Publications

The directed migration of epithelial cell collectives through coordinated movements plays a crucial role in various physiological processes and is increasingly understood at the level of large confluent monolayers. However, numerous processes rely on the migration of small groups of polarized epithelial clusters in complex environments, and their responses to external geometries remain poorly understood. To address this, we cultivate primary epithelial keratocyte tissues on adhesive microstripes to create autonomous epithelial clusters with well-defined geometries. We show that their migration efficiency is strongly influenced by the contact geometry and the orientation of cell-cell contacts with respect to the direction of migration. A combination of velocity and polarity alignment with contact regulation of locomotion in an active matter model captures quantitatively the experimental data. Furthermore, we predict that this combination of rules enables efficient navigation in complex geometries, which we confirm experimentally. Altogether, our findings provide a conceptual framework for extracting the interaction rules of active systems from their interaction with physical boundaries, as well as design principles for collective navigation in complex microenvironments. The collective migration of cell clusters is modulated by substrate geometry through a combination of velocity and polarity alignment.

JTD Keywords: Collective migration, Complex, Forces, Mode

Steroid myopathy is a clinically challenging condition exacerbated by prolonged corticosteroid use or adrenal tumors. In this study, we engineered a functional three-dimensional (3D) in vitro skeletal muscle model to investigate steroid myopathy. By subjecting our bioengineered muscle tissues to dexamethasone treatment, we reproduced the molecular and functional aspects of this disease. Dexamethasone caused a substantial reduction in muscle force, myotube diameter and induced fatigue. We observed nuclear translocation of the glucocorticoid receptor (GCR) and activation of the ubiquitin-proteasome system within our model, suggesting their coordinated role in muscle atrophy. We then examined the therapeutic potential of taurine in our 3D model for steroid myopathy. Our findings revealed an upregulation of phosphorylated AKT by taurine, effectively countering the hyperactivation of the ubiquitin- proteasomal pathway. Importantly, we demonstrate that discontinuing corticosteroid treatment was insufficient to restore muscle mass and function. Taurine treatment, when administered concurrently with corticosteroids, notably enhanced contractile strength and protein turnover by upregulating the AKT-mTOR axis. Our model not only identifies a promising therapeutic target, but also suggests combinatorial treatment that may benefit individuals undergoing corticosteroid treatment or those diagnosed with adrenal tumors.

JTD Keywords: 3d bioengineered skeletal muscle tissues, Adrenal cortex hormones, Atroph, Colocalization, Corticosteroids, Dexamethasone, Glucocorticoid-receptor, Humans, Mechanisms, Models, biological, Mtor protein, human, Muscle contraction, Muscle fibers, skeletal, Muscle strength, Muscle, skeletal, Muscular diseases, Organ size, Phosphorylation, Proteasome endopeptidase complex, Proto-oncogene proteins c-akt, Receptors, glucocorticoid, Signal transduction, Skeletal-muscle, Steroid myopathy, Steroids, Supplementation, Taurin, Taurine, Tor serine-threonine kinases, Ubiquitin

Over the past decades, the development of nanoparticles (NPs) to increase the efficiency of clinical treatments has been subject of intense research. Yet, most NPs have been reported to possess low efficacy as their actuation is hindered by biological barriers. For instance, synovial fluid (SF) present in the joints is mainly composed of hyaluronic acid (HA). These viscous media pose a challenge for many applications in nanomedicine, as passive NPs tend to become trapped in complex networks, which reduces their ability to reach the target location. This problem can be addressed by using active NPs (nanomotors, NMs) that are self-propelled by enzymatic reactions, although the development of enzyme-powered NMs, capable of navigating these viscous environments, remains a considerable challenge. Here, the synergistic effects of two NMs troops, namely hyaluronidase NMs (HyaNMs, Troop 1) and urease NMs (UrNMs, Troop 2) are demonstrated. Troop 1 interacts with the SF by reducing its viscosity, thus allowing Troop 2 to swim more easily through the SF. Through their collective motion, Troop 2 increases the diffusion of macromolecules. These results pave the way for more widespread use of enzyme-powered NMs, e.g., for treating joint injuries and improving therapeutic effectiveness compared with traditional methods. The conceptual idea of the novel approach using hyaluronidase NMs (HyaNMs) to interact with and reduce the viscosity of the synovial fluid (SF) and urease NMs (UrNMs) for a more efficient transport of therapeutic agents in joints.image

JTD Keywords: Biological barrier, Clinical research, Clinical treatments, Collective motion, Collective motion,nanomotors,nanorobots,swarming,viscous medi, Collective motions, Complex networks, Enzymatic reaction, Enzymes, Hyaluronic acid, Hyaluronic-acid,ph,viscoelasticity,adsorption,barriers,behavior,ureas, Macromolecules, Medical nanotechnology, Nano robots, Nanomotors, Nanorobots, Swarming, Synovial fluid, Target location, Viscous media, Viscous medium

