Crosstalk, Decellularization, Extracellular-matrix, Growth, Hydrogels, In-vitro, Inhibition, Mechanical-properties, Metastasis, Proliferation

Heterotopic ossification (HO) is a condition triggered by an injury leading to the formation of mature lamellar bone in extraskeletal soft tissues. Despite being a frequent complication of orthopedic and trauma surgery, brain and spinal injury, the etiology of HO is poorly understood. The aim of this study is to evaluate the hypothesis that a sustained local ionic homeostatic imbalance (SLIHI) created by mineral formation during tissue calcification modulates inflammation to trigger HO. This evaluation also considers the role SLIHI could play for the design of cell-free, drug-free osteoinductive bone graft substitutes. The evaluation contains five main sections. The first section defines relevant concepts in the context of HO and provides a summary of proposed causes of HO. The second section starts with a detailed analysis of the occurrence and involvement of calcification in HO. It is followed by an explanation of the causes of calcification and its consequences. This allows to speculate on the potential chemical modulators of inflammation and triggers of HO. The end of this second section is devoted to in vitro mineralization tests used to predict the ectopic potential of materials. The third section reviews the biological cascade of events occurring during pathological and material-induced HO, and attempts to propose a quantitative timeline of HO formation. The fourth section looks at potential ways to control HO formation, either acting on SLIHI or on inflammation. Chemical, physical, and drug-based approaches are considered. Finally, the evaluation finishes with a critical assessment of the definition of osteoinduction.

JTD Keywords: apatite, beta-tricalcium phosphate, bone, bone graft, bone morphogenetic protein, demineralized bone-matrix, experimental myositis-ossificans, extracellular calcium, heterotopic ossification, in-vitro, inflammation, multinucleated giant-cells, osteoinduction, spinal-cord-injury, total hip-arthroplasty, traumatic brain-injury, Apatite, Calcium-sensing receptor, Osteoinduction

Tuning of the crosslink density (CLD) in the rubber compounds is very crucial for optimizing the physical and mechanical properties of the ultimate rubber products. Conventionally, CLD can be measured via rheological methods such as moving die rheometer (MDR), via mechanical tests such as temperature scanning stress relaxation analysis (TSSR), or via direct swelling experiments using Flory–Rehner approach. In the current study, two novel techniques, focused ion beam - scanning electron microscopy (FIB-SEM) processing, with simultaneous energy dispersive X-ray spectrometry (EDS) mapping analysis and scanning acoustic microscopy (SAM) were combined and correlated to conventional methods on a model recipe of ethylene propylene diene monomer (EPDM) compound having different sulphur contents. Depending on the applied technique, the increase in the crosslink density with sulphur content was found to be 1.7 fold for the Flory–Rehner approach and 1.2 fold for both TSSR and MDR. It is directly monitored from the FIB-SEM-EDS analysis that the sulphur distribution and agglomeration behavior increased in line with ZnO content, which is an indirect indication of the rise in crosslink density. The impedance maps of the crosslinked samples obtained through SAM analysis revealed that the impedance of the samples increased with the increasing sulphur content, which can be attributed to higher level of crosslink density. A quantified correlation was obtained between SAM images and the crosslink density of the samples. It was shown that SAM is a promising tool for practical and non-destructive analysis for determining the formation of crosslink density of the rubbers. © The Author(s) 2022.

JTD Keywords: blends, compressibility, crosslink density, cure characteristics, ethylene propylene diene monomer, focused ion beam, mechanical-properties, morphology, natural-rubber, particles, scanning acoustic microscopy, scanning electron microscopy, sulfur, thermal-stability, vulcanization, Composite soft materials, Cross-link densities, Crosslink density, Crosslinking, Density (specific gravity), Ethylene, Ethylene propylene diene monomer, Flory-rehner, Focused ion beam - scanning electron microscopy, Focused ion beam-scanning electron microscopies, Ii-vi semiconductors, Monomers, Moving die rheometers, Physical and mechanical properties, Propylene, Relaxation analysis, Rubber, Scanning acoustic microscopy, Scanning electron microscopy, Stress relaxation, Sulfur contents, Temperature scanning stress relaxations, Zinc oxide

Emerging evidence points to coordinated action of chemical and mechanical cues during brain development. At early stages of neocortical development, angiogenic factors and chemokines such as CXCL12, ephrins, and semaphorins assume crucial roles in orchestrating neuronal migration and axon elongation of postmitotic neurons. Here we explore the intrinsic mechanical properties of the developing marginal zone of the pallium in the migratory pathways and brain distribution of the pioneer Cajal-Retzius cells. These neurons are generated in several proliferative regions in the developing brain (e.g., the cortical hem and the pallial subpallial boundary) and migrate tangentially in the preplate/marginal zone covering the upper portion of the developing cortex. These cells play crucial roles in correct neocortical layer formation by secreting several molecules such as Reelin. Our results indicate that the motogenic properties of Cajal-Retzius cells and their perinatal distribution in the marginal zone are modulated by both chemical and mechanical factors, by the specific mechanical properties of Cajal-Retzius cells, and by the differential stiffness of the migratory routes. Indeed, cells originating in the cortical hem display higher migratory capacities than those generated in the pallial subpallial boundary which may be involved in the differential distribution of these cells in the dorsal-lateral axis in the developing marginal zone.

JTD Keywords: atomic force microscopy, cajal-retzius cells, cortical development, marginal zone, mechanical cues, Atomic force microscopy, Cajal-retzius cells, Central-nervous-system, Cortical development, Cortical hem, Developing cerebral-cortex, Expression, Growth, Marginal zone, Mechanical cues, Mouse, Neuronal migration, Nogo receptor, Olfactory ensheathing cells, Tangential migration, Traction force microscopy

The influence of the properties of different solid substrates on the tethering of two antibodies, IgG1-CR3022 and IgG1-S309, which were specifically engineered for the detection of SARS-CoV-2, has been examined at the molecular level using conventional and accelerated Molecular Dynamics (cMD and aMD, respectively). Two surfaces with very different properties and widely used in immunosensors for diagnosis, amorphous silica and the most stable facet of the face-centered cubic gold structure, have been considered. The effects of such surfaces on the structure and orientation of the immobilized antibodies have been determined by quantifying the tilt and hinge angles that describe the orientation and shape of the antibody, respectively, and the dihedrals that measure the relative position of the antibody arms with respect to the surface. Results show that the interactions with amorphous silica, which are mainly electrostatic due to the charged nature of the surface, help to preserve the orientation and structure of the antibodies, especially of the IgG1-CR3022, indicating that the primary sequence of those antibodies also plays some role. Instead, short-range van der Waals interactions with the inert gold surface cause a higher degree tilting and fraying of the antibodies with respect to amorphous silica. The interactions between the antibodies and the surface also affect the correlation among the different angles and dihedrals, which increases with their strength. Overall, results explain why amorphous silica substrates are frequently used to immobilize antibodies in immunosensors. © 2022 The Authors

JTD Keywords: amorphous silica, antibody immobilization, enzyme, gol d, gold, immobilization, immunosensor, molecu l a r dynamics, molecular dynamics, protein adsorption, sars-cov-2 immunosensor, simulations, spike protein, surface interactions, target, vaccine, Amorphous silica, Antibodies, Antibody engineering, Antibody immobilization, Antibody structure, Article, Chemical detection, Computer model, Controlled study, Dihedral angle, Gold, In-silico, Molecular dynamics, Molecular levels, Molecular-dynamics, Nonhuman, Property, Sars, Sars-cov-2 immunosensor, Severe acute respiratory syndrome coronavirus 2, Silica, Silico studies, Silicon dioxide, Solid substrates, Structure analysis, Substrate chemistry, Substrates, Van der waals forces, Virus detection

One of the main limitations of in vitro studies on lung diseases is the difficulty of maintaining the type II phenotype of alveolar epithelial cells in culture. This fact has previously been related to the translocation of the mechanosensing Yes-associated protein (YAP) to the nuclei and Rho signaling pathway. In this work, we aimed to culture and subculture primary alveolar type II cells on extracellular matrix lung-derived hydrogels to assess their suitability for phenotype maintenance. Cells cultured on lung hydrogels formed monolayers and maintained type II phenotype for a longer time as compared with those conventionally cultured. Interestingly, cells successfully grew when they were subsequently cultured on a dish. Moreover, cells cultured on a plate showed the active form of the YAP protein and the formation of stress fibers and focal adhesions. The results of chemically inhibiting the Rho pathway strongly suggest that this is one of the mechanisms by which the hydrogel promotes type II phenotype maintenance. These results regarding protein expression strongly suggest that the chemical and biophysical properties of the hydrogel have a considerable impact on the transition from ATII to ATI phenotypes. In conclusion, culturing primary alveolar epithelial cells on lung ECM-derived hydrogels may facilitate the prolonged culturing of these cells, and thus help in the research on lung diseases.

JTD Keywords: adhesion, alveolar cells, expression, extracellular matrix, hydrogels, pathway, surfactant, type ii phenotype, yap, Extracellular matrix, Transplantation, Type ii phenotype

Many substances in plant extracts are known for their biological activities. These substances act in different ways, exerting overall protective effects against many diseases, especially skin disorders. However, plant extracts’ health benefits are often limited by low bioavailability. To overcome these limitations, drug delivery systems can be employed. In this study, we evaluated the antioxidant power of an ethanolic extract from Myrtus communis L. (myrtle) berries through colorimetric tests (DPPH and FRAP). The antioxidant activity was also verified by using fibroblast cell culture through cellular Reactive Oxygen Species (ROS) levels measurements. Moreover, the myrtle extract was formulated in phospholipid vesicles to improve its bioavailability and applicability. Myrtle liposomes were characterized by size, surface charge, storage stability, and entrapment efficiency; visualized by using cryo-TEM images; and assayed for cytocompatibility and anti-ROS activity. Our results suggest that myrtle liposomes were cytocompatible and improved the extract’s antioxidant power in fibroblasts, suggesting a potential skin application for these formulations and confirming that nanotechnologies could be a valid tool to enhance plant extracts’ potentialities.

JTD Keywords: antioxidant, bioactive compounds, capacity, essential oils, fibroblast, liposomes, myrtle extract, skin, Communis l., Myrtle extract, Skin

Aim: To unveil the influence of cell-matrix adhesions in the establishment of gap junction intercellular communication (GJIC) during cell condensation in chondrogenesis. Materials & methods: Previously developed nanopatterns of the cell adhesive ligand arginine-glycine-aspartic acid were used as cell culture substrates to control cell adhesion at the nanoscale. In vitro chondrogenesis of mesenchymal stem cells was conducted on the nanopatterns. Cohesion and GJIC were evaluated in cell condensates. Results: Mechanical stability and GJIC are enhanced by a nanopattern configuration in which 90% of the surface area presents adhesion sites separated less than 70 nm, thus providing an onset for cell signaling. Conclusion: Cell-matrix adhesions regulate GJIC of mesenchymal cell condensates during in vitro chondrogenesis from a threshold configuration at the nanoscale.

JTD Keywords: arginine-glycine-aspartic acid, arginine–glycine–aspartic acid, cell adhesion, condensation, dendrimer-based nanopatterning, gap junction intercellular communication, Actin, Adhesion, Arginine-glycine-aspartic acid, Cell adhesion, Collagen, Condensation, Connexin-43, Dendrimer-based nanopatterning, Dynamics, Extracellular-matrix, Fibronectin, Gap junction intercellular communication, Mesenchymal stem cells, Permeability, Phenotype, Vinculin

Collagen VI-related dystrophies (COL6-RDs) are a group of rare congenital neuromuscular dystrophies that represent a continuum of overlapping clinical phenotypes that go from the milder Bethlem myopathy (BM) to the severe Ullrich congenital muscular dystrophy, for which there is no effective treatment. Mutations in one of the three Collagen VI genes alter the incorporation of this protein into the extracellular matrix (ECM), affecting the assembly and the structural integrity of the whole fibrillar network. Clinical hallmarks of COL6-RDs are secondary to the ECM disruption and include muscle weakness, proximal joint contractures, and distal hyperlaxity. Although some traits have been identified in patients’ ECMs, a correlation between the ECM features and the clinical phenotype has not been established, mainly due to the lack of predictive and reliable models of the pathology. Herein, we engineered a new personalized pre-clinical model of COL6-RDs using cell-derived matrices (CDMs) technology to better recapitulate the complexity of the native scenario. We found that CDMs from COL6-RD patients presented alterations in ECM structure and composition, showing a significantly decreased Collagen VI secretion, especially in the more severe phenotypes, and a decrease in Fibrillin-1 inclusion. Next, we examined the Collagen VI-mediated deposition of Fibronectin in the ECM, finding a higher alignment, length, width, and straightness than in patients with COL6-RDs. Overall, these results indicate that CDMs models are promising tools to explore the alterations that arise in the composition and fibrillar architecture due to mutations in Collagen VI genes, especially in early stages of matrix organization. Ultimately, CDMs derived from COL6-RD patients may become relevant pre-clinical models, which may help identifying novel biomarkers to be employed in the clinics and to investigate novel therapeutic targets and treatments. Copyright © 2022 Almici, Chiappini, López-Márquez, Badosa, Blázquez, Caballero, Montero, Natera-de Benito, Nascimento, Roldán, Lagunas, Jiménez-Mallebrera and Samitier.

JTD Keywords: alpha-3 chain, binding, collagen vi related muscular dystrophy, decellularisation, decellularized matrices, deficiency, expression, extracellular matrix, fibroblasts, fibronectin, in vitro model, patient-derived ecms, skeletal-muscle, ullrich, Cell-derived matrices, Collagen, Collagen vi related muscular dystrophy, Decellularisation, Decellularization, Extracellular matrices, Extracellular matrix, Genes, In vitro model, In-vitro, In-vitro models, Matrix, Matrix model, Muscular dystrophy, Pathology, Patient-derived ecm, Patient-derived ecms, Pre-clinical

During development, organs reach precise shapes and sizes. Organ morphology is not always obtained through growth; a classic counterexample is the condensation of the nervous system during Drosophila embryogenesis. The mechanics underlying such condensation remain poorly understood. Here, we characterize the condensation of the embryonic ventral nerve cord (VNC) at both subcellular and tissue scales. This analysis reveals that condensation is not a unidirectional continuous process but instead occurs through oscillatory contractions. The VNC mechanical properties spatially and temporally vary, and forces along its longitudinal axis are spatially heterogeneous. We demonstrate that the process of VNC condensation is dependent on the coordinated mechanical activities of neurons and glia. These outcomes are consistent with a viscoelastic model of condensation, which incorporates time delays and effective frictional interactions. In summary, we have defined the progressive mechanics driving VNC condensation, providing insights into how a highly viscous tissue can autonomously change shape and size.

JTD Keywords: actomyosin, central nervous system, drosophila, glia, mechanics, morphogenesis, neuron, ventral nerve cord, Collagen-iv, Contraction, Forces, Gene, Glial-cells, Migration, Morphogenesis, Quantification, System, Tissue, Viscolelastic model

Research into the development of therapeutic strategies is basedmainly on animal models and cell cultures. The ability to extrapolatedata from them is limited, and research on new drugs cannot beperformed efficiently. This is especially dramatic in rare diseases,which are intrinsically very heterogeneous. The generation of ad-vanced models using tissue engineering and patient-derived cellsallows fabricating new platforms for studying pathological processesand discovering new potential drugs. Here, we developed a patient-derived 3D functional skeletal muscle for Duchenne muscular dys-trophy (DMD). DMD is the most prevalent neuromuscular diseasediagnosed during childhood. The disease is characterized by pro-gressive degeneration of skeletal and cardiac muscle caused by thelack of dystrophin protein. Although there are several molecules indrug development for DMD, there is no treatment available for pa-tients to date. By using a 3D-printed casting mold, we encapsulatedpatient-derived myogenic precursor cells in a fibrin-composite ma-trix. This platform incorporated two flexible T-shaped pillars thatprovided continuous tension to the tissue, thus allowing the orien-tation of the muscle fibers. Our 3D muscle model expressed maturemuscle markers and responded to electric pulse stimulation (EPS).Besides, contraction dynamics between DMD and control tissueswere shown to be different. Moreover, an increase of damagemarkers after EPS was observed in DMD but not in healthy tissues.Finally, the tissues will be integrated into a microfluidic device tomonitor drug administration. Eventually, the microfluidic systemwill be connected to a biosensors system for the real-time detectionof biomarkers.

JTD Keywords: Casting, Contraction dynamics, Muscular dystrophy

The severe neurological disorder epilepsy affects almost 1% of the world population. For patients who suffer from pharmacoresistant focal-onset epilepsy, electroencephalographic (EEG) recordings are essential for the localization of the brain area where seizures start. Apart from the visual inspection of the recordings, quantitative EEG signal analysis techniques proved to be useful for this purpose. Among other features, regularity versus irregularity and phase coherence versus phase independence allowed characterizing brain dynamics from the measured EEG signals. Can phase irregularities also characterize brain dynamics? To address this question, we use the univariate coefficient of phase velocity variation, defined as the ratio of phase velocity standard deviation and the mean phase velocity. Beyond that, as a bivariate measure we use the classical mean phase coherence to quantify the degree of phase locking. All phase-based measures are combined with surrogates to test null hypotheses about the dynamics underlying the signals. In the first part of our analysis, we use the Rössler model system to study our approach under controlled conditions. In the second part, we use the Bern-Barcelona EEG database which consists of focal and nonfocal signals extracted from seizure-free recordings. Focal signals are recorded from brain areas where the first seizure EEG signal changes can be detected, and nonfocal signals are recorded from areas that are not involved in the seizure at its onset. Our results show that focal signals have less phase variability and more phase coherence than nonfocal signals. Once combined with surrogates, the mean phase velocity proved to have the highest discriminative power between focal and nonfocal signals. In conclusion, conceptually simple and easy to compute phase-based measures can help to detect features induced by epilepsy from EEG signals. This holds not only for the classical mean phase coherence but even more so for univariate measures of phase irregularity. © 2022 American Physical Society.

JTD Keywords: brain, entropy, epileptogenic networks, functional connectivity, hilbert transform, seizure onset, surrogate data, synchronization, time-series, Biomedical signal processing, Brain areas, Brain dynamics, Dynamics, Electroencephalographic signals, Electroencephalography, Electrophysiology, Intracranial eeg signals, Localisation, Neurological disorders, Neurology, Phase based, Phase coherence, Signal detection, Simple++, Univariate, Velocity, World population

Large doses of movement practice have been shown to restore upper extremities' motor function in a significant subset of individuals post-stroke. However, such large doses are both difficult to implement in the clinic and highly inefficient. In addition, an important reduction in upper extremity function and use is commonly seen following rehabilitation-induced gains, resulting in “rehabilitation in vain”. For those with mild to moderate sensorimotor impairment, the limited spontaneous use of the more affected limb during activities of daily living has been previously proposed to cause a decline of motor function, initiating a vicious cycle of recovery, in which non-use and poor performance reinforce each other. Here, we review computational, experimental, and clinical studies that support the view that if arm use is raised above an effective threshold, one enters a virtuous cycle in which arm use and function can reinforce each other via self-practice in the wild. If not, one enters a vicious cycle of declining arm use and function. In turn, and in line with best practice therapy recommendations, this virtuous/vicious cycle model advocates for a paradigm shift in neurorehabilitation whereby rehabilitation be embedded in activities of daily living such that self-practice with the aid of wearable technology that reminds and motivates can enhance paretic limb use of those who possess adequate residual sensorimotor capacity. Altogether, this model points to a user-centered approach to recovery post-stroke that is tailored to the participant's level of arm use and designed to motivate and engage in self-practice through progressive success in accomplishing meaningful activities in the wild. Copyright © 2022 Ballester, Winstein and Schweighofer.

JTD Keywords: compensatory movement, computational neurorehabilitation, decision-making, individuals, learned non-use, learned nonuse, monkeys, neurorehabilitation, recovery, rehabilitation, stroke, stroke patients, wearable sensors, wrist, Arm movement, Article, Cerebrovascular accident, Clinical decision making, Clinical practice, Clinical study, Compensatory movement, Computational neurorehabilitation, Computer model, Daily life activity, Decision-making, Experimental study, Human, Induced movement therapy, Learned non-use, Musculoskeletal function, Neurorehabilitation, Paresis, Sensorimotor function, Stroke, Stroke rehabilitation, User-centered design, Vicious cycle, Virtuous cycle, Wearable sensors

Visual impairments are a critical medical hurdle to be addressed in modern society. Müller glia (MG) have regenerative potential in the retina in lower vertebrates, but not in mammals. However, in mice, in vivo cell fusion between MG and adult stem cells forms hybrids that can partially regenerate ablated neurons.We used organotypic cultures of human retina and preparations of dissociated cells to test the hypothesis that cell fusion between human MG and adult stem cells can induce neuronal regeneration in human systems. Moreover, we established a microinjection system for transplanting human retinal organoids to demonstrate hybrid differentiation.We first found that cell fusion occurs between MG and adult stem cells, in organotypic cultures of human retina as well as in cell cultures. Next, we showed that the resulting hybrids can differentiate and acquire a proto-neural electrophysiology profile when the Wnt/beta-catenin pathway is activated in the adult stem cells prior fusion. Finally, we demonstrated the engraftment and differentiation of these hybrids into human retinal organoids.We show fusion between human MG and adult stem cells, and demonstrate that the resulting hybrid cells can differentiate towards neural fate in human model systems. Our results suggest that cell fusion-mediated therapy is a potential regenerative approach for treating human retinal dystrophies.This work was supported by La Caixa Health (HR17-00231), Velux Stiftung (976a) and the Ministerio de Ciencia e Innovación, (BFU2017-86760-P) (AEI/FEDER, UE), AGAUR (2017 SGR 689, 2017 SGR 926).Published by Elsevier B.V.

JTD Keywords: cell fusion, expression, fusion, ganglion-cells, in-vitro, mouse, müller glia, neural differentiation, organoids, regeneration, retina regeneration, stem cells, stromal cells, transplantation, 4',6 diamidino 2 phenylindole, 5' nucleotidase, Agarose, Alcohol, Arpe-19 cell line, Article, Beta catenin, Beta tubulin, Bone-marrow-cells, Bromophenol blue, Buffer, Calcium cell level, Calcium phosphate, Calretinin, Canonical wnt signaling, Cd34 antigen, Cell culture, Cell fusion, Cell viability, Coculture, Complementary dna, Confocal microscopy, Cornea transplantation, Cryopreservation, Cryoprotection, Crystal structure, Current clamp technique, Dimethyl sulfoxide, Dodecyl sulfate sodium, Edetic acid, Electrophysiology, Endoglin, Fetal bovine serum, Fibroblast growth factor 2, Flow cytometry, Fluorescence activated cell sorting, Fluorescence intensity, Glyceraldehyde 3 phosphate dehydrogenase, Glycerol, Glycine, Hoe 33342, Immunofluorescence, Immunohistochemistry, Incubation time, Interleukin 1beta, Lentivirus vector, Matrigel, Mercaptoethanol, Microinjection, Mueller cell, Müller glia, N methyl dextro aspartic acid, Nerve cell differentiation, Neural differentiation, Nitrogen, Nonhuman, Organoids, Paraffin, Paraffin embedding, Paraformaldehyde, Patch clamp technique, Penicillin derivative, Phenolsulfonphthalein, Phenotype, Phosphate buffered saline, Phosphoprotein phosphatase inhibitor, Polyacrylamide gel electrophoresis, Potassium chloride, Povidone iodine, Promoter region, Proteinase inhibitor, Real time polymerase chain reaction, Receptor type tyrosine protein phosphatase c, Restriction endonuclease, Retina, Retina dystrophy, Retina regeneration, Retinol, Rhodopsin, Rna extraction, Stem cell, Stem cells, Subcutaneous fat, Tunel assay, Visual impairment, Western blotting

Decellularization procedures have been developed and optimized for the entire organ or tissue blocks, by either perfusion of decellularizing agents through the tissue’s vasculature or submerging large sections in decellularizing solutions. However, some research aims require the analysis of native as well as decellularized tissue slices side by side, but an optimal protocol has not yet been established to address this need. Thus, the main goal of this work was to develop a fast and efficient decellularization method for tissue slices—with an emphasis on lung—while attached to a glass slide. To this end, different decellularizing agents were compared for their effectiveness in cellular removal while preserving the extracellular matrix. The intensity of DNA staining was taken as an indicator of remaining cells and compared to untreated sections. The presence of collagen, elastin and laminin were quantified using immunostaining and signal quantification. Scaffolds resulting from the optimized protocol were mechanically characterized using atomic force microscopy. Lung scaffolds were recellularized with mesenchymal stromal cells to assess their biocompatibility. Some decellularization agents (CHAPS, triton, and ammonia hydroxide) did not achieve sufficient cell removal. Sodium dodecyl sulfate (SDS) was effective in cell removal (1% remaining DNA signal), but its sharp reduction of elastin signal (only 6% remained) plus lower attachment ratio (32%) singled out sodium deoxycholate (SD) as the optimal treatment for this application (6.5% remaining DNA signal), due to its higher elastin retention (34%) and higher attachment ratio (60%). Laminin and collagen were fully preserved in all treatments. The SD decellularization protocol was also successful for porcine and murine (mice and rat) lungs as well as for other tissues such as the heart, kidney, and bladder. No significant mechanical differences were found before and after sample decellularization. The resulting acellular lung scaffolds were shown to be biocompatible (98% cell survival after 72 h of culture). This novel method to decellularize tissue slices opens up new methodological possibilities to better understand the role of the extracellular matrix in the context of several diseases as well as tissue engineering research and can be easily adapted for scarce samples like clinical biopsies. Copyright © 2022 Narciso, Ulldemolins, Júnior, Otero, Navajas, Farré, Gavara and Almendros.

JTD Keywords: biocompatibility, bioscaffold recellularization, decellularization, extracellular matrix, flow, impact, lung, scaffolds, tissue slices, Ammonia, Bio-scaffolds, Biocompatibility, Biological organs, Bioscaffold recellularization, Cell removal, Cells, Collagen, Cytology, Decellularization, Dna, Dna signals, Elastin, Extracellular matrices, Extracellular matrix, Extracellular-matrix, Glycoproteins, Laminin, Lung, Mammals, Recellularization, Scaffolds (biology), Sodium deoxycholate, Sulfur compounds, Tissue, Tissue slice, Tissue slices

Remarkable progress in bioengineering over the past two decades has enabled the formulation of fundamental design principles for a variety of medical and non-medical applications. These advancements have laid the foundation for building multicellular engineered living systems (M-CELS) from biological parts, forming functional modules integrated into living machines. These cognizant design principles for living systems encompass novel genetic circuit manipulation, self-assembly, cell–cell/matrix communication, and artificial tissues/organs enabled through systems biology, bioinformatics, computational biology, genetic engineering, and microfluidics. Here, we introduce design principles and a blueprint for forward production of robust and standardized M-CELS, which may undergo variable reiterations through the classic design-build-test-debug cycle. This Review provides practical and theoretical frameworks to forward-design, control, and optimize novel M-CELS. Potential applications include biopharmaceuticals, bioreactor factories, biofuels, environmental bioremediation, cellular computing, biohybrid digital technology, and experimental investigations into mechanisms of multicellular organisms normally hidden inside the “black box” of living cells.

JTD Keywords: cell-fate specification, endothelial-cells, escherichia-coli, extracellular-matrix, gene-expression noise, nuclear hormone-receptors, pluripotent stem-cells, primitive endoderm, transcription factors, Artificial tissues, Assembly cells, Biological parts, Biological systems, Bioremediation, Blood-brain-barrier, Cell engineering, Cell/matrix communication, Design principles, Environmental technology, Functional modules, Fundamental design, Genetic circuits, Genetic engineering, Living machines, Living systems, Medical applications, Molecular biology, Synthetic biology

The characterization of newly synthesized materials is a cornerstone of all chemistry and nanotechnology laboratories. For this purpose, a wide array of analytical techniques have been standardized and are used routinely by laboratories across the globe. With these methods we can understand the structure, dynamics and function of novel molecular architectures and their relations with the desired performance, guiding the development of the next generation of materials. Moreover, one of the challenges in materials chemistry is the lack of reproducibility due to improper publishing of the sample preparation protocol. In this context, the recent adoption of the reporting standard MIRIBEL (Minimum Information Reporting in Bio–Nano Experimental Literature) for material characterization and details of experimental protocols aims to provide complete, reproducible and reliable sample preparation for the scientific community. Thus, MIRIBEL should be immediately adopted in publications by scientific journals to overcome this challenge. Besides current standard spectroscopy and microscopy techniques, there is a constant development of novel technologies that aim to help chemists unveil the structure of complex materials. Among them super-resolution microscopy (SRM), an optical technique that bypasses the diffraction limit of light, has facilitated the study of synthetic materials with multicolor ability and minimal invasiveness at nanometric resolution. Although still in its infancy, the potential of SRM to unveil the structure, dynamics and function of complex synthetic architectures has been highlighted in pioneering reports during the last few years. Currently, SRM is a sophisticated technique with many challenges in sample preparation, data analysis, environmental control and automation, and moreover the instrumentation is still expensive. Therefore, SRM is currently limited to expert users and is not implemented in characterization routines. This perspective discusses the potential of SRM to transition from a niche technique to a standard routine method for material characterization. We propose a roadmap for the necessary developments required for this purpose based on a collaborative effort from scientists and engineers across disciplines.

JTD Keywords: blinking, fluorophore, intramolecular spirocyclization, localization, nanoparticles, resolution limit, reveals, single-molecule fluorescence, stimulated-emission, Characterization techniques, Diffraction, Distributed computer systems, Environmental management, Information reporting, Material chemistry, Materials characterization, Minimum information, Optical reconstruction microscopy, Optical resolving power, Sample preparation, Structure dynamics, Structure functions, Super-resolution microscopy, Synthesized materials

The dizzying life of the homeostatic intestinal epithelium is governed by a complex interplay between fate, form, force and function. This interplay is beginning to be elucidated thanks to advances in intravital and ex vivo imaging, organoid culture, and biomechanical measurements. Recent discoveries have untangled the intricate organization of the forces that fold the monolayer into crypts and villi, compartmentalize cell types, direct cell migration, and regulate cell identity, proliferation and death. These findings revealed that the dynamic equilibrium of the healthy intestinal epithelium relies on its ability to precisely coordinate tractions and tensions in space and time. In this review, we discuss recent findings in intestinal mechanobiology, and highlight some of the many fascinating questions that remain to be addressed in this emerging field.Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.

