Publications

Silica-based nanoparticles have been extensively investigated as advanced bioimaging probes and smart nanocarriers for the development of nanomedicine because of their unique properties. The deep understanding of nanoparticle-biological system (nanobiology) interactions is an important determinant for the success of nanomedicine. In this Review, we focus on the assessment of significant nanobio interactions along with the intracellular fates of silica-based nanoparticles by advanced microscopy and link this information to their recent biomedical applications. Initially, the fundamental concepts of the dynamic interactions between nanoparticles and proteins followed by cells and key factors that influence these nanobio interactions are briefly introduced. Furthermore, the basic principles of advanced imaging modalities for the analysis of nanobio interactions in this study are described. Next, the utilization of advanced imaging for the characterization of protein coronas and monitoring the cellular internalization and intracellular trafficking/fate of individual nanoparticles are comprehensively summarized. Finally, recent biomedical applications of nanoparticles in bioimaging and drug delivery are discussed.

JTD Keywords: Adsorption, Cellular uptake, Desig, Drug-delivery, Mesoporous organosilica nanoparticles, Nanocapsules, Protein corona, Size, Superresolution microscopy, Surface-properties

Odorant discrimination mechanisms are based on the differential interactions between odorant molecules and olfactory receptors (ORs). Biohybrid sensors based on ORs described to date show selectivity towards specific versus non-specific binding of odorants, being unable to distinguish between specific ligands of different affinity. Here we disclose a method that enables odorant discrimination based on the modulation of the capacitive response of the receptor, which allows the differentiation of three high-affinity hOR1A1 agonists. We performed voltammetry and impedance measurements of the hOR1A1 receptor selectively immobilized on a gold electrode in the absence and presence of the agonists. Binding induces a decrease in the capacitive response of the receptor that is proportional to the ligand potency, reaching up to a 40% decrease for the cognate ligand dihydrojasmone, which is attributed to changes in the magnitude and orientation of the electric dipole in the receptor, thereby regulating its response to the applied electric field.

JTD Keywords: Agonist, Biosenso, Electric dipol, Electrochemical impedance spectroscopy, Impedance, Nose, Odorant receptors, Olfactory receptor, Selectivity, Specific capacitance

A multiplexed microarray chip (Immuno-mu SARS2) aiming at providing information on the prognosis of the COVID-19 has been developed. The diagnostic technology records information related to the profile of the immunological response of patients infected by the SARS-CoV-2 virus. The diagnostic technology delivers information on the avidity of the sera against 28 different peptide epitopes and 7 proteins printed on a 25 mm2 area of a glass slide. The peptide epitopes (12-15 mer) derived from structural proteins (Spike and Nucleocapsid) have been rationally designed, synthesized, and used to develop Immuno-mu SARS2 as a multiplexed and high-throughput fluorescent microarray platform. The analysis of 755 human serum samples (321 from PCR+ patients; 288 from PCR- patients; 115 from prepandemic individuals and classified as hospitalized, admitted to intensive-care unit (ICU), and exitus) from three independent cohorts has shown that the chips perform with a 98% specificity and 91% sensitivity identifying RT-PCR+ patients. Computational analysis utilized to correlate the immunological signatures of the samples analyzed indicate significant prediction rates against exitus conditions with 82% accuracy, ICU admissions with 80% accuracy, and 73% accuracy over hospitalization requirement compared to asymptomatic patients' fingerprints. The miniaturized microarray chip allows simultaneous determination of 96 samples (24 samples/slide) in 90 min and requires only 10 mu L of sera. The diagnostic approach presented for the first time here could have a great value in assisting clinicians in decision-making based on the information provided by the Immuno-mu SARS2 regarding progression of the disease and could be easily implemented in diagnostics of other infectious diseases.

JTD Keywords: Antibodies, Clinical diagnostic, Diagnosis, High-throughput, Machine learning, Microarray, Multiplexation, Nucleocapsid protein, Peptide epitopes, Sars-cov-, Sars-cov-2, Serological signature, Seroprevalence, Severity prediction, Spik

The challenge of delivering therapeutics to the central nervous system due to the restrictive nature of the blood-brain barrier (BBB) is a substantial hurdle in neuropharmacology. Our research introduces a breakthrough approach using microtubule-dependent transcytosis facilitated by novel aqueous compounds. We synthesized a series of red-emitting pyran nitrile derivatives. The molecular structure of compounds, photophysical properties, and water solubility were characterized. BBB permeability of BN1 was assessed in an in vitro BBB model. The transmembrane transport mechanism was next analyzed. The derivative was injected in the wild-type mouse for evaluation of brain penetration and biodistribution in the brain. We further investigated the potential of BN1-functionalized BBB-nonpenetrated silica nanoparticles for brain targeting. This compound demonstrated an ability to form endosomes within the phospholipid layer, thus enabling efficient penetration of the BBB via microtubule-mediated transcytosis, as evidenced in vitro model. This was further confirmed by in vivo experiments that BN1 displays the excellent BBB penetration and retained in brain parenchyma. Furthermore, BBB-impermeable mesoporous silica nanoparticle codelivery system markedly enhanced the transport efficiency to the brain in vivo by BN1-functionalized. These findings indicate that our designed aqueous molecules not only are capable of traversing the BBB but also serve as a viable new strategy for central-nervous-system-targeted drug delivery.

JTD Keywords: Blood-brain-barrier, Desig, In-vivo detection, Mesoporous silica, Probes, Silica nanoparticles

The development of 3D-printed calcium phosphate bone grafts represents a breakthrough for personalized bone tissue engineering. However, their high brittleness is a limiting factor for clinical applications. The present work aims to overcome this drawback by developing a composite ink for direct ink writing using a polycaprolactone solution (20 wt% in pyridine) as binder for a self-hardening alpha-tricalcium phosphate powder. The ink consolidation results from a two-step process. First, evaporation of the solvent gives rise to a flexible green body, in which the polymer matrix retains the loose ceramic particles. Subsequently, upon contact with water, the hydrolysis of these particles creates a continuous network of biomimetic hydroxyapatite nanocrystals, with the consequent hardening of the ink. The presence of the polymer improves the mechanical properties, increasing the flexural strength from 2 to 4 MPa and the strain energy density from 0.2 to 1.1 kJ/m3. Unlike the inks based on hydrogel binders, where the hardening reaction starts as soon as the liquid and solid components are mixed, the ink prepared with the non-aqueous binder is stable, with over 5 months shelf life, and allows the hardening reaction to be triggered at will by contact with water.

JTD Keywords: 3d printing, Bone scaffold, Calcium phosphates, Mechanical propertie, Polycaprolactone

[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

Biofabrication of three-dimensional (3D) cultures through the 3D Bioprinting technique opens new perspectives and applications of cell-laden hydrogels. However, to continue with the progress, new BioInks with specific properties must be carefully designed. In this study, we report the synthesis and 3D Bioprinting of an electroconductive BioInk made of gelatin/fibrinogen hydrogel, C2C12 mouse myoblast and 5% w/w of conductive poly (3,4-ethylenedioxythiophene) nanoparticles (PEDOT NPs). The influence of PEDOT NPs, incorporated in the cellladen BioInk, not only showed a positive effect in cells viability, differentiation and myotube functionalities, also allowed the printed constructs to behaved as BioCapacitors. Such devices were able to electrochemically store a significant amount of energy (0.5 mF/cm2), enough to self-stimulate as BioActuator, with typical contractions ranging from 27 to 38 mu N, during nearly 50 min. The biofabrication of 3D constructs with the proposed electroconductive BioInk could lead to new devices for tissue engineering, biohybrid robotics or bioelectronics.

JTD Keywords: 3d bioprinting, Animal, Animals, Bioactuator, Bioactuators, Biocapacitor, Biofabrication, Bioprinting, Biosensing techniques, C2c12 myoblasts, Cells, Chemistry, Electric conductivity, Electroconductive, Electroconductive bioink, Ethylenedioxythiophenes, Genetic procedures, Hydrogel, Hydrogels, Mice, Mouse, Pedot nps, Pedot nps,3d bioprinting,electroconductive bioink,bioactuator,biocapacito, Poly (3,4-ethylenedioxythiophene) nanoparticle, Printing, three-dimensional, Procedures, Skeletal-muscle,cytotoxicity,polymer, Synthesis (chemical), Three dimensional printing, Tissue engineering, Tissue scaffolds

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. (c) 2024 The Authors. Published by Elsevier Ltd on behalf of Acta Materialia Inc. This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ )

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

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

A new generation of fully bioresorbable metallic Zn-based alloys could be used for stenting applications; however, the initial surface degradation delays stent re-endothelialization. Thus, this work proposes a dual strategy to control the corrosion and accelerate the endothelialization of ZnMg and ZnAg biodegradable alloys. First, a stable polycaprolactone (PCL) coating is obtained and followed by its functionalization with either linear RGD (Arg-Gly-Asp) or REDV (Arg-Glu-Asp-Val) peptides or a dual peptide-based platform combining both sequences (RGD-REDV). Scratching tests showed neither delamination nor detachment of the polymeric coating. Potentiodynamic polarization (PDP) and electrochemical impedance spectroscopy (EIS) measurements confirmed the corrosion resistance after PCL coating by revealing lower current density and higher absolute impedance values. X-ray photoelectron spectroscopy (XPS) and fluorescent microscopy confirmed the correct peptide immobilization onto PCL coated Zn alloys. The functionalized samples exhibited enhanced human umbilical vein endothelial cells (HUVEC) adhesion. The higher number of adhered cells to the functionalized surfaces with the RGD-REDV platform demonstrates the synergistic effect of combining both RGD and REDV sequences. Higher corrosion resistance together with enhanced endothelialization indicates that the dual functionalization of Zn alloys with PCL and peptide-based RGD-REDV platform holds great potential to overcome the clinical limitations of current biodegradable metal stents.