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

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, Actins, Bar proteins, Cell biology, Cell membrane, Homeostasis, Human, Mechanobiology, Membrane biophysics, Mouse, Physics of living systems

Live cell actin visualization is fundamental for exploring cellular motility, cytokinesis, intracellular transport, and other correlated functions. The current imaging techniques that allow imaging of actin in its native environment are optical and electron microscopy. Such imaging techniques offer high enough resolution to investigate the ultrastructure of actin however they come at the expense of actin integrity. Inspired by the lack of suitable probes that preserve actin's integrity, we designed a cyclometalated Ir (III) complex that interacts with live cells and displays light switch behaviour upon specific actin binding. The exceptional photophysical properties of the proposed probe allow unprecedented resolution of cytoskeleton ultrastructures under stimulated emission depletion (STED) super-resolution nanoscopy. Moreover, the Ir complex enables the capability of visualizing actin polymers and periodicity under correlative light electron microscopy (CLEM) and liquid-phase electron microscopy (LPEM) at similar to 8 nm resolution.

JTD Keywords: Actin dynamics, Actin targeting, Adhesion, Cells, Clem, Fluorescent, Iridium (iii) complex, Lead, Light, Lpem, Super-resolution ultrastructures

The endosomal sorting complex required for transport (ESCRT) is a multi-protein machinery involved in several membrane remodeling processes. Different approaches have been used to resolve how ESCRT proteins scission membranes. However, the underlying mechanisms generating membrane deformations are still a matter of debate. Here, giant unilamellar vesicles, microfluidic technology, and micropipette aspiration are combined to continuously follow the ESCRT-III-mediated membrane remodeling on the single-vesicle level for the first time. With this approach, we identify different mechanisms by which a minimal set of three ESCRT-III proteins from Entamoeba histolytica reshape the membrane. These proteins modulate the membrane stiffness and spontaneous curvature to regulate bud size and generate intraluminal vesicles even in the absence of ATP. We demonstrate that the bud stability depends on the protein concentration and membrane tension. The approaches introduced here should open the road to diverse applications in synthetic biology for establishing artificial cells with several membrane compartments.© 2022 The Author(s).

JTD Keywords: bilayer, curvature, diffusion-coefficients, identification, membrane-scission, phase-diagram, reveals, sorting complex, structural basis, Biophysics, Biotechnology, Cell biology, Giant vesicles, Membranes

Folding a polymer chain into a well-defined single-chain polymeric nanoparticle (SCPN) is a fascinating approach to obtaining structured and functional nanoparticles. Like all polymeric materials, SCPNs are heterogeneous in their nature due to the polydispersity of their synthesis: the stochastic synthesis of polymer backbone length and stochastic functionalization with hydrophobic and hydrophilic pendant groups make structural diversity inevitable. Therefore, in a single batch of SCPNs, nanoparticles with different physicochemical properties are present, posing a great challenge to their characterization at a single-particle level. The development of techniques that can elucidate differences between SCPNs at a single-particle level is imperative to capture their potential applications in different fields such as catalysis and drug delivery. Here, a Nile Red based spectral point accumulation for imaging in nanoscale topography (NR-sPAINT) super-resolution fluorescence technique was implemented for the study of SCPNs at a single-particle level. This innovative method allowed us to (i) map the small-molecule binding rates on individual SCPNs and (ii) map the polarity of individual SCPNs for the first time. The SCPN designs used here have the same polymeric backbone but differ in the number of hydrophobic groups. The experimental results show notable interparticle differences in the binding rates within the same polymer design. Moreover, a marked polarity shift between the different designs is observed. Interestingly, interparticle polarity heterogeneity was unveiled, as well as an intraparticle diversity, information which has thus far remained hidden by ensemble techniques. The results indicate that the addition of hydrophobic pendant groups is vital to determine binding properties and induces single-particle polarity diversity. Overall, NR-sPAINT represents a powerful approach to quantifying the single-particle polarity of SCPNs and paves the way to relate the structural heterogeneity to functionality at the single-particle level. This provides an important step toward the aim of rationally designing SCPNs for the desired application.

JTD Keywords: binding, complex, Microscopy

It has been recently shown that electron transfer between mitochondrial cytochrome c and the cytochrome c1 subunit of the cytochrome bc1 can proceed at long-distance through the aqueous solution. Cytochrome c is thought to adjust its activity by changing the affinity for its partners via Tyr48 phosphorylation, but it is unknown how it impacts the nanoscopic environment, interaction forces, and long-range electron transfer. Here, we constrain the orientation and separation between cytochrome c1 and cytochrome c or the phosphomimetic Y48pCMF cytochrome c, and deploy an array of single-molecule, bulk, and computational methods to investigate the molecular mechanism of electron transfer regulation by cytochrome c phosphorylation. We demonstrate that phosphorylation impairs long-range electron transfer, shortens the long-distance charge conduit between the partners, strengthens their interaction, and departs it from equilibrium. These results unveil a nanoscopic view of the interaction between redox protein partners in electron transport chains and its mechanisms of regulation.© 2022. The Author(s).