JTD Keywords: crypt fission, designer matrices, differentiation, growth, gut, migration, model, scaffold, tissue mechanics, Cell migration, Cell proliferation, Ex vivo study, Human tissue, Intestine epithelium, Monolayer culture, Organoid, Review, Stem-cell, Tension, Traction therapy

The endosomal sorting complex required for transport (ESCRT) machinery mediates membrane fission reactions that exhibit a different topology from that observed in clathrin-coated vesicles. In all of the ESCRT-mediated events, the nascent vesicle buds away from the cytosol. However, ESCRT proteins are able to act upon membranes with different geometries. For instance, the formation of multivesicular bodies (MVBs) and the biogenesis of extracellular vesicles both require the participation of the ESCRT-III sub-complex, and they differ in their initial membrane geometry before budding starts: the protein complex acts either from outside the membrane organelle (causing inward budding) or from within (causing outward budding). Several studies have reconstituted the action of the ESCRT-III subunits in supported bilayers and cell-sized vesicles mimicking the geometry occurring during MVBs formation (in-bud), but extracellular vesicle budding (out-bud) mechanisms remain less explored, because of the outstanding difficulties encountered in encapsulation of functional ESCRT-III in vesicles. Here, we provide a different approach that allows the recreation of the out-bud formation, by combining giant unilamellar vesicles as a membrane model and a microinjection system. The vesicles are immobilized prior to injection via weak adhesion to the chamber coverslip, which also ensures preserving the membrane excess area required for budding. After protein injection, vesicles exhibit outward budding. The approach presented in this work can be used in the future to disentangle the mechanisms underlying ESCRT-III-mediated fission, recreating the geometry of extracellular bud production, which remains a challenge. Moreover, the microinjection methodology can be also adapted to interrogate the action of other cytosolic components on the encapsulating membranous organelle. Copyright: © 2022 The Authors.

JTD Keywords: adhesion, budding, electroformation, escrt-iii, exosomes, extracellular vesicles, giant unilamellar vesicle (guv), light, microinjection, microparticles, plasma, Adhesion, Budding, Escrt-iii, Extracellular vesicles, Giant unilamellar vesicle (guv), Membrane, Microinjection

JTD Keywords: augmentation, calcium-phosphate, expansion, fabrication, hydroxyapatite, made artificial bones, osteogenesis, reconstruction, regeneration, 3-d printing, 3d printers, 3d printing, 3d-printing, Abstracting, Additives, Biomaterial, Bone, Bone regeneration, Bone substitution, Bone tissue engineering, Ceramic, Ceramics, Clinic, Controlled drug delivery, Personalized medicines, Surgical planning, Targeted drug delivery, Three-dimensional-printing, Tricalcium phosphate scaffolds

The mechanical signals sensed by the alveolar cells through the changes in the local matrix stiffness of the extracellular matrix (ECM) are determinant for regulating cellular functions. Therefore, the study of the mechanical response of lung tissue becomes a fundamental aspect in order to further understand the mechanosensing signals perceived by the cells in the alveoli. This study is focused on the development of a finite element (FE) model of a decellularized rat lung tissue strip, which reproduces accurately the mechanical behaviour observed in the experiments by means of a tensile test. For simulating the complex structure of the lung parenchyma, which consists of a heterogeneous and non-uniform network of thin-walled alveoli, a 3D model based on a Voronoi tessellation is developed. This Voronoi-based model is considered very suitable for recreating the geometry of cellular materials with randomly distributed polygons like in the lung tissue. The material model used in the mechanical simulations of the lung tissue was characterized experimentally by means of AFM tests in order to evaluate the lung tissue stiffness on the micro scale. Thus, in this study, the micro (AFM test) and the macro scale (tensile test) mechanical behaviour are linked through the mechanical simulation with the 3D FE model based on Voronoi tessellation. Finally, a micro-mechanical FE-based model is generated from the Voronoi diagram for studying the stiffness sensed by the alveolar cells in function of two independent factors: the stretch level of the lung tissue and the geometrical position of the cells on the extracellular matrix (ECM), distinguishing between pneumocyte type I and type II. We conclude that the position of the cells within the alveolus has a great influence on the local stiffness perceived by the cells. Alveolar cells located at the corners of the alveolus, mainly type II pneumocytes, perceive a much higher stiffness than those located in the flat areas of the alveoli, which correspond to type I pneumocytes. However, the high stiffness, due to the macroscopic lung tissue stretch, affects both cells in a very similar form, thus no significant differences between them have been observed. © 2021 The Authors

JTD Keywords: rat, scaffolds, stiffness, Afm, Animal cell, Animal experiment, Animal model, Animal tissue, Article, Biological organs, Cell function, Cells, Computational geometry, Cytology, Extracellular matrices, Extracellular matrix, Extracellular-matrix, Geometry, High stiffness, Human, Lung alveolus cell type 1, Lung alveolus cell type 2, Lung parenchyma, Lung tissue, Male, Mechanical behavior, Mechanical modeling, Mechanical simulations, Mechanosensing, Model-based opc, Nonhuman, Physical model, Rat, Rigidity, Stiffness, Stiffness matrix, Tensile testing, Thin walled structures, Three dimensional finite element analysis, Tissue, Type ii, Voronoi tessellations

JTD Keywords: amino-acids, catalysis, dopant-free hydroxyapatite, electrical properties, electrophotosynthesis, nitrogen, thermally-stimulated polarization, Advanced materials, Biocompatibility, Biomedical applications, Brushite, Doped hydroxyapatites, Electric voltage, Electrical characterization, Electrochemical impedance spectroscopy, Equivalent circuits, Future perspectives, Highest temperature, Hydroxyapatite, Interfaces (materials), Material interfaces, Medical applications, Polarization, Polarization conditions, Surface-charges, Technological applications

The use of broadly neutralizing antibodies against human immunodeficiency virus type 1 (HIV-1) has been shown to be a promising therapeutic modality in the prevention of HIV infection. Understanding the b12-gp120 binding mechanism under physiological conditions may assist the development of more broadly effective antibodies. In this work, the main conformations and interactions between the receptor-binding domain (RBD) of spike glycoprotein gp120 of HIV-1 and the IgG1-b12 mAb are studied. Accelerated molecular dynamics (aMD) and ab initio hybrid molecular dynamics have been combined to determine the most persistent interactions between the most populated conformations of the antibody-antigen complex under physiological conditions. The results show the most persistent receptor-binding mapping in the conformations of the antibody-antigen interface in solution. The binding-free-energy decomposition reveals a small enhancement in the contribution played by the CDR-H3 region to the b12-gp120 interface compared to the crystal structure.

JTD Keywords: antibody, complex, functionals, gp120 envelope glycoprotein, hiv, immunodeficiency-virus, noncovalent interactions, simulations, software integration, Ab initio, Accelerated molecular dynamics, Accelerated molecular-dynamics, Antibodies, Antigens, Binding energy, Binding mechanisms, Computational studies, Crystal structure, Diseases, Free energy, Hiv infection, Human immunodeficiency virus, Molecular dynamics, Neutralizing antibodies, Physiological condition, Physiology, Receptor-binding domains, Therapeutic modality, Viruses

Despite tremendous progress in the understanding of COVID-19, mechanistic insight into immunological, disease-driving factors remains limited. We generated maVie16, a mouse-adapted SARS-CoV-2, by serial passaging of a human isolate. In silico modeling revealed how only three Spike mutations of maVie16 enhanced interaction with murine ACE2. maVie16 induced profound pathology in BALB/c and C57BL/6 mice, and the resulting mouse COVID-19 (mCOVID-19) replicated critical aspects of human disease, including early lymphopenia, pulmonary immune cell infiltration, pneumonia, and specific adaptive immunity. Inhibition of the proinflammatory cyto-kines IFN? and TNF substantially reduced immunopathology. Importantly, genetic ACE2-deficiency completely prevented mCOVID-19 development. Finally, inhalation therapy with recombinant ACE2 fully protected mice from mCOVID-19, revealing a novel and efficient treatment. Thus, we here present maVie16 as a new tool to model COVID-19 for the discovery of new therapies and show that disease severity is determined by cytokine-driven immunopathology and critically dependent on ACE2 in vivo. © Gawish et al.

JTD Keywords: covid-19 mouse model, covid-19 therapy, cytokine storm, immunology, inflammation, mavie16, mouse, mouse-adapted sars-cov-2, program, recombinant soluble ace2, tmprss2, Adaptive immunity, Angiotensin converting enzyme 2, Angiotensin-converting enzyme 2, Animal, Animal cell, Animal experiment, Animal model, Animal tissue, Animals, Apoptosis, Article, Bagg albino mouse, Breathing rate, Bronchoalveolar lavage fluid, C57bl mouse, Cell composition, Cell infiltration, Controlled study, Coronavirus disease 2019, Coronavirus spike glycoprotein, Covid-19, Cytokeratin 18, Cytokine production, Dipeptidyl carboxypeptidase, Disease model, Disease models, animal, Disease severity, Drosophila-melanogaster, Enzyme linked immunosorbent assay, Expression vector, Flow cytometry, Gamma interferon, Gene editing, Gene expression, Gene mutation, Genetic engineering, Genetics, Glycosylation, High mobility group b1 protein, Histology, Histopathology, Immune response, Immunocompetent cell, Immunology, Immunopathology, Interferon-gamma, Interleukin 2, Metabolism, Mice, inbred balb c, Mice, inbred c57bl, Mouse-adapted sars-cov-2, Myeloperoxidase, Neuropilin 1, Nonhuman, Nucleocapsid protein, Pathogenicity, Peptidyl-dipeptidase a, Pyroptosis, Recombinant soluble ace2, Renin angiotensin aldosterone system, Rna extraction, Rna isolation, Sars-cov-2, Severe acute respiratory syndrome coronavirus 2, Spike glycoprotein, coronavirus, T lymphocyte activation, Trabecular meshwork, Tumor necrosis factor, Virology, Virus load, Virus replication, Virus transmission, Virus virulence

With the currently available materials and technologies it is difficult to mimic the mechanical properties of soft living tissues. Additionally, another significant problem is the lack of information about the mechanical properties of these tissues. Alternatively, the use of phantoms offers a promising solution to simulate biological bodies. For this reason, to advance in the state-of-the-art a wide range of organs (e.g., liver, heart, kidney as well as brain) and hydrogels (e.g., agarose, polyvinyl alcohol –PVA–, Phytagel –PHY– and methacrylate gelatine –GelMA–) were tested regarding their mechanical properties. For that, viscoelastic behavior, hardness, as well as a non-linear elastic mechanical response were measured. It was seen that there was a significant difference among the results for the different mentioned soft tissues. Some of them appear to be more elastic than viscous as well as being softer or harder. With all this information in mind, a correlation between the mechanical properties of the organs and the different materials was performed. The next conclusions were drawn: (1) to mimic the liver, the best material is 1% wt agarose; (2) to mimic the heart, the best material is 2% wt agarose; (3) to mimic the kidney, the best material is 4% wt GelMA; and (4) to mimic the brain, the best materials are 4% wt GelMA and 1% wt agarose. Neither PVA nor PHY was selected to mimic any of the studied tissues. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

JTD Keywords: brain, composite hydrogel, dynamic mechanical analysis, elastography, hardness, hydrogels, in-vitro, liver, materials, mechanical-properties, mimicking, soft tissues, tissue scaffolding, viscoelasticity, warner-braztler shear test, warner–braztler shear test, Dynamic mechanical analysis, Hardness, Hydrogels, Materials, Mimicking, Soft tissues, Tissue scaffolding, Viscoelastic characterization, Viscoelasticity, Warner–braztler shear test

Mycobacterium bovis bacillus Calmette-Guérin (BCG) efficacy as an immunotherapy tool can be influenced by the genetic background or immune status of the treated population and by the BCG substrain used. BCG comprises several substrains with genetic differences that elicit diverse phenotypic characteristics. Moreover, modifications of phenotypic characteristics can be influenced by culture conditions. However, several culture media formulations are used worldwide to produce BCG. To elucidate the influence of growth conditions on BCG characteristics, five different substrains were grown on two culture media, and the lipidic profile and physico-chemical properties were evaluated. Our results show that each BCG substrain displays a variety of lipidic profiles on the outermost surface depending on the growth conditions. These modifications lead to a breadth of hydrophobicity patterns and a different ability to reduce neutral red dye within the same BCG substrain, suggesting the influence of BCG growth conditions on the interaction between BCG cells and host cells.

JTD Keywords: cell wall, efficacy, glycerol, hydrophobicity, lipid, neutral red, pdim, pgl, protein, strains, viability, virulence, Acylglycerol, Albumin, Article, Asparagine, Bacterial cell wall, Bacterial gene, Bacterium culture, Bcg vaccine, Catalase, Cell wall, Chloroform, Controlled study, Escherichia coli, Gene expression, Genomic dna, Glycerol, Glycerol monomycolate, Hexadecane, Housekeeping gene, Hydrophobicity, Immune response, Immunogenicity, Immunotherapy, Lipid, Lipid fingerprinting, Magnesium sulfate, Mercaptoethanol, Methanol, Methylglyoxal, Molybdatophosphoric acid, Mycobacterium bovis bcg, Neutral red, Nonhuman, Pdim, Petroleum ether, Pgl, Phenotype, Physical chemistry, Real time reverse transcription polymerase chain reaction, Rna 16s, Rna extraction, Rv0577, Staining, Thin layer chromatography, Unclassified drug

Drug development is an ever-growing field, increasingly requesting reliable in vitro tools to speed up early screening phases, reducing the need for animal experiments. In oral delivery, understanding the absorption pattern of a new drug in the small intestine is paramount. Classical two-dimensional (2D) in vitro models are generally too simplistic and do not accurately represent native tissues. The main goal of this work was to develop an advanced three-dimensional (3D) in vitro intestinal model to test absorption in a more reliable manner, by better mimicking the native environment. The 3D model is composed of a collagen-based stromal layer with embedded fibroblasts mimicking the intestinal lamina propria and providing support for the epithelium, composed of enterocytes and mucus-secreting cells. An endothelial layer, surrogating the absorptive capillary network, is also present. The cellular crosstalk between the different cells present in the model is unveiled, disclosing key players, namely those involved in the contraction of collagen by fibroblasts. The developed 3D model presents lower levels of P-glycoprotein (P-gp) and Multidrug Resistance Protein 2 (MRP2) efflux transporters, which are normally overexpressed in traditional Caco-2 models, and are paramount in the absorption of many compounds. This, allied with transepithelial electrical resistance (TEER) values closer to physiological ranges, leads to improved and more reliable permeability outcomes, which are observed when comparing our results with in vivo data.

JTD Keywords: 3d intestinal model, drug absorption, drug development, endothelium, hydrogel, 3d intestinal model, 3d modeling, 3d models, 3d-modeling, Alkaline-phosphatase, Animal experiments, Biopharmaceutics classification, Caco-2 cells, Cell culture, Collagen, Collagen gel, Drug absorption, Drug development, Endothelium, Fibroblasts, Glycoproteins, Hydrogel, In-vitro, Matrix metalloproteinases, Membrane-permeability, Paracellular transport, Permeability, Single-pass vs., Speed up

Introduction: After a stroke, a wide range of deficits can occur with varying onset latencies. As a result, assessing impairment and recovery are enormous challenges in neurorehabilitation. Although several clinical scales are generally accepted, they are time-consuming, show high inter-rater variability, have low ecological validity, and are vulnerable to biases introduced by compensatory movements and action modifications. Alternative methods need to be developed for efficient and objective assessment. In this study, we explore the potential of computer-based body tracking systems and classification tools to estimate the motor impairment of the more affected arm in stroke patients. Methods: We present a method for estimating clinical scores from movement parameters that are extracted from kinematic data recorded during unsupervised computer-based rehabilitation sessions. We identify a number of kinematic descriptors that characterise the patients' hemiparesis (e.g., movement smoothness, work area), we implement a double-noise model and perform a multivariate regression using clinical data from 98 stroke patients who completed a total of 191 sessions with RGS. Results: Our results reveal a new digital biomarker of arm function, the Total Goal-Directed Movement (TGDM), which relates to the patients work area during the execution of goal-oriented reaching movements. The model's performance to estimate FM-UE scores reaches an accuracy of R-2: 0.38 with an error (sigma: 12.8). Next, we evaluate its reliability (r = 0.89 for test-retest), longitudinal external validity (95% true positive rate), sensitivity, and generalisation to other tasks that involve planar reaching movements (R-2: 0.39). The model achieves comparable accuracy also for the Chedoke Arm and Hand Activity Inventory (R-2: 0.40) and Barthel Index (R-2: 0.35). Conclusions: Our results highlight the clinical value of kinematic data collected during unsupervised goal-oriented motor training with the RGS combined with data science techniques, and provide new insight into factors underlying recovery and its biomarkers.

JTD Keywords: interactive feedback, motion classification, motion sensing, multivariate regression, posture monitoring, rehabilitation, stroke, Adult, Aged, Analytic method, Arm movement, Article, Barthel index, Brain hemorrhage, Cerebrovascular accident, Chedoke arm and hand activity inventory, Clinical protocol, Cognitive defect, Computer analysis, Controlled study, Convergent validity, Correlation coefficient, Disease severity, External validity, Female, Fugl meyer assessment for the upper extremity, Functional assessment, Functional status assessment, General health status assessment, Hemiparesis, Human, Interactive feedback, Ischemic stroke, Kinematics, Major clinical study, Male, Mini mental state examination, Motion classification, Motion sensing, Motor analog scale, Movement, Multivariate regression, Muscle function, Posture monitoring, Probability, Recovery, Rehabilitation, Reliability, Retrospective study, Stroke, Stroke patient, Test retest reliability, Therapy, Total goal directed movement, Upper extremities, Upper limb, Upper-limb, Wolf motor function test

In clinical practice, when a patient is undergoing mechanical ventilation, it is important to identify the optimal moment for extubation, minimizing the risk of failure. However, this prediction remains a challenge in the clinical process. In this work, we propose a new protocol to study the extubation process, including the electromyographic diaphragm signal (diaEMG) recorded through 5-channels with surface electrodes around the diaphragm muscle. First channel corresponds to the electrode on the right. A total of 40 patients in process of withdrawal of mechanical ventilation, undergoing spontaneous breathing tests (SBT), were studied. According to the outcome of the SBT, the patients were classified into two groups: successful (SG: 19 patients) and failure (FG: 21 patients) groups. Parameters extracted from the envelope of each channel of diaEMG in time and frequency domain were studied. After analyzing all channels, the second presented maximum differences when comparing the two groups of patients, with parameters related to root mean square (p = 0.005), moving average (p = 0.001), and upward slope (p = 0.017). The third channel also presented maximum differences in parameters as the time between maximum peak (p = 0.004), and the skewness (p = 0.027). These results suggest that diaphragm EMG signal could contribute to increase the knowledge of the behaviour of respiratory system in these patients and improve the extubation process.Clinical Relevance - This establishes the characterization of success and failure patients in the extubation process. © 2021 IEEE.

JTD Keywords: classification, recognition, Airway extubation, Artificial ventilation, Clinical practices, Clinical process, Diaphragm, Diaphragm muscle, Diaphragms, Electrodes, Electromyographic, Extubation, Frequency domain analysis, Human, Humans, Maximum differences, Mechanical ventilation, New protocol, Respiration, artificial, Respiratory system, Risk of failure, Spontaneous breathing, Surface electrode, Surface emg signals, Thorax, Ventilation, Ventilator weaning

Abstract The development of nanostructured plasmonic biosensors has been widely widespread in the last years, motivated by the potential benefits they can offer in integration, miniaturization, multiplexing opportunities, and enhanced performance label-free biodetection in a wide field of applications. Between them, engineering tissues represent a novel, challenging, and prolific application field for nanostructured plasmonic biosensors considering the previously described benefits and the low levels of secreted biomarkers (?pM–nM) to detect. Here, we present an integrated plasmonic nanocrystals-based biosensor using high throughput nanostructured polycarbonate substrates. Metallic film thickness and incident angle of light for reflectance measurements were optimized to enhance the detection of antibody–antigen biorecognition events using numerical simulations. We achieved an enhancement in biodetection up to 3× as the incident angle of light decreases, which can be related to shorter evanescent decay lengths. We achieved a high reproducibility between channels with a coefficient of variation below 2% in bulk refractive index measurements, demonstrating a high potential for multiplexed sensing. Finally, biosensing potential was demonstrated by the direct and label-free detection of interleukin-6 biomarker in undiluted cell culture media supernatants from bioengineered 3D skeletal muscle tissues stimulated with different concentrations of endotoxins achieving a limit of detection (LOD) of ? 0.03 ng/mL (1.4 pM).

JTD Keywords: assay, crystals, drug, label-free biosensing, molecules, plasmonic nanostructures, sensors, skeletal muscle, tissue engineering, Biodetection, Biomarkers, Biosensors, Cell culture, Cells, Chemical detection, Histology, Interleukin-6, Interleukin6 (il6), Label free, Label-free biosensing, Muscle, Nano-structured, Nanocrystals, Plasmonic nanocrystals, Plasmonic nanostructures, Plasmonics, Polycarbonate substrates, Polycarbonates, Refractive index, Sensitivity, Skeletal muscle, Tissue engineering, Tissues engineerings

The generation of kidney organoids from human pluripotent stem cells (hPSCs) has represented a relevant scientific achievement in the organoid field. Importantly, hPSC-derived kidney organoids contain multiple nephron-like structures that exhibit some renal functional characteristics and have the capacity to respond to nephrotoxic agents. In this review, we first discuss how bioengineering approaches can help overcome current kidney organoid challenges. Next, we focus on recent works exploiting kidney organoids for drug screening and disease modeling applications. Finally, we provide a state of the art on current research toward the potential application of kidney organoids and renal cells derived from hPSCs for future renal replacement therapies.

JTD Keywords: Bioengineering, Converting enzyme-ii, Crispr/cas9 gene editing, Disease, Disease modeling, Extracellular-matrix, Generation, Human pluripotent stem cells, Kidney organoids, Kidney regeneration, Model, Mouse, Reveals, Scaffold, Transplantation

Abstract Critical dynamics, characterized by scale-free neuronal avalanches, is thought to underlie optimal function in the sensory cortices by maximizing information transmission, capacity, and dynamic range. In contrast, deviations from criticality have not yet been considered to support any cognitive processes. Nonetheless, neocortical areas related to working memory and decision-making seem to rely on long-lasting periods of ignition-like persistent firing. Such firing patterns are reminiscent of supercritical states where runaway excitation dominates the circuit dynamics. In addition, a macroscopic gradient of the relative density of Somatostatin (SST+) and Parvalbumin (PV+) inhibitory interneurons throughout the cortical hierarchy has been suggested to determine the functional specialization of low- versus high-order cortex. These observations thus raise the question of whether persistent activity in high-order areas results from the intrinsic features of the neocortical circuitry. We used an attractor model of the canonical cortical circuit performing a perceptual decision-making task to address this question. Our model reproduces the known saddle-node bifurcation where persistent activity emerges, merely by increasing the SST+/PV+ ratio while keeping the input and recurrent excitation constant. The regime beyond such a phase transition renders the circuit increasingly sensitive to random fluctuations of the inputs -i.e., chaotic-, defining an optimal SST+/PV+ ratio around the edge-of-chaos. Further, we show that both the optimal SST+/PV+ ratio and the region of the phase transition decrease monotonically with increasing input noise. This suggests that cortical circuits regulate their intrinsic dynamics via inhibitory interneurons to attain optimal sensitivity in the face of varying uncertainty. Hence, on the one hand, we link the emergence of supercritical dynamics at the edge-of-chaos to the gradient of the SST+/PV+ ratio along the cortical hierarchy, and, on the other hand, explain the behavioral effects of the differential regulation of SST+ and PV+ interneurons by neuromodulators like acetylcholine in the presence of input uncertainty.

JTD Keywords: attractor model, cortex, cortical networks, edge-of-chaos, model, nmda receptors, Attractor model, Cortical hierarchies, Decision making, Dynamics, Edge of chaos, Edge-of-chaos, High-order, Higher-order, Inhibitory interneurons, Neurons, Optimal decision making, Persistent activities, Persistent activity, Supercritical, Supercriticality

Pulmonary fibrosis (PF) is a progressive disease that disrupts the mechanical homeostasis of the lung extracellular matrix (ECM). These effects are particularly relevant in the lung context, given the dynamic nature of cyclic stretch that the ECM is continuously subjected to during breathing. This work uses an in vivo model of pulmonary fibrosis to characterize the macro-and micromechanical properties of lung ECM subjected to stretch. To that aim, we have compared the micromechanical properties of fibrotic ECM in baseline and under stretch conditions, using a novel combination of Atomic Force Microscopy (AFM) and a stretchable membrane-based chip. At the macroscale, fibrotic ECM displayed strain-hardening, with a stiffness one order of magnitude higher than its healthy counterpart. Conversely, at the microscale, we found a switch in the stretch-induced mechanical behaviour of the lung ECM from strain-hardening at physiological ECM stiffnesses to strain-softening at fibrotic ECM stiffnesses. Similarly, we observed solidification of healthy ECM versus fluidization of fibrotic ECM in response to stretch. Our results suggest that the mechanical behaviour of fibrotic ECM under stretch involves a potential built-in mechanotransduction mechanism that may slow down the progression of PF by steering resident fibroblasts away from a pro-fibrotic profile. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

JTD Keywords: atomic force microscopy, extracellular matrix, fibrosis, mechanics, mechanosensing, strain, system, viscoelasticity, Atomic force microscopy, Extracellular matrix, Fibrosis, Lung fibrosis, Mechanosensing

The extracellular matrix mechanical properties regulate processes in development, cancer, and fibrosis. Among the distinct mechanical properties, the vast majority of research has focused on the extracellular matrix's elasticity as the primary determinant of cell and tissue behavior. However, both cells and the extracellular matrix are not only elastic but also viscous. Despite viscoelasticity being a universal feature of living tissues, our knowledge of the influence of the extracellular matrix's viscoelasticity in cell behavior is limited. This mini-review describes some of the recent findings that have highlighted the role of the extracellular matrix's viscoelasticity in cell and tissue dynamics.

JTD Keywords: behavior, cell adhesion, cell mechanics, cell migration, deformability, extracellular matrix, extracellular matrix mechanics, fluidity, forces, hydrogels, mechanobiology, mechanotransduction, tissue mechanics, viscoelasticity, viscosity, Cell adhesion, Cell mechanics, Cell migration, Extracellular matrix, Extracellular matrix mechanics, Fluidity, Mechanobiology, Mechanotransduction, Migration, Tissue mechanics, Viscoelasticity, Viscosity

[Figure: see text].

JTD Keywords: activation, cell extrusion, contraction, drives, homeostasis, interface, junctions, kinase, tension, Adhesion, Article, Cell membranes, Chemical activation, Cytology, E-cadherin, Epithelial monolayers, Epithelium, Homoeostasis, Human, Mechanical instabilities, Monolayers, Morphological transformations, Morphology, Normal tissue, Oncogenics, Soft substrates, Substrates, Tissue, Tissue mechanics, Tissue morphology, Tumor development

Most morphogenetic and pathological processes are driven by cells responding to the surrounding matrix, such as its composition, architecture, and mechanical properties. Despite increasing evidence for the role of extracellular matrix (ECM) in tissue and disease development, many in vitro substitutes still fail to effectively mimic the native microenvironment. We established a novel method to produce macroscale (>1 cm) mesenchymal cell-derived matrices (CDMs) aimed to mimic the fibrotic tumor microenvironment surrounding epithelial cancer cells. CDMs are produced by human adipose mesenchymal stem cells cultured in sacrificial 3D scaffold templates of fibronectin-coated poly-lactic acid microcarriers (MCs) in the presence of macromolecular crowders. We showed that decellularized CDMs closely mimic the fibrillar protein composition, architecture, and mechanical properties of human fibrotic ECM from cancer masses. CDMs had highly reproducible composition made of collagen types I and III and fibronectin ECM with tunable mechanical properties. Moreover, decellularized and MC-free CDMs were successfully repopulated with cancer cells throughout their 3D structure, and following chemotherapeutic treatment, cancer cells showed greater doxorubicin resistance compared to 3D culture in collagen hydrogels. Collectively, these results support the use of CDMs as a reproducible and tunable tool for developing 3D in vitro cancer models.

JTD Keywords: 3d cell-derived matrices, adipose mesenchymal stem cells, collagen matrix, colorectal adenocarcinoma, cytotoxicity assay, deposition, expansion, extracellular microenvironment, extracellular-matrix, fibronectin, growth, macromolecular crowders, microcarriers, scaffolds, tissue, 3d cell-derived matrices, Adipose mesenchymal stem cells, Cytotoxicity assay, Extracellular microenvironment, Macromolecular crowders, Mesenchymal stem-cells, Microcarriers

A plethora of applications using polysaccharides have been developed in recent years due to their availability as well as their frequent nontoxicity and biodegradability. These polymers are usually obtained from renewable sources or are byproducts of industrial processes, thus, their use is collaborative in waste management and shows promise for an enhanced sustainable circular economy. Regarding the development of novel delivery systems for biotherapeutics, the potential of polysaccharides is attractive for the previously mentioned properties and also for the possibility of chemical modification of their structures, their ability to form matrixes of diverse architectures and mechanical properties, as well as for their ability to maintain bioactivity following incorporation of the biomolecules into the matrix. Biotherapeutics, such as proteins, growth factors, gene vectors, enzymes, hormones, DNA/RNA, and antibodies are currently in use as major therapeutics in a wide range of pathologies. In the present review, we summarize recent progress in the development of polysaccharide-based hydrogels of diverse nature, alone or in combination with other polymers or drug delivery systems, which have been implemented in the delivery of biotherapeutics in the pharmaceutical and biomedical fields. © 2021 American Chemical Society.

