Functionalization of polyhydroxyurethanes by α-alkylidene cyclic carbonates towards new elastomers for 3D-printed biomedical devices

The abstract is provided as supporting informatio

Different Oxidative Stress Response in Keratinocytes and Fibroblasts of Reconstructed Skin Exposed to Non Extreme Daily-Ultraviolet Radiation

Experiments characterizing the biological effects of sun exposure have usually involved solar simulators. However, they addressed the worst case scenario i.e. zenithal sun, rarely found in common outdoor activities. A non-extreme ultraviolet radiation (UV) spectrum referred as “daily UV radiation” (DUVR) with a higher UVA (320–400 nm) to UVB (280–320 nm) irradiance ratio has therefore been defined. In this study, the biological impact of an acute exposure to low physiological doses of DUVR (corresponding to 10 and 20% of the dose received per day in Paris mid-April) on a 3 dimensional reconstructed skin model, was analysed. In such conditions, epidermal and dermal morphological alterations could only be detected after the highest dose of DUVR. We then focused on oxidative stress response induced by DUVR, by analyzing the modulation of mRNA level of 24 markers in parallel in fibroblasts and keratinocytes. DUVR significantly modulated mRNA levels of these markers in both cell types. A cell type differential response was noticed: it was faster in fibroblasts, with a majority of inductions and high levels of modulation in contrast to keratinocyte response. Our results thus revealed a higher sensitivity in response to oxidative stress of dermal fibroblasts although located deeper in the skin, giving new insights into the skin biological events occurring in everyday UV exposure

Design, manufacturing and testing of a green non-isocyanate polyurethane prosthetic heart valve.

peer reviewedThe sole effective treatment for most patients with heart valve disease is valve replacement by implantation of mechanical or biological prostheses. However, mechanical valves represent high risk of thromboembolism, and biological prostheses are prone to early degeneration. In this work, we aim to determine the potential of novel environmentally-friendly non-isocyanate polyurethanes (NIPUs) for manufacturing synthetic prosthetic heart valves. Polyhydroxyurethane (PHU) NIPUs are synthesized via an isocyanate-free route, tested in vitro, and used to produce aortic valves. PHU elastomers reinforced with a polyester mesh show mechanical properties similar to native valve leaflets. These NIPUs do not cause hemolysis. Interestingly, both platelet adhesion and contact activation-induced coagulation are strongly reduced on NIPU surfaces, indicating low thrombogenicity. Fibroblasts and endothelial cells maintain normal growth and shape after indirect contact with NIPUs. Fluid-structure interaction (FSI) allows modeling of the ideal valve design, with minimal shear stress on the leaflets. Injection-molded valves are tested in a pulse duplicator and show ISO-compliant hydrodynamic performance, comparable to clinically-used bioprostheses. Poly(tetrahydrofuran) (PTHF)-NIPU patches do not show any evidence of calcification over a period of 8 weeks. NIPUs are promising sustainable biomaterials for the manufacturing of improved prosthetic valves with low thrombogenicity

Diversity of Biological Effects Induced by Longwave UVA Rays (UVA1) in Reconstructed Skin

<div><p>Despite their preponderance amongst the ultraviolet (UV) range received on Earth, the biological impacts of longwave UVA1 rays (340–400 nm) upon human skin have not been investigated so thoroughly. Nevertheless, recent studies have proven their harmful effects and involvement in carcinogenesis and immunosuppression. In this work, an i<i>n vitro</i> reconstructed human skin model was used for exploring the effects of UVA1 at molecular, cellular and tissue levels. A biological impact of UVA1 throughout the whole reconstructed skin structure could be evidenced, from morphology to gene expression analysis. UVA1 induced immediate injuries such as generation of reactive oxygen species and thymine dimers DNA damage, accumulating preferentially in dermal fibroblasts and basal keratinocytes, followed by significant cellular alterations, such as fibroblast apoptosis and lipid peroxidation. The full genome transcriptomic study showed a clear UVA1 molecular signature with the modulation of expression of 461 and 480 genes in epidermal keratinocytes and dermal fibroblasts, respectively (fold change> = 1.5 and adjusted p value<0.001). Functional enrichment analysis using GO, KEGG pathways and bibliographic analysis revealed a real stress with up-regulation of genes encoding heat shock proteins or involved in oxidative stress response. UVA1 also affected a wide panel of pathways and functions including cancer, proliferation, apoptosis and development, extracellular matrix and metabolism of lipids and glucose. Strikingly, one quarter of modulated genes was related to innate immunity: genes involved in inflammation were strongly up-regulated while genes involved in antiviral defense were severely down-regulated. These transcriptomic data were confirmed in dose-response and time course experiments using quantitative PCR and protein quantification. Links between the evidenced UVA1-induced impacts and clinical consequences of UVA1 exposure such as photo-aging, photo-immunosuppression and cancer are discussed. These early molecular events support the contribution of UVA1 to long term harmful consequences of UV exposure and underline the need of an adequate UVA1 photoprotection.</p></div

