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High mannose has previously associated with insulin resistance and cardiovascular disease (CVD). Our objective is to establish whether mannose is associated with anatomical evidence of coronary artery disease (CAD).Plasma mannose concentrations were measured by liquid chromatography/tandem mass spectrometry in a discovery cohort (n = 513) and a validation cohort (n = 221) of carefully phenotyped individuals. In both cohorts CAD was quantitated using state-of-the-art imaging techniques (coronary computed coronary tomography angiography (CCTA), invasive coronary angiography and optical coherence tomography). Information on subsequent CVD events/death was collected. Associations of mannose with angiographic variables and biomarkers were tested using univariate and multivariate regression models. Survival analysis was performed using the Kaplan-Meier estimator.Mannose was related to indices of CAD and features of plaque vulnerability. In the discovery cohort, mannose was a marker of quantity and quality of CCTA-proven CAD and subjects with a mannose level in the top quartile had a significantly higher risk of CVD events/death (p = 3.6e-5). In the validation cohort, mannose was significantly associated with fibrous cap thickness 65 μm (odds ratio = 1.32 per each 10 μmol/L mannose change [95% confidence interval, 1.05-1.65]) and was an independent predictor of death (hazard ratio for mannose≥vs 84.6 μmol/L: 4.0(95%CI, 1.4-11.3), p = 0.006).The current data add novel evidence that high mannose is a signature of CAD with a vulnerable plaque phenotype, consistently across measures of severity of vessel involvement and independent of the traditional correlates of CVD, and that it is an independent predictor of incident adverse outcomes.

Objectives This aimed at the design and production of engineered 3D scaffold prototypes using a natural polymeric bioink made of chitosan and poly-γ-glutamic acid with a specific focus on 3D-bioprinting process and on 3D framework geometry. Methods Prototypes were produced using a 3D bioprinter exploiting layer-by-layer deposition technology. The 3D scaffold prototypes were fully characterized concerning pore size and size distribution, stability in different experimental conditions, swelling capability, and human dermal fibroblasts viability. Key findings Hexagonal framework combined with biopaper allowed stabilizing the 3-layers structure during process manufacturing and during incubation in cell culture conditions. The stability of 3-layers structure was well preserved for 48 h. Crosslinking percentages of 2-layers and 3-layers prototype were 88.2 and 68.39, respectively. The swelling study showed a controlled swelling capability for 2-layers and 3-layers prototype, ∼5%. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay results showed good biocompatibility of 3-layers prototype and their suitability for preserving 48 h cell viability in 3D cultures. Moreover, a significant increment of absorbance value was measured after 48 h, demonstrating cell growth. Conclusions Bioink obtained combining chitosan and poly-γ-glutamic acid represents a good option for 3D bioprinting. A stable 3D structure was realized by layer-by-layer deposition technology; compared with other papers, the present study succeeded in using medical healthcare-grade polymers, no-toxic crosslinker, and solvents according to ICH Topic Q3C (R4).

Nowadays the most important issues for commercial SOFC systems are related to their durability and stability of performance both of which are affected by fuel feeding composition. The trace compounds contained in biogenous fuels can interact with SOFC components causing evident drops in performance and severely reducing the lifetime of the entire system. Due to the significant impact on SOFC performance from widely varying fuel inputs, the influence of trace compounds should be investigated in real case studies and practical applications. Anode deactivation and porous blocking electrodes are the main consequences that occur due to the presence of trace compounds impact. This work shows an experimental case study where the anode supported solid oxide fuel cells short stacks are fed with fuels containing trace compounds. These compounds were selected from the biological decomposition of organic matters. In addition, sulphur compounds were selected chlorine, aromatic compounds, terpenes, and carboxyl compounds were selected. The most deleterious impact on the stack was due to the sulphur action. However, chlorine compounds also affected the external part of the cell causing the stack to break due to the corrosion of the sealant.

Non-ablative and ablative fractional erbium lasers are among the most frequently used resources in dermatology for facial rejuvenation and for treating dermatological disorders. This type of erbium laser can be found at wavelengths of 1540 or 1550 nm, which are classified as non-ablative erbium glass, and at 2940 nm, classified as ablative erbium YAG. Despite the reports of their clinical benefits, few scientific studies have demonstrated the efficacy and safety of these lasers in the short or long term. In order to substantiate the effects, benefits, and safety of applying the erbium glass and erbium YAG lasers, a systematic review was carried out from August to December 2019 about studies published in the last 20 years. Randomized clinical trials in humans were considered that evaluated the efficacy, safety, and benefits of applying the fractional lasers erbium glass and erbium YAG to facial rejuvenation, skin spots, and atrophic acne scars. A total of 338 articles were identified; 76 articles remained after their titles and abstracts were read, and 42 articles were selected after removing the duplicates. After the articles were read in full, 17 of these articles were included in the systematic review (453 patients). The erbium glass and erbium YAG lasers seem promising in the short term, with minimal adverse effects; however, the long-term efficacy and safety still present limitations. Consequently, future research is needed, with better methodological standardization and a follow-up with a longer evaluation period for possible permanent adverse effects to determine the standardization and safety of therapy with erbium glass and erbium YAG lasers.