JTD Keywords: apoptosis, binding, cardiolipin, complex, dynamics, force, respiration, structural basis, tyrosine phosphorylation, Histone chaperone activity

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

Supramolecular chemistry is the quintessential backbone of all biological processes. It encompasses a wide range from the metabolic network to the self-assembled cytoskeletal network. Combining the chemical diversity with the plethora of functional depth that biological systems possess is a daunting task for synthetic chemists to emulate. The only route for approaching such a challenge lies in understanding the complex and dynamic systems through advanced analytical techniques. The supramolecular complexity that can be successfully generated and analyzed is directly dependent on the analytical treatment of the system parameters. In this review, we illustrate advanced analytical techniques that have been used to investigate various supramolecular systems including complex mixtures, dynamic self-assembly, and functional nanomaterials. The underlying theme of such an overview is not only the exceeding detail with which traditional experiments can be probed but also the fact that complex experiments can now be attempted owing to the analytical techniques that can resolve an ensemble in astounding detail. Furthermore, the review critically analyzes the current state of the art analytical techniques and suggests the direction of future development. Finally, we envision that integrating multiple analytical methods into a common platform will open completely new possibilities for developing functional chemical systems.

JTD Keywords: analytical techniques, dynamic self-assembly, high-speed afm, liquid cell tem, Analytical technique, Analytical techniques, Biological process, Chemical analysis, Chemical diversity, Complex networks, Cytoskeletal network, Dynamic self-assembly, High-speed afm, Hydrogels, In-situ, Liquid cell tem, Metabolic network, Microscopy, Nanoscale, Proteins, Self assembly, Supramolecular chemistry, Supramolecular systems, System chemistry, Systems chemistry

Cell nuclei are submitted to mechanical forces, which in turn affect nuclear and cell functions. Recent evidence shows that a crucial mechanically regulated nuclear function is nucleocytoplasmic transport, mediated by nuclear pore complexes (NPCs). Mechanical regulation occurs at two levels: first, by force application to the nucleus, which increases NPC permeability likely through NPC stretch. Second, by the mechanical properties of the transported proteins themselves, as mechanically labile proteins translocate through NPCs faster than mechanically stiff ones. In this perspective, we discuss this evidence and the associated mechanisms by which mechanics can regulate the nucleo-cytoplasmic partitioning of proteins. Finally, we analyze how mechanical regulation of nucleocytoplasmic transport can provide a systematic approach to the study of mechanobiology and open new avenues both in fundamental and applied research. (C) 2022 Author(s).

JTD Keywords: Architecture, Association, Force, Nuclear-pore complex, Pathways, Protein import, Sun1

Mechanical force controls fundamental cellular processes in health and disease, and increasing evidence shows that the nucleus both experiences and senses applied forces. Such forces can lead to the nuclear translocation of proteins, but whether force controls nucleocytoplasmic transport, and how, remains unknown. Here we show that nuclear forces differentially control passive and facilitated nucleocytoplasmic transport, setting the rules for the mechanosensitivity of shuttling proteins. We demonstrate that nuclear force increases permeability across nuclear pore complexes, with a dependence on molecular weight that is stronger for passive than for facilitated diffusion. Owing to this differential effect, force leads to the translocation of cargoes into or out of the nucleus within a given range of molecular weight and affinity for nuclear transport receptors. Further, we show that the mechanosensitivity of several transcriptional regulators can be both explained by this mechanism and engineered exogenously by introducing appropriate nuclear localization signals. Our work unveils a mechanism of mechanically induced signalling, probably operating in parallel with others, with potential applicability across signalling pathways.; Andreu et al. show that force regulates nucleocytoplasmic transport by weakening the permeability barrier of nuclear pore complexes, affecting passive and facilitated diffusion in different ways.

JTD Keywords: Activation, Inhibitor, Matrix, Mechanotransduction, Nesprins, Nucleoporins, Permeability barrier, Pore complex, Proteins, Transmission

The construction of biomembranes that faithfully capture the properties and dynamic functions of cell membranes remains a challenge in the development of synthetic cells and their application. Here a new concept for synthetic cell membranes based on the self-assembly of amphiphilic comb polymers into vesicles, termed ionic combisomes (i-combisomes) is introduced. These combs consist of a polyzwitterionic backbone to which hydrophobic tails are linked by electrostatic interactions. Using a range of microscopies and molecular simulations, the self-assembly of a library of combs in water is screened. It is discovered that the hydrophobic tails form the membrane's core and force the backbone into a rod conformation with nematic-like ordering confined to the interface with water. This particular organization resulted in membranes that combine the stability of classic polymersomes with the biomimetic thickness, flexibility, and lateral mobility of liposomes. Such unparalleled matching of biophysical properties and the ability to locally reconfigure the molecular topology of its constituents enable the harboring of functional components of natural membranes and fusion with living bacteria to “hijack” their periphery. This provides an almost inexhaustible palette to design the chemical and biological makeup of the i-combisomes membrane resulting in a powerful platform for fundamental studies and technological applications.