JTD Keywords: Binary alloys, Biodegradable metals, Bioresorbable, Cardiovascular applications, Cell adhesive peptides, Corrosion, Corrosion resistance, Corrosion resistant alloys, Corrosion resistant coatings, Degradation, Dual peptide-based platform, Electrochemical corrosion, Electrochemical impedance spectroscopy, Endothelial cells, Endothelialization, Functionalization, Functionalizations, In-vitro, Magnesium alloys, Metallics, Mg alloy, Peptides, Polycaprolactone coating, Polymer-coatings, Rgd-functionalization, Silver alloys, Stents, Surface, X ray photoelectron spectroscopy, Zinc, Zinc alloys, Zn alloys, Zn-based alloys

Polymeric membranes exhibit unique and modulate transport properties when they are properly functionalised, which make them ideal for ions transport, molecules separation and molecules interactions. The present work proposes the design and fabrication of nanostructured membranes, composed by biodegradable poly(lactic acid) (PLA) and poly(ethylene glycol) (PEG), incorporating a lipophilic molecule (cholesterol) covalently bonded, were especially designed to provide even more application opportunities in sensors field. Electrochemical studies, by means of electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and square wave voltammetry (SWV), revealed important differences regarding the functionalised and non-functionalised PLA systems. PEGcholesterol building block units showed a clear affinity with ascorbic acid (vitamin C) and Trolox (R) (a watersoluble analogue of vitamin E), both hydrophilic in nature, with a limit of detection capacity of 8.12 mu M for AA and 3.53 mu M for AA and Trolox, respectively, in aqueous salt solution. The bioinspired polymer may be used to incorporate antioxidant property that allow the design of anti-stress biosensors, electrodes for the detection of vitamin C or vitamin E in biomedical nutrition programs, among other applications.

JTD Keywords: Antioxidant molecules, Antioxidants, Application programs, Ascorbic acid, Biomimetics, C (programming language), Capacity, Chemical detection, Cholesterol, Cyclic voltammetry, Electrochemical detection, Electrochemical impedance spectroscopy, Functional polymers, Functionalized, Lactic acid, Molecules, Nanomembranes, Poly ethylene glycols, Poly lactic acid, Poly(ethylene glycol), Poly(ethyleneglycol), Poly(lactic acid), Polyethylene glycols, Vitamin-e

Osteoporotic-related fractures are among the leading causes of chronic disease morbidity in Europe and in the US. While a significant percentage of fractures can be repaired naturally, in delayed-union and non-union fractures surgical intervention is necessary for proper bone regeneration. Given the current lack of optimized clinical techniques to adequately address this issue, bone tissue engineering (BTE) strategies focusing on the development of scaffolds for temporarily replacing damaged bone and supporting its regeneration process have been gaining interest. The piezoelectric properties of bone, which have an important role in tissue homeostasis and regeneration, have been frequently neglected in the design of BTE scaffolds. Therefore, in this study, we developed novel hydroxyapatite (HAp)-filled osteoinductive and piezoelectric poly(vinylidene fluoride-co-tetrafluoroethylene) (PVDF-TrFE) nanofibers via electrospinning capable of replicating the tissue's fibrous extracellular matrix (ECM) composition and native piezoelectric properties. The developed PVDF-TrFE/HAp nanofibers had biomimetic collagen fibril-like diameters, as well as enhanced piezoelectric and surface properties, which translated into a better capacity to assist the mineralization process and cell proliferation. The biological cues provided by the HAp nanoparticles enhanced the osteogenic differentiation of seeded human mesenchymal stem/stromal cells (MSCs) as observed by the increased ALP activity, cell-secreted calcium deposition and osteogenic gene expression levels observed for the HAp-containing fibers. Overall, our findings describe the potential of combining PVDF-TrFE and HAp for developing electroactive and osteoinductive nanofibers capable of supporting bone tissue regeneration.© 2023 The Author(s). Published by National Institute for Materials Science in partnership with Taylor & Francis Group.

JTD Keywords: composites, electrospinning, hydroxyapatite, piezoelectricity, promote, pvdf, pvdf-trfe, removal, scaffolds, temperature, Bone tissue engineering, Electrospinning, Electrospun polycaprolactone, Hydroxyapatite, Piezoelectricity, Pvdf-trfe

Naso- and oropharyngeal swabs are the Center for Disease Control and Prevention (CDC) -recommended disease sampling methods for respiratory viruses. The short swabbing time for sampling by these methods may lead to variability in test results. Further, these methods are mildly invasive and can cause discomfort, tearing or gag reflexes in tested individuals. If longer sampling time is coupled with lesser patient discomfort, test reliability and patient compliance can be improved. Towards this end, we developed cationic biopolymer membranes for the electrostatic capturing of viruses in the oral cavity. Here, chemically (EDC-NHS) crosslinked uncharged chitosan (CS) nanofiber membranes were conferred either with negative surface charge by anionic poly-aspartic acid (pAsp) coating or positive charge by cationic poly-L-lysine (PLL). Consistent with our preliminary findings of dynamic light scattering (DLS) size measurements showing large agglomerates of anionic virus-like particles (VLPs) and cationic PLL in solution, a 75% increase in VLP adsorption by PLL coated CS membranes was recorded by enzyme linked immunosorbent assay (ELISA), in comparison to untreated controls. It is envisaged that the electrostatic concentration of respiratory viruses on cationic membranes can be superior alternatives to traditional swabbing in the oral cavity.

JTD Keywords: Cationic biopolymer membranes, Disease sampling, Dynamic light scattering (dls), Electrostatic capture of viruses, Enzyme linked immunosorbent assay (elisa), Magnetic beads, Virus -like particles (vlps)

The use of nanocarriers in medicine, so-called nanomedicine, is one of the most innovative strategies for targeting drugs at the action site and increasing their activity index and effectiveness. Phytomedicine is the oldest traditional method used to treat human diseases and solve health problems. The recent literature on the treatment of malaria infections using nanodelivery systems and phytodrugs or supplements has been analyzed. For the first time, in the present review, a careful look at the considerable potential of nanomedicine in promoting phytotherapeutic efficacy was done, and its key role in addressing a translation through a significant reduction of the current burden of malaria in many parts of the world has been underlined.

JTD Keywords: antiplasmodial activity, bioavailability, chloroquine, combination therapy, discovery, drug-delivery, drug-delivery systems, nanocapsules, nanomedicine, natural molecules, pharmacokinetics, phytomedicine, plasmodium-falciparum, Artemisinin-based combination therapy, Drug-delivery systems, Nanomedicine, Natural molecules, Phytomedicine, Solid lipid nanoparticles

Nanoparticles (NPs) are used to encapsulate therapeutic cargos and deliver them specifically to the target site. The intracellular trafficking of NPs dictates the NP-cargo distribution within different cellular compartments, and thus governs their efficacy and safety. Knowledge in this field is crucial to understand their biological fate and improve their rational design. However, there is a lack of methods that allow precise localization and quantification of individual NPs within distinct cellular compartments simultaneously. Here, we address this issue by proposing a correlative light and electron microscopy (CLEM) method combining direct stochastic optical reconstruction microscopy (dSTORM) and transmission electron microscopy (TEM). We aim at combining the advantages of both techniques to precisely address NP localization in the context of the cell ultrastructure. Individual fluorescently-labelled poly(lactide-co-glycolide)-poly(ethylene glycol) (PLGA-PEG) NPs were directly visualized by dSTORM and assigned to cellular compartments by TEM. We first tracked NPs along the endo-lysosomal pathway at different time points, then demonstrated the effect of chloroquine on their intracellular distribution (i.e. endosomal escape). The proposed protocol can be applied to fluorescently labelled NPs and/or cargo, including those not detectable by TEM alone. Our studies are of great relevance to obtain important information on NP trafficking, and crucial for the design of more complex nanomaterials aimed at cytoplasmic/nucleic drug delivery.

JTD Keywords: chemistry, delivery, endocytosis, endosomal escape, exocytosis, fluorescence, light, size, tomography, Cellular uptake, Chemistry, Delivery, Endocytosis, Endosomal escape, Exocytosis, Fluorescence, Light, Size, Tomography

A study of the shape memory effect on extruded polylactic acid (PLA) and polycaprolactone (PCL) blends, which were transformed into films and movable components of articulated specimens by hot pressing and 3D printing, respectively, is presented. After characterizing their chemical structure by FTIR spectroscopy and their wetta-bility, the thermal properties and mechanical response of the blends were evaluated and compared with those of neat PLA and PCL. The blends exhibited very good interfacial adhesion between the phases, even though they are immiscible polymers. The thermoresponsive shape memory effects of neat PLA, neat PCL and PLA/PCL blends with different compositions (90/30, 70/30 and 50/50 w/w%) were evaluated considering three consecutive heating-cooling cycles. Comparison of the initial permanent state geometry with the geometries achieved after each heating-cooling cycle for both films and 3D printed specimens, evidenced that the 70/30 w/w% blend exhibited the best behavior. Thus, the blends obtained with such composition showed the maximum reversibility between the temporary and permanent states (i.e. highest shape recovery capability) and shape fixing of such two states.