A critical step of pre-mRNA splicing is the recruitment of U2 snRNP to the branch point sequence of an intron. U2 snRNP conformation changes extensively during branch helix formation and several RNA-dependent-ATPases are implicated in the process. However, the molecular mechanisms involved remain to be fully dissected. We took advantage of the differential nucleotide triphosphates requirements for DExD/H-box enzymes to probe their contributions to in vitro spliceosome assembly. Both ATP and GTP hydrolysis support the formation of A-complex, indicating the activity of a DEAH-enzyme because DEAD-enzymes are selective for ATP. We immunodepleted DHX15 to assess its involvement and although splicing efficiency decreases with reduced DHX15, A-complex accumulation incongruently increases. DHX15 depletion also results in the persistence of the atypical ATP-independent interaction between U2 snRNP and a minimal substrate that is otherwise destabilized in the presence of either ATP or GTP. These results lead us to hypothesize that DHX15 plays a quality control function in U2 snRNP’s engagement with an intron. In efforts to identify the RNA target of DHX15, we determined that an extended polypyrimidine tract is not necessary for disruption of the atypical interaction between U2 snRNP and the minimal substrate. We also examined U2 snRNA by RNase H digestion and identified nucleotides in the branch binding region that become accessible with both ATP and GTP hydrolysis, again implicating a DEAH-enzyme. Together, our results demonstrate that multiple ATP-dependent rearrangements are likely involved in U2 snRNP addition to the spliceosome and that DHX15 can have an expanded role in splicing.

Shape-Memory Polymers (SMPs) are considered a kind of smart material able to modify size, shape, stiffness and strain in response to different external (heat, electric and magnetic field, water or light) stimuli including the physiologic ones such as pH, body temperature and ions concentration. The ability of SMPs is to memorize their original shape before triggered exposure and after deformation, in the absence of the stimulus, and to recover their original shape without any help. SMPs nanofibers (SMPNs) have been increasingly investigated for biomedical applications due to nanofiber’s favorable properties such as high surface area per volume unit, high porosity, small diameter, low density, desirable fiber orientation and nanoarchitecture mimicking native Extra Cellular Matrix (ECM). This review focuses on the main properties of SMPs, their classification and shape-memory effects. Moreover, advantages in the use of SMPNs and different biomedical application fields are reported and discussed.

Hyaluronic acid-based nanoparticles (HA NPs) can be used to deliver a protein cargo to cells overexpressing HA receptors such as CD44 since they combine the low toxicity of the carrier and the retention of the protein integrity with the receptor-mediated internalization. HA properties play a crucial but sometimes unclear role in managing the formation and stability of the meshwork, cell interactions, and ultimately the protein entrapment efficacy. Nowadays, microfluidic is an innovative technology that allows to overcome limits linked to the NPs production, guaranteeing reproducibility and control of individual batches. Taking advantage of this technique, in this research work, the role of HA weight average molecular weight (Mw) in NPs formation inside a microfluidic device has been specifically faced. Based on the relationship between polymer Mw and solution viscosity, a methodological approach has been proposed to ensure critical quality attributes (size of 200 nm, PDI ≤ 0.3) to NPs made by HA with different Mw (280, 540, 710 and 820 kDa). The feasibility of the protein encapsulation was demonstrated by using Myoglobin, as a model neutral protein, with an encapsulation efficiency always higher than 50%. Lastly, all NPs samples were successfully internalized by CD44-expressing cells.

Common wheat grains are characterised by low concentrations of Ca, K, and Mg, which can be partially removed with the bran during milling processes. This preliminary study investigated the effects of foliar fertilisation at the earing stage with nitrates of Ca, Mg, and K contemporarily, together with a small amount of urea and protein hydrolysate as potential carriers, in two contrasting common wheat varieties, i.e., Solehio (medium proteins content) and Vivendo (high proteins content). Based on the preliminary grain-to-straw concentration ratio of these minerals, two biofortification targets were applied in order to increase their grain contents by +20% and +40%, in comparison with untreated controls. Here, we demonstrate that the highest fertilisation dose was effective in increasing grain K by 13% and Mg by 16% in Vivendo, and Ca by 7% in Solehio, with no boosting effects of the co-formulants urea and protein hydrolysate. In addition to some qualitative benefits due to nitrates supply, negligible phytotoxicity symptoms were observed, as revealed by the NDVI vegetational index dynamics. Although the biofortification target was not fully achieved, this study firstly reports the possibility to increase at the same time Mg and K, and to a lower extent Ca in wheat grains. It is concluded that efficient multiple biofortification should consider a variety-depend response, while further studies are necessary to investigate the effects of different fertilisation timings and doses for improving the poor mineral translocation to the grains.