JTD Keywords: biodegradable dextran hydrogels, biotherapeutics, bone morphogenetic protein-2, carrageenan-based hydrogels, chitosan-based hydrogels, controlled delivery, controlled-release, cross-linked hydrogels, growth-factor delivery, hydrogels, in-vitro characterization, polysaccharides, self-healing hydrogel, stimuli-responsiveness, tissue engineering, Antibodies, Bioactivity, Biodegradability, Biomedical fields, Biomolecules, Biotherapeutics, Chemical modification, Circular economy, Controlled delivery, Controlled drug delivery, Delivery systems, Drug delivery system, Functional polymers, Hyaluronic-acid hydrogels, Hydrogels, Industrial processs, Polysaccharides, Recent progress, Renewable sources, Stimuli-responsiveness, Targeted drug delivery, Tissue engineering, Waste management

Gas chromatography—ion mobility spectrometry (GC-IMS) allows the fast, reliable, and inexpensive chemical composition analysis of volatile mixtures. This sensing technology has been successfully employed in food science to determine food origin, freshness and preventing alimentary fraud. However, GC-IMS data is highly dimensional, complex, and suffers from strong non-linearities, baseline problems, misalignments, peak overlaps, long peak tails, etc., all of which must be corrected to properly extract the relevant features from samples. In this work, a pipeline for signal pre-processing, followed by four different approaches for feature extraction in GC-IMS data, is presented. More precisely, these approaches consist of extracting data features from: (1) the total area of the reactant ion peak chromatogram (RIC); (2) the full RIC response; (3) the unfolded sample matrix; and (4) the ion peak volumes. The resulting pipelines for data processing were applied to a dataset consisting of two different quality class Iberian ham samples, based on their feeding regime. The ability to infer chemical information from samples was tested by comparing the classification results obtained from partial least-squares discriminant analysis (PLS-DA) and the samples’ variable importance for projection (VIP) scores. The choice of a feature extraction strategy is a trade-off between the amount of chemical information that is preserved, and the computational effort required to generate the data models.

JTD Keywords: authenticity, classification, electronic-nose, feature extraction, food analysis, gc-ims, headspace, least-squares, models, pld-da, pre-processing, quality, sensory analysis, wine, Feature extraction, Food analysis, Gc-ims, Hs-gc-ims, Pld-da, Pre-processing

Pseudomonas aeruginosa biofilms and the capacity of the bacterium to coexist and interact with a broad range of microorganisms have a substantial clinical impact. This review focuses on the main traits of P. aeruginosa biofilms, such as the structural composition and regulatory networks involved, placing particular emphasis on the clinical challenges they represent in terms of antimicrobial susceptibility and biofilm infection clearance. Furthermore, the ability of P. aeruginosa to grow together with other microorganisms is a significant pathogenic attribute with clinical relevance; hence, the main microbial interactions of Pseudomonas are especially highlighted and detailed throughout this review. This article also explores the infections caused by single and polymicrobial biofilms of P. aeruginosa and the current models used to recreate them under laboratory conditions. Finally, the antimicrobial and antibiofilm strategies developed against P. aeruginosa mono and multispecies biofilms are detailed at the end of this review.

JTD Keywords: aeruginosa models, antibiotic-resistance, antimicrobials, bacterial biofilms, biofilms, c-di-gmp, chronic infections, enterococcus-faecalis, extracellular dna, in-vitro, lectin pa-iil, p, p. aeruginosa models, polymicrobial, polymicrobial interactions, staphylococcus-aureus, Antimicrobials, Biofilms, Chronic infections, P. aeruginosa models, Polymicrobial, Pseudomonas aeruginosa, Urinary-tract-infection

Catalytically powered micro/nanobots (MNBs) can perform active movement by harnessing energy from in situ chemical reactions and show tremendous potential in biomedical applications. However, the development of imageable MNBs that are driven by bioavailable fuels and possess multiple therapeutic functions remains challenging. To resolve such issues, we herein propose enzyme (urease) powered liquid metal (LM) nanobots that are naturally of multiple therapeutic functions and imaging signals. The main body of the nanobot is composed of a biocompatible LM nanoparticle encapsulated by polydopamine (PDA). Urease enzyme needed for the powering and desired drug molecules, e.g., cefixime trihydrate antibiotic, are grafted on external surfaces of the PDA shell. Such a chemical composition endows the nanobots with dual-mode ultrasonic (US) and photoacoustic (PA) imaging signals and favorable photothermal effect. These LM nanobots exhibit positive chemotaxis and therefore can be collectively guided along a concentration gradient of urea for targeted transportation. When exposed to NIR light, the LM nanobots would deform and complete the function change from active drug carriers to photothermal reagents, to achieve synergetic antibacterial treatment by both photothermal and chemotherapeutic effects. The US and PA properties of the LM nanoparticle can be used to not only track and monitor the active movement of the nanobots in a microfluidic vessel model but also visualize their dynamics in the bladder of a living mouse in vivo. To conclude, the LM nanobots demonstrated herein represent a proof-of-concept therapeutic nanosystem with multiple biomedical functionalities, providing a feasible tool for preclinical studies and clinical trials of MNB-based imaging-guided therapy.

JTD Keywords: cell, chemo-photothermal therapy, chemotaxis, image tracking, liquid metal nanobots, nanomotors, tracking, Chemo-photothermal therapy, Chemotaxis, Image tracking, Liquid metal nanobots, Nanomotors

Abstract Purpose of the Review This review aims to summarize the current knowledge of the extracellular matrix remodeling during hepatic fibrosis. We discuss the diverse interactions of the extracellular matrix with hepatic cells and the surrounding matrix in liver fibrosis, with the focus on the molecular pathways and the mechanisms that regulate extracellular matrix remodeling. Recent Findings The extracellular matrix not only provides structure and support for the cells, but also controls cell behavior by providing adhesion signals and by acting as a reservoir of growth factors and cytokines. Summary Hepatic fibrosis is characterized by an excessive accumulation of extracellular matrix. During fibrogenesis, the natural remodeling process of the extracellular matrix varies, resulting in the excessive accumulation of its components, mainly collagens. Signals released by the extracellular matrix induce the activation of hepatic stellate cells, which are the major source of extracellular matrix and most abundant myofibroblasts in the liver. Graphical abstract

JTD Keywords: collagen, extracellular matrix, hepatic stellate cell, liver fibrosis, metalloproteinases, Tgf-?1, Tgf-β1

Cell response to force regulates essential processes in health and disease. However, the fundamental mechanical variables that cells sense and respond to remain unclear. Here we show that the rate of force application (loading rate) drives mechanosensing, as predicted by a molecular clutch model. By applying dynamic force regimes to cells through substrate stretching, optical tweezers, and atomic force microscopy, we find that increasing loading rates trigger talin-dependent mechanosensing, leading to adhesion growth and reinforcement, and YAP nuclear localization. However, above a given threshold the actin cytoskeleton softens, decreasing loading rates and preventing reinforcement. By stretching rat lungs in vivo, we show that a similar phenomenon may occur. Our results show that cell sensing of external forces and of passive mechanical parameters (like tissue stiffness) can be understood through the same mechanisms, driven by the properties under force of the mechanosensing molecules involved. Cells sense mechanical forces from their environment, but the precise mechanical variable sensed by cells is unclear. Here, the authors show that cells can sense the rate of force application, known as the loading rate, with effects on YAP nuclear localization and cytoskeletal stiffness remodelling.

JTD Keywords: Actin cytoskeleton, Actin filament, Actin-filament, Adhesion, Animal, Animals, Atomic force microscopy, Breathing, Cell, Cell adhesion, Cell culture, Cell nucleus, Cells, cultured, Cytoplasm, Extracellular-matrix, Fibroblast, Fibroblasts, Fibronectin, Frequency, Gene knockdown, Gene knockdown techniques, Genetics, Germfree animal, Integrin, Intracellular signaling peptides and proteins, Knockout mouse, Lung, Male, Mechanotransduction, Mechanotransduction, cellular, Metabolism, Mice, Mice, knockout, Microscopy, atomic force, Mouse, Optical tweezers, Paxillin, Physiology, Primary cell culture, Pxn protein, mouse, Rat, Rats, Rats, sprague-dawley, Respiration, Signal peptide, Softening, Specific pathogen-free organisms, Sprague dawley rat, Stress, Substrate, Substrate rigidity, Talin, Talin protein, mouse, Tln2 protein, mouse, Traction, Transmission, Ultrastructure, Yap1 protein, rat

Mesenchymal stromal cell (MSC)-based cell therapy in acute respiratory diseases is based on MSC secretion of paracrine factors. Several strategies have proposed to improve this are being explored including pre-conditioning the MSCs prior to administration. We here propose a strategy for improving the therapeutic efficacy of MSCs based on cell preconditioning by growing them in native extracellular matrix (ECM) derived from the lung. To this end, a bioink with tunable stiffness based on decellularized porcine lung ECM hydrogels was developed and characterized. The bioink was suitable for 3D culturing of lung-resident MSCs without the need for additional chemical or physical crosslinking. MSCs showed good viability, and contraction assays showed the existence of cell–matrix interactions in the bioprinted scaffolds. Adhesion capacity and length of the focal adhesions formed were increased for the cells cultured within the lung hydrogel scaffolds. Also, there was more than a 20-fold increase of the expression of the CXCR4 receptor in the 3D-cultured cells compared to the cells cultured in plastic. Secretion of cytokines when cultured in an in vitro model of lung injury showed a decreased secretion of pro-inflammatory mediators for the cells cultured in the 3D scaffolds. Moreover, the morphology of the harvested cells was markedly different with respect to conventionally (2D) cultured MSCs. In conclusion, the developed bioink can be used to bioprint structures aimed to improve preconditioning MSCs for therapeutic purposes.

JTD Keywords: 3d bioprinting, acute lung injury, adhesion, collagen, differentiation, dimension, elastic properties, extracellular matrix, hydrogels, in-vitro, mechanical-properties, mesenchymal stromal cells, microenvironment, potentiate, tissue engineering, 3d bioprinting, Acute lung injury, Extracellular matrix, Hydrogels, Mesenchymal stromal cells, Stem-cells, Tissue engineering

The functionalization of nanoparticles with functional moieties is a key strategy to achieve cell targeting in nanomedicine. The interplay between size and ligand number is crucial for the formulation performance and needs to be properly characterized to understand nanoparticle structure-activity relations. However, there is a lack of methods able to measure both size and ligand number at the same time and at the single particle level. Here, we address this issue by introducing a correlative light and electron microscopy (CLEM) method combining super-resolution microscopy (SRM) and transmission electron microscopy (TEM) imaging. We apply our super-resCLEM method to characterize the relationship between size and ligand number and density in PLGA-PEG nanoparticles. We highlight how heterogeneity found in size can impact ligand distribution and how a significant part of the nanoparticle population goes completely undetected in the single-technique analysis. Super-resCLEM holds great promise for the multiparametric analysis of other parameters and nanomaterials.

JTD Keywords: cellular uptake, correlative light and electron microscopy (clem), density, electron microscopy (em), functionalization, heterogeneity, nanomedicine, nanoparticles, pegylation, plga, progress, quantification, size, Correlative light and electron microscopy (clem), Electron microscopy (em), Heterogeneity, Nanomedicine, Nanoparticles, Physicochemical characterization, Super-resolution microscopy (srm)

The characterization of polymeric materials is key towards the understanding of structure–activity relations and therefore for the rational design of novel and improved materials for a myriad of applications. Many microscopy techniques are currently used, with electron microscopy, fluorescence microscopy, and atomic force microscopy being the most relevant. In this perspective paper, we discuss the use of correlative imaging, that is, the combination of multiple imaging methodologies on the same sample, in the field of polymeric materials. This innovative approach is emerging as a powerful tool to unveil the structure and functional properties of biological and synthetic structures. Here we discuss the possibilities of correlative imaging and highlight their potential to answer open questions in polymer science.

JTD Keywords: correlative imaging, electron microscopy, material characterization, resolution microscopy, super‐, Atomic force microscopy, Correlative imaging, Electron microscopy, Material characterization, Super-resolution microscopy

Polymer nanocomposite materials based on metallic nanowires are widely investigated as transparent and flexible electrodes or as stretchable conductors and dielectrics for biosensing. Here we show that Scanning Dielectric Microscopy (SDM) can map the depth distribution of metallic nanowires within the nanocomposites in a non-destructive way. This is achieved by a quantitative analysis of sub-surface electrostatic force microscopy measurements with finite-element numerical calculations. As an application we determined the three-dimensional spatial distribution of ?50 nm diameter silver nanowires in ?100 nm-250 nm thick gelatin films. The characterization is done both under dry ambient conditions, where gelatin shows a relatively low dielectric constant, ?r ? 5, and under humid ambient conditions, where its dielectric constant increases up to ?r ? 14. The present results show that SDM can be a valuable non-destructive subsurface characterization technique for nanowire-based nanocomposite materials, which can contribute to the optimization of these materials for applications in fields such as wearable electronics, solar cell technologies or printable electronics. © The Royal Society of Chemistry.

JTD Keywords: composite, constant, electrodes, mode, nanostructures, objects, progress, subsurface, tomography, Composite materials, Dielectric materials, Electric force microscopy, Electrostatic force, Force microscopy, Low dielectric constants, Nanocomposites, Numerical calculation, Polymer nanocomposite, Printable electronics, Scanning dielectric microscopy, Silver nanowires, Solar cell technology, Stretchable conductors, Subsurface characterizations, Transparent electrodes, Wearable technology

The human olfactory system remains one of the most challenging biological systems to replicate. Humans use it without thinking, where it can equally offer protection from harm and bring enjoyment in equal measure. It is the system’s ability to detect and analyze complex odors, without the need for specialized infra-structure, that is the envy of many scientists. The field of artificial olfaction has recruited and stimulated interdisciplinary research and commercial development for several applications that include malodor measurement, medical diagnostics, food and beverage quality, environment and security. Over the last century, innovative engineers and scientists have been focused on solving a range of problems associated with measurement and control of odor. The IEEE Sensors Journal has published Special Issues on olfaction in 2002 and 2012. Here we continue that coverage. In this article, we summarize early work in the 20th Century that served as the foundation upon which we have been building our odor-monitoring instrumental and measurement systems. We then examine the current state of the art that has been achieved over the last two decades as we have transitioned into the 21st Century. Much has been accomplished, but great progress is needed in sensor technology, system design, product manufacture and performance standards. In the final section, we predict levels of performance and ubiquitous applications that will be realized during in the mid to late 21st Century.

JTD Keywords: air-quality, breath analysis, calibration transfer, chemical sensor arrays, chemosensor arrays, drift compensation, electronic nose, gas sensors, headspace sampling, machine learning, machine olfaction, odor detection, plume structure, voc analysis, Artificial olfaction, Electrodes, Electronic nose, Electronic nose technology, Headspace sampling, Instruments, Machine learning, Machine olfaction, Monitoring, Odor detection, Olfactory, Sensor phenomena and characterization, Sensors, Temperature sensors, Voc analysis

Background & Aims: Prognostic models of cirrhosis underestimate disease severity for patients with cirrhosis and ascites. Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix protein linked to hepatic neoangiogenesis and fibrogenesis. We investigated ascites MFAP4 as a predictor of transplant-free survival in patients with cirrhosis and ascites. Methods: A dual-centre observational study of patients with cirrhosis and ascites recruited consecutively in relation to a paracentesis was carried out. Patients were followed up for 1 year, until death or liver transplantation (LTx). Ascites MFAP4 was tested with the model for end-stage liver disease (MELD-Na), CLIF Consortium Acute Decompensation (CLIF-C AD), and Child-Pugh score in Cox regression models. Results: Ninety-three patients requiring paracentesis were included. Median ascites MFAP4 was 29.7 U/L [22.3–41.3], and MELD-Na was 19 [16–23]. A low MELD-Na score (<20) was observed in 49 patients (53%). During follow-up, 20 patients died (22%), and 6 received LTx (6%). High ascites MFAP4 (>29.7 U/L) was associated with 1-year transplant-free survival (p = 0.002). In Cox regression, ascites MFAP4 and MELD-Na independently predicted 1-year transplant-free survival (hazard ratio [HR] = 0.97, p = 0.03, and HR = 1.08, p = 0.01, respectively). Ascites MFAP4 and CLIF-C AD also predicted survival independently (HR = 0.96, p = 0.02, and HR = 1.05, p = 0.03, respectively), whereas only ascites MFAP4 did, controlling for the Child-Pugh score (HR = 0.97, p = 0.03, and HR = 1.18, p = 0.16, respectively). For patients with MELD-Na <20, ascites MFAP4 but not ascites protein predicted 1-year transplant-free survival (HR 0.91, p = 0.02, and HR = 0.94, p = 0.17, respectively). Conclusions: Ascites MFAP4 predicts 1-year transplant-free survival in patients with cirrhosis and ascites. In patients with low MELD-Na scores, ascites MFAP4, but not total ascites protein, significantly predicted 1-year transplant-free survival. Lay summary: Patients with cirrhosis who have fluid in the abdomen, ascites, are at an increased risk of death and in need for liver transplantation. Our study identified patients with ascites and a poor prognosis by measuring microfibrillar associated protein 4 (MFAP4), a protein present in the abdominal fluid. Patients with low levels of the MFAP4 protein are at particularly increased risk of death or liver transplantation, suggesting that clinical care should be intensified in this group of patients. © 2021 The Authors

JTD Keywords: biomarker, clif-c ad, clif consortium acute decompensation, cps, child-pugh score, crp, c-reactive protein, ct, computed tomography, decompensated, ecm, extracellular matrix, fibrosis, fluid protein, gfr, glomerular filtration rate, hr, hazard ratio, inr, internationalised normal ratio, liver disease, liver-cirrhosis, ltx, liver transplantation, markers, meld-na, model for end-stage liver disease, mfap4, microfibrillar associated protein 4, mortality, nash, non-alcoholic steatohepatitis, natural-history, prognosis, risk-factors, sbp, spontaneous bacterial peritonitis, scores, stage, Biomarker, Decompensated, Egfr, estimated gfr, Fibrosis, Liver disease, Mortality, Prognosis, Spontaneous bacterial peritonitis

Significance: The incidence of chronic wounds is increasing due to our aging population and the augment of people afflicted with diabetes. With the extended knowledge on the biological mechanisms underlying these diseases, there is a novel influx of medical technologies into the conventional wound care market. Recent Advances: Several nanotechnologies have been developed demonstrating unique characteristics that address specific problems related to wound repair mechanisms. In this review, we focus on the most recently developed nanotechnology-based therapeutic agents and evaluate the efficacy of each treatment in in vivo diabetic models of chronic wound healing. Critical Issues: Despite the development of potential biomaterials and nanotechnology-based applications for wound healing, this scientific knowledge is not translated into an increase of commercially available wound healing products containing nanomaterials. Future Directions: Further studies are critical to provide insights into how scientific evidences from nanotechnology-based therapies can be applied in the clinical setting.

JTD Keywords: chronic, diabetes, liposomes, nanofibers, nanoparticles, Chronic, Chronic wound, Diabetes, Diabetic wound, Diabetic-rats, Dressings, Drug mechanism, Extracellular-matrix, Growth-factor, Human, In-vitro, Liposome, Liposomes, Mesenchymal stem-cells, Metal nanoparticle, Nanofiber, Nanofibers, Nanofibrous scaffolds, Nanoparticles, Nanotechnology, Nonhuman, Polyester, Polymer, Polysaccharide, Priority journal, Protein, Review, Self assembled protein nanoparticle, Silk fibroin, Skin wounds, Wound healing, Wound healing promoting agent

© 2020 The Authors. Bioengineering & Translational Medicine published by Wiley Periodicals LLC on behalf of The American Institute of Chemical Engineers. Biodistribution studies are essential in drug carrier design and translation, and radiotracing provides a sensitive quantitation for this purpose. Yet, for biodegradable formulations, small amounts of free-label signal may arise prior to or immediately after injection in animal models, causing potentially confounding biodistribution results. In this study, we refined a method to overcome this obstacle. First, we verified free signal generation in animal samples and then, mimicking it in a controllable setting, we injected mice intravenously with a radiolabeled drug carrier formulation (125I-antibody/3DNA) containing a known amount of free radiolabel (125I), or free 125I alone as a control. Corrected biodistribution data were obtained by separating the free radiolabel from blood and organs postmortem, using trichloroacetic acid precipitation, and subtracting the confounding signal from each tissue measurement. Control free 125I-radiolabel was detected at ≥85% accuracy in blood and tissues, validating the method. It biodistributed very heterogeneously among organs (0.6–39 %ID/g), indicating that any free 125I generated in the body or present in an injected formulation cannot be simply corrected to the free-label fraction in the original preparation, but the free label must be empirically measured in each organ. Application of this method to the biodistribution of 125I-antibody/3DNA, including formulations directed to endothelial target ICAM-1, showed accurate classification of free 125I species in blood and tissues. In addition, this technique rendered data on the in vivo degradation of the traced agents over time. Thus, this is a valuable technique to obtain accurate measurements of biodistribution using 125I and possibly other radiotracers.

JTD Keywords: biodistribution data correction, degradation, drug delivery carriers, free label, in vivo biodistribution, radiotracing, trichloroacetic acid precipitation, Biodistribution data correction, Degradation, Drug delivery carriers, Free label, In vivo biodistribution, Radiotracing, Trichloroacetic acid precipitation

Bioinspired hybrid soft robots that combine living and synthetic components are an emerging field in the development of advanced actuators and other robotic platforms (i.e., swimmers, crawlers, and walkers). The integration of biological components offers unique characteristics that artificial materials cannot precisely replicate, such as adaptability and response to external stimuli. Here, we present a skeletal muscle–based swimming biobot with a three-dimensional (3D)–printed serpentine spring skeleton that provides mechanical integrity and self-stimulation during the cell maturation process. The restoring force inherent to the spring system allows a dynamic skeleton compliance upon spontaneous muscle contraction, leading to a cyclic mechanical stimulation process that improves the muscle force output without external stimuli. Optimization of the 3D-printed skeletons is carried out by studying the geometrical stiffnesses of different designs via finite element analysis. Upon electrical actuation of the muscle tissue, two types of motion mechanisms are experimentally observed: directional swimming when the biobot is at the liquid-air interface and coasting motion when it is near the bottom surface. The integrated compliant skeleton provides both the mechanical self-stimulation and the required asymmetry for directional motion, displaying its maximum velocity at 5 hertz (800 micrometers per second, 3 body lengths per second). This skeletal muscle–based biohybrid swimmer attains speeds comparable with those of cardiac-based biohybrid robots and outperforms other muscle-based swimmers. The integration of serpentine-like structures in hybrid robotic systems allows self-stimulation processes that could lead to higher force outputs in current and future biomimetic robotic platforms. Copyright © 2021 The Authors, some rights reserved;

JTD Keywords: actuators, design, fabrication, mechanics, mems, myotubes, platform, tissue, 3d printers, Agricultural robots, Biological components, Biomimetic processes, Electrical actuation, Geometrical stiffness, Intelligent robots, Liquefied gases, Liquid-air interface, Mechanical integrity, Mechanical stimulation, Muscle, Muscle contractions, Phase interfaces, Robotics, Serpentine, Springs (components), Threedimensional (3-d)

Asthma is a widespread disease characterized by chronic airway inflammation. It causes substantial disability, impaired quality of life, and avoidable deaths around the world. The main treatment for asthmatic patients is the administration of corticosteroids, which improves the quality of life; however, prolonged use of corticosteroids interferes with extracellular matrix elements. Therefore, cell-based therapies are emerging as a novel therapeutic contribution to tissue regeneration for lung diseases. This study aimed to summarize the advancements in cell therapy involving mesenchymal stromal cells, extracellular vesicles, and immune cells such as T-cells in asthma. Our findings provide evidence that the use of mesenchymal stem cells, their derivatives, and immune cells such as T-cells are an initial milestone to understand how emergent cell-based therapies are effective to face the challenges in the development, progression, and management of asthma, thus improving the quality of life.

JTD Keywords: asthma treatments, cell-based therapies, chronic lung diseases, extracellular vesicles, immune cells, mesenchymal stromal cells, Asthma treatments, Cell-based therapies, Chronic lung diseases, Extracellular vesicles, Immune cells, Mesenchymal stromal cells

In this work beclomethasone dipropionate was loaded into liposomes and hyalurosomes modified with mucin to improve the ability of the payload to counteract the oxidative stress and involved damages caused by cigarette smoke in the airway. The vesicles were prepared by dispersing all components in the appropriate vehicle and sonicating them, thus avoiding the use of organic solvents. Unilamellar and bilamellar vesicles small in size (~117 nm), homogeneously dispersed (polydispersity index lower than 0.22) and negatively charged (~−11 mV), were obtained. Moreover, these vesicle dispersions were stable for five months at room temperature (~25 C). In vitro studies performed using the Next Generation Impactor confirmed the suitability of the formulations to be nebulized as they were capable of reaching the last stages of the impactor that mimic the deeper airways, thus improving the deposition of beclomethasone in the target site. Further, biocompatibility studies performed by using 16HBE bronchial epithelial cells confirmed the high biocompatibility and safety of all the vesicles. Among the tested formulations, only mucin-hyalurosomes were capable of effectively counteracting the production of reactive oxygen species (ROS) induced by cigarette smoke extract, suggesting that this formulation may represent a promising tool to reduce the damaging effects of cigarette smoke in the lung tissues, thus reducing the pathogenesis of cigarette smoke-associated diseases such as chronic obstructive pulmonary disease, emphysema, and cancer. ◦

JTD Keywords: 16hbe cells, beclomethasone, cigarette smoke extract, mucin, oxidative stress, phospholipid vesicles, pulmonary delivery, 16hbe cells, Beclomethasone, Cigarette smoke extract, Mucin, Oxidative stress, Phospholipid vesicles, Pulmonary delivery

The puropse of this study was to evaluate associations of cisterna chyli (CCh) diameter with portal hemodynamics and the influence of TIPS-creation in cirrhotic patients. 93 cirrhotic patients (57 male, mean age 59 years) received CT prior to TIPS-creation. 38/93 additionally underwent post-interventional CT. CCh-diameter was measured. After categorization into patients with and without large venous collaterals (i.e. > 6 mm), data were analyzed regarding associations between CCh-diameter, clinical and portal-hemodynamic parameters and diameter-changes after TIPS-creation. Patient survival post-TIPS was analyzed. Median portosystemic pressure-gradient decreased from 20 to 9 mmHg after TIPS-creation. Large venous collaterals were observed in 59 patients. In 69/93 patients (74.2%) the CCh was detectable. Mean pre-interventional diameter was 9.4 ± 2.7 mm (large collaterals: 8.7 ± 2.0 mm, no large collaterals: 10.7 ± 3.2 mm, p = 0.003). CCh-diameter correlated strongly with pre-TIPS portal-pressure (Rs = 0.685, p = 0.0001), moderately with portosystemic-gradient (Rs = 0.524, p = 0.006), liver shear-wave-elastography (Rs = 0.597, p = 0.004) and spleen size (Rs = 0.501, p = 0.01) in patients without large collaterals, but not in patients with large collaterals. Post-TIPS CCh-diameter decreased significantly from 10.2 ± 2.8 mm to 8.3 ± 3.0 mm (p < 0.001). Patients without a detectable CCh on CT survived significantly shorter. The diameter of the CCh is associated with portal-pressure and decreases after TIPS-creation in cirrhotic patients, reflecting a portal decompression mechanism via the lymphatic system. Lack of larger central lymphatics detectable on CT may be associated with shorter survival.

JTD Keywords: circulation, cisterna chyli, fluid, hepatic cirrhosis, shear-wave elastography, thoracic-duct, vessels, Significant portal-hypertension

In the last year the COVID19 pandemic clearly illustrated the potential threat that viruses pose to our society. The characterization of viral structures and the identification of key proteins involved in each step of the cycle of infection are crucial to develop treatments. However, the small size of viruses, invisible under conventional fluorescence microscopy, make it difficult to study the organization of protein clusters within the viral particle. The applications of super-resolution microscopy have skyrocketed in the last years, converting this group into one of the leading techniques to characterize viruses and study the viral infection in cells, breaking the diffraction limit by achieving resolutions up to 10 nm using conventional probes such as fluorescent dyes and proteins. There are several super-resolution methods available and the selection of the right one it is crucial to study in detail all the steps involved in the viral infection, quantifying and creating models of infection for relevant viruses such as HIV-1, Influenza, herpesvirus or SARS-CoV-1. Here we review the use of super-resolution microscopy (SRM) to study all steps involved in the viral infection and antiviral design. In light of the threat of new viruses, these studies could inspire future assays to unveil the viral mechanism of emerging viruses and further develop successful antivirals against them.

JTD Keywords: antivirals, characterization, imaging, super-resolution, virus, Antivirals, Characterization, Imaging, Super-resolution, Virus

© 2021 National Academy of Sciences. All rights reserved. Electrophysiological studies in rodents show that active navigation enhances hippocampal theta oscillations (4–12 Hz), providing a temporal framework for stimulus-related neural codes. Here we show that active learning promotes a similar phase coding regime in humans, although in a lower frequency range (3–8 Hz). We analyzed intracranial electroencephalography (iEEG) from epilepsy patients who studied images under either volitional or passive learning conditions. Active learning increased memory performance and hippocampal theta oscillations and promoted a more accurate reactivation of stimulus-specific information during memory retrieval. Representational signals were clustered to opposite phases of the theta cycle during encoding and retrieval. Critically, during active but not passive learning, the temporal structure of intracycle reactivations in theta reflected the semantic similarity of stimuli, segregating conceptually similar items into more distant theta phases. Taken together, these results demonstrate a multilayered mechanism by which active learning improves memory via a phylogenetically old phase coding scheme.

JTD Keywords: active learning, dynamics, gamma-power, hippocampus, intracranial eeg, movement, navigation, neural phase coding, oscillations, representations, retrieval, rhythm, theta oscillations, toolbox, Active learning, Theta oscillations, Working-memory

In recent years considerable progress has been made in the development of faithful procedures for the differentiation of human pluripotent stem cells (hPSCs). An important step in this direction has also been the derivation of organoids. This technology generally relies on traditional three-dimensional culture techniques that exploit cell-autonomous self-organization responses of hPSCs with minimal control over the external inputs supplied to the system. The convergence of stem cell biology and bioengineering offers the possibility to provide these stimuli in a controlled fashion, resulting in the development of naturally inspired approaches to overcome major limitations of this nascent technology. Based on the current developments, we emphasize the achievements and ongoing challenges of bringing together hPSC organoid differentiation, bioengineering and ethics. This Review underlines the need for providing engineering solutions to gain control of self-organization and functionality of hPSC-derived organoids. We expect that this knowledge will guide the community to generate higher-grade hPSC-derived organoids for further applications in developmental biology, drug screening, disease modelling and personalized medicine. This Review provides an overview of bioengineering technologies that can be harnessed to facilitate the culture, self-organization and functionality of human pluripotent stem cell-derived organoids.