JTD Keywords: 3d printing, Fibers, Films, Poly(lactic acid), Polycaprolactone, Polylactic acid, Polymer, Shape fixing, Shape-memory polymers, Unimolecular micelles

Biodegradable periodic mesoporous organosilica nanoparticles (nanoPMOs) are widely used as responsive drug delivery platforms for targeted chemotherapy of cancer. However, the evaluation of their properties such as surface functionality and biodegradability is still challenging, which has a significant impact on the efficiency of chemotherapy. In this study, we have applied direct stochastic optical reconstruction microscopy (dSTORM), a single-molecule super-resolution microscopy technique, to quantify the degradation of nanoPMOs triggered by glutathione and the multivalency of antibody-conjugated nanoPMOs. Subsequently, the effect of these properties on cancer cell targeting, drug loading and release capability, and anticancer activity is also studied. Due to the higher spatial resolution at the nanoscale, dSTORM imaging is able to reveal the structural properties (i.e., size and shape) of fluorescent and biodegradable nanoPMOs. The quantification of nanoPMOs' biodegradation using dSTORM imaging demonstrates their excellent structure-dependent degradation behavior at a higher glutathione concentration. The surface functionality of anti-M6PR antibody-conjugated nanoPMOs as quantified by dSTORM imaging plays a key role in prostate cancer cell labeling: oriented antibody is more effective than random ones, while high multivalency is also effective. The higher biodegradability and cancer cell-targeting properties of nanorods conjugated with oriented antibody (EAB4H) effectively deliver the anticancer drug doxorubicin to cancer cells, exhibiting potent anticancer effects.

JTD Keywords: drug-delivery, nanocapsules, nanomaterials, nanomedicine, release, Silica nanoparticles

JTD Keywords: alignment, bioresorbable stents, cells, design, electrospinning, everolimus, impact, in-vitro, poly(l-lactic-co-e-caprolactone), proliferation, release, sirolimus, Scaffold topography, Solvent-cast direct-writing

Polymeric spherical nucleic acids comprise pH-sensitive, polymer-conjugated oligonucleotides that self-assemble into nanoparticles with the ability to escape endosomes, overcoming a major obstacle in nucleic acid delivery.

JTD Keywords: c9orf72, cellular uptake, dna, encapsulation, expansion, ph, stability, trafficking, vesicles, Hexanucleotide repeat

JTD Keywords: 4-ethylenedioxythiophene), carbon nanoparticles, conductive hydrogel, manganese oxide poly(3, sensors, supercapacitor, supercapacitors, temperature sensor, Alginate, Carbon nanotubes

The second-line antileishmanial compound pentamidine is administered intramuscularly or, preferably, by intravenous infusion, with its use limited by severe adverse effects, including diabetes, severe hypoglycemia, myocarditis and renal toxicity. We sought to test the potential of phospholipid vesicles to improve the patient compliance and efficacy of this drug for the treatment of leishmaniasis by means of aerosol therapy. The targeting to macrophages of pentamidine-loaded liposomes coated with chondroitin sulfate or heparin increased about twofold (up to ca. 90%) relative to noncoated liposomes. The encapsulation of pentamidine in liposomes ameliorated its activity on the amastigote and promastigote forms of Leishmania infantum and Leishmania pifanoi, and it significantly reduced cytotoxicity on human umbilical endothelial cells, for which the concentration inhibiting 50% of cell viability was 144.2 ± 12.7 µM for pentamidine-containing heparin-coated liposomes vs. 59.3 ± 4.9 µM for free pentamidine. The deposition of liposome dispersions after nebulization was evaluated with the Next Generation Impactor, which mimics human airways. Approximately 53% of total initial pentamidine in solution reached the deeper stages of the impactor, with a median aerodynamic diameter of ~2.8 µm, supporting a partial deposition on the lung alveoli. Upon loading pentamidine in phospholipid vesicles, its deposition in the deeper stages significantly increased up to ~68%, and the median aerodynamic diameter decreased to a range between 1.4 and 1.8 µm, suggesting a better aptitude to reach the deeper lung airways in higher amounts. In all, nebulization of liposome-encapsulated pentamidine improved the bioavailability of this neglected drug by a patient-friendly delivery route amenable to self-administration, paving the way for the treatment of leishmaniasis and other infections where pentamidine is active.

JTD Keywords: aerosol therapy, delivery-systems, drug encapsulation, drugs, ex-vivo models, formulation, leishmania infantum, leishmania pifanoi, leishmaniasis, liposomes, macrophages, miltefosine, pentamidine, pharmacology, pulmonary absorption, visceral leishmaniasis, Aerosol therapy, Amphotericin-b treatment, Drug encapsulation, Leishmania infantum, Leishmania pifanoi, Leishmaniasis, Liposomes, Pentamidine

In addition of a key catecholamine neurotransmitter, dopamine is is the metabolite predominantly produced by specific types of tumors (e.g. paragangliomas and neuroblastomas), which cannot be diagnosed using conven-tional sensitive tests. Within this context, development of flexible electrochemical sensors to monitor dopamine levels in physiological fluids for the early diagnosis and control of diseases related to abnormal levels of such compound, is necessary. In this work, a flexible self-supported membrane, which acts directly as electrode, has been developed to detect dopamine. The membrane consists of three nanoperforated polylactic acid (PLA) layers, which provide flexibility and mechanical integrity, separated by two layers of an electroactive copolymer, which are obtained by electrochemical copolymerization of 3,4-ethylenedioxythiophene and aniline. The sensitivity and detection limit provided by the electroactive copolymer, which is accessible to dopamine molecules through the nanoperforations of the PLA outer layers, is 1.846 mu A/(cm2.mu M) and 1.7 mu M, respectively, in a urea-rich environments that mimics urine. These values allow us to propose the self-standing flexible electrodes devel-oped in this study for the detection of dopamine in patients affected by paragangliomas and neuroblastomas tumors, which typically present dopamine concentrations between 2 and 7 mu M.

JTD Keywords: 4-ethylenedioxythiophene), Conducting polymer, Electrochemical sensor, Electrodes, Hydrogels, Poly(3, Polyaniline, Polylactic acid, Selective detection, Sensors, Supercapacitors

JTD Keywords: capacitive deionization, chitosan-based hydrogels, composite, desalination, n-isopropylacrylamide, poly(n-isopropylacrylamide), polymers, swelling behavior, thermosensitive hydrogels, walled carbon nanotubes, water cleaning, water evaporation, Biomedical sensors, Critical solution temperature

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

This work presents the analytical physical modeling of undoped organic metal-electrolyte-semiconductor (OMES) capacitors in the framework of the Nernst-Planck-Poisson theory, including the presence of compact interfacial layers. This work derives an exact analytical solution, up to a quadrature, for the stationary electric potential and charge density distributions in both the semiconductor film and the electrolyte solution, and from them the sheet semiconductor charge and the stationary differential capacitance are obtained as a function of the applied voltage. The dependence of these magnitudes on the physical device parameters, like the ionic concentration of the electrolyte, the capacitance of the interfacial compact layers and the injected hole density is then analyzed. This work shows that ionic diffusive effects in the electrolyte can play an important role in the device response, inducing a broadening of the transition from the weak to the strong accumulation regimes. This fact can make that the strong accumulation regime is not achieved in OMES within the usual voltage operation range of these devices. The analytical solution is validated by means of finite element numerical calculations. The implications of the results obtained on the physics of electrolyte gated organic field effect transistors (EGOFETs) are discussed.

JTD Keywords: Analytical model, Equivalent-circuit model, Metal electrolyte semiconductor capacitors, Metal insulator semiconductor capacitors, Organic devices

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

Chronic obstructive pulmonary disease (COPD) requires a multifactorial assessment, evaluating the airflow limitation and symptoms of the patients. The 6-min walk test (6MWT) is commonly used to evaluate the functional exercise capacity in these patients. This study aims to propose a novel predictive model of the major 6MWT outcomes for COPD assessment, without physical performance measurements.Cardiopulmonary and clinical parameters were obtained from fifty COPD patients. These parameters were used as inputs of a Bayesian network (BN), which integrated three multivariate models including the 6-min walking distance (6MWD), the maximum HR (HRmax) after the walking, and the HR decay 3 min after (HRR3). The use of BN allows the assessment of the patients' status by predicting the 6MWT outcomes, but also inferring disease severity parameters based on actual patient's 6MWT outcomes.Firstly, the correlation obtained between the estimated and actual 6MWT measures was strong (R = 0.84, MAPE = 8.10% for HRmax) and moderate (R = 0.58, MAPE = 15.43% for 6MWD and R = 0.58, MAPE = 32.49% for HRR3), improving the classical methods to estimate 6MWD. Secondly, the classification of disease severity showed an accuracy of 78.3% using three severity groups, which increased up to 84.4% for two defined severity groups.We propose a powerful two-way assessment tool for COPD patients, capable of predicting 6MWT outcomes without the need for an actual walking exercise. This model-based tool opens the way to implement a continuous monitoring system for COPD patients at home and to provide more personalized care.Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.