Biocompatible thermosetting hydrogels are attractive compounds either for the delivery of drugs or as medical devices for tissue regeneration purposes. In this work Poly(Gamma-Glutamic Acid) (G-PGA) and G-PGA based hydrogels were explored. The hypothesis was to evaluate G-PGA and crosslinking agents combinations in order to obtain thermosetting hydrogels suitable for injection. Attention focused on hydrogels that can form by ionic interaction with compounds bearing amine groups, and/or by interaction with polymers such as chitosan and symmetric triblock copolymer (polyethylene glycol-polypropylene glycol- polyethylene glycol). Polymer solutions and related hydrogels characterization involved: rheologic behavior, osmolarity, syringeability and injectability. Results showed that G-PGA solutions syringeability was always acceptable, while injectability test did not meet aspiration by 22G needle. Osmolarity value suitable for injection could be obtain by adding buffering solutions. Only ternary blends made of chitosan, sodium beta-glyceroposphate and G-PGA, or G-PGA, Pluronic F68 and l -lysine, with fixed compositions, resulted in thermosetting gels. The obtained thermosetting gels were not reversible.

Schistosomiasis is a major neglected tropical disease with more than 200 million infections annually. Despite only one drug, praziquantel, being available, the drug pipeline against schistosomiasis is empty, and drug screening tools have limitations. We evaluated the potential of human liver microtissues (hLiMTs) in antischistosomal drug discovery. Because hLiMTs express all human P450 enzymes, they are an excellent tool to evaluate compounds’ bioinactivation, bioactivation, and toxicity. To validate the metabolic conversion capacity of hLiMTs, we first quantified (R)- and (S)-praziquantel and the main metabolite trans-OH-praziquantel following incubation with 0.032–50 μM (0.01–15.62 μg/mL) praziquantel for up to 72 h by a validated LC-MS/MS method. We cocultured hLiMTs with newly transformed schistosomula (NTS) and evaluated the antischistosomal activity and cytotoxicity of three prodrugs terfenadine, tamoxifen citrate, and flutamide. HLiMTs converted 300–350 ng (R)-praziquantel within 24 h into trans-OH-praziquantel. We observed changes in the IC50 values for terfenadine, flutamide, and tamoxifen citrate in comparison to the standard NTS assay in vitro. Cytotoxicity was observed at high concentrations of flutamide and tamoxifen citrate. An in vitro platform containing hLiMTs could serve as an advanced drug screening tool for Schistosoma mansoni, providing information on reduced or increased activity and toxicity. ISSN:2373-8227

α-synuclein is a small protein that is mainly expressed in the synaptic terminals of nervous tissue. Although its implication in neurodegeneration is well established, the physiological role of α-synuclein remains elusive. Given its involvement in the modulation of synaptic transmission and the emerging role of microtubules at the synapse, the current study aimed at investigating whether α-synuclein becomes involved with this cytoskeletal component at the presynapse. We first analyzed the expression of α-synuclein and its colocalization with α-tubulin in murine brain. Differences were found between cortical and striatal/midbrain areas, with substantia nigra pars compacta and corpus striatum showing the lowest levels of colocalization. Using a proximity ligation assay, we revealed the direct interaction of α-synuclein with α-tubulin in murine and in human brain. Finally, the previously unexplored interaction of the two proteins in vivo at the synapse was disclosed in murine striatal presynaptic boutons through multiple approaches, from confocal spinning disk to electron microscopy. Collectively, our data strongly suggest that the association with tubulin/microtubules might actually be an important physiological function for α-synuclein in the synapse, thus suggesting its potential role in a neuropathological context.

Zonal modeling is a common technique for the numerical certification of fire-extinguishing systems, however it is not valid to simulate the complex physical phenomena that occurs near the agent injection. We present a multi-scale method for the accurate generation of inflow boundary conditions valid for zonal modeling based on the description of the phase change of a rapidly depressurised mist of a fire suppression system. The generation of accurate boundary conditions includes the characterization of the injection of the fire suppression agent from atomization to evaporation and mixing. The multi-scale methodology is based on the use of a high fidelity multiphase conservative level set LES for the characterization of the nozzle to develop an empirical model for primary breakup. Secondly, a low fidelity particle-based method with phase change and unsteady RANS is used for parametric studies. This multi-scale approach requires an affordable computational effort. The multi-scale methodology is tested in a system consisting of a pressurised fire extinguishing agent (Novec-1230) that is injected into the ambient through a nozzle that produces the atomization of the agent. The accuracy of the developed approach is compared with the experimental data. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the grant agreement No 785549 (FireExtintion: H2020-CS2-CFP06-2017-01).

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can infect a broad range of human tissues by using the host receptor angiotensin-converting enzyme 2 (ACE2). Individuals with comorbidities associated with severe COVID-19 display higher levels of ACE2 in the lungs compared to those without comorbidities, and conditions such as cell stress, elevated glucose levels and hypoxia may also increase the expression of ACE2. Here, we showed that patients with Barrett's esophagus (BE) have a higher expression of ACE2 in BE tissues compared to normal squamous esophagus, and that the lower pH associated with BE may drive this increase in expression. Human primary monocytes cultured in reduced pH displayed increased ACE2 expression and higher viral load upon SARS-CoV-2 infection. We also showed in two independent cohorts of 1,357 COVID-19 patients that previous use of proton pump inhibitors is associated with 2- to 3-fold higher risk of death compared to those not using the drugs. Our work suggests that pH has a great influence on SARS-CoV-2 Infection and COVID-19 severity.