JTD Keywords: Differentiation, Embryonic-tissues, Extracellular-matrix, In-vitro, Kidney organoids, Model, Neural-tube, Pluripotent stem-cells, Reconstitution, Self-organization

© 2020 The Authors Although the concept of selective delivery has been postulated over 100 years ago, no targeted nanomedicine has been clinically approved so far. Nanoparticles modified with targeting ligands to promote the selective delivery of therapeutics towards a specific cell population have been extensively reported. However, the rational design of selective particles is still challenging. One of the main reasons for this is the lack of quantitative theoretical and experimental understanding of the interactions involved in cell targeting. In this review, we discuss new theoretical models and experimental methods that provide a quantitative view of targeting. We show the new advancements in multivalency theory enabling the rational design of super-selective nanoparticles. Furthermore, we present the innovative approaches to obtain key targeting parameters at the single-cell and single molecule level and their role in the design of targeting nanoparticles. We believe that the combination of new theoretical multivalent design and experimental methods to quantify receptors and ligands aids in the rational design and clinical translation of targeted nanomedicines.

JTD Keywords: binding-kinetics, biological identity, biomolecular corona, blood-brain-barrier, drug-delivery, gold nanoparticles, multivalency, nanotechnology, protein corona, quantitative characterization, rational design, super-selectivity, superresolution microscopy, tumor heterogeneity, Ligand-receptor interactions, Multivalency, Nanotechnology, Quantitative characterization, Rational design, Super-selectivity

© 2021 by the authors. Licensee MDPI, Basel, Switzerland. Achieving intracellular delivery of protein therapeutics within cells remains a significant challenge. Although custom formulations are available for some protein therapeutics, the development of non‐toxic delivery systems that can incorporate a variety of active protein cargo and maintain their stability, is a topic of great relevance. This study utilized ionic polyphosphazenes (PZ) that can assemble into supramolecular complexes through non‐covalent interactions with different types of protein cargo. We tested a PEGylated graft copolymer (PZ‐PEG) and a pyrrolidone containing linear derivative (PZ‐PYR) for their ability to intracellularly deliver FITC‐avidin, a model protein. In endothelial cells, PZ‐PYR/protein exhibited both faster internalization and higher uptake levels than PZ‐PEG/protein, while in cancer cells both polymers achieved similar uptake levels over time, although the internalization rate was slower for PZ‐PYR/protein. Uptake was mediated by endocytosis through multiple mechanisms, PZ‐PEG/avidin colocalized more profusely with endo-lysosomes, and PZ‐PYR/avidin achieved greater cytosolic delivery. Consequently, a PZ‐PYR-delivered anti‐F‐actin antibody was able to bind to cytosolic actin filaments without needing cell permeabilization. Similarly, a cell‐impermeable Bax‐BH3 peptide known to induce apoptosis, decreased cell viability when complexed with PZ‐PYR, demonstrating endo‐lysosomal escape. These biodegradable PZs were non‐toxic to cells and represent a promising platform for drug delivery of protein therapeutics.

JTD Keywords: cytosolic delivery, cytotoxicity, delivery of apoptotic peptides, endosomal escape, intracellular delivery of antibody, intracellular protein delivery, Cytosolic delivery, Cytotoxicity, Delivery of apoptotic peptides, Endosomal escape, Intracellular delivery of antibody, Intracellular protein delivery, Polyphosphazene polymers

Osteosarcoma is the most common primary bone tumor, and its first line of treatment presents a high failure rate. The 5-year survival for children and teenagers with osteosarcoma is 70% (if diagnosed before it has metastasized) or 20% (if spread at the time of diagnosis), stressing the need for novel therapies. Recently, cold atmospheric plasmas (ionized gases consisting of UV-Vis radiation, electromagnetic fields and a great variety of reactive species) and plasma-treated liquids have been shown to have the potential to selectively eliminate cancer cells in different tumors through an oxidative stress-dependent mechanism. In this work, we review the current state of the art in cold plasma therapy for osteosarcoma. Specifically, we emphasize the mechanisms unveiled thus far regarding the action of plasmas on osteosarcoma. Finally, we review current and potential future approaches, emphasizing the most critical challenges for the development of osteosarcoma therapies based on this emerging technique.

JTD Keywords: cancer stem cells, cold atmospheric plasma, osteosarcoma, oxidative stress, plasma treated liquids, reactive oxygen and nitrogen species, Antineoplastic activity, Antineoplastic agent, Cancer chemotherapy, Cancer stem cell, Cancer stem cells, Cancer surgery, Cancer survival, Cell therapy, Cold atmospheric plasma, Cold atmospheric plasma therapy, Electromagnetism, Human, In vitro study, Intracellular signaling, Oncogene, Osteosarcoma, Oxidative stress, Plasma treated liquids, Reactive nitrogen species, Reactive oxygen and nitrogen species, Reactive oxygen metabolite, Review, Tumor microenvironment

© 2020 Elsevier Inc. Severe asthma is associated with an increased airway smooth muscle (ASM) mass and altered composition of the extracellular matrix (ECM). Studies have indicated that ECM-ASM cell interactions contribute to this remodeling and its limited reversibility with current therapy. Three-dimensional matrices allow the study of complex cellular responses to different stimuli in an almost natural environment. Our goal was to obtain acellular bronchial matrices and then develop a recellularization protocol with ASM cells. We studied equine bronchi as horses spontaneously develop a human asthma-like disease. The bronchi were decellularized using Triton/Sodium Deoxycholate. The obtained scaffolds retained their anatomical and histological properties. Using immunohistochemistry and a semi-quantitative score to compare native bronchi to scaffolds revealed no significant variation for matrixial proteins. DNA quantification and electrophoresis revealed that most DNA was 29.6 ng/mg of tissue ± 5.6, with remaining fragments of less than 100 bp. Primary ASM cells were seeded on the scaffolds. Histological analysis of the recellularizations showed that ASM cells migrated and proliferated primarily in the decellularized smooth muscle matrix, suggesting a chemotactic effect of the scaffolds. This is the first report of primary ASM cells preferentially repopulating the smooth muscle matrix layer in bronchial matrices. This protocol is now being used to study the molecular interactions occurring between the asthmatic ECMs and ASM to identify effectors of asthmatic bronchial remodeling.

JTD Keywords: 2d, airway smooth muscle cells, asthma, decellularization, disease, elastin, extracellular matrix, lung scaffolds, migration, peptide, recellularization, tissues, Airway smooth muscle cells, Asthma, Culture-systems, Decellularization, Extracellular matrix, Recellularization

Marine ancestors of freshwater sponges had to undergo a series of physiological adaptations to colonize harsh and heterogeneous limnic environments. Besides reduced salinity, river-lake systems also have calcium concentrations far lower than seawater. Cell adhesion in sponges is mediated by calcium-dependent multivalent self-interactions of sulfated polysaccharide components of membrane-bound proteoglycans named aggregation factors. Cells of marine sponges require seawater average calcium concentration (10 mM) to sustain adhesion promoted by aggregation factors. We demonstrate here that the freshwater sponge Spongilla alba can thrive in a calcium-poor aquatic environment and that their cells are able to aggregate and form primmorphs with calcium concentrations 40-fold lower than that required by marine sponges cells. We also find that their gemmules need calcium and other micronutrients to hatch and generate new sponges. The sulfated polysaccharide purified from S. alba has sulfate content and molecular size notably lower than those from marine sponges. Nuclear magnetic resonance analyses indicated that it is composed of a central backbone of non- and 2-sulfated α- and β-glucose units decorated with branches of α-glucose. Assessments with atomic force microscopy/single-molecule force spectroscopy show that S. alba glucan requires 10-fold less calcium than sulfated polysaccharides from marine sponges to self-interact efficiently. Such an ability to retain multicellular morphology with low environmental calcium must have been a crucial evolutionary step for freshwater sponges to successfully colonize inland waters.

JTD Keywords: Carbohydrate interactions, Evolutionary adaptation, Porifera, Proteoglycans, Sulfated polysaccharides

Alcoholism is one of the leading and increasingly prevalent reasons of liver associated morbidity and mortality worldwide. Alcoholic hepatitis (AH) constitutes a severe disease with currently no satisfying treatment options. Lipoxin A4 (LXA4), a 15-lipoxygenase (ALOX15)-dependent lipid mediator involved in resolution of inflammation, showed promising pre-clinical results in the therapy of several inflammatory diseases. Since inflammation is a main driver of disease progression in alcoholic hepatitis, we investigated the impact of endogenous ALOX15-dependent lipid mediators and exogenously applied LXA4 on AH development. A mouse model for alcoholic steatohepatitis (NIAAA model) was tested in Alox12/15+/+ and Alox12/15−/− mice, with or without supplementation of LXA4. Absence of Alox12/15 aggravated parameters of liver disease, increased hepatic immune cell infiltration in AH, and elevated systemic neutrophils as a marker for systemic inflammation. Interestingly, i.p. injections of LXA4 significantly lowered transaminase levels only in Alox12/15−/− mice and reduced hepatic immune cell infiltration as well as systemic inflammatory cytokine expression in both genotypes, even though steatosis progressed. Thus, while LXA4 injection attenuated selected parameters of disease progression in Alox12/15−/− mice, its beneficial impact on immunity was also apparent in Alox12/15+/+ mice. In conclusion, pro-resolving lipid mediators may be beneficial to reduce inflammation in alcoholic hepatitis.

JTD Keywords: Alcoholic hepatitis, Arachidonate 12/15-lipoxygenase (Alox12/15), Lipoxin A4, Resolution of inflammation, Specialized pro-resolving lipid mediators (SPMs)

Understanding cell/material interactions is essential to design functional cell-responsive materials. While the scientific literature abounds with formulations of biomimetic materials, only a fraction of them focused on mechanisms of the molecular interactions between cells and material. To provide new knowledge on the strategies for materials/cell recognition and binding, supramolecular benzene-1,3,5-tricarboxamide copolymers bearing benzoxaborole moieties are anchored on the surface of human erythrocytes via benzoxaborole/sialic-acid binding. This interaction based on both dynamic covalent and non-covalent chemistries is visualized in real time by means of total internal reflection fluorescence microscopy. Exploiting this imaging method, we observe that the functional copolymers specifically interact with the cell surface. An optimal fiber affinity towards the cells as a function of benzoxaborole concentration demonstrates the crucial role of multivalency in these cell/material interactions.

JTD Keywords: Boronic acid, Cell/material interactions, Multivalency, Red blood cells, Supramolecular polymers

Tumor cell-surface markers are usually overexpressed or mutated protein receptors for which spatiotemporal regulation differs between and within cancers. Single-molecule fluorescence imaging can profile individual markers in different cellular contexts with molecular precision. However, standard single-molecule imaging methods based on overexpressed genetically encoded tags or cumbersome probes can significantly alter the native state of receptors. We introduce a live-cell points accumulation for imaging in nanoscale topography (PAINT) method that exploits aptamers as minimally invasive affinity probes. Localization and tracking of individual receptors are based on stochastic and transient binding between aptamers and their targets. We demonstrated single-molecule imaging of a model tumor marker (EGFR) on a panel of living cancer cells. Affinity to EGFR was finely tuned by rational engineering of aptamer sequences to define receptor motion and/or native receptor density.

JTD Keywords: Aptamers, Cell-surface receptors, Live-cell imaging, PAINT, Single-molecule tracking

Conventional drug solubilization strategies limit the understanding of the full potential of poorly water-soluble drugs during drug screening. Here, we propose a screening approach in which poorly water-soluble drugs are entrapped in poly(2-(methacryloyloxyethyl phosphorylcholine)-poly(2-(diisopropylaminoethyl methacryate) (PMPC-PDPA) polymersomes (POs) to enhance drug solubility and facilitate intracellular delivery. By using a human pediatric glioma cell model, we demonstrated that PMPC-PDPA POs mediated intracellular delivery of cytotoxic and epigenetic drugs by receptor-mediated endocytosis. Additionally, when delivered in combination, drug-loaded PMPC-PDPA POs triggered both an enhanced drug efficacy and synergy compared to that of a conventional combinatorial screening. Hence, our comprehensive synergy analysis illustrates that our screening methodology, in which PMPC-PDPA POs are used for intracellular codelivery of drugs, allows us to identify potent synergistic profiles of anticancer drugs.

JTD Keywords: Combination therapy, Drug screening, Drug solubilization, Intracellular drug delivery, Polymeric nanoparticles, Synergy analysis

Intracellular invasion is an advantageous mechanism used by pathogens to evade host defense and antimicrobial therapy. In patients, the intracellular microbial lifestyle can lead to infection persistence and recurrence, thus worsening outcomes. Lung infections caused by Pseudomonas aeruginosa, especially in cystic fibrosis (CF) patients, are often aggravated by intracellular invasion and persistence of the pathogen. Proliferation of the infectious species relies on a continuous deoxyribonucleotide (dNTP) supply, for which the ribonucleotide reductase enzyme (RNR) is the unique provider. The large genome plasticity of P. aeruginosa and its ability to rapidly adapt to different environments are challenges for studying the pathophysiology associated with this type of infection. Using different reference strains and clinical isolates of P. aeruginosa independently combined with alveolar (A549) and bronchial (16HBE14o- and CF-CFBE41o-) epithelial cells, we analyzed host–pathogen interactions and intracellular bacterial persistence with the aim of determining a cell type-directed infection promoted by the P. aeruginosa strains. The oscillations in cellular toxicity and oxygen consumption promoted by the intracellular persistence of the strains were also analyzed among the different infectious lung models. Significantly, we identified class II RNR as the enzyme that supplies dNTPs to intracellular P. aeruginosa. This discovery could contribute to the development of RNR-targeted strategies against the chronicity occurring in this type of lung infection. Overall our study demonstrates that the choice of bacterial strain is critical to properly study the type of infectious process with relevant translational outcomes.

JTD Keywords: Pseudomonas aeruginosa, Intracellular persistence, Lung, Epithelial cells, Clinical isolates, Host-pathogen interactions, Intracellular lifestyle, Chronic infections, Cystic fibrosis, Ribonucleotide reductase

Regenerative medicine and disease models have evolved in recent years from two to three dimensions, providing in vitro constructs that are more similar to in vivo tissues. By mimicking native tissues, cell-derived matrices (CDMs) have emerged as new modifiable extracellular matrices for a variety of tissues, allowing researchers to study basic cellular processes in tissue-like structures, test tissue regeneration approaches, and model disease development. In this review, different fabrication techniques and characterization methods of CDMs are presented and examples of their application in cell behavior studies, tissue regeneration, and disease models are provided. In addition, future guidelines and perspectives in the field of CDMs are discussed.

JTD Keywords: 3D models, Biomaterials, Cell-derived matrices, Extracellular matrix, Personalized therapies

Among several factors behind drug resistance evolution in malaria is the challenge of administering overall doses that are not toxic for the patient but that, locally, are sufficiently high to rapidly kill the parasites. Thus, a crucial antimalarial strategy is the development of drug delivery systems capable of targeting antimalarial compounds to Plasmodium with high specificity. In the present study, extracellular vesicles (EVs) have been evaluated as a drug delivery system for the treatment of malaria. EVs derived from naive red blood cells (RBCs) and from Plasmodium falciparum-infected RBCs (pRBCs) were isolated by ultrafiltration followed by size exclusion chromatography. Lipidomic characterization showed that there were no significant qualitative differences between the lipidomic profiles of pRBC-derived EVs (pRBC-EVs) and RBC-derived EVs (RBC-EVs). Both EVs were taken up by RBCs and pRBCs, although pRBC-EVs were more efficiently internalized than RBC-EVs, which suggested their potential use as drug delivery vehicles for these cells. When loaded into pRBC-EVs, the antimalarial drugs atovaquone and tafenoquine inhibited in vitro P. falciparum growth more efficiently than their free drug counterparts, indicating that pRBC-EVs can potentially increase the efficacy of several small hydrophobic drugs used for the treatment of malaria.

JTD Keywords: Antimalarial drugs, Drug delivery, Extracellular vesicles, Malaria, Plasmodium falciparum

This paper describes the design of a linear phase low-pass differentiator filter with a finite impulse response (FIR) for extracting transient features of gas sensor signals (the so-called “bouts”). The detection of these bouts is relevant for estimating the distance of a gas source in a turbulent plume. Our current proposal addresses the shortcomings of previous ‘bout’ estimation methods, namely: (i) they were based in non-causal digital filters precluding real time operation, (ii) they used non-linear phase filters leading to waveform distortions and (iii) the smoothing action was achieved by two filters in cascade, precluding an easy tuning of filter performance. The presented method is based on a low-pass FIR differentiator, plus proper post-processing, allowing easy algorithmic implementation for real-time robotic exploration. Linear phase filters preserve signal waveform in the bandpass region for maximum reliability concerning both ‘bout’ detection and amplitude estimation. As a case study, we apply the proposed filter to predict the source distance from recordings obtained with metal oxide (MOX) gas sensors in a wind tunnel. We first perform a joint optimization of the cut-off frequency of the filter and the bout amplitude threshold, for different wind speeds, uncovering interesting relationships between these two parameters. We demonstrate that certain combinations of parameters can reduce the prediction error to 8 cm (in a distance range of 1.45 m) improving previously reported performances in the same dataset by a factor of 2.5. These results are benchmarked against traditional source distance estimators such as the mean, variance and maximum of the response. We also study how the length of the measurement window affects the performance of different signal features, and how to select the filter parameters to make the predictive models more robust to changes in wind speed. Finally, we provide a MATLAB implementation of the bout detection algorithm and all analysis code used in this study.

JTD Keywords: Gas sensors, Differentiator, Low pass filter, Metal oxide semiconductor, MOX sensors, Signal processing, Feature extraction, Gas source localization, Robotics

Tackling the first stages of the chondrogenic commitment is essential to drive chondrogenic differentiation to healthy hyaline cartilage and minimize hypertrophy. During chondrogenesis, the extracellular matrix continuously evolves, adapting to the tissue adhesive requirements at each stage. Here, we take advantage of previously developed nanopatterns, in which local surface adhesiveness can be precisely tuned, to investigate its effects on prechondrogenic condensation. Fluorescence live cell imaging, immunostaining, confocal microscopy and PCR analysis are used to follow the condensation process on the nanopatterns. Cell tracking parameters, condensate morphology, cell–cell interactions, mechanotransduction and chondrogenic commitment are evaluated in response to local surface adhesiveness. Results show that only condensates on the nanopatterns of high local surface adhesiveness are stable in culture and able to enter the chondrogenic pathway, thus highlighting the importance of controlling cell–substrate adhesion in the tissue engineering strategies for cartilage repair.

JTD Keywords: Dendrimer, Nanopatterning, RGD, Mesenchymal cell condensation, Cell–cell interactions, YAP, Chondrogenesis

Galleria mellonella larvae are an alternative in vivo model that has been extensively used to study the virulence and pathogenicity of different bacteria due to its practicality and lack of ethical constraints. However, the larvae possess intrinsic autofluorescence that obstructs the use of fluorescent proteins to study bacterial infections, hence better methodologies are needed. Here, we report the construction of a promoter probe vector with bioluminescence expression as well as the optimization of a total bacterial RNA extraction protocol to enhance the monitoring of in vivo infections. By employing the vector to construct different gene promoter fusions, variable gene expression levels were efficiently measured in G. mellonella larvae at various time points during the course of infection and without much manipulation of the larvae. Additionally, our optimized RNA extraction protocol facilitates the study of transcriptional gene levels during an in vivo infection. The proposed methodologies will greatly benefit bacterial infection studies as they can contribute to a better understanding of the in vivo infection processes and pathogen–mammalian host interactions.

JTD Keywords: Galleria mellonella, P. aeruginosa, Hemolymph, Hemocytes, Bioluminescence, Promoter probe vector, Optimized RNA extraction, Ribonucleotide reductases

Peripheral nerve injuries, including motor neuron axonal injury, often lead to functional impairments. Current therapies are mostly limited to surgical intervention after lesion, yet these interventions have limited success in restoring functionality. Current activity-based therapies after axonal injuries are based on trial-error approaches in which the details of the underlying cellular and molecular processes are largely unknown. Here we show the effects of the modulation of both neuronal and muscular activity with optogenetic approaches to assess the regenerative capacity of cultured motor neuron (MN) after lesion in a compartmentalized microfluidic-assisted axotomy device. With increased neuronal activity, we observed an increase in the ratio of regrowing axons after injury in our peripheral-injury model. Moreover, increasing muscular activity induces the liberation of leukemia inhibitory factor and glial cell line-derived neurotrophic factor in a paracrine fashion that in turn triggers axonal regrowth of lesioned MN in our 3D hydrogel cultures. The relevance of our findings as well as the novel approaches used in this study could be useful not only after axotomy events but also in diseases affecting MN survival.

JTD Keywords: Neuromuscular junction, Microfluidics, Axotomy, Paracrine signaling

Background: Alcoholic liver disease (ALD) is a public health concern that is the cause of half of all cirrhosis-related deaths. Early detection of fibrosis, ideally in the precirrhotic stage, is a key strategy for improving ALD outcomes and for preventing progression to cirrhosis. Previous studies identified the blood-borne marker human microfibrillar-associated protein 4 (MFAP4) as a biomarker for detection of hepatitis C virus (HCV)-related fibrosis. Aim: To evaluate the diagnostic accuracy of MFAP4 to detect ALD-induced fibrosis. Method: We performed a prospective, liver biopsy-controlled study involving 266 patients with prior or current alcohol overuse. Patients were split into a training and a validation cohort. Results: MFAP4 was present in fibrotic hepatic tissue and serum MFAP4 levels increased with fibrosis grade. The area under the receiver operating characteristic curve (AUROC) for detection of cirrhosis was 0.91 (95% CI 0.85-0.96) in the training cohort and 0.91 (95% CI 0.79-1.00) in the validation cohort. For detection of advanced fibrosis, the AUROC was 0.88 (95% CI 0.81-0.94) in the training cohort and 0.92 (95% CI 0.83-1.00) in the validation cohort. The diagnostic accuracy did not differ between MFAP4 and the enhanced liver fibrosis (ELF) test or transient elastography (TE) in an intention-to-diagnose analysis. MFAP4 did not predict hepatic decompensation in a time-to-decompensation analysis in a subgroup of patients with cirrhosis. Conclusion: MFAP4 is a novel biomarker that can detect ALD-related fibrosis with high accuracy.

JTD Keywords: Biomarker, Cirrhosis, Extracellular matrix protein, Liver biopsy, Non-invasive testing

Background & Aims: Renal function assessed by creatinine is a key prognostic factor in cirrhotic patients. However, creatinine is influenced by several factors, rendering interpretation difficult in some situations. This is especially important in early stages of renal dysfunction where renal impairment might not be accompanied by an increase in creatinine. Other parameters, such as cystatin C (CysC) and beta‐trace protein (BTP), have been evaluated to fill this gap. However, none of these studies have considered the role of the patient's sex. The present study analysed CysC and BTP to evaluate their prognostic value and differentiate them according to sex. Patients and methods: CysC and BTP were measured in 173 transjugular intrahepatic portosystemic shunt (TIPS)-patients from the NEPTUN-STUDY(NCT03628807) and analysed their relationship with mortality and sex. Propensity score for age, MELD, etiology and TIPS indication was used. Results: Cystatin C and BTP showed excellent correlations with creatinine values at baseline and follow-up. CysC was an independent predictor of overall mortality (HR = 1.66(1.33-2.06)) with an AUC of 0.75 and identified a cut-off of 1.55 mg/L in the whole cohort. Interestingly, CysC was significantly lower in females, also after propensity score matching. In males, the only independent predictor was the creatinine level (HR = 1.54(1.25-1.58)), while in females CysC levels independently predicted mortality (HR = 3.17(1.34-7.52)). Conclusion: This study demonstrates for the first time that in TIPS-patients creatinine predicts mortality in males better than in females, whereas CysC is a better predictor of mortality in females. These results may influence future clinical decisions on therapeutic options for example, allocation for liver transplantation in TIPS-patients.

JTD Keywords: Beta-trace protein, Cirrhosis, Cystatin C, Portal hypertension, Renal function

In the past decade, society has experienced notable growth in a variety of technological areas. However, the Fourth Industrial Revolution has not been embraced yet. Industry 4.0 imposes several challenges which include the necessity of new architectural models to tackle the uncertainty that open environments represent to cyber-physical systems (CPS). Waste Electrical and Electronic Equipment (WEEE) recycling plants stand for one of such open environments. Here, CPSs must work harmoniously in a changing environment, interacting with similar and not so similar CPSs, and adaptively collaborating with human workers. In this paper, we support the Distributed Adaptive Control (DAC) theory as a suitable Cognitive Architecture for managing a recycling plant. Specifically, a recursive implementation of DAC (between both single-agent and large-scale levels) is proposed to meet the expected demands of the European Project HR-Recycler. Additionally, with the aim of having a realistic benchmark for future implementations of the recursive DAC, a micro-recycling plant prototype is presented.

JTD Keywords: Cognitive architecture, Distributed Adaptive Control, Recycling plant, Navigation, Motor control, Human-Robot Interaction

We present a new computational framework for modeling moral decision-making in artificial agents based on the notion of ‘Machine Morality as Cooperation’. This framework integrates recent advances from cross-disciplinary moral decision-making literature into a single architecture. We build upon previous work outlining cognitive elements that an artificial agent would need for exhibiting latent morality, and we extend it by providing a computational realization of the cognitive architecture of such an agent. Our work has implications for cognitive and social robotics. Recent studies in human neuroimaging have pointed to three different decision-making processes, Pavlovian, model-free and model-based, that are defined by distinct neural substrates in the brain. Here, we describe how computational models of these three cognitive processes can be implemented in a single cognitive architecture by using the distributed and hierarchical organization proposed by the DAC theoretical framework. Moreover, we propose that a pro-social drive to cooperate exists at the Pavlovian level that can also bias the rest of the decision system, thus extending current state-of-the-art descriptive models based on harm-aversion.

JTD Keywords: Morality, Moral decision-making, Computational models, Cognitive architectures, Cognitive robotics, Human-robot interaction

This pilot study investigates the role of social sensorimotor contingencies as exhibited from a humanoid robot to allow mutual understanding and social entrainment in a group social activity. The goal is to evaluate whether sensorimotor contingencies can lead to transparent and understandable interactions while we explore the dimension of personality. We propose the task of taking a selfie with a robot and a group of humans as the benchmark to evaluate the social sensorimotor contingencies displayed. We have constructed two models of interaction with an introverted and extroverted robot. We also seek to address the gap in research in context and personality of social sensorimotor contingencies in HRI and contribute to the field of personality in social robotics by determining what type of behaviour of the robot attracts certain personalities in humans in group settings. Although the sample size was small, and there were no significant differences between conditions, results suggest that the expression of sensorimotor contingencies can lead to successful coupling and interactions.

JTD Keywords: Human-robot interaction, Personality, Social robots, Social sensorimotor contingencies

Modern optical sensors coupled with time-lapse microscopy devices and dedicated software tools allow the miniaturization of laboratories for biological experiments leading to the Organ-On-Chip (OoC) framework. OoCs allow performing massive measurements on a large number of cells under the assumption of reproducibility conditions, permitting to investigate the cell dynamics in terms of motility and shape changes over time. In this work, we present the OoC platform used in a preliminary study of the Rare Haemolytic Anaemia (RHA) disease, a group of rare diseases characterized by haemolysis, which is the premature loss of red blood cells (RBCs). Preliminary results demonstrate the effectiveness of shape measurement for the diagnosis of RHA.

JTD Keywords: Anaemia diagnosis, Cell tracking, Plasticity measurement, Time-lapse microscopy

Tissue elasticity is a critical regulator of cell behavior in normal and diseased conditions like fibrosis and cancer. Since the extracellular matrix (ECM) is a major regulator of tissue elasticity and function, several ECM-based models have emerged in the last decades, including in vitro endogenous ECM, decellularized tissue ECM and ECM hydrogels. The development of such models has urged the need to quantify their elastic properties particularly at the nanometer scale, which is the relevant length scale for cell-ECM interactions. For this purpose, the versatility of atomic force microscopy (AFM) to quantify the nanomechanical properties of soft biomaterials like ECM models has emerged as a very suitable technique. In this chapter we provide a detailed protocol on how to assess the Young's elastic modulus of ECM models by AFM, discuss some of the critical issues, and provide troubleshooting guidelines as well as illustrative examples of AFM measurements, particularly in the context of cancer.

JTD Keywords: 3D ECM hydrogels, Atomic force microscopy, Decellularized tissue, Elastic modulus, Endogenous ECM, Extracellular matrix

The composition and architecture of the extracellular matrix (ECM) and their dynamic alterations, play an important regulatory role on numerous cellular processes. Cells embedded in 3D scaffolds show phenotypes and morphodynamics reminiscent of the native scenario. This is in contrast to flat environments, where cells display artificial phenotypes. The structural and biomolecular properties of the ECM are critical in regulating cell behavior via mechanical, chemical and topological cues, which induce cytoskeleton rearrangement and gene expression. Indeed, distinct ECM architectures are encountered in the native stroma, which depend on tissue type and function. For instance, anisotropic geometries are associated with ECM degradation and remodeling during tumor progression, favoring tumor cell invasion. Overall, the development of innovative in vitro ECM models of the ECM that reproduce the structural and physicochemical properties of the native scenario is of upmost importance to investigate the mechanistic determinants of tumor dissemination. In this chapter, we describe an extremely versatile technique to engineer three-dimensional (3D) matrices with controlled architectures for the study of pathophysiological processes in vitro. To this aim, a confluent culture of “sacrificial” fibroblasts was seeded on top of microfabricated guiding templates to induce the 3D ECM growth with specific isotropic or anisotropic architectures. The resulting matrices, and cells seeded on them, recapitulated the structure, composition, phenotypes and morphodynamics typically found in the native scenario. Overall, this method paves the way for the development of in vitro ECMs for pathophysiological studies with potential clinical relevance.

JTD Keywords: Extracellular matrix, Cell-derived matrix, 3D model, Biomimicry, Anisotropy

This book constitutes the proceedings of the )th International Conference on Biomimetic and Biohybrid Systems, Living Machines 2020, held in Freiburg, Germany, in July 2020. Due to COVID-19 pandemic the conference was held virtually. The 32 full and 7 short papers presented in this volume were carefully reviewed and selected from 45 submissions. They deal with research on novel life-like technologies inspired by the scientific investigation of biological systems, biomimetics, and research that seeks to interface biological and artificial systems to create biohybrid systems.