JTD Keywords: amphiphilic comb polymers, bottom-up synthetic biology, hybrid vesicles, polyelectrolyte-surfactant complexes, polymersomes, synthetic biomembranes, Amphiphilic comb polymers, Biomimetics, Bottom-up synthetic biology, Hybrid vesicles, Hydrophobic and hydrophilic interactions, Liposomes, Polyelectrolyte-surfactant complexes, Polymers, Polymersomes, Synthetic biomembranes, Vesicle fusion, Water

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

Membrane fission triggered by the endosomal sorting complex required for transport (ESCRT) is an important process observed in several pathogenic and non-pathogenic cellular events. From a synthetic-biology viewpoint, ESCRT proteins represent an interesting machinery for the construction of cell mimetic sub-compartments produced by fission. Since their discovery, the studies on ESCRT-III-mediated action, have mainly focused on protein dynamics, ignoring the role of lipid organization and membrane phase state. Recently, it has been suggested that membrane buds formed by the action of ESCRT-III are generated from transient microdomains in endosomal membranes. However, the interplay between membrane domain formation and ESCRT remodeling pathways has not been investigated. Here, giant unilamellar vesicles made of ternary lipid mixtures, either homogeneous in phase or exhibiting liquid-ordered/liquid-disordered phase coexistence, were employed as a model membrane system. These vesicles were incubated with purified recombinant ESCRT-III proteins from the parasite Entamoeba histolytica. In homogeneous membranes, we observe that EhVps32 can trigger domain formation while EhVps20 preferentially co-localizes in the liquid disordered phase. The addition of EhVps24 appears to induce the formation of intraluminal vesicles produced from the liquid-ordered phase. In phase separated membranes, the intraluminal vesicles are also generated from the liquid-ordered phase and presumably emerge from the phase boundary region. Our findings reinforce the hypothesis that ESCRT-mediated remodeling depends on the membrane phase state. Furthermore, the obtained results point to a potential synthetic biology approach for establishing eukaryotic mimics of artificial cells with microcompartments of specific membrane composition, which can also differ from that of the mother vesicle.

JTD Keywords: cell-membranes, coexistence, complex, escrt-iii, fission, guvs, lipid domains, lipid rafts, membrane fission, microcompartments, microscopy, phase separation, plasma-membrane, protein microarrays, structural basis, ternary mixtures, Escrt-iii, Giant unilamellar vesicles, Guvs, Lipid domains, Membrane fission, Microcompartments, Phase separation, Ternary mixtures

Biological cognition is based on the ability to autonomously acquire knowledge, or epistemic autonomy. Such self-supervision is largely absent in artificial neural networks (ANN) because they depend on externally set learning criteria. Yet training ANN using error backpropagation has created the current revolution in artificial intelligence, raising the question of whether the epistemic autonomy displayed in biological cognition can be achieved with error backpropagation-based learning. We present evidence suggesting that the entorhinal–hippocampal complex combines epistemic autonomy with error backpropagation. Specifically, we propose that the hippocampus minimizes the error between its input and output signals through a modulatory counter-current inhibitory network. We further discuss the computational emulation of this principle and analyze it in the context of autonomous cognitive systems. © 2021 Elsevier Ltd

JTD Keywords: computational model, dentate gyrus, error backpropagation, granule cells, grid cells, hippocampus, inhibition, input, neural-networks, neurons, transformation, Artificial intelligence, Artificial neural network, Back propagation, Backpropagation, Brain, Cognitive systems, Counter current, Error back-propagation, Error backpropagation, Errors, Expressing interneurons, Hippocampal complex, Hippocampus, Human experiment, Input and outputs, Learning, Mammal, Mammalian hippocampus, Mammals, Neural networks, Nonhuman, Review, Self-supervised learning

© 2020 Elsevier Inc. Correlative light and electron microscopy (CLEM) entails a group of multimodal imaging techniques that are combined to pinpoint to the location of fluorescently labeled molecules in the context of their ultrastructural cellular environment. Here we describe a detailed workflow for STORM-CLEM, in which STochastic Optical Reconstruction Microscopy (STORM), an optical super-resolution technique, is correlated with transmission electron microscopy (TEM). This protocol has the advantage that both imaging modalities have resolution at the nanoscale, bringing higher synergies on the information obtained. The sample is prepared according to the Tokuyasu method followed by click-chemistry labeling and STORM imaging. Then, after heavy metal staining, electron microscopy imaging is performed followed by correlation of the two images. The case study presented here is on intracellular pathogens, but the protocol is versatile and could potentially be applied to many types of samples.