Polymeric tubular vascular grafts represent a likely alternative to autologous vascular grafts for treating peripheral artery occlusive disease. This preliminary research study applied cutting-edge electrospinning technique for manufacturing prototypes with diameter ≤ 6 mm and based on biocompatible and biodegradable polymers such as polylactide-polycaprolactone, polylactide-co-glycolide and polyhydroxyethylmethacrylate combined in different design approaches (layering and blending). Samples were characterized about fiber morphology, diameter, size distribution, porosity, fluid uptake capability, and mechanical properties. Biocompatibility and cell interaction were evaluated by in vitro test. Goal of this preliminary study was to discriminate among the prototypes and select which composition and design approach could better suit tissue regeneration purposes. Results showed that electrospinning technique is suitable to obtain grafts with a diameter 6 mm and thickness between 140 ± 7-175 ± 4 μm. Scanning electron microscopy analysis showed fibers with suitable micrometric diameters and pore size between 5 and 35 μm. polyhydroxyethylmethacrylate provided high hydrophilicity (≃ 100°) and optimal cell short term proliferation (cell viability ≃ 160%) in accordance with maximum fluid uptake ability (300-350%). Moreover, addition of polyhydroxyethylmethacrylate lowered suture retention strength at value 1 N. Prototypes obtaining combining polylactide-co-glycolide and polylactide-coglycolide/ polyhydroxyethylmethacrylate with polylactide-polycaprolactone in a bilayered structure showed optimal mechanical behavior resembling native bovine vessel.

Electrospun scaffolds made of nano- and micro-fibrous non-woven mats from biodegradable polymers have been intensely investigated in recent years. In this field, polymer-based materials are broadly used for biomedical applications since they can be managed in high scale, easily shaped, and chemically changed to tailor their specific biologic properties. Nonetheless polymeric materials can be reinforced with inorganic materials to produce a next-generation composite with improved properties. Herein, the role of graphene nanoplatelets (GNPs) on electrospun poly-l-lactide-co-poly-&epsilon, caprolactone (PLA&ndash, PCL, 70:30 molar ratio) fibers was investigated. Microfibers of neat PLA&ndash, PCL and with different amounts of GNPs were produced by electrospinning and they were characterized for their physicochemical and biologic properties. Results showed that GNPs concentration notably affected the fibers morphology and diameters distribution, influenced PLA&ndash, PCL chain mobility in the crystallization process and tuned the mechanical and thermal properties of the electrospun matrices. GNPs were also liable of slowing down copolymer degradation rate in simulated physiological environment. However, no toxic impurities and degradation products were pointed out up to 60 d incubation. Furthermore, preliminary biologic tests proved the ability of the matrices to enhance fibroblast cells attachment and proliferation probably due to their unique 3D-interconnected structure.

This in vitro study evaluated cell viability and metabolism, nitric oxide release and production of two chemokines and one cytokine by cultured human dental pulp fibroblasts (HDPF) in contact with two glass ionomer cements (Ketac Molar-KM and Vitrebond-VB), Single Bond (SB) and calcium hydroxide (Dycal-DY). Cultures of HDPF were established by means of an explant technique. The specimens were prepared under sterile conditions and in disks measuring 5 mm x 2 mm obtained from a prefabricated mold and placed on a permeable membrane to avoid direct contact with the cells. Cytotoxicity was assessed by Trypan Blue exclusion method and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Nitric oxide release in cell supernatant was detected by the Griess Method whereas stromal derived factor-1 alpha (SDF-1α or CXCL12), chemokine (C-X-C motif) ligand 8 [Interleukin 8 (IL-8 or CXCL8)] and interleukin-6 (IL-6) were detected by ELISA. RT-qPCR was employed for gene expression analysis. Statistical analyses were performed by One-way ANOVA followed by Tukey’s post hoc test for materials independent of the time, and Two-way ANOVA followed by Bonferroni correction test for the comparisons between materials and experimental time (p

Poly(methyl-methacrylate) (PMMA) is a biocompatible and non-biodegradable polymer widely used as biomedical material. PMMA microcapsules with suitable dimension and porosity range are proposed to encapsulate live cells useful for tissue regeneration purposes. The aim of this work was to evaluate the feasibility of producing cell-loaded PMMA microcapsules through &ldquo, high efficiency vibrational technology&rdquo, (HEVT). Preliminary studies were conducted to set up the process parameters for PMMA microcapsules production and human dermal fibroblast, used as cell model, were encapsulated in shell/core microcapsules. Microcapsules morphometric analysis through optical microscope and scanning electron microscopy highlighted that uniform microcapsules of 1.2 mm with circular surface pores were obtained by HEVT. Best process conditions used were as follows: frequency of 200 Hz, voltage of 750 V, flow rate of core solution of 10 mL/min, and flow rate of shell solution of 0.5 bar. Microcapsule membrane allowed permeation of molecules with low and medium molecular weight up to 5900 Da and prevented diffusion of high molecular weight molecules (11,000 Da). The yield of the process was about 50% and cell encapsulation efficiency was 27% on total amount. The cell survived and growth up to 72 h incubation in simulated physiologic medium was observed.