JTD Keywords: Artificial intelligence, Soft robotics, Biomimetics, Insect navigation, Synthetic nervous system, Computer vision, Bio-inspired materials, Visual homing, Locomotion+, Image processing, Intelligent robots, Human-robot interaction, Machine learning, Snake robot, Mobile robots, Robotic systems, Drosophila, Robots, Sensors, Signal processing

Integrins, and integrin-mediated adhesions, have long been recognized to provide the main molecular link attaching cells to the extracellular matrix (ECM) and to serve as bidirectional hubs transmitting signals between cells and their environment. Recent evidence has shown that their combined biochemical and mechanical properties also allow integrins to sense, respond to and interact with ECM of differing properties with exquisite specificity. Here, we review this work first by providing an overview of how integrin function is regulated from both a biochemical and a mechanical perspective, affecting integrin cell-surface availability, binding properties, activation or clustering. Then, we address how this biomechanical regulation allows integrins to respond to different ECM physicochemical properties and signals, such as rigidity, composition and spatial distribution. Finally, we discuss the importance of this sensing for major cell functions by taking cell migration and cancer as examples.

JTD Keywords: Cell migration, Extracellular matrix, Integrins, Mechanotransduction, Single-molecule biophysics

Biological function requires cell-cell adhesions to tune their cohesiveness; for instance, during the opening of new fluid-filled cavities under hydraulic pressure. To understand the physical mechanisms supporting this adaptability, we develop a stochastic model for the hydraulic fracture of adhesive interfaces bridged by molecular bonds. We find that surface tension strongly enhances the stability of these interfaces by controlling flaw sensitivity, lifetime, and optimal architecture in terms of bond clustering. We also show that bond mobility embrittles adhesions and changes the mechanism of decohesion. Our study provides a mechanistic background to understand the biological regulation of cell-cell cohesion and fracture.

JTD Keywords: Biomimetic & bio-inspired materials, Cell adhesion, Fracture, Self-healing

Background & Aims: Remodelling of extracellular matrix is crucial in progressive liver fibrosis. Collagen type III desposition has been shown in acute decompensation. Extratracellular matrix is compiled of deposition of various components. The role of basement membrane collagen type IV in advanced cirrhosis and acute decompensation is unclear and investigated in this study. Methods: Patients with decompensated cirrhosis from the prospective NEPTUN cohort (ClinicalTrials.gov Identifier: NCT03628807), who underwent transjugular intrahepatic portosystemic shunt procedure were included. Clinical and laboratory parameters, PRO-C4 and C4M levels were measured in blood samples from portal and hepatic veins just before transjugular intrahepatic portosystemic shunt placement. Results: Levels of C4M and PRO-C4 are significantly lower in patients with massive ascites and impaired renal sodium excretion. C4M and PRO-C4 show gender-specific profiles with significantly lower levels in females compared to males. Females with higher C4M levels show higher mortality. By contrast, males with higher C4M levels show lower mortality. In multivariate Cox regression analysis, C4M is an independent predictor of survival in female patients. Conclusion: This study shows that markers of collagen type IV remodelling do not accumulate in severe renal dysfunction. Although collagen type IV degradation markers derive from the liver, portal venous C4M levels are relevant for survival. Moreover, it demonstrates that circulating C4M shows gender-specific profiles, which can independently predict survival in female patients with decompensated cirrhosis.

JTD Keywords: ACLF, Acute decompensation, Acute-on-chronic liver failure, Cirrhosis, Collagen type IV, Extracellular matrix remodelling, Gender, Liver, Portal hypertension, Transjugular intrahepatic portosystemic shunt

Adult zebrafish, in contrast to mammals, are able to regenerate their hearts in response to injury or experimental amputation. Our understanding of the cellular and molecular bases that underlie this process, although fragmentary, has increased significantly over the last years. However, the role of the extracellular matrix (ECM) during zebrafish heart regeneration has been comparatively rarely explored. Here, we set out to characterize the ECM protein composition in adult zebrafish hearts, and whether it changed during the regenerative response. For this purpose, we first established a decellularization protocol of adult zebrafish ventricles that significantly enriched the yield of ECM proteins. We then performed proteomic analyses of decellularized control hearts and at different times of regeneration. Our results show a dynamic change in ECM protein composition, most evident at the earliest (7 days post-amputation) time-point analyzed. Regeneration associated with sharp increases in specific ECM proteins, and with an overall decrease in collagens and cytoskeletal proteins. We finally tested by atomic force microscopy that the changes in ECM composition translated to decreased ECM stiffness. Our cumulative results identify changes in the protein composition and mechanical properties of the zebrafish heart ECM during regeneration.

JTD Keywords: Animal models, Atomic force microscopy, Cardiovascular disease, Cardiovascular function or biology, Developmental biology, Extracellular matrix, Heart regeneration, Proteomic analysis

The mechanical properties of the extracellular matrix (ECM)–a complex, 3D, fibrillar scaffold of cells in physiological environments–modulate cell behavior and can drive tissue morphogenesis, regeneration, and disease progression. For simplicity, it is often convenient to assume these properties to be time-invariant. In living systems, however, cells dynamically remodel the ECM and create time-dependent local microenvironments. Here, we show how cell-generated contractile forces produce substantial irreversible changes to the density and architecture of physiologically relevant ECMs–collagen I and fibrin–in a matter of minutes. We measure the 3D deformation profiles of the ECM surrounding cancer and endothelial cells during stages when force generation is active or inactive. We further correlate these ECM measurements to both discrete fiber simulations that incorporate fiber crosslink unbinding kinetics and continuum-scale simulations that account for viscoplastic and damage features. Our findings further confirm that plasticity, as a mechanical law to capture remodeling in these networks, is fundamentally tied to material damage via force-driven unbinding of fiber crosslinks. These results characterize in a multiscale manner the dynamic nature of the mechanical environment of physiologically mimicking cell-in-gel systems.

JTD Keywords: Collagens, Fibrin, Extracellular matrix, Cross-linking, Cell physiology, Deformation, Fluorescence imaging, Cell biology

Objectives: To investigate the clinical potential of fat-free muscle area (FFMA) to predict outcome in patients with liver-predominant metastatic colorectal cancer (mCRC) undergoing radioembolization (RE) with 90Yttrium microspheres. Methods: Patients with mCRC who underwent RE in our center were included in this retrospective study. All patients received liver magnetic resonance imaging including standard T2-weighted images. The total erector spinae muscle area and the intramuscular adipose tissue area were measured at the level of the origin of the superior mesenteric artery and subtracted to calculate FFMA. Cutoff values for definition of low FFMA were 3644 mm2 in men and 2825 mm2 in women. The main outcome was overall survival (OS). For survival analysis, the Kaplan-Meier method and Cox regressions comparing various clinic-oncological parameters which potentially may affect OS were performed. Results: Seventy-seven patients (28 female, mean age 60 ± 11 years) were analyzed. Mean time between MRI and the following RE was 17 ± 31 days. Median OS after RE was 178 days. Patients with low FFMA had significantly shortened OS compared to patients with high FFMA (median OS: 128 vs. 273 days, p = 0.017). On multivariate Cox regression analysis, OS was best predicted by FFMA (hazard ratio (HR) 2.652; p < 0.001). Baseline bilirubin (HR 1.875; p = 0.030), pattern of tumor manifestation (HR 1.679; p = 0.001), and model of endstage liver disease (MELD) score (HR 1.164; p < 0.001) were also significantly associated with OS. Conclusions: FFMA was associated with OS in patients receiving RE for treatment of mCRC and might be a new prognostic biomarker for survival prognosis.

JTD Keywords: Brachytherapy, Colorectal cancer, Magnetic resonance imaging, Sarcopenia

Sizing natural or engineered single nanoscale objects is fundamental in many areas of science and technology. To achieve it several advanced microscopic techniques have been developed, mostly based on electron and scanning probe microscopies. Still for soft and poorly adhered samples the existing techniques face important challenges. Here, we propose an alternative method to size single nanoscale objects based on the measurement of its electric polarization. The method is based on Electrostatic Force Microscopy measurements combined with a specifically designed multiparameter quantification algorithm, which gives the physical dimensions (height and width) of the nanoscale object. The proposed method is validated with ~50 nm diameter silver nanowires, and successfully applied to ~10 nm diameter bacterial polar flagella, an example of soft and poorly adhered nanoscale object. We show that an accuracy comparable to AFM topographic imaging can be achieved. The main advantage of the proposed method is that, being based on the measurement of long-range polarization forces, it can be applied without contacting the sample, what is key when considering poorly adhered and soft nanoscale objects. Potential applications of the proposed method to a wide range of nanoscale objects relevant in Material, Life Sciences and Nanomedicine is envisaged.

JTD Keywords: Characterization and analytical techniques, Imaging techniques

Metallic implants have some limitations related to bioactivity and bacteria colonization leading to infections. In this regard, calcium phosphate coatings can be used as carrier for drug delivery in order to improve the mentioned drawbacks. The present work proposes the introduction of an antibacterial agent in the course of a pulsed and reverse pulsed electrodeposition. Calcium phosphate coatings were prepared in 30 min using different pulse waveforms (unipolar-bipolar), current densities (2–5 mA/cm2) and temperatures (40–60 °C). Mechanical stability of the as-coated surfaces was studied in order to select the optimal electrodeposition conditions. Subsequently, selected coatings were loaded with an antiseptic agent, chlorhexidine digluconate (CHX), via a single-step co-deposition procedure. CHX concentration added to the electrolyte was adjusted to 3 mM based on the antibacterial efficacy of the loaded coatings evaluated in vitro with Staphylococcus aureus and Escherichia coli bacteria strains. Whereas the same chlorhexidine concentration was added to the electrolyte, results showed that the amount of CHX loaded was different for each condition while release kinetics was maintained. The results of this work demonstrate that a pulsed co-deposition strategy has great potential to modulate local delivery of antibacterial agents such as chlorhexidine digluconate, which may prevent early phase infections of metallic implants after insertion.

JTD Keywords: Antibacterial agent, Calcium phosphate, Characterization, Coating, Pulse electrodeposition, Titanium

Concentrations down to 3 nM of the rhS100A4 protein, associated with human tumor development, have been detected in undiluted urine using an integrated sensor based on microring resonators in the emerging Al2O3 photonic platform. The fabricated microrings were designed for operation in the C-band (λ = 1565 nm) and exhibited a high-quality factor in air of 3.2 × 105. The bulk refractive index sensitivity of the devices was ~100 nm/RIU (for TM polarization) with a limit of detection of ~10−6 RIU. A surface functionalization protocol was developed to allow for the selective binding of the monoclonal antibodies designed to capture the target biomarker to the surface of the Al2O3 microrings. The detection of rhS100A4 proteins at clinically relevant concentrations in urine is a big milestone towards the use of biosensors for the screening and early diagnosis of different cancers. Biosensors based on this microring technology can lead to portable, multiplexed and easy-to-use point of care devices.

JTD Keywords: Distributed feedback lasers, Effective refractive index, Laser coupling, Polarization maintaining fibers, Refractive index, Scanning electron microscopy

Cyanide based silver electroplating is a low-cost reliable and well-established process for metal deposition. However, delicate handling during the process is needed because of the high toxicity of cyanide, for the persons and the environment. Uracil based silver electrodeposition got the attention of this field, because of its low cost and non-toxic nature. However, little is known about the silver complexation with uracil and the process behind the silver electroplating. In this work, we studied a hitherto unknown phenomenon on the diverse structure’s formation of silver uracil coordination complex due to the presence of different alkaline counterions. The distinct structuration of this complex clearly impacts on the efficiency and deposition yields of silver electroplating. We demonstrate the unknown key role that play hydroxide counterions in the uracil-silver coordination, and the different molecular structures created on the basis of the used counterion. The hydroxide counterion determines monomeric and polymeric complex formation with silver, which affects the solubility of the uracil silver complex and its subsequent electrodeposition. The different molecular complexes were characterized by FT-IR, UV–vis, DRUV–vis and multi-nuclear NMR spectroscopy and the silver electrodeposition by cyclic voltammetry and TOF-SIMS. This study sheds some light in the improvement of silver electroplating process

JTD Keywords: Coordination complex, Electrometallization, Electroplating, Metal complex, Silver electrodeposition, Uracil

This protocol uses natural type I collagen to generate three-dimensional (3-D) hydrogel for monitoring and analyzing the axonal growth. The protocol is centered on culturing small pieces of embryonic or early postnatal rodent brains inside a 3-D hydrogel formed by the rat tail tendon-derived type I collagen with specific porosity. Tissue pieces are cultured inside the hydrogel and confronted to specific brain fragments or genetically-modified cell aggregates to produce and secrete molecules suitable for creating a gradient inside the porous matrix. The steps of this protocol are simple and reproducible but include critical steps to be considered carefully during its development. Moreover, the behavior of growing axons can be monitored and analyzed directly using a phase-contrast microscope or mono/multiphoton fluorescence microscope after fixation by immunocytochemical methods.

JTD Keywords: 3-D hydrogel cultures, Axonal growth, Cell transfection, Chemoattraction, Chemorepulsion, Embryonic nervous system, Issue 148, Neuroscience, Tissue explants

Hypoxia is a common characteristic of many solid tumors that has been associated with tumor aggressiveness. Limited diffusion of oxygen generates a gradient of oxygen availability from the blood vessel to the interstitial space and may underlie the recruitment of macrophages fostering cancer progression. However, the available data based on the recruitment of circulating cells to the tumor microenvironment has been so far carried out by conventional co-culture systems which ignore the hypoxic gradient between the vessel to the tumor interstitium. Here, we have designed a novel easy-to-build cell culture device that enables evaluation of cellular cross-talk and cell migration while they are being simultaneously exposed to different oxygenation environments. As a proof-of-concept of the potential role of differential oxygenation among interacting cells we have evaluated the activation and recruitment of macrophages in response to hypoxic melanoma, breast, and kidney cancer cells. We found that hypoxic melanoma and breast cancer cells co-cultured with normoxic macrophages enhanced their directional migration. By contrast, hypoxic kidney cells were not able to increase their recruitment. We also identified well-described hypoxia-induced pathways which could contribute in the immune cell recruitment (VEGFA and PTGS2 genes). Moreover, melanoma and breast cancer increased their proliferation. However, oxygenation levels affected neither kidney cancer cell proliferation nor gene expression, which in turn resulted in no significant changes in macrophage migration and polarization. Therefore, the cell culture device presented here provides an excellent opportunity for researchers to reproduce the in vivo hypoxic gradients in solid tumors and to study their role in recruiting circulating cells to the tumor in specific types of cancer.

JTD Keywords: Hypoxia gradient, Macrophage motility, Models of host-tumor interactions, Novel assay technology, Tumor progression

Facial expressions are salient social features that crucial in communication, and humans are capable of reading the messages faces convey and the emotions they display. Robots that interact with humans will need to employ similar communication channels for successful interactions. Here, we focus on the readability of the facial expressions of a humanoid robot. We conducted an online survey where participants evaluated emotional stimuli and assessed the robot’s expressions. Results suggest that the robot’s facial expressions are correctly recognised and the appraisal of the robots expressive elements are consistent with the literature.

JTD Keywords: Emotion recognition, Facial expressions, Human-robot interaction

This paper explores which signal features of a gas sensor are optimum for assessing the proximity to a gas source in an open environment. Specifically, we compare three statistical descriptors of the signal (mean, variance and maximum response) against the 'bout' frequency, a feature computed in the derivative of the response. The experimental setup includes a generator of turbulent plumes and a sensing board composed of three metal oxide (MOX) sensors of different types. The main conclusion is that the maximum response is the most robust feature across the three sensors. The 'bout' frequency can be very sensitive to an additional parameter (the noise threshold).

JTD Keywords: Feature extraction, Gas plume, Gas sensors, Gas source localization, MOX, Signal processing

Estimating the affective state of a user during a computer task traditionally relies on either subjective reports or analysis of physiological signals, facial expressions, and other measures. These methods have known limitations, can be intrusive and may require specialized equipment. An alternative would be employing a ubiquitous device of everyday use such as a standard keyboard. Here we investigate if we can infer the emotional state of a user by analyzing their typing patterns. To test this hypothesis, we asked 400 participants to caption a set of emotionally charged images taken from a standard database with known ratings of arousal and valence. We computed different keystroke pattern dynamics, including keystroke duration (dwell time) and latency (flight time). By computing the mean value of all of these features for each image, we found a statistically significant negative correlation between dwell times and valence, and between flight times and arousal. These results highlight the potential of using keystroke dynamics to estimate the affective state of a user in a non-obtrusive way and without the need for specialized devices.

JTD Keywords: Feature extraction, Correlation, Keyboards, Task analysis, Statistical analysis, Affective computing, Standards, Keystroke, Keyboard, Typing, Arousal, Valence, Affect

This book constitutes the proceedings of the 8th International Conference on Biomimetic and Biohybrid Systems, Living Machines 2019, held in Nara, Japan, in July 2019. The 26 full and 16 short papers presented in this volume were carefully reviewed and selected from 45 submissions. They deal with research on novel life-like technologies inspired by the scientific investigation of biological systems, biomimetics, and research that seeks to interface biological and artificial systems to create biohybrid systems.

JTD Keywords: Artificial intelligence, Biomimetics, Computer architecture, Human robot interaction, Human-Computer Interaction (HCI), Humanoid robot, Image processing, Learning algorithms, Mobile robots, Multipurpose robots, Neural networks, Quadruped robots, Reinforcement learning, Robot learning, Robotics, Robots, Sensor, Sensors, Swarm robotics, User interfaces

Endocytosis and vesicular trafficking are cellular processes that regulate numerous functions required to sustain life. From a translational perspective, they offer avenues to improve the access of therapeutic drugs across cellular barriers that separate body compartments and into diseased cells. However, the fact that many factors have the potential to alter these routes, impacting our ability to effectively exploit them, is often overlooked. Altered vesicular transport may arise from the molecular defects underlying the pathological syndrome which we aim to treat, the activity of the drugs being used, or side effects derived from the drug carriers employed. In addition, most cellular models currently available do not properly reflect key physiological parameters of the biological environment in the body, hindering translational progress. This article offers a critical overview of these topics, discussing current achievements, limitations and future perspectives on the use of vesicular transport for drug delivery applications.

JTD Keywords: Cellular vesicles, Vesicle fusion, Fission and intracellular trafficking, Drug delivery systems and nanomedicines, Transcytosis and endocytosis of drugs carriers, Disease effects on vesicular trafficking, Drug effects on vesicular trafficking, Role of the biological environment

Generating complex, human-like behaviour in a humanoid robot like the iCub requires the integration of a wide range of open source components and a scalable cognitive architecture. Hence, we present the iCub-HRI library which provides convenience wrappers for components related to perception (object recognition, agent tracking, speech recognition, touch detection), object manipulation (basic and complex motor actions) and social interaction (speech synthesis, joint attention) exposed as a C++ library with bindings for Java (allowing to use iCub-HRI within Matlab) and Python. In addition to previously integrated components, the library allows for simple extension to new components and rapid prototyping by adapting to changes in interfaces between components. We also provide a set of modules which make use of the library, such as a high-level knowledge acquisition module and an action recognition module. The proposed architecture has been successfully employed for a complex human-robot interaction scenario involving the acquisition of language capabilities, execution of goal-oriented behaviour and expression of a verbal narrative of the robot's experience in the world. Accompanying this paper is a tutorial which allows a subset of this interaction to be reproduced. The architecture is aimed at researchers familiarising themselves with the iCub ecosystem, as well as expert users, and we expect the library to be widely used in the iCub community.

JTD Keywords: Robotics, iCub Humanoid, YARP, Software architecture, C++, Python, Java, Human-robot interaction

Fibrosis development was initially conceived as an incessant progressive condition. Nowadays, it has become evident that fibrotic tissue undergoes a continuous two-way process: fibrogenesis and fibrinolysis, characterizing the remodeling of extracellular matrix (ECM). However, in established fibrosis, this two-way process is tipped towards fibrogenesis and this leads to a self-perpetuating accumulation of ECM, a distinct metabolic unit, together with other cells and processes promoting fibrosis deposition. Several mechanisms promote fibrosis regression, such as degradation of ECM, infiltration of restorative macrophages, prevention of the epithelial-mesenchymal transition of hepatocytes, restoration of the liver sinusoidal endothelial cells’ differentiation phenotype, and reversion to quiescence, apoptosis and senescence of hepatic stellate cells (HSC). Hence, fibrosis is the result of an unbalanced two-way process of matrix remodeling. At the late stage of the disease, antifibrotic interventions could become necessary to reverse self-perpetuating fibrogenesis and accelerate regression of fibrosis even if cause and cofactors of hepatic injury have been eliminated. This review outlines some of the important mechanisms leading towards regression of liver fibrosis.

JTD Keywords: Hepatic stellate cells, Extracellular matrix, remodeling, Rho-associated kinases, Janus kinases

Mucosal administration of drugs and drug delivery systems has gained increasing interest. However, nanoparticles intended to protect and deliver drugs to epithelial surfaces require transport through the surface-lining mucus. Translation from bench to bedside is particularly challenging for mucosal administration since a variety of parameters will influence the specific barrier properties of the mucus including the luminal fluids, the microbiota, the mucus composition and clearance rate, and the condition of the underlying epithelia. Besides, after administration, nanoparticles interact with the mucosal components, forming a biomolecular corona that modulates their behavior and fate after mucosal administration. These interactions are greatly influenced by the nanoparticle properties, and therefore different designs and surface-engineering strategies have been proposed. Overall, it is essential to evaluate these biomolecule-nanoparticle interactions by complementary techniques using complex and relevant mucus barrier matrices.

JTD Keywords: Nanoparticle formulation strategies, Corona formation, Digestive tract, Respiratory tract, Luminal content, Methodologies, Analysis

Biomaterials can interact with cells directly, that is, by direct contact of the cells with the material surface, or indirectly, through soluble species that can be released to or uptaken from the surrounding fluids. However, it is difficult to characterise the relevance of this fluid-mediated interaction separately from the topography and composition of the substrate, because they are coupled variables. These fluid-mediated interactions are amplified in the case of highly reactive calcium phosphates (CaPs) such as biomimetic calcium deficient hydroxyapatite (CDHA), particularly in static in vitro cultures. The present work proposes a strategy to decouple the effect of ion exchange from topographical features by adjusting the volume ratio between the cell culture medium and biomaterial (VCM/VB). Increasing this ratio allowed mitigating the drastic ionic exchanges associated to the compositional changes experienced by the material exposed to the cell culture medium. This strategy was validated using rat mesenchymal stem cells (rMSCs) cultured on CDHA and beta-tricalcium phosphate (β-TCP) discs using different VCM/VB ratios. Whereas in the case of β-TCP the cell response was not affected by this ratio, a significant effect on cell adhesion and proliferation was found for the more reactive CDHA. The ionic exchange, produced by CDHA at low VCM/VB, altered cell adhesion due to the reduced number of focal adhesions, caused cell shrinkage and further rMCSs apoptosis. This was mitigated when using a high VCM/VB, which attenuated the changes of calcium and phosphate concentrations in the cell culture medium, resulting in rMSCs spreading and a viability over time. Moreover, rMSCs showed an earlier expression of osteogenic genes on CDHA compared to sintered β-TCP when extracellular calcium fluctuations were reduced. Statement of Significance: Fluid mediated interactions play a significant role in the bioactivity of calcium phosphates. Ionic exchange is amplified in the case of biomimetic hydroxyapatite, which makes the in vitro characterisation of cell-material interactions especially challenging. The present work proposes a novel and simple strategy to explore the mechanisms of interaction of biomimetic and sintered calcium phosphates with mesenchymal stem cells. The effects of topography and ion exchange are analysed separately by modifying the volume ratio between cell culture medium and biomaterial. High ionic fluctuations interfered in the maturation of focal adhesions, hampering cell adhesion and leading to increased apoptosis and reduced proliferation rate.

JTD Keywords: Calcium phosphates, Mesenchymal stem cells, Intracellular calcium, Cell adhesion

Extracellular matrix (ECM) proteins are key mediators of cell/material interactions. The surface density and conformation of these proteins adsorbed on the material surface influence cell adhesion and the cellular response. We have previously shown that subtle variations in surface chemistry lead to drastic changes in the conformation of adsorbed fibronectin (FN). On poly(ethyl acrylate) (PEA), FN unfolds and displays domains for cell adhesion and FN-FN interaction, whereas on poly(methyl acrylate) (PMA) – with only one methyl group less – FN remains globular as it is in solution. The effect of the strength of the protein/material interaction in cell response, and its relation to protein density and conformation, has received limited attention so far. In this work, we used FN-functionalized AFM cantilevers to evaluate, via force spectroscopy, the strength of interaction between fibronectin and the underlying polymer which controls FN conformation (PEA and PMA). We found that the strength of FN/PEA interaction is significantly higher than FN/PMA, which limits the mobility of FN layer on PEA, reduces the ability of cells to mechanically reorganize FN and then leads to enhanced proteolysis and degradation of the surrounding matrix with compromised cell viability. By contrast, both PEA and PMA support cell adhesion when FN density is increased and also in the presence of serum or other serum proteins, including vitronectin (VN) and bovine serum albumin (BSA), which provide a higher degree of mobility to the matrix. Statement of Significance: The identification of parameters influencing cell response is of paramount importance for the design of biomaterials that will act as synthetic scaffolds for cells to anchor, grow and, eventually, become specialised tissues. Cells interact with materials through an intermediate layer of proteins adsorbed on the material surface. It is known that the density and conformation of these proteins determine cell behaviour. Here we show that the strength of protein/material interactions, which has received very limited attention so far, is key to understand the cellular response to biomaterials. Very strong protein/material interactions reduce the ability of cells to mechanically reorganize proteins at the material interface which results in enhanced matrix degradation, leading ultimately to compromised cell viability.

JTD Keywords: Fibronectin adsorption, Fibronectin remodeling, Protein mobility, Protein-material interaction strength

There is growing recognition that the mechanical interactions between cells and their local extracellular matrix (ECM) are central regulators of tissue development, homeostasis, repair and disease progression. The unique ability of atomic force microscopy (AFM) to probe quantitatively mechanical properties and forces at the nanometer or micrometer scales in all kinds of biological samples has been instrumental in the recent advances in cell and tissue mechanics. In this review we illustrate how AFM has provided important insights on our current understanding of the mechanobiology of cells, ECM and cell-ECM bidirectional interactions, particularly in the context of soft acinar tissues like the mammary gland or pulmonary tissue. AFM measurements have revealed that intrinsic cell micromechanics is cell-type specific, and have underscored the prominent role of β1 integrin/FAK(Y397) signaling and the actomyosin cytoskeleton in the mechanoresponses of both parenchymal and stromal cells. Moreover AFM has unveiled that the micromechanics of the ECM obtained by tissue decellularization is unique for each anatomical compartment, which may support both its specific function and cell differentiation. AFM has also enabled identifying critical mechanoregulatory proteins involved in branching morphogenesis (MMP14) and acinar differentiation (α3β1 integrin), and has clarified the role of altered tissue mechanics and architecture in a variety of pathologic conditions. Critical technical issues of AFM mechanical measurements like tip geometry effects are also discussed.

JTD Keywords: Atomic force microscopy, Beta1 integrin, Elastic modulus, Extracellular matrix, Morphogenesis, Tissue decellularization

N-methyl-D-aspartate receptors (NMDARs) respond to glutamate to allow the influx of calcium ions and the signaling to the mitogen-activated protein kinase (MAPK) cascade. Both MAPK- and Ca2+-mediated events are important for both neurotransmission and neural cell function and fate. Using a heterologous expression system, we demonstrate that NMDAR may interact with the EF-hand calcium-binding proteins calmodulin, calneuron-1, and NCS1 but not with caldendrin. NMDARs were present in primary cultures of both neurons and microglia from cortex and hippocampus. Calmodulin in microglia, and calmodulin and NCS1 in neurons, are necessary for NMDA-induced MAP kinase pathway activation. Remarkably, signaling to the MAP kinase pathway was blunted in primary cultures of cortical and hippocampal neurons and microglia from wild-type animals by proteins involved in neurodegenerative diseases: α-synuclein, Tau, and p-Tau. A similar blockade by pathogenic proteins was found using samples from the APPSw,Ind transgenic Alzheimer’s disease model. Interestingly, a very marked increase in NMDAR–NCS1 complexes was identified in neurons and a marked increase of both NMDAR–NCS1 and NMDAR–CaM complexes was identified in microglia from the transgenic mice. The results show that α-synuclein, Tau, and p-Tau disrupt the signaling of NMDAR to the MAPK pathway and that calcium sensors are important for NMDAR function both in neurons and microglia. Finally, it should be noted that the expression of receptor–calcium sensor complexes, specially those involving NCS1, is altered in neural cells from APPSw,Ind mouse embryos/pups.

JTD Keywords: Alzheimer’s disease, Calmodulin, Calneuron-1, Caldendrin, NCS1, Extracellular signal-regulated kinase, Glutamate receptor, Proximity ligation assay

JTD Keywords: Extracellular vesicles, Hemozoin, Malaria, Nanomedicine, Plasmodium, Red blood cell, Targeted drug delivery

This paper introduces a cognitive architecture for a humanoid robot to engage in a proactive, mixed-initiative exploration and manipulation of its environment, where the initiative can originate from both the human and the robot. The framework, based on a biologically-grounded theory of the brain and mind, integrates a reactive interaction engine, a number of state-of-the art perceptual and motor learning algorithms, as well as planning abilities and an autobiographical memory. The architecture as a whole drives the robot behavior to solve the symbol grounding problem, acquire language capabilities, execute goal-oriented behavior, and express a verbal narrative of its own experience in the world. We validate our approach in human-robot interaction experiments with the iCub humanoid robot, showing that the proposed cognitive architecture can be applied in real time within a realistic scenario and that it can be used with naive users.

JTD Keywords: Autobiographical Memory., Biology, Cognition, Cognitive Robotics, Computer architecture, Distributed Adaptive Control, Grounding, Human-Robot Interaction, Humanoid robots, Robot sensing systems, Symbol Grounding

Observing biochemical processes within living cell is imperative for biological and medical research. Fluoresce imaging is widely used for intracellular sensing of cell membranes, nuclei, lysosomes, and pH. Electrochemical assays have been proposed as an alternative to fluorescence-based assays because of excellent analytical features of electrochemical devices. Notably, thanks to the rapid progress of micro/nanotechnologies and electrochemical techniques, intracellular electrochemical sensing is making rapid progress, leading to a successful detection of intracellular components. Such insight can provide a deep understanding of cellular biological processes and, ultimately, define the human healthy and diseased states. In this review, we present an overview of recent research progress in intracellular electrochemical sensing. We focus on two main topics, electrochemical extraction of cytosolic contents from cells and intracellular electrochemical sensing in situ.