JTD Keywords: 6mwt, bayesian networks, copd, distance, exercise capacity, physical capacity, reference equations, severity, survival, wearables, 6mwt, Bayes theorem, Bayesian networks, Copd, Exercise test, Exercise tolerance, Heart-rate recovery, Humans, Physical capacity, Physical functional performance, Pulmonary disease, chronic obstructive, Walking, Wearables

Abstract Nerves are a component of the tumor microenvironment contributing to cancer progression, but the role of cells from nerves in facilitating cancer invasion remains poorly understood. Here we show that Schwann cells (SCs) activated by cancer cells collectively function as Tumor Activated Schwann cell Tracks (TASTs) that promote cancer cell migration and invasion. Non-myelinating SCs form TASTs and have cell gene expression signatures that correlate with diminished survival in patients with pancreatic ductal adenocarcinoma. In TASTs, dynamic SCs form tracks that serve as cancer pathways and apply forces on cancer cells to enhance cancer motility. These SCs are activated by c-Jun, analogous to their reprogramming during nerve repair. This study reveals a mechanism of cancer cell invasion that co-opts a wound repair process and exploits the ability of SCs to collectively organize into tracks. These findings establish a novel paradigm of how cancer cells spread and reveal therapeutic opportunities.

JTD Keywords: dissemination, escape, mechanisms, progression, Perineural invasion

Type 1 Diabetes results from autoimmune response elicited against β-cell antigens. Nowadays, insulin injections remain the leading therapeutic option. However, injection treatment fails to emulate the highly dynamic insulin release that β-cells provide. 3D cell-laden microspheres have been proposed during the last years as a major platform for bioengineering insulin-secreting constructs for tissue graft implantation and a model for in vitro drug screening platforms. Current microsphere fabrication technologies have several drawbacks: the need for an oil phase containing surfactants, diameter inconsistency of the microspheres, and high time-consuming processes. These technologies have widely used alginate for its rapid gelation, high processability, and low cost. However, its low biocompatible properties do not provide effective cell attachment. This study proposes a high-throughput methodology using a 3D bioprinter that employs an ECM-like microenvironment for effective cell-laden microsphere production to overcome these limitations. Crosslinking the resulting microspheres with tannic acid prevents collagenase degradation and enhances spherical structural consistency while allowing the diffusion of nutrients and oxygen. The approach allows customization of microsphere diameter with extremely low variability. In conclusion, a novel bio-printing procedure is developed to fabricate large amounts of reproducible microspheres capable of secreting insulin in response to extracellular glucose stimuli.© 2022 The Authors. Advanced Materials Technologies published by Wiley‐VCH GmbH.

JTD Keywords: 3d bioprinter, beta-cell, biomaterial, collagen, encapsulation, mechanics, microspheres, survival, 3d bioprinter, ?-cell, Advanced material technologies, Biocompatibility, Cell encapsulations, Cells, Collagen, Cross-linking, Cytology, Drug delivery, Encapsulation, Fabrication, Flavonoids, Gelation, In-vitro, Insulin injections, Insulin release, Microspheres, Tannic acid, Tannins, Throughput, Tissue grafts, Type 1 diabetes, Β‐cell

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

Poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) enhance the delivery of therapeutic enzymes for replacement therapy of lysosomal storage disorders. Previous studies examined NPs encapsulating or coated with enzymes, but these formulations have never been compared. We examined this using hyaluronidase (HAse), deficient in mucopolysaccharidosis IX, and acid sphingomyelinase (ASM), deficient in types A–B Niemann–Pick disease. Initial screening of size, PDI, ζ potential, and loading resulted in the selection of the Lactel II co-polymer vs. Lactel I or Resomer, and Pluronic F68 surfactant vs. PVA or DMAB. Enzyme input and addition of carrier protein were evaluated, rendering NPs having, e.g., 181 nm diameter, 0.15 PDI, −36 mV ζ potential, and 538 HAse molecules encapsulated per NP. Similar NPs were coated with enzyme, which reduced loading (e.g., 292 HAse molecules/NP). NPs were coated with targeting antibodies (> 122 molecules/NP), lyophilized for storage without alterations, and acceptably stable at physiological conditions. NPs were internalized, trafficked to lysosomes, released active enzyme at lysosomal conditions, and targeted both peripheral organs and the brain after i.v. administration in mice. While both formulations enhanced enzyme delivery compared to free enzyme, encapsulating NPs surpassed coated counterparts (18.4- vs. 4.3-fold enhancement in cells and 6.2- vs. 3-fold enhancement in brains), providing guidance for future applications.

JTD Keywords: active enzymes, encapsulation, enhanced delivery, enzyme therapeutics, formulation parameters, icam-1 targeting, icam-1-targeted nanocarriers, in vivo biodistribution, in-vitro, lysosomal delivery, model, oral delivery, plga nanoparticles, poly(lactic-co-glycolic acid) nanoparticles, protein therapeutics, surface loading, Acid sphingomyelinase, Animals, Encapsulation, Enzyme replacement therapy, Enzyme therapeutics, Icam-1 targeting, In vivo biodistribution, Lysosomal delivery, Lysosomes, Mice, Nanoparticles, Poly(lactic-co-glycolic acid) nanoparticles, Polymers, Surface loading, Surface-active agents

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

0

JTD Keywords: cancer, cells, cellular basis, delivery, encapsulation, in-vitro, inflammation, macrophage, methotrexate, pathogenesis, polymersome, polymersomes, synoviocyte, Arthritis, Rheumatoid-arthritis

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

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

Nowadays, liquid biopsy represents one of the most promising techniques for early diagnosis, monitoring, and therapy screening of cancer. This novel methodology includes, among other techniques, the isolation, capture, and analysis of circulating tumor cells (CTCs). Nonetheless, the identification of CTC from whole blood is challenging due to their extremely low concentration (1-100 per ml of whole blood), and traditional methods result insufficient in terms of purity, recovery, throughput and/or viability of the processed sample. In this context, the development of microfluidic devices for detecting and isolating CTCs offers a wide range of new opportunities due to their excellent properties for cell manipulation and the advantages to integrate and bring different laboratory processes into the microscale improving the sensitivity, portability, reducing cost and time. This chapter explores current and recent microfluidic approaches that have been developed for the analysis and detection of CTCs, which involve cell capture methods based on affinity binding and label-free methods and detection based on electrical, chemical, and optical sensors. All the exposed technologies seek to overcome the limitations of commercial systems for the analysis and isolation of CTCs, as well as to provide extended analysis that will allow the development of novel and more efficient diagnostic tools.

JTD Keywords: Cancer detection, Cancer diagnosis, Cancer-cells, Capture, Chip, Circulating tumor cells, Enrichment, Label-free isolation, Liquid biopsy, Microchannel, Microfluidics, Separation, Ultra-fast

JTD Keywords: biocompatibility, bioresorbable stents, degradation, mechanical-properties, poly(epsilon-caprolactone), poly-l-lactic acid, polylactide, radiopacity, thermogel, x-ray imaging, Barium sulfate, Biocompatibility, Bioresorbable, Bioresorbable scaffolds, Bioresorbable stent, Bioresorbable stents, Blood vessels, Computerized tomography, Controlled drug delivery, Coronary heart disease, Direct-writing, Endothelial cells, Fabrication strategies, Injection molding, Lactic acid, Poly-l-lactic acid, Poly-l-lactic acids, Radiopacity, Scaffolds (biology), Solvent cast, Solvent-cast direct-writing, Solvents, Stents, Struts, Sulfur compounds, Targeted drug delivery, X-ray imaging

Nanowire-based nanocomposite materials are being developed as transparent and flexible electrodes or as stretchable conductors and dielectrics for biosensing. Here, we theoretically investigate the use of scanning dielectric microscopy (SDM) to characterize these materials in a nondestructive way, with a special focus on the achievable spatial resolution and the possibility of detection of the capacitive coupling between nearby nanowires. Numerical calculations with models involving single and multiple buried nanowires have been performed. We demonstrate that the capacitance gradient spread function of a single buried nanowire consists of a modified Lorenzianan with a cubic decay. We show that the achievable spatial resolution can be determined with good accuracy with the help of this spread function. It is shown that, in general, the spatial resolution worsens when any system parameter decreases the maximum of the nanowire spread function or increases its width, or both. Finally, we show that SDM measurements are also sensitive to the capacitive coupling between nearby nanowires. This latter result is of utmost relevance since the macroscopic electric properties of nanowire nanocomposites largely depend on the electric interaction between nearby nanowires. The present results show that SDM can be a valuable nondestructive subsurface characterization technique for nanowire nanocomposite materials.

JTD Keywords: depth, electrodes, nanocomposites, nanowires, sdm, spatial resolution, subsurface, tomography, Capacitive coupling, Force microscopy, Nanocomposites, Nanowires, Sdm, Spatial resolution, Subsurface

The epidemic spread of many viral infections is mediated by the environmental conditions and influenced by the ambient humidity. Single virus particles have been mainly visualized by atomic force microscopy (AFM) in liquid conditions, where the effect of the relative humidity on virus topography and surface cannot be systematically assessed. In this work, we employed multi-frequency AFM, simultaneously with standard topography imaging, to study the nanoscale wetting of individual Tobacco Mosaic virions (TMV) from ambient relative humidity to water condensation (RH > 100%). We recorded amplitude and phase vs. distance curves (APD curves) on top of single virions at various RH and converted them into force vs. distance curves. The high sensitivity of multifrequency AFM to visualize condensed water and sub-micrometer droplets, filling gaps between individual TMV particles at RH > 100%, is demonstrated. Dynamic force spectroscopy allows detecting a thin water layer of thickness ⁓1 nm, adsorbed on the outer surface of single TMV particles at RH < 60%.