In this study, an innovative microfluidics-based method was developed for one-step synthesis of hyaluronic acid (HA)-based nanoparticles (NPs), by exploiting polyelectrolytic interactions between HA and chitosan (CS), in order to improve reliability, reproducibility and possible scale-up of the NPs preparation. The on-chip synthesis, using a staggered herringbone micromixer, allowed to produce HA/CS NPs with tailored-made size and suitable for both parenteral (117.50 &plusmn, 4.51 nm) and loco-regional (349.15 &plusmn, 38.09 nm) administration, mainly composed by HA (more than 85% wt) with high negative surface charge (&lt, &minus, 20 mV). HA/CS NPs were successfully loaded with a challenging water-insoluble molecule, Everolimus (EVE), an FDA- and EMA-approved anticancer drug able to lead to cell cycle arrest, reduced angiogenesis and promotion of apoptosis. HA/CS NPs resulted to be massively internalized in CD44+ human mesenchymal stem cells via CD44 receptor-mediated endocytosis. HA/CS NPs selectiveness towards CD44 was highlighted by blocking CD44 receptor by anti-CD44 primary antibody and by comparison to CS-based NPs cellular uptake. Eventually, high effectiveness in inhibiting cell proliferation was demonstrated on-chip synthetized EVE loaded HA/CS NPs by tracking in vitro DNA synthesis.

This study evaluated in vitro cell viability and metabolism, nitric oxide release and production of chemokines by cultured human dental pulp fibroblasts (DPF) under contact with HEMA and Single Bond. Cultures of DPF were established by means of an explant technique. Once plated, cells were kept under contact with increasing concentrations of HEMA (10, 100 and 1000 nM) or Single Bond (SB) [10-fold serially diluted in culture medium (10-4, 10-3 and 10-2 v/v)] and also with polymerized SB components. Cytotoxicity was assessed by Trypan Blue exclusion method and MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. Nitric oxide release on cell supernatant was detected by Griess Method whereas chemokines (CXCL12 and CXCL8) were detected by ELISA. RT-qPCR was employed for chemokines gene expression analysis. Cytotoxic tests showed significant differences for SB 10-2. None of the tested materials significantly altered NO levels. Protein levels of CXCL12 were significantly decreased only by HEMA. On the other hand, while CXCL12 mRNA remained unaltered, gene expression of CXCL8 had significant decrease with all materials, except for polymerized SB. In conclusion, Single Bond and HEMA at various concentrations, decreased expression and production of molecules involved in inflammatory processes and, therefore, the use of adhesive systems such as pulp capping materials must be viewed with caution due to its large cytotoxic effect when in close contact with the pulp.

Ivermectin (IVM) is a poorly water-soluble drug, which can be used for the treatment of several parasitic human infections (e.g. tropical diseases). The aim of this work was to prepare and characterize biodegradable s.c. implants, potentially enabling 6 months controlled IVM release. The implants were prepared by compressing mixtures of IVM and α-Tocopherol (TCP)-loaded microparticles based on poly-D, l -lactide or poly-e-caprolactone together with sucrose and magnesium stearate. Fifty mg implants containing 7 mg IVM were sterilized by gamma irradiation. The systems were characterized using DSC, FTIR, GPC, EPR, a crushing test, and an in vitro release tests. Results showed that implants were physically stable and they were more stable towards ionizing radiation than the corresponding microspheres. IVM release from the implants and corresponding microspheres slowed down upon irradiation in all cases. In vitro drug release from poly-D, l -lactide implants was slower than from poly-e-caprolactone implants (e.g., 25% after 2 months compared to 100% after 15 d). Mixing poly-D, l -lactide and poly-e-caprolactone based microparticles at a ratio of 1:1 (weight:weight) led to 70% IVM release after 6 months.

Electrospinning is a quite consolidated technique to produce polymer nanofibrous matrices whose nanostructured morphology received great interest for application in the biomedical field i.e. in manufacturing matrices for tissue regeneration. Purpose of the work to design electrospun matrices made of Poly- l -lactide-co-poly-e-caprolactone (PLA-PCL) 70:30 M ratio and their thoroughly physico-chemical and functional characterization. The ultimate goal of the research work is to obtain electrospun matrices suitable for circular substitution of esophageal defects. However the paper deals with very preliminary investigation mainly on electrospinning process and physical-chemical characterization of the electrospun matrices. The investigation on electrospinning process conditions involves polymer starting solution concentration between 20% w/v and 25% w/v, viscosity and surface tension, but also process parameters of electrospinning apparatus in order to optimize them at achieving homogeneous and reproducible nanofibers. The optimized electrospun matrices are characterized for their cytocompatibility, morphology (SEM analysis), polymer molecular structure in the solid state (FT-IR analysis) and thermal behavior (DSC analysis) and mechanical properties. In vitro degradation test is performed to evaluate electrospun matrix biodegradability. Results show good cytocompatibility for all polymer concentrations and electrospinning times. Moreover, the electrospun matrices biodegradation is slower with respect to matrices made by solvent casting method. The behavior is related to modifications in the polymer solid state and to polymer chains structure.