JTD Keywords: Micro/nanoelectrode, Analytical electrochemistry, Intracellular sensing, Cell analysis

We aimed to biomechanically evaluate the effect of the supraspinatus tendon on tuberosity stability using two different reverse shoulder arthroplasty (RSA) models for complex proximal humeral fractures (PHFs).

JTD Keywords: Tuberosity reconstruction, Reverse shoulder arthroplasty, Supraspinatus, Cadaveric study, Rotator cuff excision, Complex proximal humeral fractures

In this paper, we argue that the future of Artificial Intelligence research resides in two keywords: integration and embodiment. We support this claim by analyzing the recent advances in the field. Regarding integration, we note that the most impactful recent contributions have been made possible through the integration of recent Machine Learning methods (based in particular on Deep Learning and Recurrent Neural Networks) with more traditional ones (e.g. Monte-Carlo tree search, goal babbling exploration or addressable memory systems). Regarding embodiment, we note that the traditional benchmark tasks (e.g. visual classification or board games) are becoming obsolete as state-of-the-art learning algorithms approach or even surpass human performance in most of them, having recently encouraged the development of first-person 3D game platforms embedding realistic physics. Building on this analysis, we first propose an embodied cognitive architecture integrating heterogeneous subfields of Artificial Intelligence into a unified framework. We demonstrate the utility of our approach by showing how major contributions of the field can be expressed within the proposed framework. We then claim that benchmarking environments need to reproduce ecologically-valid conditions for bootstrapping the acquisition of increasingly complex cognitive skills through the concept of a cognitive arms race between embodied agents.

JTD Keywords: Cognitive Architectures, Embodied Artificial Intelligence, Evolutionary Arms Race, Unified Theories of Cognition

The components of a Living Machine must be integrated into a functioning whole, which requires a detailed understanding of the architecture of living machines. This chapter starts with a conceptual and historical analysis which from Plato brings us to nineteenth-century neuroscience and early concepts of the layered structure of nervous systems. These concepts were further captured in the cognitive behaviorism of Tolman and came to full fruition in the cognitive revolution of the second half of the twentieth century. Verschure subsequently describes the most relevant proposals of cognitive architectures followed by an overview of the few proposals stemming from modern neuroscience on the architecture of the brain. Subsequently, we will look at contemporary contenders that mediate between cognitive and brain architecture. An important challenge to any model of cognitive architectures is how to benchmark it. Verschure proposes the Unified Theories of Embodied Minds (UTEM) benchmark which advances from Newell’s classic Unified Theories of Cognition benchmark.

JTD Keywords: Architecture, Mind, Brain, Organization, System, Virtualization, Abstraction layers

This chapter takes the view that emotions of living machines can be seen from the perspective of self-regulation and appraisal. We will first look at the pragmatic needs to endow machines with emotions and subsequently describe some of the historical background of the science of emotions and its different interpretations and links to affective neuroscience. Subsequently, we argue that emotions can be cast in terms of self-regulation where they provide for a descriptor of the state of the homeostatic processes that maintain the relationship between the agent and its internal and external environment. We augment the notion of homeostasis with that of allostasis which signifies a change from stability through a fixed equilibrium to stability through continuous change. The chapter shows how this view can be used to create complex living machines where emotions are anchored in the need fulfillment of the agent, in this case considering both utilitarian and epistemic needs.

JTD Keywords: Emotion, Motivation, Needs, Appraisal, Self-regulation, Homeostasis, Allostasis, Human–robot interaction, James–Lange theory

Within the research in Molecular Biology, one important field along the years has been the analyses on how prokaryotes regulate the expression of their genes and what the consequences of these activities are. Prokaryotes have attracted the interests of researchers not only because the processes taking place in their world are important to cells, but also because many of the effects often can be readily measured, both at the single cell level and in large populations. Contributing to the interest of the present topic is the fact that modulation of gene activity involves the sensing of intra- and inter-cellular conditions, DNA binding and DNA dynamics, and interaction with the replication/transcription machinery of the cell. All of these processes are fundamental to the operation of a biological entity and they condition its lifestyle. Further, the discoveries achieved in the bacterial world have been of ample use in eukaryotes. In addition to the fundamental interest of understanding modulation of prokaryotic lifestyle by DNA-binding proteins, there is an added interest from the healthcare point of view. As it is well-known the antibiotic-resistance strains of pathogenic bacteria are a major world problem, so that there is an urgent need of innovative approaches to tackle it. Human and animal infectious diseases impose staggering costs worldwide in terms of loss of human life and livestock, diminished productivity, and the heavy economic burden of disease. The global dimension of international trade, personal travel, and population migration expands at an ever-accelerating rate. This increasing mobility results in broader and quicker dissemination of bacterial pathogens and in rapid spread of antibiotic resistance. The majority of the newly acquired resistances are horizontally spread among bacteria of the same or different species by processes of lateral (horizontal) gene transfer, so that discovery of new antibiotics is not the definitive solution to fighting infectious diseases. There is an absolute need of finding novel alternatives to the “classical” approach to treat infections by bacterial pathogens, and these new ways must include the exploration and introduction of novel antibacterials, the development of alternative strategies, and the finding of novel bacterial targets. However, all these approaches will result in a stalemate if we, researchers, are not able to achieve a better understanding of the mechanistic processes underlying bacterial gene expression. It is, then, imperative to continue gaining insight into the basic mechanisms by which bacterial cells regulate the expression of their genes. That is why our Research Topic hosted by Frontiers in Molecular Biosciences was timely, and the output of it offers novel and up-to-date points of view to the “simple” bacterial world.

JTD Keywords: DNA-protein interactions, Gene regulation in Prokaryotes, Replication control, Regulation of Bacterial Gene Expression, Global Regulatory Networks

Directed cell migration is essential for many biological processes, such as embryonic development or cancer progression. Cell contractility and adhesion to the extracellular matrix are known to regulate cell locomotion machinery. However, the cross-talk between extrinsic and intrinsic factors at the molecular level on the biophysical mechanism of three dimensional (3D)-directed cell migration is still unclear. In this work, a novel physiologically relevant in vitro model of the extracellular microenvironment is used to reveal how the topological anisotropy of the extracellular matrix synergizes with actomyosin contractility to modulate directional cell migration morphodynamics. This study shows that cells seeded on polarized 3D matrices display asymmetric protrusion morphodynamics and in-vivo-like phenotypes. It is found that matrix anisotropy significantly enhances cell directionality, but strikingly, not the invasion distance of cells. In Rho-inhibited cells, matrix anisotropy counteracts the lack of actomyosin-driven forces to stabilize cell directionality suggesting a myosin-II-independent mechanism for cell guidance. Finally, this study shows that on isotropic 3D environments, cell directionality is independent of actomyosin contractility. Altogether, this study provides novel quantitative data on the biomechanical regulation of directional cell motion and shows the important regulatory role of matrix anisotropy and actomyosin forces to guide cell migration in 3D microenvironments.

JTD Keywords: Anisotropy, Directed cell migration, Extracellular matrices, Migration modes, Three dimensional microenvironments

The interaction of cells with their natural environment influences a large variety of cellular phenomena, including cell adhesion, proliferation, and migration. The complex extracellular matrix network has challenged the attempts to replicate in vitro the heterogeneity of the cell environment and has threatened, in general, the relevance of in vitro studies. In this work, we describe a new and extremely versatile approach to generate native-like extracellular matrices with controlled morphologies for the in vitro study of cellular processes. This general approach combines the confluent culture of fibroblasts with microfabricated guiding templates to direct the three-dimensional growth of well-defined extracellular networks which recapitulate the structural and biomolecular complexity of features typically found in vivo. To evaluate its performance, we studied fundamental cellular processes, including cell cytoskeleton organization, cell-matrix adhesion, proliferation, and protrusions morphodynamics. In all cases, we found striking differences depending on matrix architecture and, in particular, when compared to standard two-dimensional environments. We also assessed whether the engineered matrix networks influenced cell migration dynamics and locomotion strategy, finding enhanced migration efficiency for cells seeded on aligned matrices. Altogether, our methodology paves the way to the development of high-performance models of the extracellular matrix for potential applications in tissue engineering, diagnosis, or stem-cell biology.

JTD Keywords: Biomimetics, Cell migration, Engineered cell-derived matrices, Extracellular matrix, In vitro model

Engineering dynamic stem cell niche-like environment offers opportunity to obtain better control of the fate of stem cells. We identified, for the first time, that periodic changes in the adhesive environment of human adipose derived mesenchymal stem cells (ADSCs) alters dramatically their asymmetric division but not their ability for symmetric renewal. Hereby, we used smart thermo-responsive polymer (PNIPAM) to create a dynamic adhesive environment for ADSCs by applying periodic temperature cycles to perturb adsorbed adhesive proteins to substratum interaction. Cumulative population doubling time (CPDT) curves showed insignificant decline in the symmetric cell growth studied for up to 13th passages accompanied with small changes in the overall cell morphology and moderately declined fibronectin (FN) matrix deposition probably as a functional consequence of ADSCs ageing. However, a substantial alteration in the differentiation potential of ADSCs from both early and late passages (3rd and 14th, respectively) was found when the cells were switched to osteogenic differentiation conditions. This behavior was evidenced by the significantly altered alkaline phosphatase activity and Ca deposition (Alizarin red) assayed at 3, 14 and 21 day in comparison to the control samples of regular TC polystyrene processed under same temperature settings.

JTD Keywords: Cell ageing, Dynamic adhesive environment, Extracellular matrix, Mesenchymal stem cells, PNIPAM, Stem cell niche, Symmetric and asymmetric cell growth, Thermo-cycling, Thermo-responsive polymer

The underlying mechanisms by which extracellular matrix (ECM) mechanics influences cell and tissue function remain to be elucidated because the events associated with this process span size scales from tissue to molecular level. Furthermore, ECM has an extremely complex hierarchical 3D structure and the load distribution is highly dependent on the architecture and mechanical properties of ECM. In the present study, the macro- and microscale mechanical properties of collagen gel were studied. Dynamic rheological testing was performed to study the macroscale mechanical properties of collagen gel. The microscale mechanical properties of collagen gel were measured using optical magnetic twisting cytometry (OMTC). Ferromagnetic beads embedded in the matrix were used as mechanical probes. Our study on the multiscale mechanical properties of collage matrix suggests several interesting differences between macro and microscale mechanical properties originated from the scales of measurements. At the macroscopic scale, storage and loss modulus increase with collagen concentrations. Nonaffine collagen fibril structural network deformation plays an important role in determining the macroscopic mechanical properties of the collagen matrix. At the microscopic scale, however, the local mechanical properties are less sensitive to changes in collagen concentration because of the more immediate/direct deformation of collagen fibrils in the OMTC measurements through forces exerted by locally attached ferromagnetic beads. The loss modulus is more affected by the local interstitial fluid environment, leading to a rather dramatic increase in viscosity with frequency, especially at higher frequencies (>10 Hz). A finite element model was developed to study the geometric factors in the OMTC measurements when the collagen matrix was considered to be hyperelastic. Our results show that the geometric factors are dependent on collagen concentration, or the stiffness of matrix, when nonlinear material properties of the matrix are considered, and thus interpretation of the apparent modulus from OMTC measurements should be conducted carefully.

JTD Keywords: Keywords: collagen, Extracellular matrix, Geometric factor, Nonaffine deformation, Optical magnetic twisting cytometry

Graft polymer surfactants provide very good colloidal stability because of strong steric repulsions between adsorbed surfactant films. The elastic and adhesion properties of adsorbed hydrophobically modified inulin polymer surfactant (INUTEC NRA) have been directly measured using Atomic Force Microscopy (AFM) measurements. For this purpose, poly(methyl methacrylate/butyl acrylate), P(MMA/BuA), latexes prepared in the presence of the hydrophobically modified inulin (INUTEC NRA) were used. These latexes (diameter 118 nm and polydispersity index of 1.05) showed a very high colloidal stability in water and in the presence of electrolyte (up to 0.2 mol dm−3 KBr). The latexes were deposited on mica, which was silanated to enhance the adhesion of the latex particles to the surface. A silicon nitride tip with approximately 10 nm diameter that also contained an adsorbed layer of surfactant was used in the AFM apparatus. The tip was allowed to approach, contact thereafter the particles with an applied force of 12.5 nN, and finally detach from the film. Both elastic (Young’s) modulus of the film and adhesion force were studied. The results showed that the adsorbed surfactant films are highly elastic and their elastic modulus and adhesion force did not change significantly with the presence of Na2SO4 up to 0.05 mol dm−3. The high elastic contribution to the steric interaction ensures strong repulsion between the latex particles both in water and at high electrolyte concentrations. In addition, the lack of dependence of adhesion force on electrolyte concentration ensures uniform deposition of the latex particles on a flat substrate as for example in coating applications. These results show the advantages of using a graft polymer surfactant for enhancing the stability of particle suspensions, as illustrated in previous investigations.

JTD Keywords: AFM, Colloidal stability, Interaction forces, Steric repulsion

Elucidating the factors that direct the spatio-temporal organization of evolving tissues is one of the primary purposes in the study of development. Various propositions claim to have been important contributions to the understanding of the mechanical properties of cells and tissues in their spatiotemporal organization in different developmental and morphogenetic processes. However, due to the lack of reliable and accessible tools to measure material properties and tensional parameters in vivo, validating these hypotheses has been difficult. Here we present methods employing atomic force microscopy (AFM) and particle tracking with the aim of quantifying the mechanical properties of the intact zebrafish embryo yolk cell during epiboly. Epiboly is an early conserved developmental process whose study is facilitated by the transparency of the embryo. These methods are simple to implement, reliable, and widely applicable since they overcome intrusive interventions that could affect tissue mechanics. A simple strategy was applied for the mounting of specimens, AFM recording, and nanoparticle injections and tracking. This approach makes these methods easily adaptable to other developmental times or organisms.

JTD Keywords: Developmental Biology, Zebrafish, Yolk, Atomic Force Microscopy, Cortical Tension, Microrheology, Nanoparticle tracking

The increasing recognition that tissue elasticity is an important regulator of cell behavior in normal and pathologic conditions such as fibrosis and cancer has driven the development of cell culture substrata with tunable elasticity. Such development has urged the need to quantify the elastic properties of these cell culture substrata particularly at the nanometer scale, since this is the relevant length scale involved in cell-extracellular matrix (ECM) mechanical interactions. To address this need, we have exploited the versatility of atomic force microscopy to quantify the elastic properties of a variety of cell culture substrata used in mechanobiology studies, including floating collagen gels, ECM-coated polyacrylamide gels, and decellularized tissue sections. In this review we summarize major findings in this field from our group within the context of the state-of-the-art in the field, and provide a critical discussion on the applicability and complementarity of currently available cell culture assays with tunable elasticity. In addition, we briefly describe how the limitations of these assays provide opportunities for future research, which is expected to continue expanding our understanding of the mechanobiological aspects that support both normal and diseased conditions.

JTD Keywords: 3D culture, Atomic force microscopy, Elastic modulus, Extracellular matrix, Polyacrylamide

Regenerative medicine is a relatively new field with new requirements for smart materials, where composites will have a strong role to play. The new paradigm of regenerative medicine and tissue engineering requires biomaterials with high specificity, where physical and chemical properties are duly tailored and combined with appropriate mechanical and degradation features in order to trigger specific cell events and functions involved in the regenerative process. In this chapter, the chemical, physical, and biological elements that have to be targeted by biocomposites in regenerative medicine are described.

JTD Keywords: Biocomposite, Regenerative medicine, Tissue engineering, Scaffolds, Cell/material interactions

Portal hypertension either develops due to progressive liver fibrosis or is the consequence of vascular liver diseases such as portal vein thrombosis or non-cirrhotic portal hypertension. This chapter focuses on different rodent models of liver fibrosis with portal hypertension and also in few non-cirrhotic portal hypertension models. Importantly, after the development of portal hypertension, the proper assessment of drug effects in the portal and systemic circulation should be discussed. The last part of the chapter is dedicated in these techniques to assess the in vivo hemodynamics and the ex vivo techniques of the isolated liver perfusion and vascular contractility.

JTD Keywords: Aortic ring contraction, Bile duct ligation, Carbon tetrachloride, Colored microsphere technique, High-fat diet, Isolated in situ liver perfusion, Methionine-choline-deficient diet, Partial portal vein ligation, Portal hypertension

Global regulators play an essential role in the adaptation of bacterial cells to specific niches. Bacterial pathogens thriving in the tissues and organs of their eukaryotic hosts are a well-studied example. Some of the proteins that recognize local DNA structures rather than specific nucleotide sequences act as global modulators in many bacteria, both Gram-negative and -positive. To this class of regulators belong the H-NS-like proteins, mainly identified in γ-Proteobacteria, and the MgaSpn-like proteins identified in Firmicutes. H-NS and MgaSpn from Escherichia coli and Streptococcus pneumoniae, respectively, neither have sequence similarity nor share structural domains. Nevertheless, they display common features in their interaction with DNA, namely: (i) they bind to DNA in a non-sequence-specific manner, (ii) they have a preference for intrinsically curved DNA regions, and (iii) they are able to form multimeric complexes on linear DNA. Using DNA fragments from the hemolysin operon regulatory region of the E. coli plasmid pHly152, we show in this work that MgaSpn is able to recognize particular regions on extended H-NS binding sites. Such regions are either located at or flanked by regions of potential bendability. Moreover, we show that the regulatory region of the pneumococcal P1623B promoter, which is recognized by MgaSpn, contains DNA motifs that are recognized by H-NS. These motifs are adjacent to regions of potential bendability. Our results suggest that both regulatory proteins recognize similar structural characteristics of DNA.

JTD Keywords: Global transcriptional regulators, Nucleoid-associated proteins, Mga/AtxA family, Protein-DNA interactions, DNA bendability

Genome editing on human pluripotent stem cells (hPSCs) together with the development of protocols for organ decellularization opens the door to the generation of autologous bioartificial hearts. Here we sought to generate for the first time a fluorescent reporter human embryonic stem cell (hESC) line by means of Transcription activator-like effector nucleases (TALENs) to efficiently produce cardiomyocyte-like cells (CLCs) from hPSCs and repopulate decellularized human heart ventricles for heart engineering. In our hands, targeting myosin heavy chain locus (MYH6) with mCherry fluorescent reporter by TALEN technology in hESCs did not alter major pluripotent-related features, and allowed for the definition of a robust protocol for CLCs production also from human induced pluripotent stem cells (hiPSCs) in 14 days. hPSCs-derived CLCs (hPSCs-CLCs) were next used to recellularize acellular cardiac scaffolds. Electrophysiological responses encountered when hPSCs-CLCs were cultured on ventricular decellularized extracellular matrix (vdECM) correlated with significant increases in the levels of expression of different ion channels determinant for calcium homeostasis and heart contractile function. Overall, the approach described here allows for the rapid generation of human cardiac grafts from hPSCs, in a total of 24 days, providing a suitable platform for cardiac engineering and disease modeling in the human setting.

JTD Keywords: Cardiac function, Extracellular matrix, Gene targeting, Pluripotent stem cells

Aim: Explore the use of polymeric micelles in the development of powders intended for pulmonary delivery of biopharmaceuticals, using insulin as a model protein. Materials & methods: Formulations were assessed in vitro for aerosolization properties and in vivo for efficacy and safety using a streptozotocin-induced diabetic rat model. Results: Powders presented good aerosolization properties like fine particle fraction superior to 40% and a mass median aerodynamic diameter inferior of 6 μm. Endotracheally instilled powders have shown a faster onset of action than subcutaneous administration of insulin at a dose of 10 IU/kg, with pharmacological availabilities up to 32.5% of those achieved by subcutaneous route. Additionally, micelles improved the hypoglycemic effect of insulin. Bronchoalveolar lavage screening for toxicity markers (e.g., lactate dehydrogenase, cytokines) revealed no signs of lung inflammation and cytotoxicity 14 days postadministration. Conclusion: Developed powders showed promising safety and efficacy characteristics for the systemic delivery of insulin by pulmonary administration.

JTD Keywords: Fine particle fraction, Inhalation, Insulin, In vivo, Pharmacological availability, Polymeric micelles, Pulmonary toxicity

The way cells are organized within a tissue dictates how they sense and respond to extracellular signals, as cues are received and interpreted based on expression and organization of receptors, downstream signaling proteins, and transcription factors. Part of this microenvironmental context is the result of forces acting on the cell, including forces from other cells or from the cellular substrate or basement membrane. However, measuring forces exerted on and by cells is difficult, particularly in an in vivo context, and interpreting how forces affect downstream cellular processes poses an even greater challenge. Here, we present a simple method for monitoring and analyzing forces generated from cell collectives. We demonstrate the ability to generate traction force data from human embryonic stem cells grown in large organized epithelial sheets to determine the magnitude and organization of cell-ECM and cell-cell forces within a self-renewing colony. We show that this method can be used to measure forces in a dynamic hESC system and demonstrate the ability to map intracolony protein localization to force organization.

JTD Keywords: Epiblast, Human embryonic stem cells, Mechanotransduction, Monolayer stress microscopy, Self-organization, Traction force

Differentiating normal from adventitious respiratory sounds (RS) is a major challenge in the diagnosis of pulmonary diseases. Particularly, continuous adventitious sounds (CAS) are of clinical interest because they reflect the severity of certain diseases. This study presents a new classifier that automatically distinguishes normal sounds from CAS. It is based on the multi-scale analysis of instantaneous frequency (IF) and envelope (IE) calculated after ensemble empirical mode decomposition (EEMD). These techniques have two major advantages over previous techniques: high temporal resolution is achieved by calculating IF-IE and a priori knowledge of signal characteristics is not required for EEMD. The classifier is based on the fact that the IF dispersion of RS signals markedly decreases when CAS appear in respiratory cycles. Therefore, CAS were detected by using a moving window to calculate the dispersion of IF sequences. The study dataset contained 1494 RS segments extracted from 870 inspiratory cycles recorded from 30 patients with asthma. All cycles and their RS segments were previously classified as containing normal sounds or CAS by a highly experienced physician to obtain a gold standard classification. A support vector machine classifier was trained and tested using an iterative procedure in which the dataset was randomly divided into training (65%) and testing (35%) sets inside a loop. The SVM classifier was also tested on 4592 simulated CAS cycles. High total accuracy was obtained with both recorded (94.6% ± 0.3%) and simulated (92.8% ± 3.6%) signals. We conclude that the proposed method is promising for RS analysis and classification.

JTD Keywords: Diseases, Dispersion, Empirical mode decomposition, Feature extraction, Informatics, Support vector machines

Purpose: To assess, with finite element (FE) calculations, whether immediate weight bearing would be possible after surgical stabilization either with cannulated screws or with a locking plate in a split fracture of the lateral tibial plateau (LTP). Methods: A split fracture of the LTP was recreated in a FE model of a human tibia. A three-dimensional FE model geometry of a human femur-tibia system was obtained from the VAKHUM project database, and was built from CT images from a subject with normal bone morphologies and normal alignment. The mesh of the tibia was reconverted into a geometry of NURBS surfaces. A split fracture of the lateral tibial plateau was reproduced by using geometrical data from patient radiographs. A locking screw plate (LP) and a cannulated screw (CS) systems were modelled to virtually reduce the fracture and 80 kg static body-weight was simulated. Results: While the simulated body-weight led to clinically acceptable interfragmentary motion, possible traumatic bone shear stresses were predicted nearby the cannulated screws. With a maximum estimation of about 1.7 MPa maximum bone shear stresses, the Polyax system might ensure more reasonable safety margins. Conclusions: Split fractures of the LTP fixed either with locking screw plate or cannulated screws showed no clinically relevant IFM in a FE model. The locking screw plate showed higher mechanical stability than cannulated screw fixation. The locking screw plate might also allow full or at least partial weight bearing under static posture at time zero.

JTD Keywords: Bone fixation, Finite element, Fracture fixation, Interfragmentary motion, Tibial plateau fractures, Weight bearing

Changes in the left ventricle function produce alternans in the hemodynamic and electric behavior of the cardiovascular system. A total of 49 cardiomyopathy patients have been studied based on the blood pressure signal (BP), and were classified according to the left ventricular ejection fraction (LVEF) in low risk (LR: LVEF>35%, 17 patients) and high risk (HR: LVEF≤35, 32 patients) groups. We propose to characterize these patients using a linear and a nonlinear methods, based on the spectral estimation and the recurrence plot, respectively. From BP signal, we extracted each systolic time interval (STI), upward systolic slope (BPsl), and the difference between systolic and diastolic BP, defined as pulse pressure (PP). After, the best subset of parameters were obtained through the sequential feature selection (SFS) method. According to the results, the best classification was obtained using a combination of linear and nonlinear features from STI and PP parameters. For STI, the best combination was obtained considering the frequency peak and the diagonal structures of RP, with an area under the curve (AUC) of 79%. The same results were obtained when comparing PP values. Consequently, the use of combined linear and nonlinear parameters could improve the risk stratification of cardiomyopathy patients.

JTD Keywords: Feature extraction, Blood pressure, Heart rate, Estimation, Data mining, Covariance matrices, Hospitals

Aging population is a major concern that is reflected in the increase of chronic diseases. Heart Failure (HF) is one of the most common chronic diseases of elderly people that is punctuated with acute episodes, which result in hospitalization. The periodic modulation of the amplitude of the breathing pattern is proved to be one of the multiple symptoms of an acute episode, and thus, the features extracted from its characterization contribute in the improvement of the first diagnosis of the clinical practice. The main objective of this study is to evaluate if the features extracted from the breathing pattern along with common clinical variables are reliable enough to detect Periodic Breathing (PB). A dataset of 44 elderly patients containing clinical information and a short record of electrocardiogram and respiratory flow signal was used to train two machine learning classification methods: Support Vector Machine (SVM) and Linear Discriminant Analysis (LDA). All the available clinical parameters within the dataset along with the parameters characterizing the respiratory pattern were used to classify the observations into two groups. SVM classification was optimized and performed using a = -8 and C = 10.04 giving an accuracy of 88.2 % sensitivity of 90 % and specificity of 85.7 % Similar results were achieved with LDA classifying with an accuracy of 82.4 %, a sensitivity of 81.8% and specificity of 83.3 % PB has been accurately detected using both classifiers.

JTD Keywords: Support vector machines, Feature extraction, Training, Senior citizens, Standards, Training data, Hospitals

Adherent cells exert traction forces on their substrate, and these forces play important roles in biological functions such as mechanosensing, cell differentiation and cancer invasion. The method of choice to assess these active forces is traction force microscopy (TFM). Despite recent advances, TFM remains highly sensitive to measurement noise and exhibits limited spatial resolution. To improve the resolution and noise robustness of TFM, here we adapt techniques from compressed sensing (CS) to the reconstruction of the traction field from the substrate displacement field. CS enables the recovery of sparse signals at higher resolution from lower resolution data. Focal adhesions (FAs) of adherent cells are spatially sparse implying that traction fields are also sparse. Here we show, by simulation and by experiment, that the CS approach enables circumventing the Nyquist-Shannon sampling theorem to faithfully reconstruct the traction field at a higher resolution than that of the displacement field. This allows reaching state-of-the-art resolution using only a medium magnification objective. We also find that CS improves reconstruction quality in the presence of noise. Statement of Significance A great scientific advance of the past decade is the recognition that physical forces determine an increasing list of biological processes. Traction force microscopy which measures the forces that cells exert on their surroundings has seen significant recent improvements, however the technique remains sensitive to measurement noise and severely limited in spatial resolution. We exploit the fact that the force fields are sparse to boost the spatial resolution and noise robustness by applying ideas from compressed sensing. The novel method allows high resolution on a larger field of view. This may in turn allow better understanding of the cell forces at the multicellular level, which are known to be important in wound healing and cancer invasion.

JTD Keywords: Compressed sensing, High resolution, Traction force microscopy

Many cell therapies rely on the ability of mesenchymal stromal cells (MSCs) to diffuse and localize throughout the target tissue-such as tumoral and ischemic tissues-, in response to specific cytokine signals, rather than being concentrated at the site of implantation. Therefore, it is fundamental to engineer biomaterial carriers as reservoirs, from which cells can migrate, possibly in a controlled manner. In this work, microcarriers (μCs) made of polylactic acid are characterized as MSC delivery vehicles capable of modulating key chemotactic pathways. The effect of different functionalization strategies on MSC migratory behavior from the μCs is studied in vitro in relation to SDF-1α/CXCR4 axis,-a major actor in MSC recruitment, chemotaxis and homing. Collagen and arginine-glycine-aspartic acid (RGD) peptides were either covalently grafted or physisorbed on μC surface. While stable covalent modifications promoted better cell adhesion and higher proliferation compared to physisorption, the functionalization method of the μCs also affected the cells migratory behavior in response to SDF-1α (CXCL12) stimulation. Less stable coatings (physisorbed) showed sensibly higher number of migrating cells than covalent collagen/RGD coatings. The combination of physic-chemical cues provided by protein/peptide functionalization and stimuli induced by 3D culture on μCs improved MSC expression of CXCR4, and exerted a control over cell migration, a condition suitable to promote cell homing after transplantation in vivo. These are key findings to highlight the impact of surface modification approaches on chemokine-triggered cell release, and allow designing biomaterials for efficient and controlled cell delivery to damaged tissues.