JTD Keywords: amplitude-modulation am-afm, atomic-force microscopy, capillary, force reconstruction, multifrequency afm, nanoscale wetting, persistence, reconstruction, relative-humidity, surfaces, tobacco mosaic virus (tmv), tobamovirus, transmission, water, Amplitude-modulation am-afm, Force reconstruction, Humidity, Microscopy, atomic force, Multifrequency afm, Nanoscale wetting, Tobacco mosaic virus, Tobacco mosaic virus (tmv), Tobacco mosaic virus (tmv), nanoscale wetting, Tobacco-mosaic-virus, Virion, Water, Wettability

In vitro research for the study of type 2 diabetes (T2D) is frequently limited by the availability of a functional model for islets of Langerhans. To overcome the limitations of obtaining pancreatic islets from different sources, such as animal models or human donors, immortalized cell lines as the insulin-producing INS1E beta-cells have appeared as a valid alternative to model insulin-related diseases. However, immortalized cell lines are mainly used in flat surfaces or monolayer distributions, not resembling the spheroid-like architecture of the pancreatic islets. To generate islet-like structures, the use of scaffolds appeared as a valid tool to promote cell aggregations. Traditionally-used hydrogel encapsulation methods do not accomplish all the requisites for pancreatic tissue engineering, as its poor nutrient and oxygen diffusion induces cell death. Here, we use cryogelation technology to develop a more resemblance scaffold with the mechanical and physical properties needed to engineer pancreatic tissue. This study shows that carboxymethyl cellulose (CMC) cryogels prompted cells to generate beta-cell clusters in comparison to gelatin-based scaffolds, that did not induce this cell organization. Moreover, the high porosity achieved with CMC cryogels allowed us to create specific range pseudoislets. Pseudoislets formed within CMC-scaffolds showed cell viability for up to 7 d and a better response to glucose over conventional monolayer cultures. Overall, our results demonstrate that CMC-scaffolds can be used to control the organization and function of insulin-producing beta-cells, representing a suitable technique to generate beta-cell clusters to study pancreatic islet function.

JTD Keywords: biomaterial, cryogel, pancreatic islets, scaffold, tissue engineering, ?-cell, Animals, Architecture, Beta-cell, Beta-cell heterogeneity, Biomaterial, Carboxymethyl cellulose, Cell culture, Cell death, Cell engineering, Cell organization, Cells, Cellulose, Cryogel, Cryogels, Cytoarchitecture, Delivery, Diabetes mellitus, type 2, Encapsulation methods, Gelation, Gene-expression, Humans, Immortalized cells, Insulin, Insulin secretory responses, Islets of langerhans, Islets of langerhans transplantation, Mechanical and physical properties, Monolayer culture, Monolayers, Pancreatic islets, Pancreatic tissue, Pancreatic-islets, Proliferation, Scaffold, Scaffolds, Scaffolds (biology), Size, Tissue, Tissue engineering, Tissue scaffolds, Β-cell

Breast cancer is the most frequent type of cancer and the major cause of mortality in women. The rapid development of various therapeutic options has led to the improvement of treatment outcomes; nevertheless, one-third of estrogen receptor (ER)-positive patients relapse due to cancer cell acquired resistance. Here, we use dynamic BH3 profiling (DBP), a functional predictive assay that measures net changes in apoptotic priming, to find new effective treatments for ER+ breast cancer. We observed anti-apoptotic adaptations upon treatment that pointed to metronomic therapeutic combinations to enhance cytotoxicity and avoid resistance. Indeed, we found that the anti-apoptotic proteins BCL-xL and MCL-1 are crucial for ER+ breast cancer cells resistance to therapy, as they exert a dual inhibition of the pro-apoptotic protein BIM and compensate for each other. In addition, we identified the AKT inhibitor ipatasertib and two BH3 mimetics targeting these anti-apoptotic proteins, S63845 and A-1331852, as new potential therapies for this type of cancer. Therefore, we postulate the sequential inhibition of both proteins using BH3 mimetics as a new treatment option for refractory and relapsed ER+ breast cancer tumors.

JTD Keywords: apoptosis, bh3 mimetics, cell-line, chemotherapy, classification, dbp, death, er+ breast cancer, fulvestrant, her2, inhibitor, kinase, pik3ca, priming, resistance, targeted therapies, Apoptosis, Bh3 mimetics, Dbp, Endocrine therapy, Er plus breast cancer, Er+ breast cancer, Priming, Resistance, Targeted therapies

Amphiphilic block co-polymer nanoparticles are interesting candidates for drug delivery as a result of their unique properties such as the size, modularity, biocompatibility and drug loading capacity. They can be rapidly formulated in a nanoprecipitation process based on self-assembly, resulting in kinetically locked nanostructures. The control over this step allows us to obtain nanoparticles with tailor-made properties without modification of the co-polymer building blocks. Furthermore, a reproducible and controlled formulation supports better predictability of a batch effectiveness in preclinical tests. Herein, we compared the formulation of PLGA-PEG nanoparticles using the typical manual bulk mixing and a microfluidic chip-assisted nanoprecipitation. The particle size tunability and controllability in a hydrodynamic flow focusing device was demonstrated to be greater than in the manual dropwise addition method. We also analyzed particle size and encapsulation of fluorescent compounds, using the common bulk analysis and advanced microscopy techniques: Transmission Electron Microscopy and Total Internal Reflection Microscopy, to reveal the heterogeneities occurred in the formulated nanoparticles. Finally, we performed in vitro evaluation of obtained NPs using MCF-7 cell line. Our results show how the microfluidic formulation improves the fine control over the resulting nanoparticles, without compromising any appealing property of PLGA nanoparticle. The combination of microfluidic formulation with advanced analysis methods, looking at the single particle level, can improve the understanding of the NP properties, heterogeneities and performance.

JTD Keywords: controlled-release, doxorubicin, encapsulation, functional nanoparticles, nanoprecipitation, pharmacokinetics, polymeric nanoparticles, shape, surface-chemistry, Breast neoplasms, Drug carriers, Drug delivery systems, Female, Humans, In-vitro, Mcf-7 cells, Microfluidics, Nanoparticles, Polyesters, Polyethylene glycol-poly(lactide-co-glycolide), Polyethylene glycols, Polymers

Liposomes are widely used as drug delivery carriers and as cell model systems. Here, we measure the dielectric properties of individual liposomes adsorbed on a metal electrode by in-liquid scanning dielectric microscopy in force detection mode. From the measurements the lamellarity of the liposomes, the separation between the lamellae and the specific capacitance of the lipid bilayer can be obtained. As application we considered the case of non-extruded DOPC liposomes with radii in the range ~ 100–800 nm. Uni-, bi- and tri-lamellar liposomes have been identified, with the largest population corresponding to bi-lamellar liposomes. The interlamellar separation in the bi-lamellar liposomes is found to be below ~ 10 nm in most instances. The specific capacitance of the DOPC lipid bilayer is found to be ~ 0.75 µF/cm2 in excellent agreement with the value determined on solid supported planar lipid bilayers. The lamellarity of the DOPC liposomes shows the usual correlation with the liposome's size. No correlation is found, instead, with the shape of the adsorbed liposomes. The proposed approach offers a powerful label-free and non-invasive method to determine the lamellarity and dielectric properties of single liposomes. [Figure not available: see fulltext.].

JTD Keywords: constant, force, lamellarity, liposomes, membrane capacitance, model, nanoscale, scanning dielectric microscopy, Lamellarity, Liposomes, Membrane capacitance, Nanoscale, Polarization properties, Scanning dielectric microscopy

Mapping the dielectric properties of cells with nanoscale spatial resolution can be an im-portant tool in nanomedicine and nanotoxicity analysis, which can complement structural and mechanical nanoscale measurements. Recently we have shown that dielectric constant maps can be obtained on dried fixed cells in air environment by means of scanning dielectric force volume mi-croscopy. Here, we demonstrate that such measurements can also be performed in the much more challenging case of fixed cells in liquid environment. Performing the measurements in liquid media contributes to preserve better the structure of the fixed cells, while also enabling accessing the local dielectric properties under fully hydrated conditions. The results shown in this work pave the way to address the nanoscale dielectric imaging of living cells, for which still further developments are required, as discussed here.