JTD Keywords: cells, click-chemistry, complex, correlative light and electron microscopy, cycloaddition, ligation, localization, proteins, resolution limit, single molecule localization microscopy, stochastic optical reconstruction microscopy (storm), storm, super-resolution microscopy, tokuyasu cryo-sectioning, tool, Click-chemistry, Correlative light and electron microscopy, Fluorescent-probes, Single molecule localization microscopy, Stochastic optical reconstruction microscopy (storm), Super-resolution microscopy, Tokuyasu cryo-sectioning, Transmission electron microscopy

© 2020 American Chemical Society. The performance of supramolecular nanocarriers as drug delivery systems depends on their stability in the complex and dynamic biological media. After administration, nanocarriers are challenged by physiological barriers such as shear stress and proteins present in blood, endothelial wall, extracellular matrix, and eventually cancer cell membrane. While early disassembly will result in a premature drug release, extreme stability of the nanocarriers can lead to poor drug release and low efficiency. Therefore, comprehensive understanding of the stability and assembly state of supramolecular carriers in each stage of delivery is the key factor for the rational design of these systems. One of the main challenges is that current 2D in vitro models do not provide exhaustive information, as they fail to recapitulate the 3D tumor microenvironment. This deficiency in the 2D model complexity is the main reason for the differences observed in vivo when testing the performance of supramolecular nanocarriers. Herein, we present a real-time monitoring study of self-assembled micelles stability and extravasation, combining spectral confocal microscopy and a microfluidic cancer-on-a-chip. The combination of advanced imaging and a reliable 3D model allows tracking of micelle disassembly by following the spectral properties of the amphiphiles in space and time during the crucial steps of drug delivery. The spectrally active micelles were introduced under flow and their position and conformation continuously followed by spectral imaging during the crossing of barriers, revealing the interplay between carrier structure, micellar stability, and extravasation. Integrating the ability of the micelles to change their fluorescent properties when disassembled, spectral confocal imaging and 3D microfluidic tumor blood vessel-on-a-chip resulted in the establishment of a robust testing platform suitable for real-time imaging and evaluation of supramolecular drug delivery carrier's stability.

JTD Keywords: cancer-on-a-chip, complex, delivery, endothelial-cells, in-vitro, microfluidic, model, nanoparticle, penetration, shear-stress, stability, supramolecular, Cancer-on-a-chip, Cell-culture, Micelle, Microfluidic, Nanoparticle, Stability, Supramolecular

We propose a method to control chimera states in a ring-shaped network of nonlocally coupled phase oscillators. This method acts exclusively on the network’s connectivity. Using the idea of a pacemaker oscillator, we investigate which is the minimal action needed to control chimeras. We implement the pacemaker choosing one oscillator and making its links unidirectional. Our results show that a pacemaker induces chimeras for parameters and initial conditions for which they do not form spontaneously. Furthermore, the pacemaker attracts the incoherent part of the chimera state, thus controlling its position. Beyond that, we find that these control effects can be achieved with modifications of the network’s connectivity that are less invasive than a pacemaker, namely, the minimal action of just modifying the strength of one connection allows one to control chimeras.

JTD Keywords: Complex networks, Oscillators, Spatiotemporal phenomena

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

Engineering approaches allow biological structures and behaviours to be reconstituted in vitro. A biologist and a physicist discuss the potential and limitations of this bottom-up philosophy in providing insights into complex biological processes.

JTD Keywords: Biophysics, Complexity, Engineering

The grand quest for a scientific understanding of consciousness has given rise to many new theoretical and empirical paradigms for investigating the phenomenology of consciousness as well as clinical disorders associated to it. A major challenge in this field is to formalize computational measures that can reliably quantify global brain states from data. In particular, information-theoretic complexity measures such as integrated information have been proposed as measures of conscious awareness. This suggests a new framework to quantitatively classify states of consciousness. However, it has proven increasingly difficult to apply these complexity measures to realistic brain networks. In part, this is due to high computational costs incurred when implementing these measures on realistically large network dimensions. Nonetheless, complexity measures for quantifying states of consciousness are important for assisting clinical diagnosis and therapy. This article is meant to serve as a lookup table of measures of consciousness, with particular emphasis on clinical applicability. We consider both, principle-based complexity measures as well as empirical measures tested on patients. We address challenges facing these measures with regard to realistic brain networks, and where necessary, suggest possible resolutions. We address challenges facing these measures with regard to realistic brain networks, and where necessary, suggest possible resolutions.

JTD Keywords: Consciousness in the Clinic, Computational neuroscience, Complexity measures, Clinical Neuroscience, Measures of consciousness

We study two-layer networks of identical phase oscillators. Each individual layer is a ring network for which a non-local intra-layer coupling leads to the formation of a chimera state. The number of oscillators and their natural frequencies is in general different across the layers. We couple the phases of individual oscillators in one layer to the phase of the mean field of the other layer. This coupling from the mean field to individual oscillators is done in both directions. For a sufficient strength of this interlayer coupling, the phases of the mean fields lock across the two layers. In contrast, both layers continue to exhibit chimera states with no locking between the phases of individual oscillators across layers, and the two mean field amplitudes remain uncorrelated. Hence, the networks’ mean fields show phase synchronization which is analogous to the one between low-dimensional chaotic oscillators. The required coupling strength to achieve this mean field phase synchronization increases with the mismatches in the network sizes and the oscillators’ natural frequencies.