JTD Keywords: Cell therapy, Chemotaxis, ECM (extracellular matrix), Mesenchymal stromal cells, Surface modification

Objective - Marfan's syndrome is characterized by the formation of ascending aortic aneurysms resulting from altered assembly of extracellular matrix microfibrils and chronic tissue growth factor (TGF)-β signaling. TGF-β is a potent regulator of the vascular smooth muscle cell (VSMC) phenotype. We hypothesized that as a result of the chronic TGF-β signaling, VSMC would alter their basal differentiation phenotype, which could facilitate the formation of aneurysms. This study explores whether Marfan's syndrome entails phenotypic alterations of VSMC and possible mechanisms at the subcellular level. Approach and Results - Immunohistochemical and Western blotting analyses of dilated aortas from Marfan patients showed overexpression of contractile protein markers (α-smooth muscle actin, smoothelin, smooth muscle protein 22 alpha, and calponin-1) and collagen I in comparison with healthy aortas. VSMC explanted from Marfan aortic aneurysms showed increased in vitro expression of these phenotypic markers and also of myocardin, a transcription factor essential for VSMC-specific differentiation. These alterations were generally reduced after pharmacological inhibition of the TGF-β pathway. Marfan VSMC in culture showed more robust actin stress fibers and enhanced RhoA-GTP levels, which was accompanied by increased focal adhesion components and higher nuclear localization of myosin-related transcription factor A. Marfan VSMC and extracellular matrix measured by atomic force microscopy were both stiffer than their respective controls. Conclusions - In Marfan VSMC, both in tissue and in culture, there are variable TGF-β-dependent phenotypic changes affecting contractile proteins and collagen I, leading to greater cellular and extracellular matrix stiffness. Altogether, these alterations may contribute to the known aortic rigidity that precedes or accompanies Marfan's syndrome aneurysm formation.

JTD Keywords: Actin, Aortic aneurysms, Aortic stiffness, Extracellular matrix, Focal adhesion, Myocardin, RhoA, TGF-β

Olfactory ensheathing cell (OEC) transplantation emerged some years ago as a promising therapeutic strategy to repair injured spinal cord. However, inhibitory molecules are present for long periods of time in lesioned spinal cord, inhibiting both OEC migration and axonal regrowth. Two families of these molecules, chondroitin sulphate proteoglycans (CSPG) and myelin-derived inhibitors (MAIs), are able to trigger inhibitory responses in lesioned axons. Mounting evidence suggests that OEC migration is inhibited by myelin. Here we demonstrate that OEC migration is largely inhibited by CSPGs and that inhibition can be overcome by the bacterial enzyme Chondroitinase ABC. In parallel, we have generated a stable OEC cell line overexpressing the Nogo receptor (NgR) ectodomain to reduce MAI-associated inhibition in vitro and in vivo. Results indicate that engineered cells migrate longer distances than unmodified OECs over myelin or oligodendrocyte-myelin glycoprotein (OMgp)-coated substrates. In addition, they also show improved migration in lesioned spinal cord. Our results provide new insights toward the improvement of the mechanisms of action and optimization of OEC-based cell therapy for spinal cord lesion.

JTD Keywords: Olfactory ensheathing cells, Traction force microscopy, Chondroitin sulphate proteoglycans, Cell migration, Nogo receptor ectodomain

Calcium signaling participates in different cellular processes leading to cell migration. TRPV4, a non-selective cation channel that responds to mechano-osmotic stimulation and heat, is also involved in cell migration. However, the mechanistic involvement of TRPV4 in cell migration is currently unknown. We now report that expression of the mutant channel TRPV4-121AAWAA (lacking the phosphoinositide-binding site 121KRWRK125 and the response to physiological stimuli) altered HEK293 cell migration. Altered migration patterns included periods of fast and persistent motion followed by periods of stalling and turning, and the extension of multiple long cellular protrusions. TRPV4-WT overexpressing cells showed almost complete loss of directionality with frequent turns, no progression, and absence of long protrusions. Traction microscopy revealed higher tractions forces in the tail of TRPV4-121AAWAA than in TRPV4-WT expressing cells. These results are consistent with a defective and augmented tail retraction in TRPV4-121AAWAA- and TRPV4-WT-expressing cells, respectively. The activity of calpain, a protease implicated in focal adhesion (FA) disassembly, was decreased in TRPV4-121AAWAA compared with TRPV4-WT-expressing cells. Consistently, larger focal adhesions were seen in TRPV4-121AAWAA compared with TRPV4-WT-expressing HEK293 cells, a result that was also reproduced in T47D and U87 cells. Similarly, overexpression of the pore-dead mutant TRPV4-M680D resumed the TRPV4-121AAWAA phenotype presenting larger FA. The migratory phenotype obtained in HEK293 cells overexpressing TRPV4-121AAWAA was mimicked by knocking-down TRPC1, a cationic channel that participates in cell migration. Together, our results point to the participation of TRPV4 in the dynamics of trailing adhesions, a function that may require the interplay of TRPV4 with other cation channels or proteins present at the FA sites.

JTD Keywords: Calcium, Calpain, Focal adhesion, Migration, Traction forces, TRPV4

Introduction. Selection of a biomaterial-based scaffold that mimics native myocardial extracellular matrix (ECM) architecture can facilitate functional cell attachment and differentiation. Although decellularized myocardial ECM accomplishes these premises, decellularization processes may variably distort or degrade ECM structure. Materials and methods. Two decellularization protocols (DP) were tested on porcine heart samples (epicardium, mid myocardium and endocardium). One protocol, DP1, was detergent-based (SDS and Triton X-100), followed by DNase I treatment. The other protocol, DP2, was focused in trypsin and acid with Triton X-100 treatments. Decellularized myocardial scaffolds were reseeded by embedding them in RAD16-I peptidic hydrogel with adipose tissue-derived progenitor cells (ATDPCs). Results. Both protocols yielded acellular myocardial scaffolds (~82% and ~94% DNA reduction for DP1 and DP2, respectively). Ultramicroscopic assessment of scaffolds was similar for both protocols and showed filamentous ECM with preserved fiber disposition and structure. DP1 resulted in more biodegradable scaffolds (P = 0.04). Atomic force microscopy revealed no substantial ECM stiffness changes post-decellularization compared to native tissue. The Young’s modulus did not differ between heart layers (P = 0.69) or decellularization protocols (P = 0.15). After one week, recellularized DP1 scaffolds contained higher cell density (236 ± 106 and 98 ± 56 cells/mm2 for recellularized DP1 and DP2 scaffolds, respectively; P = 0.04). ATDPCs in both DP1 and DP2 scaffolds expressed the endothelial marker isolectin B4, but only in the DP1 scaffold ATDPCs expressed the cardiac markers GATA4, connexin43 and cardiac troponin T. Conclusions. In our hands, DP1 produced myocardial scaffolds with higher cell repopulation and promotes ATDPCs expression of endothelial and cardiomyogenic markers.

JTD Keywords: Acellular myocardial scaffold, Adipose tissue-derived progenitor cells, Decellularization protocols, Extracellular matrix, Myocardial infarction, Recellularization

This work investigates the performance of cardiorespiratory analysis detecting periodic breathing (PB) in chest wall recordings in mountaineers climbing to extreme altitude. The breathing patterns of 34 mountaineers were monitored unobtrusively by inductance plethysmography, ECG and pulse oximetry using a portable recorder during climbs at altitudes between 4497 and 7546 m on Mt. Muztagh Ata. The minute ventilation (VE) and heart rate (HR) signals were studied, to identify visually scored PB, applying time-varying spectral, coherence and entropy analysis. In 411 climbing periods, 30–120 min in duration, high values of mean power (MPVE) and slope (MSlopeVE) of the modulation frequency band of VE, accurately identified PB, with an area under the ROC curve of 88 and 89 %, respectively. Prolonged stay at altitude was associated with an increase in PB. During PB episodes, higher peak power of ventilatory (MPVE) and cardiac (MP LF HR ) oscillations and cardiorespiratory coherence (MP LF Coher ), but reduced ventilation entropy (SampEnVE), was observed. Therefore, the characterization of cardiorespiratory dynamics by the analysis of VE and HR signals accurately identifies PB and effects of altitude acclimatization, providing promising tools for investigating physiologic effects of environmental exposures and diseases.

JTD Keywords: High-altitude periodic breathing, Cardiorespiratory characterization, Time-varying spectral analysis, Acclimatization, Hypoxia

In computer-assisted beating heart surgeries, accurate tracking of the heart's motion is of huge importance and there is a continuous need to eliminate any source of error that might disturb the tracking process. One source of error is the specular reflection that appears on the glossy surface of the heart. In this paper, we propose a robust solution for the detection and removal of specular highlights. A hybrid color attributes and wavelet based edge projection approach is applied to accurately identify the affected regions. These regions are then recovered using a dynamic search-based inpainting with adaptive windowing. Experimental results demonstrate the precision and efficiency of the proposed method. Moreover, it has a real-time performance and can be generalized to various other applications.

JTD Keywords: Heart, Image color analysis, Image edge detection, Surgery, Tracking, Wavelet transforms

In this paper, we propose a new approach to domestic action recognition based on a set of features which describe the relation between poses and movements of both hands. These features represent a set of basic actions in a kitchen in terms of the mimics of the hand movements, without needing information of the objects present in the scene. They address specifically the intra-class dissimilarity problem, which occurs when the same action is performed in different ways. The goal is to create a generic methodology that enables a robotic assistant system to recognize actions related to daily life activities and then, be endowed with a proactive behavior. The proposed system uses depth and color data acquired from a Kinect-style sensor and a hand tracking system. We analyze the relevance of the proposed hand-based features using a state-space search approach. Finally, we show the effectiveness of our action recognition approach using our own dataset.

JTD Keywords: Histograms, Joints, Robot sensing systems, Thumb, Tracking, Human activity recognition, Disable and elderly assistance

Abstract Fundamental biological processes including morphogenesis and tissue repair require cells to migrate collectively. In these processes, epithelial or endothelial cells move in a cooperative manner coupled by intercellular junctions. Ultimately, the movement of these multicellular systems occurs through the generation of cellular forces, exerted either on the substrate via focal adhesions (cell–substrate forces) or on neighboring cells through cell–cell junctions (cell–cell forces). Quantitative measurements of multicellular forces and kinematics with cellular or subcellular resolution have become possible only in recent years. In this chapter, we describe some of these techniques, which include particle image velocimetry to map cell velocities, traction force microscopy to map forces exerted by cells on the substrate, and monolayer stress microscopy to map forces within and between cells. We also describe experimental protocols to perform these measurements. The combination of these techniques with high-resolution imaging tools and molecular perturbations will lead to a better understanding of the mechanisms underlying collective cell migration in health and disease.

JTD Keywords: Collective cell migration, Monolayer stress microscopy, Traction force microscopy

Pose-based features have demonstrated to outperform low-levelappearance features in human action recognition. New RGB-D cameras provide locations of human joints with which geometric correspondences can be easily calculated. In this article, a new geometric correspondence between joints called Trisarea feature is presented. It is defined as the area of the triangle formed by three joints. Relevant triangles describing human pose are identified and it is shown how the variation over time of the selected Trisarea features constitutes a descriptor of human action. Experimental results show a comparison with other methods and demonstrate how this Trisarea-based representation can be applied to human action recognition.

JTD Keywords: Action descriptor, Action recognition, Pose-based feature

Xenopus tadpoles are an emerging model for developmental, genetic and behavioral studies. A small size, optical accessibility of most of their organs, together with a close genetic and structural relationship to humans make them a convenient experimental model. However, there is only a limited toolset available to measure behavior and organ function of these animals at medium or high-throughput. Herein, we describe an imaging-based platform to quantify body and autonomic movements of Xenopus tropicalis tadpoles of advanced developmental stages. Animals alternate periods of quiescence and locomotor movements and display buccal pumping for oxygen uptake from water and rhythmic cardiac movements. We imaged up to 24 animals in parallel and automatically tracked and quantified their movements by using image analysis software. Animal trajectories, moved distances, activity time, buccal pumping rates and heart beat rates were calculated and used to characterize the effects of test compounds. We evaluated the effects of propranolol and atropine, observing a dose-dependent bradycardia and tachycardia, respectively. This imaging and analysis platform is a simple, cost-effective high-throughput in vivo assay system for genetic, toxicological or pharmacological characterizations.

JTD Keywords: Xenopus tropicalis, Animal behavior, Cardiac imaging, Motion analysis, Animal tracking, Hhigh-throughput in vivo assay

The electric polarizability of DNA, represented by the dielectric constant, is a key intrinsic property that modulates DNA interaction with effector proteins. Surprisingly, it has so far remained unknown owing to the lack of experimental tools able to access it. Here, we experimentally resolved it by detecting the ultraweak polarization forces of DNA inside single T7 bacteriophages particles using electrostatic force microscopy. In contrast to the common assumption of low-polarizable behavior like proteins (εr ~ 2–4), we found that the DNA dielectric constant is ~ 8, considerably higher than the value of ~ 3 found for capsid proteins. State-of-the-art molecular dynamic simulations confirm the experimental findings, which result in sensibly decreased DNA interaction free energy than normally predicted by Poisson–Boltzmann methods. Our findings reveal a property at the basis of DNA structure and functions that is needed for realistic theoretical descriptions, and illustrate the synergetic power of scanning probe microscopy and theoretical computation techniques.

JTD Keywords: Atomic force microscopy, Atomistic simulations, DNA packaging, DNA-ligand binding, Poisson-Boltzmann equation, capsid protein, DNA, double stranded DNA, amino acid composition, article, atomic force microscopy, bacteriophage, bacteriophage T7, dielectric constant, dipole, DNA binding, DNA packaging, DNA structure, electron microscopy, ligand binding, nonhuman, polarization, priority journal, protein analysis, protein DNA interaction, scanning probe microscopy, static electricity, virion, virus capsid, virus particle, atomic force microscopy, atomistic simulations, DNA packaging, DNA-ligand binding, Poisson-Boltzmann equation, Bacteriophage T7, Capsid, Cations, Dielectric Spectroscopy, DNA, DNA, Viral, DNA-Binding Proteins, Electrochemical Techniques, Ligands, Microscopy, Atomic Force, Models, Chemical, Nuclear Proteins

In bone regeneration, silicon-based calcium phosphate glasses (Bioglasses) have been widely used since the 1970s. However, they dissolve very slowly because of their high amount of Si (SiO2 > 45%). Recently, our group has found that calcium ions released by the degradation of glasses in which the job of silicon is done by just 5% of TiO2 are effective angiogenic promoters, because of their stimulation of a cell-membrane calcium sensing receptor (CaSR). Based on this, other focused tests on angiogenesis have found that Bioglasses also have the potential to be angiogenic promoters even with high contents of silicon (80%); however, their slow degradation is still a problem, as the levels of silicon cannot be decreased any lower than 45%. In this work, we propose a new generation of hybrid organically modified glasses, ormoglasses, that enable the levels of silicon to be reduced, therefore speeding up the degradation process. Using electrospinning as a faithful way to mimic the extracellular matrix (ECM), we successfully produced hybrid fibrous mats with three different contents of Si (40, 52, and 70%), and thus three different calcium ion release rates, using an ormoglass–polycaprolactone blend approach. These mats offered a good platform to evaluate different calcium release rates as osteogenic promoters in an in vivo subcutaneous environment. Complementary data were collected to complement Ca2+ release analysis, such as stiffness evaluation by AFM, ζ-potential, morphology evaluation by FESEM, proliferation and differentiation analysis, as well as in vivo subcutaneous implantations. Material and biological characterization suggested that compositions of organic/inorganic hybrid materials with a Si content equivalent to 40%, which were also those that released more calcium, were osteogenic. They also showed a greater ability to form blood vessels. These results suggest that Si-based ormoglasses can be considered an efficient tool for calcium release modulation, which could play a key role in the angiogenic promoting process.

JTD Keywords: Biological materials, Blood vessels, Calcium, Electrospinning, Glass, Hybrid materials, Silicon oxides, Sol-gel process, Sol-gels, Angiogenesis, Biological characterization, Calcium phosphate glass, Calcium-sensing receptors, Degradation process, Extracellular matrices, Organic/inorganic hybrid materials, ormoglasses, Silicon

Bone is the main store of calcium and progenitor cells in the body. During the resorption process, the local calcium concentration reaches 8-40 mM, and the surrounding cells are exposed to these fluctuations in calcium. This stimulus is a signal that is detected through the calcium sensing receptor (CaSR), which modulates chemotactic and proliferative G protein-dependent signaling pathways. The objective of the present work is to evaluate the roles of extracellular calcium ([Ca2+]o) and the CaSR in osteoinduction. Rat bone marrow mesenchymal stromal cells (rBMSCs) were stimulated with 10 mM of Ca2+. Several experiments were conducted to demonstrate the effect of [Ca2+]o on chemotaxis, proliferation and differentiation on the osteoblastic lineage. It was found that [Ca2+]o induces rBMSCs to migrate and proliferate in a concentration-dependent manner. Real-time polymerase chain reaction and immunofluorescence also revealed that 10 mM Ca2+ stimulates overexpression of osteogenic markers in rBMSCs, including alkaline phosphatase (ALP), bone sialoprotein, collagen Ia1 and osteocalcin. Functional assays determining ALP activity and mineralization tests both corroborate the increased expression of these markers in rBMSCs stimulated with Ca2+. Moreover, CaSR blockage inhibited the cellular response to stimulation with high concentrations of [Ca2+]o, revealing that the CaSR is a key modulator of these cellular responses.

JTD Keywords: Calcium sensing receptor (CaSR), Extracellular calcium, Mesenchymal stromal cells (MSCs), Osteoinduction, Regenerative medicine

Infarcted hearts are macroscopically stiffer than healthy organs. Nevertheless, although cell behavior is mediated by the physical features of the cell niche, the intrinsic micromechanical properties of healthy and infarcted heart extracellular matrix (ECM) remain poorly characterized. Using atomic force microscopy, we studied ECM micromechanics of different histological regions of the left ventricle wall of healthy and infarcted mice. Hearts excised from healthy (n = 8) and infarcted mice (n = 8) were decellularized with sodium dodecyl sulfate and cut into 12 μm thick slices. Healthy ventricular ECM revealed marked mechanical heterogeneity across histological regions of the ventricular wall with the effective Young's modulus ranging from 30.2 ± 2.8 to 74.5 ± 8.7 kPa in collagen- and elastin-rich regions of the myocardium, respectively. Infarcted ECM showed a predominant collagen composition and was 3-fold stiffer than collagen-rich regions of the healthy myocardium. ECM of both healthy and infarcted hearts exhibited a solid-like viscoelastic behavior that conforms to two power-law rheology. Knowledge of intrinsic micromechanical properties of the ECM at the length scale at which cells sense their environment will provide further insight into the cell-scaffold interplay in healthy and infarcted hearts.

JTD Keywords: Atomic force microscopy, Extracellular matrix, Heart scaffold, Nanoindentation, Viscoelasticity

We have evaluated the behavior of single-walled carbon nanohorns as a sorbent for headspace and direct immersion (micro)solid phase extraction using volatile organic compounds (VOCs) as model analytes. The conical carbon nanohorns were first oxidized in order to increase their solubility in water and organic solvents. A microporous hollow polypropylene fiber served as a mechanical support that provides a high surface area for nanoparticle retention. The extraction unit was directly placed in the liquid sample or the headspace of an aqueous standard or a water sample to extract and preconcentrate the VOCs. The variables affecting extraction have been optimized. The VOCs were then identified and quantified by GC/MS. We conclude that direct immersion of the fiber is the most adequate method for the extraction of VOCs from both liquid samples and headspace. Detection limits range from 3.5 to 4.3 ng L-1 (excepted for toluene with 25 ng L-1), and the precision (expressed as relative standard deviation) is between 3.9 and 9.6 %. The method was applied to the determination of toluene, ethylbenzene, various xylene isomers and styrene in bottled, river and tap waters, and the respective average recoveries of spiked samples are 95.6, 98.2 and 86.0 %.

JTD Keywords: (Micro)solid phase extraction, Nanotechnology, Oxidized single-walled carbon nanohorns, Volatiles compounds, Waters

Lung disease models are useful to study how cell engraftment, proliferation and differentiation are modulated in lung bioengineering. The aim of this work was to characterize the local stiffness of decellularized lungs in aged and fibrotic mice. Mice (2- and 24-month old; 14 of each) with lung fibrosis (N=20) and healthy controls (N=8) were euthanized after 11 days of intratracheal bleomycin (fibrosis) or saline (controls) infusion. The lungs were excised, decellularized by a conventional detergent-based (sodium-dodecyl sulfate) procedure and slices of the acellular lungs were prepared to measure the local stiffness by means of atomic force microscopy. The local stiffness of the different sites in acellular fibrotic lungs was very inhomogeneous within the lung and increased according to the degree of the structural fibrotic lesion. Local stiffness of the acellular lungs did not show statistically significant differences caused by age. The group of mice most affected by fibrosis exhibited local stiffness that were ~2-fold higher than in the control mice: from 27.2±1.64 to 64.8±7.1. kPa in the alveolar septa, from 56.6±4.6 to 99.9±11.7. kPa in the visceral pleura, from 41.1±8.0 to 105.2±13.6. kPa in the tunica adventitia, and from 79.3±7.2 to 146.6±28.8. kPa in the tunica intima. Since acellular lungs from mice with bleomycin-induced fibrosis present considerable micromechanical inhomogeneity, this model can be a useful tool to better investigate how different degrees of extracellular matrix lesion modulate cell fate in the process of organ bioengineering from decellularized lungs.

JTD Keywords: Ageing, Atomic force microscopy, Decellularization, Lung fibrosis, Tissue engineering, Atomic force microscopy, Biological organs, Peptides, Sodium dodecyl sulfate, Sodium sulfate, Tissue engineering, Ageing, Decellularization, Extracellular matrices, Healthy controls, Inhomogeneities, Lung fibrosis, Micro-mechanical, Statistically significant difference, Mammals, bleomycin, adventitia, animal experiment, animal model, article, atomic force microscopy, bleomycin-induced pulmonary fibrosis, cell fate, controlled study, extracellular matrix, female, intima, lung alveolus, lung fibrosis, lung mechanics, mechanical probe, microenvironment, mouse, nonhuman, pleura, priority journal, rigidity, tissue engineering

Lung bioengineering using decellularized organ scaffolds is a potential alternative for lung transplantation. Clinical application will require donor scaffold sterilization. As gamma-irradiation is a conventional method for sterilizing tissue preparations for clinical application, the aim of this study was to evaluate the effects of lung scaffold sterilization by gamma irradiation on the mechanical properties of the acellular lung when subjected to the artificial ventilation maneuvers typical within bioreactors. Twenty-six mouse lungs were decellularized by a sodium dodecyl sulfate detergent protocol. Eight lungs were used as controls and 18 of them were submitted to a 31kGy gamma irradiation sterilization process (9 kept frozen in dry ice and 9 at room temperature). Mechanical properties of acellular lungs were measured before and after irradiation. Lung resistance (RL) and elastance (EL) were computed by linear regression fitting of recorded signals during mechanical ventilation (tracheal pressure, flow and volume). Static (Est) and dynamic (Edyn) elastances were obtained by the end-inspiratory occlusion method. After irradiation lungs presented higher values of resistance and elastance than before irradiation: RL increased by 41.1% (room temperature irradiation) and 32.8% (frozen irradiation) and EL increased by 41.8% (room temperature irradiation) and 31.8% (frozen irradiation). Similar increases were induced by irradiation in Est and Edyn. Scanning electron microscopy showed slight structural changes after irradiation, particularly those kept frozen. Sterilization by gamma irradiation at a conventional dose to ensure sterilization modifies acellular lung mechanics, with potential implications for lung bioengineering.

JTD Keywords: Gamma irradiation, Lung bioengineering, Lung decellularization, Organ scaffold, Pulmonary mechanics, Decellularization, Gamma irradiation, Mouse lung, Pulmonary mechanics, dodecyl sulfate sodium, animal tissue, Article, artificial ventilation, bioengineering, bioreactor, compliance (physical), controlled study, freezing, gamma irradiation, lung, lung mechanics, lung resistance, male, mouse, nonhuman, room temperature, scanning electron microscopy, tissue scaffold, trachea pressure

In this study gelatin (Gel) modified with calcium phosphate nanoparticles (SG5) and polycaprolactone (PCL) were used to prepare a 3D bi-layer scaffold by collecting electrospun PCL and gelatin/SG5 fibers separately in the same collector. The objective of this study was to combine the desired properties of PCL and Gel/SG5 in the same scaffold in order to enhance mineralization, thus improving the ability of the scaffold to bond to the bone tissue. The scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and the wide angle X-ray diffraction (WAXD) measurements confirmed that SG5 nanoparticles were successfully incorporated into the fibrous gelatin matrix. The composite Gel/SG5/PCL scaffold exhibited more enhanced mechanical properties than individual Gel and Gel/SG5 scaffolds. The presence of SG5 nanoparticles accelerated the nucleation and growth of apatite crystals on the surface of the composite Gel/SG5/PCL scaffold in simulated body fluid (SBF). The osteoblast response in vitro to developed electrospun scaffolds (PCL and Gel/SG5/PCL) was investigated by using normal human primary NHOst cell lines. NHOst cell culture studies showed that higher alkaline phosphatase (ALP) activity and better mineralization were obtained in the case of composite materials than in pure PCL scaffolds. The mechanically strong PCL scaffold served as a skeleton, while the Gel/SG5 fibers facilitated cell spreading and mineralization of the scaffold.

JTD Keywords: Bilayer fibrous scaffold, Ceramic nanoparticles, Electrospinning, Gelatin, Polycaprolactone, Biomechanics, Bone, Calcium phosphate, Cell culture, Electrospinning, Fourier transform infrared spectroscopy, Mechanical properties, Mineralogy, Nanoparticles, Phosphatases, Polycaprolactone, Scanning electron microscopy, X ray diffraction, Polycaprolactone, Alkaline phosphatase activity, Bone tissue engineering, Calcium phosphate nanoparticles, Ceramic nanoparticles, Fibrous scaffolds, Gelatin, Simulated body fluids, Wide-angle x-ray diffraction, Electrospuns, Scaffolds (biology), Electrospinning

Achieving high quality 3D-printed structures requires establishing the right printing conditions. Finding processing conditions that satisfy both the fabrication process and the final required scaffold properties is crucial. This work stresses the importance of studying the outcome of the plasticizing effect of PEG on PLA-based blends used for the fabrication of 3D-direct-printed scaffolds for tissue engineering applications. For this, PLA/PEG blends with 5, 10 and 20% (w/w) of PEG and PLA/PEG/bioactive CaP glass composites were processed in the form of 3D rapid prototyping scaffolds. Surface analysis and differential scanning calorimetry revealed a rearrangement of polymer chains and a topography, wettability and elastic modulus increase of the studied surfaces as PEG was incorporated. Moreover, addition of 10 and 20% PEG led to non-uniform 3D structures with lower mechanical properties. In vitro degradation studies showed that the inclusion of PEG significantly accelerated the degradation rate of the material. Results indicated that the presence of PEG not only improves PLA processing but also leads to relevant surface, geometrical and structural changes including modulation of the degradation rate of PLA-based 3D printed scaffolds.

JTD Keywords: 3D-printing, Polylactic acid, Rapid prototyping, Scaffold, Surface characterization

Lung decellularization is based on the use of physical, chemical, or enzymatic methods to break down the integrity of the cells followed by a treatment to extract the cellular material from the lung scaffold. The aim of this study was to characterize the mechanical changes throughout the different steps of lung decellularization process. Four lungs from mice (C57BL/6) were decellularized by using a conventional protocol based on sodium dodecyl sulfate. Lungs resistance (RL) and elastance (EL) were measured along decellularization steps and were computed by linear regression fitting of tracheal pressure, flow, and volume during mechanical ventilation. Transients differences found were more distinct in an intermediate step after the lungs were rinsed with deionized water and treated with 1% SDS, whereupon the percentage of variation reached approximately 80% for resistance values and 30% for elastance values. In conclusion, although a variation in extracellular matrix stiffness was observed during the decellularization process, this variation can be considered negligible overall because the resistance and elastance returned to basal values at the final decellularization step.

JTD Keywords: Lung bioengineering, Lung decellularization, Organ scaffold, dodecyl sulfate sodium, animal tissue, article, artificial ventilation, compliance (physical), controlled study, enzyme chemistry, extracellular matrix, female, flow, lung, lung decellularization, lung pressure, lung resistance, mouse, nonhuman, positive end expiratory pressure, priority journal, rigidity, tissue engineering, trachea pressure

In this work we report the fabrication of Langmuir and Langmuir-Blodgett (LB) films of a substituted ZnPc (octakis(oxyoctyl)phthalocyanine of zinc), and their characterization by means of several techniques. These characterization techniques include surface pressure (π-A) and surface potential (ΔV-A) isotherms as well as UV-vis Reflection spectroscopy and Brewster Angle Microscopy (BAM) for the films at the air-water interface together with UV-vis absorption and IR spectroscopies and Atomic Force Microscopy (AFM) for the LB films. The π-A and ΔV-A isotherms and BAM images indicate a phase transition at a surface pressure of ca. 9 mN/m and a multilayer formation at surface pressures around 19-20 mN/m; at a surface pressure around 27 mN/m a disordered collapse of the film occurs. In addition, AFM images of LB films at π = 10 mN/m and π = 20 mN/m show a monomolecular and a multilayered film, respectively. The comparison of the UV-vis spectrum of ZnPc in solution, the reflection spectra of the Langmuir films and UV-vis spectra of LB films reveals a significant reduction in the Q band intensity for the films, indicative of an organization of ZnPc in the Langmuir and LB films versus the random distribution in solution. The UV-vis Reflection spectra are also consistent with multilayer formation at surface pressures around 19-20 mN/m. The relative intensities of the IR spectrum bands change from the KBr pellet to the LB film which is also attributable to orientation effects in the film. Cyclic voltammetric experiments of LB films incorporating the ZnPc derivative show peaks that can be correlated with redox processes occurring in the phthalocyanine ring. A small but significant influence of the surface pressure and the number of deposited layers in the electrochemical behaviour is observed. The electrochemical response of cast films exhibits some differences with respect to that of LB films which have been attributed to their different molecular organizations.

JTD Keywords: Atomic Force Microscopy, Electrochemistry, Langmuir-Blodgett, Multilayers, Optical spectroscopy techniques, Zinc phthalocyanine, Atomic force microscopy, Electrochemistry, Interfaces (materials), Isotherms, Multilayers, Nitrogen compounds, Optical multilayers, Organic polymers, Zinc compounds, Brewster angle microscopy, Characterization techniques, Electrochemical behaviour, Langmuir and langmuir-blodgett films, Langmuir-blodgett, Optical spectroscopy techniques, UV-Vis Reflection Spectroscopy, Zinc phthalocyanines, Langmuir Blodgett films

Due to the increasing elderly population and the extensive number of comorbidities that affect them, studies are required to determine future increments in admission to emergency departments. Some of these studies could focus on the relation between chronic diseases and breathing pattern in elderly patients. Variations in the fractal properties of respiratory signals can be associated with several diseases. To determine the relationship between these variations and breathing patterns, and to quantify the fractal properties of respiratory flow signals, we estimated the Hurst exponent (H). Detrended fluctuation analysis (DFA) and discrete wavelet transform-based estimation (DWTE) methods were applied. The estimation methods were analyzed using simulated data series generated by fractional Gaussian noise. 43 elderly patients (19 patients with a non-periodic breathing pattern - nPB, and 24 patients with a periodic breathing pattern - PB) were studied. The results were evaluated according to the length of data and the number of averaged data series used to obtain a good estimation. The DWTE method estimated the respiratory flow signals better than the DFA method, and obtained Hurst values clustered by group. We found significant differences in the H exponent (p = 0.002) between PB and nPB patients, which showed different behavior in the fractal properties.