JTD Keywords: atomic force microscopy (afm), capacitance, constant, dielectric properties, electrostatic force microscopy (efm), functional microscopy, nanoscale, scanning dielectric microscopy (sdm), Atomic force microscopy (afm), Dielectric properties, Dielectrophoretic separation, Electrostatic force microscopy (efm), Functional micros-copy, Functional microscopy, Scanning dielectric microscopy (sdm), Scanning probe microscopy (spm)

Purpose: This paper aims to explore how the cross-fertilization of knowledge and technologies in EU-funded research projects, including serious games and gamification, is influenced by the following variables: multidisciplinarity, knowledge base and organizations (number and diversity). The interrelation of actors and projects form a network of innovation. The largest contribution to cross-fertilization comes from the multidisciplinary nature of projects and the previous knowledge and technology of actors. The analysis draws on the understanding of how consortia perform as an innovation network, what their outcomes are and what capabilities are needed to reap value. Design/methodology/approach: All the research projects including serious games and/or gamification, funded by the EU-Horizon 2020 work programme, have been analyzed to test the hypotheses in this paper. The study sample covers the period between 2014 and 2016 (June), selecting the 87 research projects that comprised 519 organizations as coordinators and participants, and 597 observations – because more organizations participate in more than one project. The data were complemented by documentary and external database analysis. Findings: To create cross-fertilization of knowledge and technologies, the following emphasis should be placed on projects: partners concern various disciplines; partners have an extensive knowledge base for generating novel combinations and added-value technologies; there is a diverse typology of partners with unique knowledge and skills; and there is a limited number of organizations not too closely connected to provide cross-fertilization. Research limitations/implications: First, the database sample covers a period of 30 months. The authors’ attention was focused on this period because H2020 prioritized for the first time the serious games and gamification with two specific calls (ICT-21–14 and ICT-24–16) and, second, for the explosion of projects including these technologies in the past years (Adkins, 2017). These facts can be understood as a way to push the research to higher technology readiness levels (TRLs) and introducing the end-user in the co-creation and co-development along the value chain. Second, an additional limitation makes reference to the European focus of the projects, missing strong regional initiatives not identified and studied. Originality/value: This paper has attempted to explore and define theoretically and empirically the characteristics found in the cross-fertilization of collaborative research projects, emphasizing which variables, and how, need to be stimulated to benefit more multidisciplinary consortia and accelerate the process of innovation. © 2021, Manel González-Piñero, Cristina Páez-Avilés, Esteve Juanola-Feliu and Josep Samitier.

JTD Keywords: absorptive-capacity, business model, cross-fertilization of knowledge, diversity, front-end, impact, innovation systems, knowledge management, management research, science, social networks, team, technology, Cross-fertilization of knowledge, Innovation, Knowledge management, Management research, Research-and-development, Technology

© 2021 Wiley-VCH GmbH One limitation of wearable electronics, and at the same time a challenge, is the lack of energy storage devices with multiple functionalities produced using clean and environmental-friendly strategies. Here, a multifunctional conductive hydrogel containing poly(3,4-ethylenedioxythiophene) (PEDOT) and alginate is fabricated, to be used as electrodes in supercapacitors, by applying water-mediated self-assembly and polymerization processes at room temperature. The interpenetration of both polymers allows the combination of flexibility and self-healing properties within the same hydrogel together with the intrinsic biocompatibility and sustainability of such materials. Initially, PEDOT:polystyrene sulfonate and alginate aqueous solutions are mixed in two different proportions (1:1 and 1:3) and ionically crosslinked with CaCl2. Subsequently, re-interpenetration of poly(hydroxymethyl-3,4-ethylenedioxythiophene) by anodic polymerization in CaCl2 aqueous solution is achieved. Re-interpenetrated 1:3 PEDOT/alginate hydrogels show excellent capacitance values (35 mF cm−2) and good capacitance retention. In addition, the electrochemical properties are not significantly changed after many cutting/self-healing cycles as observed by cyclic voltammetry. Therefore, this sustainably produced hydrogel shows promising properties for use in wearable energy storage devices.

JTD Keywords: flexibility, pedot:pss-alginate hydrogels, self-healing, sustainability, Electrochemical supercapacitors, Flexibility, Pedot:pss-alginate hydrogels, Self-healing, Sustainability

Flexible hydrogels are attracting significant interest as solid-like electrolytes for energy storage devices, especially for supercapacitors, because of their lightweight and anti-deformation features. Here, we present a comparative study of four ionic conductive hydrogels derived from biopolymers and doped with 0.1 M NaCl. More specifically, such hydrogels are constituted by κcarrageenan (κC), carboxymethyl cellulose (CMC), poly-γ-glutamic acid (PGGA) or a phenylalaninecontaining polyesteramide (PEA). After examining the morphology and the swelling ratio of the four hydrogels, which varies between 483% and 2356%, their electrical and capacitive behaviors were examined using electrochemical impedance spectroscopy. Measurements were conducted on devices where a hydrogel film was sandwiched between two identical poly(3,4-ethylenedioxythiophene) electrodes. The bulk conductivity of the prepared doped hydrogels is 76, 48, 36 and 34 mS/cm for PEA, PGGA, κC and CMC, respectively. Overall, the polyesteramide hydrogel exhibits the most adequate properties (i.e., low electrical resistance and high capacitance) to be used as semi-solid electrolyte for supercapacitors, which has been attributed to its distinctive structure based on the homogeneous and abundant distribution of both micro-and nanopores. Indeed, the morphology of the polyestermide hydrogel reduces the hydrogel resistance, enhances the transport of ions, and results in a better interfacial contact between the electrodes and solid electrolyte. The correlation between the supercapacitor performance and the hydrogel porous morphology is presented as an important design feature for the next generation of light and flexible energy storage devices for wearable electronics.

JTD Keywords: biopolymers, electrochemical impedance spectroscopy, flexible hydrogels, supercapacitor, Biopolymers, Electrochemical impedance spectroscopy, Flexible hydrogels, Supercapacitor

© 2020 Wiley-VCH GmbH In recent years, magnetism has gained an enormous amount of interest among researchers for actuating different sizes and types of bio/soft robots, which can be via an electromagnetic-coil system, or a system of moving permanent magnets. Different actuation strategies are used in robots with magnetic actuation having a number of advantages in possible realization of microscale robots such as bioinspired microrobots, tetherless microrobots, cellular microrobots, or even normal size soft robots such as electromagnetic soft robots and medical robots. This review provides a summary of recent research in magnetically actuated bio/soft robots, discussing fabrication processes and actuation methods together with relevant applications in biomedical area and discusses future prospects of this way of actuation for possible improvements in performance of different types of bio/soft robots.

JTD Keywords: capsule endoscope, controlled propulsion, conventional gastroscopy, digital microfluidics, guided capsule, liquid-metal, magnetic drug delivery, magnetic microrobots, magnetically guided capsule endoscopy, magnetotactic bacteria, nanoscribe ip-dip, navigation system, Gallium-indium egain, Magnetic bioinspired micromanipulation, Magnetic drug delivery, Magnetic microrobots, Magnetically guided capsule endoscopy, Magnetotactic bacteria

© 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

Degenerative cartilage pathologies are nowadays a major problem for the world population. Factors such as age, genetics or obesity can predispose people to suffer from articular cartilage degeneration, which involves severe pain, loss of mobility and consequently, a loss of quality of life. Current strategies in medicine are focused on the partial or total replacement of affected joints, physiotherapy and analgesics that do not address the underlying pathology. In an attempt to find an alternative therapy to restore or repair articular cartilage functions, the use of bioengineered tissues is proposed. In this study we present a three-dimensional (3D) bioengineered platform combining a 3D printed polycaprolactone (PCL) macrostructure with RAD16-I, a soft nanofibrous self-assembling peptide, as a suitable microenvironment for human mesenchymal stem cells’ (hMSC) proliferation and differentiation into chondrocytes. This 3D bioengineered platform allows for long-term hMSC culture resulting in chondrogenic differentiation and has mechanical properties resembling native articular cartilage. These promising results suggest that this approach could be potentially used in articular cartilage repair and regeneration.

JTD Keywords: 3D printing, Chondrogenic differentiation, Polycaprolactone, RAD16-I self-assembling peptide

Simultaneous drug release and monitoring using a single polymeric platform represents a significant advance in the utilization of biomaterials for therapeutic use. Tracking drug release by real-time electrochemical detection using the same platform is a simple way to guide the dosage of the drug, improve the desired therapeutic effect, and reduce the adverse side effects. The platform developed in this work takes advantage of the flexibility and loading capacity of hydrogels, the mechanical strength of microfibers, and the capacity of conducting polymers to detect the redox properties of drugs. The engineered platform is prepared by assembling two spin-coated layers of poly-γ-glutamic acid hydrogel, loaded with poly(3,4-ethylenedioxythiophene) (PEDOT) microparticles, and separated by a electrospun layer of poly-ε-caprolactone microfibers. Loaded PEDOT microparticles are used as reaction nuclei for the polymerization of poly(hydroxymethyl-3,4-ethylenedioxythiophene) (PHMeDOT), that semi-interpenetrate the whole three layered system while forming a dense network of electrical conduction paths. After demonstrating its properties, the platform is loaded with levofloxacin and its release monitored externally by UV–vis spectroscopy and in situ by using the PHMeDOT network. In situ real-time electrochemical monitoring of the drug release from the engineered platform holds great promise for the development of multi-functional devices for advanced biomedical applications.