Potential off-label therapeutic role of N-acetylcysteine (N-Ac) was recently demonstrated in the treatment of diastrophic dysplasia (DTD) using mutant mice; its main drawback is the rapid clearance from blood due to the liver metabolism. Our goal was to investigate the potential of polyethylene glycol polylactide-co-glycolide block copolymer (PLGA-PEG)-based nanoparticles (NPs) in order to improve in vivo biodistribution performances and N-Ac pharmacokinetic profile after subcutaneous administration in mice. Results suggest that N-Ac can be effectively loaded into NPs (about 99 μg/mg NPs) using a suitably optimized nanoprecipitation method. Thanks to the good physical characteristics (mean diameter100 nm, zeta potential about -8 mV) NPs can reach skeletal tissue in particular femoral head and proximal tibia epiphysis at the sixth hour after injection, remaining in the tissues till 24 h. Furthermore, pharmacokinetic study revealed a sustained N-Ac concentration in plasma with a peak concentration of 2.48 ± 1.72 μM at the 24th hour after injection. Overall, results highlight the actual interest of N-Ac-loaded PLGA-PEG NPs as useful platform for N-Ac parenteral administration.

Histone deacetylase 6 (HDAC6) is a peculiar HDAC isoform whose expression and functional alterations have been correlated with a variety of pathologies such as autoimmune disorders, neurodegenerative diseases, and cancer. It is primarily a cytoplasmic protein, and its deacetylase activity is focused mainly on nonhistone substrates such as tubulin, heat shock protein (HSP)90, Foxp3, and cortactin, to name a few. Selective inhibition of HDAC6 does not show cytotoxic effects in healthy cells, normally associated with the inhibition of Class I HDAC isoforms. Here, we describe the design and synthesis of a new class of potent and selective HDAC6 inhibitors that bear a pentaheterocyclic central core. These compounds show a remarkably low toxicity both in vitro and in vivo and are able to increase the function of regulatory T cells (Tregs) at well-tolerated concentrations, suggesting a potential clinical use for the treatment of degenerative, autoimmune diseases and for organ transplantation.

Although type-2-induced (T2-induced) epithelial dysfunction is likely to profoundly alter epithelial differentiation and repair in asthma, the mechanisms for these effects are poorly understood. A role for specific mucins, heavily N-glycosylated epithelial glycoproteins, in orchestrating epithelial cell fate in response to T2 stimuli has not previously been investigated. Levels of a sialylated MUC4β isoform were found to be increased in airway specimens from asthmatic patients in association with T2 inflammation. We hypothesized that IL-13 would increase sialylation of MUC4β, thereby altering its function and that the β-galactoside α-2,6-sialyltransferase 1 (ST6GAL1) would regulate the sialylation. Using human biologic specimens and cultured primary human airway epithelial cells (HAECs),we demonstrated that IL-13 increases ST6GAL1-mediated sialylation of MUC4β and that both were increased in asthma, particularly in sputum supernatant and/or fresh isolated HAECs with elevated T2 biomarkers. ST6GAL1-induced sialylation of MUC4β altered its lectin binding and secretion. Both ST6GAL1 and MUC4β inhibited epithelial cell proliferation while promoting goblet cell differentiation. These in vivo and in vitro data provide strong evidence for a critical role for ST6GAL1-induced sialylation of MUC4β in epithelial dysfunction associated with T2-high asthma, thereby identifying specific sialylation pathways as potential targets in asthma.

The Na+/K+/Cl- cotransporter-1 (NKCC1) and the K+/Cl- cotransporter-2 (KCC2) set the transmembrane Cl- gradient in the brain, and are implicated in epileptogenesis. We studied the postnatal distribution of NKCC1 and KCC2 in wild-type (WT) mice, and in a mouse model of sleep-related epilepsy, carrying the mutant β2-V287L subunit of the nicotinic acetylcholine receptor (nAChR). In WT neocortex, immunohistochemistry showed a wide distribution of NKCC1 in neurons and astrocytes. At birth, KCC2 was localized in neuronal somata, whereas at subsequent stages it was mainly found in the somatodendritic compartment. The cotransporters' expression was quantified by densitometry in the transgenic strain. KCC2 expression increased during the first postnatal weeks, while the NKCC1 amount remained stable, after birth. In mice expressing β2-V287L, the KCC2 amount in layer V of prefrontal cortex (PFC) was lower than in the control littermates at postnatal day 8 (P8), with no concomitant change in NKCC1. Consistently, the GABAergic excitatory to inhibitory switch was delayed in PFC layer V of mice carrying β2-V287L. At P60, the amount of KCC2 was instead higher in mice bearing the transgene. Irrespective of genotype, NKCC1 and KCC2 were abundantly expressed in the neuropil of most thalamic nuclei since birth. However, KCC2 expression decreased by P60 in the reticular nucleus, and more so in mice expressing β2-V287L. Therefore, a complex regulatory interplay occurs between heteromeric nAChRs and KCC2 in postnatal forebrain. The pathogenetic effect of β2-V287L may depend on altered KCC2 amounts in PFC during synaptogenesis, as well as in mature thalamocortical circuits.