JTD Keywords: Chaos, Complex networks, Oscillators, Synchronisation

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

A major challenge in computational and systems neuroscience concerns the quantification of information processing at various scales of the brain’s anatomy. In particular, using human intracranial recordings, the question we ask in this paper is: How can we estimate the informational complexity of the brain given the complex temporal nature of its dynamics? To address this we work with a recent formulation of network integrated information that is based on the Kullback-Leibler divergence between the multivariate distribution on the set of network states versus the corresponding factorized distribution over its parts. In this work, we extend this formulation for temporal networks and then apply it to human brain data obtained from intracranial recordings in epilepsy patients. Our findings show that compared to random re-wirings of the data, functional connectivity networks, constructed from human brain data, score consistently higher in the above measure of integrated information. This work suggests that temporal integrated information may indeed be a good starting point as a future measure of cognitive complexity.

JTD Keywords: Brain networks, Complexity measures, Computational neuroscience, Functional connectivity

In the absence of a first-principles definition, the concept of intelligence is often specified in terms of its phenomenological functions as a capacity or ability to solve problems autonomously. Whenever an agent, biological or artificial, possesses this ability, it is considered intelligent, otherwise not. While this description serves as a useful correlate of intelligence, it is far from a principled explanation that provides a general, yet precise definition along with predictions of mechanisms leading to intelligent behavior. We do not want an explanation to depend on any functionality that itself might be a consequence of intelligence. A possible conceptualization of a function-free approach might be to formulate the concept in terms of dynamical information complexity. This constitute a first step towards a statistical mechanics theory of intelligence. In this paper, we outline the steps towards a physics-based definition of intelligence.

JTD Keywords: Complexity, Information Theory, Physics of Intelligence

We have developed a new liquid chromatography-electrospray ionization tandem mass spectrometry methodology based on 2-picolylamine derivatization and positive ion mode detection for abscisic acid (ABA) identification. The selected reaction leads to the formation of an amide derivative which contains a highly active pyridyl group. The enhanced ionization allows for a 700-fold increase over commonly monitored unmodified ABA, which in turn leads to excellent limits of detection and quantification values of 0.03 and 0.15 ng mL-1, respectively. This method has been validated in the highly complex matrix of a red blood cell extract. In spite of the high sensitivity achieved, ABA could not be detected in Plasmodium falciparum-infected red blood cells, suggesting that, if present, it will be found either in ultratrace amounts or as brief bursts at defined time points within the intraerythrocytic cycle and/or in the form of a biosynthetic analogue.

JTD Keywords: Abscisic acid, Apicomplexa, Liquid chromatography-electrospray ionization tandem mass spectrometry, Malaria, Picolylamine, Plasmodium falciparum

Controlling the spin of electrons in nanoscale electronic devices is one of the most promising topics aiming at developing devices with rapid and high density information storage capabilities. The interface magnetism or spinterface resulting from the interaction between a magnetic molecule and a metal surface, or vice versa, has become a key ingredient in creating nanoscale molecular devices with novel functionalities. Here, we present a single-molecule wire that displays large (>10000%) conductance switching by controlling the spin-dependent transport under ambient conditions (room temperature in a liquid cell). The molecular wire is built by trapping individual spin crossover FeII complexes between one Au electrode and one ferromagnetic Ni electrode in an organic liquid medium. Large changes in the single-molecule conductance (>100-fold) are measured when the electrons flow from the Au electrode to either an α-up or a β-down spin-polarized Ni electrode. Our calculations show that the current flowing through such an interface appears to be strongly spin-polarized, thus resulting in the observed switching of the single-molecule wire conductance. The observation of such a high spin-dependent conductance switching in a single-molecule wire opens up a new door for the design and control of spin-polarized transport in nanoscale molecular devices at room temperature.

JTD Keywords: Density functional calculations, Magnetoresistance, Single-molecule junctions, Spin orbit coupling, Spin-crossover complexes, Spinterface, STM break-junction

Lysosomal storage disorders are currently treated by enzyme replacement therapy (ERT) through the direct administration of the unprotected recombinant protein to the patients. Herein we present an ionically cross-linked polyelectrolyte complex (PEC) composed of trimethyl chitosan (TMC) and α-galactosidase A (GLA), the defective enzyme in Fabry disease, with the capability of directly targeting endothelial cells by incorporating peptide ligands containing the RGD sequence. We assessed the physicochemical properties, cytotoxicity, and hemocompatibility of RGD-targeted and untargeted PECs, the uptake by endothelial cells and the intracellular activity of PECs in cell culture models of Fabry disease. Moreover, we also explored the effect of different freeze-drying procedures in the overall activity of the PECs. Our results indicate that the use of integrin-binding RGD moiety within the PEC increases their uptake and the efficacy of the GLA enzyme, while the freeze-drying allows the activity of the therapeutic protein to remain intact. Overall, these results highlight the potential of TMC-based PECs as a highly versatile and feasible drug delivery system for improving the ERT of lysosomal storage disorders.