JTD Keywords: Biosensors, Conducting polymers, Drug delivery, Poly-γ-glutamic acid, Poly-ε-caprolactone

Background: Severe hepatic steatosis shows a high prevalence and contributes to morbidity and mortality in human immunodeficiency virus (HIV) infected patients. Known risk factors include obesity, dyslipidaemia and features of metabolic syndrome. Fibroblast growth factor 21 (FGF-21) is involved with hepatic glucose and lipid metabolism. This study aimed to evaluate FGF-21 as a biomarker for severe hepatic steatosis in non-obese HIV-infected patients. Methods: This is a prospective, cross-sectional, monocentric study including HIV-infected out-patients. Hepatic steatosis was measured via controlled attenuation parameter (CAP) using FibroScan 502 touch (ECHOSENS, France). Severe hepatic steatosis was defined at CAP ≥ 253 dB/m. Peripheral blood samples were collected and plasma was analysed for FGF-21. Demographic and clinical characteristics were collected from patient's health records. Results: In total, 73 non-obese HIV-monoinfected patients were included in this study. Prevalence of severe hepatic steatosis was 41%. Patients with severe hepatic steatosis showed significantly higher levels of FGF-21. Univariate analysis revealed FGF-21, BMI, hyperlipidaemia, ALT levels and arterial hypertension as significant, while multivariate analysis showed only FGF-21, arterial hypertension and ALT levels as significant independent risk factors for severe hepatic steatosis. Conclusion: This study presents FGF-21 as an independent and stronger predictor of severe hepatic steatosis than blood lipids in HIV-infected patients. Moreover, arterial hypertension and ALT levels predict severe steatosis even in non-obese HIV-monoinfected patients. Furthermore, this study supports existing metabolic risk factors and expands them to non-obese HIV-infected patients.

JTD Keywords: BMI, CAP, Dyslipidaemia, FGF-21, Fibroscan, HIV, Hyperlipidaemia, Liver, NAFLD, NASH, Steatosis

Understanding how polyprotic compounds distribute within liposome (LP) suspensions is of major importance to design effective drug delivery strategies. Advances in this research field led to the definition of LP-based active drug encapsulation methods driven by transmembrane pH gradients with evidenced efficacy in the management of cancer and infectious diseases. An accurate modeling of membrane-solution drug partitioning is also fundamental when designing drug delivery systems for poorly endocytic cells, such as red blood cells (RBCs), in which the delivered payloads rely mostly on the passive diffusion of drug molecules across the cell membrane. Several experimental models have been proposed so far to predict the partitioning of polyprotic basic/acid drugs in artificial membranes. Nevertheless, the definition of a model in which the membrane-solution partitioning of each individual drug microspecies is studied relative to each other is still a topic of ongoing research. We present here a novel experimental approach based on mathematical modeling of drug encapsulation efficiency (EE) data in liposomal systems by which microspecies-specific partition coefficients are reported as a function of pH and phospholipid compositions replicating the RBC membrane in a simple and highly translatable manner. This approach has been applied to the study of several diprotic basic antimalarials of major clinical importance (quinine, primaquine, tafenoquine, quinacrine, and chloroquine) describing their respective microspecies distribution in phosphatidylcholine-LP suspensions. Estimated EE data according to the model described here closely fitted experimental values with no significant differences obtained in 75% of all pH/lipid composition-dependent conditions assayed. Additional applications studied include modeling drug EE in LPs in response to transmembrane pH gradients and lipid bilayer asymmetric charge, conditions of potential interest reflected in our previously reported RBC-targeted antimalarial nanotherapeutics.

JTD Keywords: Distribution coefficient, Liposomal systems, Malaria therapy, Nanomedicine, Partition coefficient, PH-controlled drug encapsulation, Polyprotic drug, Targeted drug delivery

Ambroxol is a pharmacological chaperone (PC) for Gaucher disease that increases lysosomal activity of misfolded β-glucocerebrosidase (GCase) while displaying a safe toxicological profile. In this work, different poly(ε-caprolactone) (PCL)-based systems are developed to regulate the sustained release of small polar drugs in physiological environments. For this purpose, ambroxol is selected as test case since the encapsulation and release of PCs using polymeric scaffolds have not been explored yet. More specifically, ambroxol is successfully loaded in electrospun PCL microfibers, which are subsequently coated with additional PCL layers using dip-coating or spin-coating. The time needed to achieve 80% release of loaded ambroxol increases from ≈15 min for uncoated fibrous scaffolds to 3 days and 1 week for dip-coated and spin-coated systems, respectively. Furthermore, it is proven that the released drug maintains its bioactivity, protecting GCase against induced thermal denaturation.

JTD Keywords: Electrospinning, Gaucher's disease, Lysosomal storage disorders, Misfolding diseases, Poly(ε-caprolactone), Polyester, Release regulation

Fluid deformable surfaces are ubiquitous in cell and tissue biology, including lipid bilayers, the actomyosin cortex or epithelial cell sheets. These interfaces exhibit a complex interplay between elasticity, low Reynolds number interfacial hydrodynamics, chemistry and geometry, and govern important biological processes such as cellular traffic, division, migration or tissue morphogenesis. To address the modelling challenges posed by this class of problems, in which interfacial phenomena tightly interact with the shape and dynamics of the surface, we develop a general continuum mechanics and computational framework for fluid deformable surfaces. The dual solid–fluid nature of fluid deformable surfaces challenges classical Lagrangian or Eulerian descriptions of deforming bodies. Here, we extend the notion of arbitrarily Lagrangian–Eulerian (ALE) formulations, well-established for bulk media, to deforming surfaces. To systematically develop models for fluid deformable surfaces, which consistently treat all couplings between fields and geometry, we follow a nonlinear Onsager formalism according to which the dynamics minimizes a Rayleighian functional where dissipation, power input and energy release rate compete. Finally, we propose new computational methods, which build on Onsager’s formalism and our ALE formulation, to deal with the resulting stiff system of higher-order partial differential equations. We apply our theoretical and computational methodology to classical models for lipid bilayers and the cell cortex. The methods developed here allow us to formulate/simulate these models in their full three-dimensional generality, accounting for finite curvatures and finite shape changes.

JTD Keywords: Capsule/cell dynamics, Computational methods, Membranes

Prevalence and risk factors for hepatic steatosis (HS) in the human immunodeficiency virus (HIV)-positive population of western countries are controversially discussed and potentially confounded by coinfection with viral hepatitis. Significant HS (more than 10% of hepatocytes) can be accurately assessed using controlled attenuation parameter (CAP) determination. Aim of this study was to assess prevalence and factors associated with significant HS in HIV monoinfected patients. A total of 364 HIV-infected patients (289 monoinfected) were included in this prospective, cross-sectional study. All patients underwent CAP determination. Steatosis was classified as S1 (significant steatosis) with CAP > 238 dB/m, S2 with CAP > 260 dB/m, and S3 with CAP > 292 dB/m. Multivariable logistic regression analyses were performed to assess the factors associated with HS in this cohort. Significant HS was detected in 118 monoinfected patients (149 in the total cohort). In the total cohort as well as in the monoinfected patients alone, HS grade distribution showed a similar pattern (S1:29%, S2:34%, and S3:37%). Interestingly, patients with HS had a longer history of HIV infection and combined antiretroviral therapy (cART). Interalia, age, gender, ethnicity, and metabolic factors were strongly associated with HS, while body mass index (BMI), triglyceride, and glycated hemoglobin (HbA1c) levels were independently associated with significant HS. HS is highly prevalent among HIV monoinfected patients. Although metabolic risk factors, such as obesity and poorly controlled diabetes, are independently associated with HS in HIV monoinfected patients, cART and control of HIV seem to play an indirect role in the development of HS, probably through the return-to-health effect.

JTD Keywords: CAP, cART, HIV monoinfection, liver injury, NAFLD

Bioactive nanocomposite scaffolds with cell-adhesive surface have excellent bone regeneration capacities. Fibronectin (FN)-immobilized nanobioactive glass (nBG)/polycaprolactone (PCL) (FN-nBG/PCL) scaffolds with an open pore architecture were generated by a robotic-dispensing technique. The surface immobilization level of FN was significantly higher on the nBG/PCL scaffolds than on the PCL scaffolds, mainly due to the incorporated nBG that provided hydrophilic chemical-linking sites. FN-nBG/PCL scaffolds significantly improved cell responses, including initial anchorage and subsequent cell proliferation. Although further in-depth studies on cell differentiation and the in vivo animal responses are required, bioactive nanocomposite scaffolds with cell-favoring surface are considered to provide promising three-dimensional substrate for bone regeneration.

JTD Keywords: Bone scaffolds, Cell response, Fibronectin, Nanobioactive glass, Nanocomposites, Polycaprolactone, Bone, Cell proliferation, Cells, Cytology, Glass, Nanocomposites, Polycaprolactone, Robotics, Bone scaffolds, Bone tissue engineering, Cell response, Fibronectin, Fibronectin immobilizations, Nano bioactive glass, Nanocomposite scaffolds, Three-dimensional substrates, Scaffolds (biology)

JTD Keywords: antimalarial, heparin, magic bullet, malaria, nanomedicine, nanotechnology, nanovector, Plasmodium, polymers, targeted drug delivery, chloroquine, immunoliposome, liposome, nanoparticle, solid lipid nanoparticle, Anopheles, antimalarial activity, drug delivery system, drug efficacy, erythrocyte, human, IC50, malaria, malaria control, nanoencapsulation, nonhuman, pathophysiology, Plasmodium, Review

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

Heparin had been demonstrated to have antimalarial activity and specific binding affinity for Plasmodium-infected red blood cells (pRBCs) vs. non-infected erythrocytes. Here we have explored if both properties could be joined into a drug delivery strategy where heparin would have a dual role as antimalarial and as a targeting element of drug-loaded nanoparticles. Confocal fluorescence and transmission electron microscopy data show that after 30. min of being added to living pRBCs fluorescein-labeled heparin colocalizes with the intracellular parasites. Heparin electrostatically adsorbed onto positively charged liposomes containing the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane and loaded with the antimalarial drug primaquine was capable of increasing three-fold the activity of encapsulated drug in Plasmodium falciparum cultures. At concentrations below those inducing anticoagulation of mouse blood in vivo, parasiticidal activity was found to be the additive result of the separate activities of free heparin as antimalarial and of liposome-bound heparin as targeting element for encapsulated primaquine. From the Clinical Editor: Malaria remains an enormous global public health concern. In this study, a novel functionalized heparin formulation used as drug delivery agent for primaquine was demonstrated to result in threefold increased drug activity in cell cultures, and in a murine model it was able to provide these benefits in concentrations below what would be required for anticoagulation. Further studies are needed determine if this approach is applicable in the human disease as well.