A biodegradable composite scaffold, useful as tridimensional bone regenerative scaffold (3D) and antibiotic delivery system (DDs), is here formulated and characterized. The device, called as 3D-DDS, is an injectable or moldable scaffold based on chitosan chloride crosslinked with glycerol 2-phosphate disodium salt and laden with bovine bone substitute (BBs) granules and gentamicin loaded polylactide-co-glycolide-co-poly ethilenglycol (PLGA-PEG) microparticles. 3D-DDS characterization results show that microparticles are homogeneously distributed in the composite matrix, and do not interfere with scaffold stability. The lyophilized 3D-DDS has high rehydration ability and is able to restrain physically bovine bone granules and microparticles up to 45 days in in vitro simulated conditions. Bactericidal action is high in the first 4 h and it accounts for superimposable effect of gentamicin release and bacteriostatic chitosan properties; sustained gentamicin bactericidal effect is evident up to 14 days.

This study was aimed at investigating the effect of the fluoride content (added as NaF) on the in vitro bioactivity of an experimental calcium silicate-based cement (wTC-Bi) obtained from white Portland cement. To this purpose, wTC-Bi and fluoride-doped wTC-Bi cements (i.e. FTC-Bi and F10TC-Bi with fluoride contents of 1% and 10% w/w, respectively) were aged in Dulbecco's Phosphate Buffered Saline (DPBS) and were comparatively analysed by micro-Raman and IR spectroscopy to investigate the presence of deposits on the surface of the cements and the composition changes of the cement as a function of the storage time. Commercial White ProRoot MTA was analyzed as reference. All the tested cements showed the formation of a calcium phosphate deposit already after 5 h of soaking. Fluoride-doped cements demonstrated a higher bioactivity than the undoped wTC-Bi cement. This result was explained in relation to the different solubility of the deposit formed on the cements: a B-type carbonated apatite on the undoped cements and a less soluble fluoride containing B-type carbonated apatite on the fluoride-doped cements. The NaF content was found to influence the apatite forming ability; actually, the cement richer in NaF, i.e. F10TC-Bi showed a lower bioactivity than FTC-Bi, which contained only 1% w/w of NaF. This result may be explained in relation to the lower hydration rate of the former, which showed the formation of lower amounts of CSH, ettringite and portlandite phases.

In a process known as overt degranulation, mast cells can release all at once a diverse array of products that are preformed and present within cytoplasmic granules. This occurs typically within seconds of stimulation by environmental factors and allergens. These potent, preformed mediators (ie, histamine, heparin, serotonin, and serine proteases) are responsible for the acute symptoms experienced in allergic conditions such as allergic conjunctivitis, allergic rhinitis, allergy-induced asthma, urticaria, and anaphylaxis. Yet, there is reason to believe that the actions of mast cells are important when they are not degranulating. Mast cells release preformed mediators and inflammatory cytokines for periods after degranulation and even without degranulating at all. Mast cells are consistently seen at sites of chronic inflammation, including nonallergic inflammation, where they have the ability to temper inflammatory processes and shape tissue morphology. Mast cells can trigger actions and chemotaxis in other important immune cells (eg, eosinophils and the newly discovered type 2 innate lymphocytes) that then make their own contributions to inflammation and disease. In this review, we will discuss the many known and theorized contributions of mast cells to allergic diseases, focusing on several prototypical allergic respiratory and skin conditions: asthma, chronic rhinosinusitis, aspirin-exacerbated respiratory disease, allergic conjunctivitis, atopic dermatitis, and some of the more common medication hypersensitivity reactions. We discuss traditionally accepted roles that mast cells play in the pathogenesis of each of these conditions, but we also delve into new areas of discovery and research that challenge traditionally accepted paradigms.

The androgen receptor (AR) plays a key role in progression to metastatic castration-resistant prostate cancer (mCRPC). Despite the recent progress in targeting persistent AR activity with the next-generation hormonal therapies (abiraterone acetate and enzalutamide), resistance to these agents limits therapeutic efficacy for many patients. Several explanations for response and/or resistance to abiraterone acetate and enzalutamide are emerging, but growing interest is focusing on importance of AR splice variants (AR-Vs) and in particular of AR-V7. Increasing evidences highlight the concept that variant expression could be used as a potential predictive biomarker and a therapeutic target in advanced prostate cancer. Therefore, understanding the mechanisms of treatment resistance or sensitivity can help to achieve a more effective management of mCRPC, increasing clinical outcomes and representing a promising and engaging area of prostate cancer research.