JTD Keywords: Discrete wavelet transforms, Diseases, Estimation, Fractals, Modulation, Senior citizens, Time series analysis

Robotic-assisted minimally invasive surgery has demonstrated its benefits in comparison with traditional procedures. However, one of the major drawbacks of current robotic system approaches is the lack of force feedback. Apart from space restrictions, the main problems of using force sensors are their high cost and the biocompatibility. In this work a proposal based on Vision Based Force Measurement is presented, in which the deformation mapping of the tissue is obtained using the `2−Regularized Optimization class, and the force is estimated via a recurrent neural network that has as inputs the kinematic variables and the deformation mapping. Moreover, the capability of RNN for predicting time series is used in order to deal with tool occlusions. The highlights of this proposal, according to the results, are: knowledge of material properties are not necessary, there is no need of adding extra sensors and a good trade-off between accuracy and efficiency has been achieved.

JTD Keywords: Force estimation, Regularized optimization, Deformable tracking, Recurrent neural network

Each bone of the skeleton constantly undergoes modeling during life to help it to adapt to changing biomechanical forces as well as remodeling to remove old bone and replace it with new, mechanically stronger bone to help preserve bone strength. Bone remodeling involves the removal of mineralized bone by osteoclasts, followed by the formation of bone matrix through the osteoblasts that subsequently become mineralized. All these assets make bone a suitable model for regeneration. Bone tissue can be grossly divided into inorganic mineral material (mostly HA), and organic material from cells and the extracellular matrix. This chapter outlines some of the bone diseases such as osteoporosis and Paget's disease. Bone can be considered as a biphasic composite material, with two phases: one the mineral and the other collagen. This combination confers better mechanical properties on the tissue than each component itself.

JTD Keywords: Bone biology, Bone cells, Bone diseases, Bone extracellular matrix, Bone mechanics, Bone remodeling, Bone tissue regeneration, Skeleton

Stroke is a major cause of disability, usually causing hemiplegic damage on the motor abilities of the patient. Stroke rehabilitation seeks restoring normal motion on the affected limb. However, normality’ of movements is usually assessed by clinical and functional tests, without considering how the motor system responds to therapy. We hypothesized that electromyographic (EMG) recordings could provide useful information for evaluating the outcome of rehabilitation from a neuromuscular perspective. Four healthy subjects were asked to perform 14 different functional movements simulating the action of reaching over a table. Each movement was defined according to the starting and target positions that the subject had to connect using linear trajectories. Bipolar recordings of EMG signals were taken from biceps and triceps muscles, and spectral and temporal characteristics were extracted for each movement. Using pattern recognition techniques we found that only two EMG channels were sufficient to accurately determine the spatial characteristics of motor activity: movement direction, length and execution zone. Our results suggest that muscles may fire in a patterned way depending on the specific characteristics of the movement and that EMG signals may codify such detailed information. These findings may be of great value to quantitatively assess post-stroke rehabilitation and to compare the neuromuscular activity of the affected and unaffected limbs, from a physiological perspective. Furthermore, disturbed movements could be characterized in terms of the muscle function to identify, which is the spatial characteristic that fails, e.g. movement direction, and guide personalized rehabilitation to enhance the training of such characteristic.

JTD Keywords: EMG, Movement spatial characteristics, Pattern recognition, Stroke rehabilitation, Upper-limb

This work introduces an interactive robotic system for assistance, conceived to tackle some of the challenges that domestic environments impose. The system is organized into a network of heterogeneous components that share both physical and logical functions to perform complex tasks. It consists of several robots for object manipulation, an advanced vision system that supplies in-formation about objects in the scene and human activity, and a spatial augmented reality interface that constitutes a comfortable means for interacting with the system. A first analysis based on users' experiences confirms the importance of having a friendly user interface. The inclusion of context awareness from visual perception enriches this interface allowing the robotic system to become a flexible and proactive assistant.

JTD Keywords: Accessibility, Activity Recognition, Ambient Intelligence, Human-Robot Interaction, Robot Assistance, Augmented reality, Complex networks, Computer vision, User interfaces, Accessibility, Activity recognition, Ambient intelligence, Domestic environments, Heterogeneous component, Interactive robotics, Robot assistance, Spatial augmented realities, Human assistance, Robotics

JTD Keywords: Optogenetics, Genetically encoded biosensors, Chloride imaging, pH monitoring, Intracellular Ca, Optopharmacology, Photochromic ligands, Photoswitchable tethered ligands

Control of membrane traffic: Photoswitchable inhibitors of protein-protein interactions were applied to photoregulate clathrin-mediated endocytosis (CME) in living cells. Traffic light (TL) peptides acting as "stop" and "go" signals for membrane traffic can be used to dissect the role of CME in receptor internalization and in cell growth, division, and differentiation.

JTD Keywords: Clathrin-mediated endocytosis, Optopharmacology, Peptides, Photoswitches, Protein-protein interactions

STUDY QUESTION Is the attachment of biofunctionalized polysilicon barcodes to the outer surface of the zona pellucida an effective approach for the direct tagging and identification of cultured embryos? SUMMARY ANSWER The results achieved provide a proof of concept for a direct embryo tagging system using biofunctionalized polysilicon barcodes, which could help to minimize the risk of mismatching errors (mix-ups) in human assisted reproduction technologies. WHAT IS KNOWN ALREADY Even though the occurrence of mix-ups is rare, several cases have been reported in fertility clinics around the world. Measures to prevent the risk of mix-ups in human assisted reproduction technologies are therefore required. STUDY DESIGN, SIZE, DURATION Mouse embryos were tagged with 10 barcodes and the effectiveness of the tagging system was tested during fresh in vitro culture (n=140) and after embryo cryopreservation (n = 84). Finally, the full-term development of tagged embryos was evaluated (n =105). PARTICIPANTS/ MATERIALS, SETTING, METHODS Mouse pronuclear embryos were individually rolled over wheat germ agglutinin-biofunctionalized polysilicon barcodes to distribute them uniformly around the ZONA PELLUCIDA surface. Embryo viability and retention of barcodes were determined during 96 h of culture. The identification of tagged embryos was performed every 24 h in an inverted microscope and without embryo manipulation to simulate an automatic reading procedure. Full-term development of the tagged embryos was assessed after their transfer to pseudo-pregnant females. To test the validity of the embryo tagging system after a cryopreservation process, tagged embryos were frozen at the 2-cell stage using a slow freezing protocol, and followed in culture for 72 h after thawing. MAIN RESULTS AND THE ROLE OF CHANCE Neither the in vitro or in vivo development of tagged embryos was adversely affected. The tagging system also proved effective during an embryo cryopreservation process. Global identification rates higher than 96 and 92% in fresh and frozen-thawed tagged embryos, respectively, were obtained when simulating an automatic barcode reading system, although these rates could be increased to 100% by simply rotating the embryos during the reading process. LIMITATIONS, REASONS FOR CAUTION The direct embryo tagging developed here has exclusively been tested in mouse embryos. Its effectiveness in other species, such as the human, is currently being tested. WIDER IMPLICATIONS OF THE FINDINGS The direct embryo tagging system developed here, once tested in human embryos, could provide fertility clinics with a novel tool to reduce the risk of mix-ups in human assisted reproduction technologies. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by Spanish Ministry of Education and Science (TEC2011-29140-C03) and by the Generalitat de Catalunya (2009SGR-00282).

JTD Keywords: Assisted reproductive technologies (ART), Biofunctionalization, Embryo tagging, Mix-ups, Traceability

In bone tissue engineering, the composition of the ionic extracellular environment (IEE) can determine both cellular fate and a biomaterial's development and performance. Therefore, precise control of the IEE and a perfect understanding of the dynamic changes that it can be subject to due to cellular activity is highly desired. To achieve this, we initially monitored how two standard osteoblast-like cell models that expressed either high or low alkaline phosphatase activity - SAOS-2 and MG63 cells, respectively - affected the extracellular concentrations of calcium and phosphate during long-term cultures. It was observed that cellular influence on the IEE varied greatly between the two models and could be linked to the capacity of cells to deposit calcium in the extracellular matrix. Miniaturized ion-selective electrodes that could allow for real-time monitoring of calcium in a minimally invasive way were then constructed. The electrodes were characterized in standard in vitro cell culture environments, prior to being successfully applied for periods of 24h, to record the dynamics of cell-induced deposition of calcium in the extracellular matrix, while using osteogenic media of either high or low concentrations of phosphate. As a result, this study provides the background and technological means for the non-destructive evaluation of the IEE in vitro and allows for the optimization and development of better models of bone tissue construction.

JTD Keywords: Extracellular ions, Ion-selective electrode, MG63, Mineralization, Osteoblasts, Saos-2, Sensor, Tissue engineering

Classification algorithms with unbalanced datasets tend to produce high predictive accuracy over the majority class, but poor predictive accuracy over the minority class. This problem is very common in biomedical data mining. This paper introduces a Support Vector Machine (SVM)-based optimized feature selection method, to select the most relevant features and maintain an accurate and well-balanced sensitivity–specificity result between unbalanced groups. A new metric called the balance index (B) is defined to implement this optimization. The balance index measures the difference between the misclassified data within each class. The proposed optimized feature selection is applied to the classification of patients' weaning trials from mechanical ventilation: patients with successful trials who were able to maintain spontaneous breathing after 48 h and patients who failed to maintain spontaneous breathing and were reconnected to mechanical ventilation after 30 min. Patients are characterized through cardiac and respiratory signals, applying joint symbolic dynamic (JSD) analysis to cardiac interbeat and breath durations. First, the most suitable parameters (C+,C−,σ) are selected to define the appropriate SVM. Then, the feature selection process is carried out with this SVM, to maintain B lower than 40%. The best result is obtained using 6 features with an accuracy of 80%, a B of 18.64%, a sensitivity of 74.36% and a specificity of 82.42%.

JTD Keywords: Support vector machines, Balance index, Sensitivity-specificity balance, Cardiorespiratory interaction, Joint symbolic dynamics, Feature selection, Weaning procedure

In this work, hybrid materials consisting on a vinylester matrix simultaneaously reinforced with jute woven fabrics and flyash particles were prepared. The tensile response and the fracture and failure behavior of these hybrid composites were investigated. Thermal stability of these materials was also studied. The aim was to obtain an environmentally friendly hybrid material with a good balance of tensile and fracture properties at relatively low cost. The effect of a novel treatment for the jute fabrics on the hybrids mechanical and fracture properties was investigated. The best balance of tensile and fracture properties was obtained for the hybrid consisting of fabrics treated with alkali under stress and fly ashes which also exhibited relatively high thermal stability.

JTD Keywords: Natural fibers, Fly ash, Hybrid composite, Mechanical properties, Fracture

Assessment of the dynamic interactions between cardiovascular signals can provide valuable information that improves the understanding of cardiovascular control. Heart rate variability (HRV) analysis is known to provide information about the autonomic heart rate modulation mechanism. Using the HRV signal, we aimed to obtain parameters for classifying patients with and without chronic heart failure (CHF), and with periodic breathing (PB), non-periodic breathing (nPB), and Cheyne-Stokes respiration (CSR) patterns. An electrocardiogram (ECG) and a respiratory flow signal were recorded in 36 elderly patients: 18 patients with CHF and 18 patients without CHF. According to the clinical criteria, the patients were classified into the follow groups: 19 patients with nPB pattern, 7 with PB pattern, 4 with Cheyne-Stokes respiration (CSR), and 6 non-classified patients (problems with respiratory signal). From the HRV signal, parameters in the time and frequency domain were calculated. Frequency domain parameters were the most discriminant in comparisons of patients with and without CHF: P_Tot (p = 0.02), P_LF (p = 0.022) and fp_HF (p = 0.021). For the comparison of the nPB vs. CSR patients groups, the best parameters were RMSSD (p = 0.028) and SDSD (p = 0.028). Therefore, the parameters appear to be suitable for enhanced diagnosis of decompensated CHF patients and the possibility of developed periodic breathing and a CSR pattern.

JTD Keywords: cardiovascular system, diseases, electrocardiography, frequency-domain analysis, geriatrics, medical signal processing, patient diagnosis, pneumodynamics, signal classification, Cheyne-Stokes respiration patterns, ECG, autonomic heart rate modulation mechanism, cardiovascular control, cardiovascular signals, chronic heart failure, decompensated CHF patients, dynamic interaction assessment, elderly patients, electrocardiogram, enhanced diagnosis, frequency domain parameters, heart rate variability analysis, patient classification, periodic breathing, respiratory flow signal recording, Electrocardiography, Frequency modulation, Frequency-domain analysis, Heart rate variability, Senior citizens, Standards

Biological olfaction outperforms chemical instrumentation in specificity, response time, detection limit, coding capacity, time stability, robustness, size, power consumption, and portability. This biological function provides outstanding performance due, to a large extent, to the unique architecture of the olfactory pathway, which combines a high degree of redundancy, an efficient combinatorial coding along with unmatched chemical information processing mechanisms. The last decade has witnessed important advances in the understanding of the computational primitives underlying the functioning of the olfactory system. EU Funded Project NEUROCHEM (Bio-ICT-FET- 216916) has developed novel computing paradigms and biologically motivated artefacts for chemical sensing taking inspiration from the biological olfactory pathway. To demonstrate this approach, a biomimetic demonstrator has been built featuring a large scale sensor array (65K elements) in conducting polymer technology mimicking the olfactory receptor neuron layer, and abstracted biomimetic algorithms have been implemented in an embedded system that interfaces the chemical sensors. The embedded system integrates computational models of the main anatomic building blocks in the olfactory pathway: The olfactory bulb, and olfactory cortex in vertebrates (alternatively, antennal lobe and mushroom bodies in the insect). For implementation in the embedded processor an abstraction phase has been carried out in which their processing capabilities are captured by algorithmic solutions. Finally, the algorithmic models are tested with an odour robot with navigation capabilities in mixed chemical plumes.

JTD Keywords: Antennal lobes, Artificial olfaction, Computational neuroscience, Olfactory bulbs, Plume tracking, Abstracting, Actuators, Algorithms, Biomimetic processes, Chemical sensors, Conducting polymers, Data processing, Flavors, Odors, Robots, Smart sensors, Embedded systems

Ischemic and dilated cardiomyopathy are associated with disorders of myocardium. Using the blood pressure (BP) signal and the values of the ventricular ejection fraction, we obtained parameters for stratifying cardiomyopathy patients as low- and high-risk. We studied 48 cardiomyopathy patients characterized by NYHA ≥2: 19 patients with dilated cardiomyopathy (DCM) and 29 patients with ischemic cardiomyopathy (ICM). The left ventricular ejection fraction (LVEF) percentage was used to classify patients in low risk (LR: LVEF > 35%, 17 patients) and high risk (HR: LVEF ≤ 35%, 31 patients) groups. From the BP signal, we extracted the upward systolic slope (BP_sl), the difference between systolic and diastolic BP (BPA), and systolic time intervals (STI). When we compared the LR and HR groups in the time domain analysis, the best parameters were standard deviation (SD) of 1=STI, kurtosis (K) of BP_sl, and K of BPA. In the frequency domain analysis, very low frequency (VLF) and high frequency (HF) bands showed statistically significant differences in comaprisons of LR and HR groups. The area under the curve of power spectral density was the best parameter in all classifications, and particularly in the very-low-and high- frequency bands (p <; 0.001). These parameters could help to improve the risk stratification of cardiomyopathy patients.

JTD Keywords: blood pressure measurement, cardiovascular system, diseases, medical disorders, medical signal processing, statistical analysis, time-domain analysis, BP signal, HR groups, LR groups, blood pressure signal analysis, cardiomyopathy patients, diastolic BP, dilated cardiomyopathy, frequency domain analysis, high-frequency bands, ischemic cardiomyopathy, left ventricular ejection fraction, low-frequency bands, myocardium disorders, patient characterization, power spectral density curve, standard deviation, statistical significant differences, systolic BP, systolic slope, systolic time intervals, time domain analysis, ventricular ejection fraction, Abstracts, Databases, Parameter extraction, Telecommunication standards, Time-frequency analysis

One of the most challenging problems in intensive care is still the process of discontinuing mechanical ventilation, called weaning process. Both an unnecessary delay in the discontinuation process and a weaning trial that is undertaken too early are undesirable. In this study, we analyzed respiratory pattern variability using the respiratory volume signal of patients submitted to two different levels of pressure support ventilation (PSV), prior to withdrawal of the mechanical ventilation. In order to characterize the respiratory pattern, we analyzed the following time series: inspiratory time, expiratory time, breath duration, tidal volume, fractional inspiratory time, mean inspiratory flow and rapid shallow breathing. Several autoregressive modeling techniques were considered: autoregressive models (AR), autoregressive moving average models (ARMA), and autoregressive models with exogenous input (ARX). The following classification methods were used: logistic regression (LR), linear discriminant analysis (LDA) and support vector machines (SVM). 20 patients on weaning trials from mechanical ventilation were analyzed. The patients, submitted to two different levels of PSV, were classified as low PSV and high PSV. The variability of the respiratory patterns of these patients were analyzed. The most relevant parameters were extracted using the classifiers methods. The best results were obtained with the interquartile range and the final prediction errors of AR, ARMA and ARX models. An accuracy of 95% (93% sensitivity and 90% specificity) was obtained when the interquartile range of the expiratory time and the breath duration time series were used a LDA model. All classifiers showed a good compromise between sensitivity and specificity.

JTD Keywords: autoregressive moving average processes, feature extraction, medical signal processing, patient care, pneumodynamics, signal classification, support vector machines, time series, ARX, autoregressive modeling techniques, autoregressive models with exogenous input, autoregressive moving average model, breath duration time series, classification method, classifier method, discontinuing mechanical ventilation, expiratory time, feature extraction, final prediction errors, fractional inspiratory time, intensive care, interquartile range, linear discriminant analysis, logistic regression analysis, mean inspiratory flow, patient respiratory volume signal, pressure support level, pressure support ventilation, rapid shallow breathing, respiratory pattern variability characterization, support vector machines, tidal volume, weaning trial, Analytical models, Autoregressive processes, Biological system modeling, Estimation, Support vector machines, Time series analysis, Ventilation

This work investigates the very low frequency (VLF) modulation of QRS slopes and heart rate variability (HRV). Electrocardiogram (ECG) and respiratory flow signal were acquired from patients with dilated cardiomyopathy and ischemic cardiomyopathy. HRV as well as the upward QRS slope (I_US) and downward QRS slope (I_DS) were extracted from the ECG. The relation between HRV and QRS slopes in the VLF band was measured using ordinary coherence in 5-minute segments. Partial coherence was then used to remove the influence that respiration simultaneously exerts on HRV and QRS slopes. A statistical threshold was determined, below which coherence values were considered not to represent a linear relation. 7 out of 276 segments belonging to 5 out of 29 patients for I_US and 10 segments belonging to 5 patients for I_DS presented a VLF modulation in QRS slopes, HRV and respiration. In these segments spectral coherence was statistically significant, while partial coherence decreased, indicating that the coupling HRV and QRS slopes was related to respiration. 4 segments had a partial coherence value below the threshold for I_US, 3 segments for I_DS. The rest of the segments also presented a notable decrease in partial coherence, but still above the threshold, which means that other non-linearly effects may also affect this modulation.

JTD Keywords: diseases, electrocardiography, feature extraction, medical signal processing, pneumodynamics, statistical analysis, ECG, QRS slopes, cardiomyopathy patients, dilated cardiomyopathy, electrocardiogram, feature extraction, heart rate variability, ischemic cardiomyopathy, ordinary coherence, partial coherence value, respiration, respiratory flow signal acquisition, spectral coherence, statistical threshold, time 5 min, very low frequency modulation, Coherence, Educational institutions, Electrocardiography, Frequency modulation, Heart rate variability

Rat model of Obstructive Sleep Apnea (OSA) is a realistic approach for studying physiological mechanisms involved in sleep. Rats are usually anesthetized and autonomic nervous system (ANS) could be blocked. This study aimed to assess the effect of anesthesia on ANS activity during OSA episodes. Seven male Sprague-Dawley rats were anesthetized intraperitoneally with urethane (1g/kg). The experiments were conducted applying airway obstructions, simulating 15s-apnea episodes for 15 minutes. Five signals were acquired: respiratory pressure and flow, SaO2, ECG and photoplethysmography (PPG). In total, 210 apnea episodes were studied. Normalized power spectrum of Pulse Rate Variability (PRV) was analyzed in the Low Frequency (LF) and High Frequency (HF) bands, for each episode in consecutive 15s intervals (before, during and after the apnea). All episodes showed changes in respiratory flow and SaO₂ signal. Conversely, decreases in the amplitude fluctuations of PPG (DAP) were not observed. Normalized LF presented extremely low values during breathing (median=7,67%), suggesting inhibition of sympathetic system due to anesthetic effect. Subtle increases of LF were observed during apnea. HRV and PPG analysis during apnea could be an indirect tool to assess the effect and deep of anesthesia.

JTD Keywords: electrocardiography, fluctuations, medical disorders, medical signal detection, medical signal processing, neurophysiology, photoplethysmography, pneumodynamics, sleep, ECG, SaO₂ flow, SaO₂ signal, airway obstructions, amplitude fluctuations, anesthesia effects, anesthetized nervous system, autonomic evaluation, autonomic nervous system, breathing, heart rate variability, high-frequency bands, low-frequency bands, male Sprague-Dawley rats, normalized power spectrum, obstructive sleep apnea, photoplethysmography, physiological mechanisms, pulse rate variability, rat model, respiratory flow, respiratory pressure, signal acquisition, sympathetic system inhibition, time 15 min, time 15 s, Abstracts, Atmospheric modeling, Computational modeling, Electrocardiography, Rats, Resonant frequency

Some of the most common clinical problems in elderly patients are related to diseases of the cardiac and respiratory systems. Elderly patients often have altered breathing patterns, such as periodic breathing (PB) and Cheyne-Stokes respiration (CSR), which may coincide with chronic heart failure. In this study, we used the envelope of the respiratory flow signal to characterize respiratory patterns in elderly patients. To study different breathing patterns in the same patient, the signals were segmented into windows of 5 min. In oscillatory breathing patterns, frequency and time-frequency parameters that characterize the discriminant band were evaluated to identify periodic and non-periodic breathing (PB and nPB). In order to evaluate the accuracy of this characterization, we used a feature selection process, followed by linear discriminant analysis. 22 elderly patients (7 patients with PB and 15 with nPB pattern) were studied. The following classification problems were analyzed: patients with either PB (with and without apnea) or nPB patterns, and patients with CSR versus PB, CSR versus nPB and PB versus nPB patterns. The results showed 81.8% accuracy in the comparisons of nPB and PB patients, using the power of the modulation peak. For the segmented signal, the power of the modulation peak, the frequency variability and the interquartile ranges provided the best results with 84.8% accuracy, for classifying nPB and PB patients.

JTD Keywords: cardiovascular system, diseases, feature extraction, geriatrics, medical signal processing, oscillations, pneumodynamics, signal classification, time-frequency analysis, Cheyne-Stokes respiration, apnea, cardiac systems, chronic heart failure, classification problems, discriminant band, diseases, elderly patients, feature selection process, frequency variability, interquartile ranges, linear discriminant analysis, nonperiodic breathing, oscillatory breathing pattern, periodic breathing, respiratory How signal, respiratory systems, signal segmentation, time 5 min, time-frequency parameters, Accuracy, Aging, Frequency modulation, Heart, Senior citizens, Time-frequency analysis

Even if the performance of a given ventilator has been evaluated in the laboratory under very well controlled conditions, inappropriate maintenance and lack of long-term stability and accuracy of the ventilator sensors may lead to ventilation errors in actual clinical practice. The aim of this study was to evaluate the actual performances of ventilators during clinical routines. A resistance (7.69 cmH(2)O/L/s) - elastance (100 mL/cmH(2)O) test lung equipped with pressure, flow, and oxygen concentration sensors was connected to the Y-piece of all the mechanical ventilators available for patients in four intensive care units (ICUs; n = 66). Ventilators were set to volume-controlled ventilation with tidal volume = 600 mL, respiratory rate = 20 breaths/minute, positive end-expiratory pressure (PEEP) = 8 cmH(2)O, and oxygen fraction = 0.5. The signals from the sensors were recorded to compute the ventilation parameters. The average standard deviation and range (min-max) of the ventilatory parameters were the following: inspired tidal volume = 607 36 (530-723) mL, expired tidal volume = 608 36 (530-728) mL, peak pressure = 20.8 2.3 (17.2-25.9) cmH(2)O, respiratory rate = 20.09 0.35 (19.5-21.6) breaths/minute, PEEP = 8.43 0.57 (7.26-10.8) cmH(2)O, oxygen fraction = 0.49 0.014 (0.41-0.53). The more error-prone parameters were the ones related to the measure of flow. In several cases, the actual delivered mechanical ventilation was considerably different from the set one, suggesting the need for improving quality control procedures for these machines.

JTD Keywords: Equipment and supplies, Medical devices, Intravenous, Quality assurance, Health care quality assessment, Ventilator accuracy, Ventilation error

Label-free detection of the material composition of nanoparticles could be enabled by the quantification of the nanoparticles’ inherent dielectric response to an applied electric field. However, the sensitivity of dielectric nanoscale objects to geometric and non-local effects makes the dielectric response extremely weak. Here we show that electrostatic force microscopy with sub-piconewton resolution can resolve the dielectric constants of single dielectric nanoparticles without the need for any reference material, as well as distinguish nanoparticles that have an identical surface but different inner composition. We unambiguously identified unlabelled ~10unm nanoparticles of similar morphology but different low-polarizable materials, and discriminated empty from DNA-containing virus capsids. Our approach should make the in situ characterization of nanoscale dielectrics and biological macromolecules possible.

JTD Keywords: Biological materials, Nanoscale materials, Characterisation and analytical techniques, Computation, modelling and theory

Newly generated olfactory receptor axons grow from the peripheral to the central nervous system aided by olfactory ensheathing cells (OECs). Thus, OEC transplantation has emerged as a promising therapy for spinal cord injuries and for other neural diseases. However, these cells do not present a uniform population, but instead a functionally heterogeneous population that exhibits a variety of responses including adhesion, repulsion, and crossover during cell–cell and cell–matrix interactions. Some studies report that the migratory properties of OECs are compromised by inhibitory molecules and potentiated by chemical gradients. Here, we demonstrated that rodent OECs express all the components of the Nogo receptor complex and that their migration is blocked by myelin. Next, we used cell tracking and traction force microscopy to analyze OEC migration and its mechanical properties over myelin. Our data relate the decrease of traction force of OEC with lower migratory capacity over myelin, which correlates with changes in the F-actin cytoskeleton and focal adhesion distribution. Lastly, OEC traction force and migratory capacity is enhanced after cell incubation with the Nogo receptor inhibitor NEP1-40.

JTD Keywords: Ensheathing glia, Traction force, microscopy, Migration, Myelin-associated inhibitors

Rapid and inexpensive development of electrochemical sensors with good exploitation potential may be produced using a polymer as a substrate material. However, fabrication of polymer-based sensors is challenging. Using photolithography and etching of gold-coated poly(ethylene-2,6-naphthalate) (PEN), we have succeeded in fabricating disk-shaped and interdigitated microelectrodes (uEs). The electrodes have an excellent adhesion to the polymer and are encapsulated using a novel room-temperature process, applicable for low-cost, high-throughput fabrication. The PEN surface has been characterized in respect of wettability, surface energy and surface roughness. Finally, the electrodes give stable and reproducible electrochemical impedance spectroscopy and cyclic voltammetry responses, using the redox couple ferrocyanide and ruthenium hexamine. The results demonstrate the robustness and functionality of the polymer-based sensor platform with minimum feature sizes of 6 um.

JTD Keywords: Poly(ethylene naphthalate), Photolithography, Microelectrodes, Interdigitated electrodes, Electrochemical characterization, Electrochemical sensor

This article reports on the research and development of a cutting-edge biomedical device for continuous in-vivo glucose monitoring. This entirely public-funded process of technological innovation has been conducted at the University of Barcelona within a context of converging technologies involving the fields of medicine, physics, chemistry, biology, telecommunications, electronics and energy. The authors examine the value chain and the market challenges faced by in-vivo implantable biomedical devices based on nanotechnologies. In so doing, they trace the process from the point of applied research to the final integration and commercialization of the product, when the social rate of return from academic research can be estimated. Using a case-study approach, the paper also examines the high-tech activities involved in the development of this nano-enabled device and describes the technology and innovation management process within the value chain conducted in a University-Hospital-Industry-Administration-Citizens framework. Here, nanotechnology is seen to represent a new industrial revolution, boosting the biomedical devices market. Nanosensors may well provide the tools required for investigating biological processes at the cellular level in vivo when embedded into medical devices of small dimensions, using biocompatible materials, and requiring reliable and targeted biosensors, high speed data transfer, safely stored data, and even energy autonomy.

JTD Keywords: Biomedical device, Diabetes, Innovation management, Nanobiosensor, Nanotechnology, Research commercialization, Technology transfer, Academic research, Applied research, Barcelona, Biocompatible materials, Biological process, Biomedical analysis, Biomedical devices, Cellular levels, Converging technologies, Glucose monitoring, High-speed data transfer, Implantable biomedical devices, Implantable devices, In-vivo, Industrial revolutions, Innovation management, Medical Devices, Nanobiosensor, Rate of return, Research and development, Technological innovation, Value chains, Biological materials, Biomedical engineering, Biosensors, Commerce, Data transfer, Earnings, Engineering education, Glucose, Implants (surgical), Industrial research, Innovation, Medical problems, Nanosensors, Nanotechnology, Technology transfer, Equipment