JTD Keywords: Heparin, Liposomes, Malaria, Plasmodium, Targeted drug delivery, Heparin, Malaria, Plasmodium, Red blood cell, Targeted drug delivery, Liposomes, 1,2 dioleoyl 3 trimethylammoniopropane, fluorescein, heparin, liposome, nanoparticle, primaquine, adsorption, animal experiment, anticoagulation, antimalarial activity, Article, binding affinity, confocal microscopy, controlled study, drug targeting, encapsulation, erythrocyte, female, fluorescence microscopy, human, human cell, in vivo study, liposomal delivery, mouse, nonhuman, Plasmodium falciparum, transmission electron microscopy

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

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

We demonstrate the possibility to manipulate 25 nm radius gold nanoparticles in the three spatial dimensions with an atomic force microscope with the use of electroenhanced capillary forces. We show that an enhanced water-bridge can be electrostatically induced between a conducting probe and a metallic nanoparticle by the application of a voltage pulse, which is able to exert a pulling capillary force on the nanoparticle strong enough to detach it from the substrate. The nanoparticle can then be moved, attached to the probe, and placed back to the desired location on the substrate simply by contacting it.

JTD Keywords: Atomic force microscopy, Capillarity, Gold, Nanoparticles, Nanotechnology

Cognitive impairment is one of the main consequences of obstructive sleep apnea (OSA) and is usually attributed in part to the oxidative stress caused by intermittent hypoxia in cerebral tissues. The presence of oxygen-reactive species in the brain tissue should be produced by the deoxygenation-reoxygenation cycles which occur at tissue level during recurrent apneic events. However, how changes in arterial blood oxygen saturation (SpO(2)) during repetitive apneas translate into oxygen partial pressure (PtO2) in brain tissue has not been studied. The objective of this study was to assess whether brain tissue is partially protected from intermittently occurring interruption of O-2 supply during recurrent swings in arterial SpO(2) in an animal model of OSA. Methods: Twenty-four male Sprague-Dawley rats (300-350 g) were used. Sixteen rats were anesthetized and noninvasively subjected to recurrent obstructive apneas: 60 apneas/h, 15 s each, for 1 h. A control group of 8 rats was instrumented but not subjected to obstructive apneas. PtO2 in the cerebral cortex was measured using a fast-response oxygen microelectrode. SpO(2) was measured by pulse oximetry. The time dependence of arterial SpO(2) and brain tissue PtO2 was carried out by Friedman repeated measures ANOVA. Results: Arterial SpO(2) showed a stable periodic pattern (no significant changes in maximum [95.5 +/- 0.5%; m +/- SE] and minimum values [83.9 +/- 1.3%]). By contrast, brain tissue PtO2 exhibited a different pattern from that of arterial SpO(2). The minimum cerebral cortex PtO2 computed during the first apnea (29.6 +/- 2.4 mmHg) was significantly lower than baseline PtO2 (39.7 +/- 2.9 mmHg; p = 0.011). In contrast to SpO(2), the minimum and maximum values of PtO2 gradually increased (p < 0.001) over the course of the 60 min studied. After 60 min, the maximum (51.9 +/- 3.9 mmHg) and minimum (43.7 +/- 3.8 mmHg) values of PtO2 were significantly greater relative to baseline and the first apnea dip, respectively. Conclusions: These data suggest that the cerebral cortex is partially protected from intermittently occurring interruption of O-2 supply induced by obstructive apneas mimicking OSA.

JTD Keywords: Near-infrared spectroscopy, Sleep-apnea, Iintermittent hypoxia, Cerebral oxygenation, Oxidative stress, Blood-flow, Rat, Apoptosis, Inflammation, Hypercapnia

We present the experimental demonstration of low-frequency dielectric constant imaging of single-layer supported biomembranes at the nanoscale. The dielectric constant image has been quantitatively reconstructed by combining the thickness and local capacitance obtained using a scanning force microscope equipped with a sub-attofarad low-frequency capacitance detector. This work opens new possibilities for studying bioelectric phenomena and the dielectric properties of biological membranes at the nanoscale.

JTD Keywords: Atomic-force microscopy, Nnear-field microscopy, Purple membrane, Scanning capacitance, Biological-systems, Fluid, Spectroscopy, Resolution, Proteins, Dynamics

Horizontal acquisition of DNA by bacteria dramatically increases genetic diversity and hence successful bacterial colonization of several niches, including the human host. A relevant issue is how this newly acquired DNA interacts and integrates in the regulatory networks of the bacterial cell. The global modulator H-NS targets both core genome and HGT genes and silences gene expression in response to external stimuli such as osmolarity and temperature. Here we provide evidence that H-NS discriminates and differentially modulates core and HGT DNA. As an example of this, plasmid R27-encoded H-NS protein has evolved to selectively silence HGT genes and does not interfere with core genome regulation. In turn, differential regulation of both gene lineages by resident chromosomal H-NS requires a helper protein: the Hha protein. Tight silencing of HGT DNA is accomplished by H-NS-Hha complexes. In contrast, core genes are modulated by H-NS homoligomers. Remarkably, the presence of Hha-like proteins is restricted to the Enterobacteriaceae. In addition, conjugative plasmids encoding H-NS variants have hitherto been isolated only from members of the family. Thus, the H-NS system in enteric bacteria presents unique evolutionary features. The capacity to selectively discriminate between core and HGT DNA may help to maintain horizontally transmitted DNA in silent form and may give these bacteria a competitive advantage in adapting to new environments, including host colonization.

JTD Keywords: 2A strain 2457T, Escherichia-Coli, Salmonella-Enterica, Protein, DNA, Expression, Binding, HHA, Shigella, Plasmid

The cytoskeleton is a complex polymer network that regulates the structural stability of living cells. Although the cytoskeleton plays a key role in many important cell functions, the mechanisms that regulate its mechanical behaviour are poorly understood. Potential mechanisms include the entropic elasticity of cytoskeletal filaments, glassy-like inelastic rearrangements of cross-linking proteins and the activity of contractile molecular motors that sets the tensional stress (prestress) borne by the cytoskeleton filaments. The contribution of these mechanisms can be assessed by studying how cell mechanics depends on temperature. The aim of this work was to elucidate the effect of temperature on cell mechanics using atomic force microscopy. We measured the complex shear modulus (G*) of human alveolar epithelial cells over a wide frequency range (0.1-25.6 Hz) at different temperatures (13-37 degrees C). In addition, we probed cell prestress by mapping the contractile forces that cells exert on the substrate by means of traction microscopy. To assess the role of actomyosin contraction in the temperature-induced changes in G* and cell prestress, we inhibited the Rho kinase pathway of the myosin light chain phosphorylation with Y-27632. Our results show that with increasing temperature, cells become stiffer and more solid-like. Cell prestress also increases with temperature. Inhibiting actomyosin contraction attenuated the temperature dependence of G* and prestress. We conclude that the dependence of cell mechanics with temperature is dominated by the contractile activity of molecular motors.

JTD Keywords: Membrane Stress Failure, Frog Skeletal-Muscle, Extracellular-Matrix, Glass-Transition, Energy Landscape, Actin-Filaments, Living Cell, Single, Traction, Cytoskeleton

A hexavalent chromium-selective sensor, based on polymeric membranes containing trioctylphosphine oxide (TOPO) deposited on a Si/Sio(2)/Si3N4 structure, has been developed. The ion-sensitivity of TOPO was investigated by capacitance measurements (C-V) and electrochemical impedance spectroscopy. A quasi-nernstian response for Cr2O72- exchange is shown. Selectivity coefficients and detection limits of Cr(VI) in the presence of interfering anions were determined experimentally using the fixed interference method. A detection limit of 10(-5) M of Cr(VI) is obtained even in presence of sulphate and chloride ions.

JTD Keywords: Hexavalent chromium, Trioctylphosphine oxide, EIS, Siloprene membrane, Capacitance-voltage

Novel medical and biological applications are driving increased interest in the fabrication of micropipette or micro-dispensers. Reduced volume samples and drug dosages are prime motivators in this effort. We have combined microfabrication technology with ion beam milling techniques to successfully produce cantilever-type polysilicon micro-dispensers with 3D enclosed microchannels. The microfabrication technology described here allows for the designing of nozzles with multiple shapes. The contribution of ion beam milling has had a large impact on the fabrication process and on further customizing shapes of nozzles and inlet ports. Functionalization tests were conducted to prove the viability of ion beam-fabricated micro-dispensers. Self-assembled monolayers were successfully formed when a gold surface was patterned with a thiol solution dispensed by the fabricated micro-dispensers.

JTD Keywords: Dip-pen nanolithography, Silicon, Deposition, Microneedles, Delivery, Arrays, Polysilicon, Capillary, Systems, Gene