The flavonoid metal-insecticide [Mg(hesp)2(phen)], denominated MgHP, has high potential for controlling agricultural pests. If applied in large scale, it may reach aquatic ecosystems and be harmful to the biota. This study evaluated the effects of MgHP in the gills of the Neotropical fish, Prochilodus lineatus by determining the activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione s-transferase (GST), and the levels of glutathione (GSH) and lipid peroxidation (LPO) after 24 and 96 h exposure to 0, 1, 10, 100 and 1000 μg L−1. The histopathological changes with emphases to mitochondria-rich cells (MRC) were evaluated as well. After 24 h exposure the enzyme activities and the GSH and LPO levels were unchanged however, after 96 h exposure to high MgHP concentration (1000 μg L−1), the GST activity and GSH levels increased. Oxidative stress measured as LPO levels did not occur after MgHP exposure in both periods. Gill tissue alterations increased after MgHP exposure to 10, 100 and 1000 μg L−1. Cellular atrophy, pillar cells changes, filament epithelium hyperplasia and hypertrophy, lamellar epithelium hyperplasia were the most frequent histopathology. MRC in the filament epithelium decreased after exposure to 24 h and increased after 96 h indicating possible transitory osmo-ionic disruption. P. lineatus exhibited high tolerance to MgHP. The increased GST activity and GSH levels after 96 h exposure suggested possible MgHP accumulation and concentration- and time-dependent response. Histopathology in the gills of exposed fish occurred at high MgHP concentrations. These results suggested that the MgHP into water, at high concentrations, affect the gills by changing GST activity, GSH levels and histology being useful biomarkers for MgHP water contamination.

Electrospinning is known to be an effective and straightforward technique to fabricate polymer non woven matrices made of nano and microfibers. Micro patterned morphology of electrospun matrices results to be outmost advantageous in the biomedical field, since it is able to mimic extracellular matrix (ECM), and favors cell adhesion and proliferation. Controlling electrospun fibers alignment is crucial for the regenerative purposes of certain tissues, such as neuronal and vascular. In this study we investigated the impact of electrospinning process parameters on fiber alignment in tubular nanofibrous matrices made of Poly ( l -lactide-co-e-caprolactone) (PLA-PCL); a Design of Experiments (DoE) approach is here proposed in order to statistically set up the process parameters. The DoE was studied keeping constants the previously set material and environmental parameters; voltage, flow rate and mandrel rotating speed were the process parameters here investigated as variables. Orientation analysis was based on ImageJ and plugin Orientation J analysis of SEM images. The results show that voltage combined with flow rate has significant impact on electrospun fiber orientation, and the greatest orientation is achieved when all the three input parameters (voltage, flow rate and mandrel rotation speed) are at their maximum value.

The microfluidic technique has emerged as a promising tool to accelerate the clinical translation of nanoparticles, and its application affects several aspects, such as the production of nanoparticles and the in vitro characterization in the microenvironment, mimicking in vivo conditions. This review covers the general aspects of the microfluidic technique and its application in several fields, such as the synthesis, recovering, and samples analysis of nanoparticles, and in vitro characterization and their in vivo application. Among these, advantages in the production of polymeric nanoparticles in a well-controlled, reproducible, and high-throughput manner have been highlighted, and detailed descriptions of microfluidic devices broadly used for the synthesis of polysaccharide nanoparticles have been provided. These nanoparticulate systems have drawn attention as drug delivery vehicles over many years; nevertheless, their synthesis using the microfluidic technique is still largely unexplored. This review deals with the use of the microfluidic technique for the synthesis of polysaccharide nanoparticles; evaluating features of the most studied polysaccharide drug carriers, such as chitosan, hyaluronic acid, and alginate polymers. The critical assessment of the most recent research published in literature allows us to assume that microfluidics will play an important role in the discovery and clinical translation of nanoplatforms.

Bronchiolitis obliterans syndrome (BOS), caused by lung allograft-derived mesenchymal cells&rsquo, abnormal proliferation and extracellular matrix deposition, is the main cause of lung allograft rejection. In this study, a mild one-step ionotropic gelation method was set up to nanoencapsulate the everolimus, a key molecule in allograft organ rejection prevention, into hyaluronic acid-decorated chitosan-based nanoparticles. Rationale was the selective delivery of everolimus into lung allograft-derived mesenchymal cells, these cells are characterized by the CD44-overexpressing feature, and hyaluronic acid has proven to be a natural selective CD44-targeting moiety. The optimal process conditions were established by a design of experiment approach (full factorial design) aiming at the control of the nanoparticle size (&le, 200 nm), minimizing the size polydispersity (PDI 0.171 &plusmn, 0.04), and at the negative &zeta, potential maximization (&minus, 30.9 mV). The everolimus was successfully loaded into hyaluronic acid-decorated chitosan-based nanoparticles (95.94 &plusmn, 13.68 &mu, g/100 mg nanoparticles) and in vitro released in 24 h. The hyaluronic acid decoration on the nanoparticles provided targetability to CD44-overexpressing mesenchymal cells isolated from bronchoalveolar lavage of BOS-affected patients. The mesenchymal cells&rsquo, growth tests along with the nanoparticles uptake studies, at 37 &deg, C and 4 &deg, C, respectively, demonstrated a clear improvement of everolimus inhibitory activity when it is encapsulated in hyaluronic acid-decorated chitosan-based nanoparticles, ascribable to their active uptake mechanism.