JTD Keywords: Enzyme replacement therapy, Fabry disease, Lysosomal delivery, Nanomedicine, Polyelectrolyte complexes, Trimethyl chitosan, α-galactosidase A

Biocompatible quercetin nanovesicles were developed by coating polyethylene glycol-containing vesicles with chitosan and nutriose, aimed at targeting the colon. Uncoated and coated vesicles were prepared using hydrogenated soy phosphatidylcholine and quercetin, a potent natural anti-inflammatory and antioxidant drug. Physicochemical characterization was carried out by light scattering, cryogenic microscopy and X-ray scattering, the results showing that vesicles were predominantly multilamellar and around 130 nm in size. The in vitro release of quercetin was investigated under different pH conditions simulating the environment of the gastrointestinal tract, and confirmed that the chitosan/nutriose coating improved the gastric resistance of vesicles, making them a potential carrier system for colon delivery. The preferential localization of fluorescent vesicles in the intestine was demonstrated using the In Vivo FX PRO Imaging System. Above all, a marked amelioration of symptoms of 2,4,6-trinitrobenzenesulfonic acid-induced colitis was observed in animals treated with quercetin-loaded coated vesicles, favoring the restoration of physiological conditions. Therefore, quercetin-loaded chitosan/nutriose-coated vesicles can represent a valuable therapeutic tool for the treatment of chronic intestinal inflammatory diseases, and presumably a preventive system, due to the synergic action of antioxidant quercetin and beneficial prebiotic effects of the chitosan/nutriose complex.

JTD Keywords: Chitosan/nutriose complex, Colon targeting, Phospholipid vesicles, Quercetin, Rat colitis

We present a procedure for calibrated complex impedance measurements and dielectric quantification with scanning microwave microscopy. The calibration procedure works in situ directly on the substrate with the specimen of interest and does not require any specific calibration sample. In the workflow tip-sample approach curves are used to extract calibrated complex impedance values and to convert measured S11 reflection signals into sample capacitance and resistance images. The dielectric constant of thin dielectric SiO2 films were determined from the capacitance images and approach curves using appropriate electrical tip-sample models and the εr value extracted at f = 19.81 GHz is in good agreement with the nominal value of εr ∼ 4. The capacitive and resistive material properties of a doped Si semiconductor sample were studied at different doping densities and tip-sample bias voltages. Following a simple serial model the capacitance-voltage spectroscopy curves are clearly related to the semiconductor depletion zone while the resistivity is rising with falling dopant density from 20 Ω to 20 kΩ. The proposed procedure of calibrated complex impedance measurements is simple and fast and the accuracy of the results is not affected by varying stray capacitances. It works for nanoscale samples on either fully dielectric or highly conductive substrates at frequencies between 1 and 20 GHz.

JTD Keywords: Complex impedance, Dielectric constant, Nanotechnology: calibration, Resistivity, Scanning microwave microscopy

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

In this work we report the fabrication and characterization of a label-free impedimetric immunosensor based on a silicon nitride (Si 3N 4) surface for the specific detection of human serum albumin (HSA) proteins. Silicon nitride provides several advantages compared with other materials commonly used, such as gold, and in particular in solid-state physics for electronic-based biosensors. However, few Si 3N 4-based biosensors have been developed; the lack of an efficient and direct protocol for the integration of biological elements with silicon-based substrates is still one of its the main drawbacks. Here, we use a direct functionalization method for the direct covalent binding of monoclonal anti-HSA antibodies on an aldehyde-functionalized Si-p/SiO 2/Si 3N 4 structure. This methodology, in contrast with most of the protocols reported in literature, requires less chemical reagents, it is less time-consuming and it does not need any chemical activation. The detection capability of the immunosensor was tested by performing non-faradaic electrochemical impedance spectroscopy (EIS) measurements for the specific detection of HSA proteins. Protein concentrations within the linear range of 10 -13-10 -7M were detected, showing a sensitivity of 0.128ΩμM -1 and a limit of detection of 10 -14M. The specificity of the sensor was also addressed by studying the interferences with a similar protein, bovine serum albumin. The results obtained show that the antibodies were efficiently immobilized and the proteins detected specifically, thus, establishing the basis and the potential applicability of the developed silicon nitride-based immunosensor for the detection of proteins in real and more complex samples.

JTD Keywords: Aldehyde, Electrochemical impedance spectroscopy, Human serum albumin, Immunosensor, Silicon nitride, Bovine serum albumins, Chemical reagents, Complex samples, Covalent binding, Detection capability, Electrochemical impedance, Electrochemical impedance spectroscopy measurements, Functionalizations, Human serum albumins, Impedimetric immunosensors, Label free, Limit of detection, Linear range, Protein concentrations, Silicon-based, Specific detection, Aldehydes

Current analytical instrumentation and continuous sensing can provide huge amounts of data. Automatic signal processing and information evaluation is needed to overcome drowning in data. Today, statistical techniques are typically used to analyse and extract information from continuous signals. However, it is very interesting to note that biology (insects and vertebrates) has found alternative solutions for chemical sensing and information processing. This is a brief introduction to the developments in the European Project: Bio-ICT NEUROCHEM: Biologically Inspired Computation for Chemical Sensing (grant no. 216916) Fp7 project devoted to biomimetic olfactory systems.

JTD Keywords: Signal processing, Chemioception, Neural nets, Computational complexity

Surface features at the micro and nanometre scale have been shown to influence and even determine cell behaviour and cytoskeleton organization through direct mechanotransductive pathways. Much less is known about the function and internal distribution of organelles of cells grown on topographically modified surfaces. In this study, the nanoimprint lithography technique was used to manufacture poly(methyl methacrylate) (PMMA) sheets with a variety of features in the micrometre size range. Normal rat kidney (NRK) fibroblasts were cultured on these substrates and immunofluorescence staining assays were performed to visualize cell adhesion, the organization of the cytoskeleton and the morphology and subcellular positioning of the Golgi complex. The results show that different topographic features at the micrometric scale induce different rearrangements of the cell cytoskeleton, which in turn alter the positioning and morphology of the Golgi complex. Microposts and microholes alter the mechanical stability of the Golgi complex by modifying the actin cytoskeleton organization leading to the compaction of the organelle. These findings prove that physically modified surfaces are a valuable tool with which to study the dynamics of cell cytoskeleton organization and its subsequent repercussion on internal cell organization and associated function.

JTD Keywords: Actin stress fibers, Golgi-complex, Focal adhesions, Cytoskeletal organization, Osteoblast adhesion, Mammalian-cells, Micron-scale, Nanoscale, Dynamics, Rho

P>Lesioned axons do not regenerate in the adult mammalian CNS, owing to the over-expression of inhibitory molecules such as myelin-derived proteins or chondroitin sulphate proteoglycans. In order to overcome axon inhibition, strategies based on extrinsic and intrinsic treatments have been developed. For myelin-associated inhibition, blockage with NEP1-40, receptor bodies or IN-1 antibodies has been used. In addition, endogenous blockage of cell signalling mechanisms induced by myelin-associated proteins is a potential tool for overcoming axon inhibitory signals. We examined the participation of glycogen synthase kinase 3 beta (GSK3 beta) and extracellular-related kinase (ERK) 1/2 in axon regeneration failure in lesioned cortical neurons. We also investigated whether pharmacological blockage of GSK3 beta and ERK1/2 activities facilitates regeneration after myelin-directed inhibition in two models: (i) cerebellar granule cells and (ii) lesioned entorhino-hippocampal pathway in slice cultures, and whether the regenerative effects are mediated by Nogo Receptor 1 (NgR1). We demonstrate that, in contrast to ERK1/2 inhibition, the pharmacological treatment of GSK3 beta inhibition strongly facilitated regrowth of cerebellar granule neurons over myelin independently of NgR1. Finally, these regenerative effects were corroborated in the lesioned entorhino-hippocampal pathway in NgR1-/- mutant mice. These results provide new findings for the development of new assays and strategies to enhance axon regeneration in injured cortical connections.

JTD Keywords: Axon inhibition, Nogo Receptor complex, Organotypic slice cultures, Pharmacological treatment

Statistical analysis, power spectral density, and Lempel Ziv complexity, are used in a multi-parameter approach to analyze four temporal series obtained from the Electrocardiographic and Respiratory Flow signals of 126 patients on weaning trials. In which, 88 patients belong to successful group (SG), and 38 patients belong to failure group (FG), i.e. failed to maintain spontaneous breathing during trial. It was found that mean values of cardiac inter-beat and breath durations give higher values for SG than for FG; Kurtosis coefficient of the spectrum of the rapid shallow breathing index is higher for FG; also Lempel Ziv complexity mean values associated with the respiratory flow signal are bigger for FG. Patients were then classified with a pattern recognition neural network, obtaining 80% of correct classifications (81.6% for FG and 79.5% for SG).

JTD Keywords: Electrocardiography, Medical signal processing, Neural nets, Pattern recognition, Pneumodynamics, Signal classification, Statistical analysis, ECG, Kurtosis coefficient, Lempel Ziv complexity, Breath durations, Cardiac interbeat durations, Electrocardiography, Multiparameter analysis, Pattern recognition neural network, Power spectral density, Respiratory flow signals, Signal classification, Spontaneous breathing, Statistical analysis, Weaning trials