Overall analysis of gene expression in reconstructed skin exposed to UVA1, using microarray.

<p>Triplicates of reconstructed skins were unexposed (control) or exposed to 40 J/cm² UVA1. Six hours later, a full genome transcriptomic study was conducted using Affymetrix microarray in fibroblasts (F) and keratinocytes (K), separately, for the 3 control reconstructed skins (samples F1-F3 and K1-K3) and for the 3 UVA1 exposed reconstructed skins (samples F4-F6 and K4-K6). <b>A</b>: Hierarchical clustering based on all probe set normalized expression data, using Ward's method and correlation distance. Y-axis of dendrogram represents the linkage distance that separates singletons or clusters. Height at which two clusters are merged in dendrogram reflects distance of the two clusters. <b>B</b>: Fold change comparison plot between fibroblasts and keratinocytes depicting number of significantly modulated probe sets 6 hours after exposure to 40J/cm² UVA1. Each probe set is plotted. On y-axis: log of fold change value in keratinocytes; on x-axis: log of fold change value in fibroblasts. In keratinocytes and in fibroblasts, 502 and 494 probe sets were differentially expressed between UVA1 exposed and control reconstructed skins (fold-change >1.5 or <0.67, Adjp<0.001), respectively. Blue circles (n = 107) represent probe sets modulated in both keratinocytes and fibroblasts. Green (n = 395) and red (n = 387) circles represent probe sets modulated only in keratinocytes and in fibroblasts, respectively. Grey circles (n = 2787): un-modulated probe sets. <b>C</b>: Heat map showing relative expression levels of the probe sets differentially expressed between UVA1-exposed and control samples (fold-change threshold >1.5 or <0.67 and Adjp value<0.001). Two-dimensional hierarchical clustering was carried out with the 502 and 494 probe sets differentially expressed between UVA1 exposed and control samples in keratinocytes and fibroblasts, respectively. Euclidean distance and Ward's method, based on normalized log2-transformed gene expression value relative to median value of each row were used <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0105263#pone.0105263-Eisen1" target="_blank">[85]</a>. Each row represents a probe set, each column represents one sample. Red, high expression. Black, median expression. Green, low expression.</p

ROS and lipid peroxidation detection in reconstructed skin exposed to UVA1.

<p>ROS assay was performed using sections of reconstructed skin after DCFH-DA probe incorporation and UVA1. Bracket and arrows indicated the fluorescent keratinocytes and fibroblasts, respectively, in UVA1-exposed samples. None of them were detected in unexposed reconstructed skin. The dotted line indicates dermal epidermal junction (<b>A</b>). Levels of DCFH-DA probe fluorescence in reconstructed skin after UVA1 exposure. AU, arbitrary units (<b>B</b>). Distance between dermal epidermal junction and the deepest positive DCFH-DA cells. Indicated values correspond to the mean of 6 measurements in each experimental condition (<b>C</b>). 8-isoprostane amount in culture medium of reconstructed skin 24 hours after UVA1 exposure (<b>D</b>). ^*, mean value significantly different from mean value at 0 J/cm²; ^a, mean value significantly different from mean value at 10 J/cm²; ^§, mean value significantly different from mean value at 20 J/cm²; ^#, mean value significantly different from mean value at 30 J/cm² (p<0.05, Student's t test).</p

Levels of secreted proteins in culture medium of reconstructed skin exposed to UVA1.

<p>Culture media were taken at 48 hours post UVA1 exposure and used to measure the amount of extracellular matrix remodeling proteins (matrix metalloproteinases, MMPs and GDF15), pro-inflammatory proteins (IL-6, GM-CSF/CSF2, CCL20 and GDF15), the HGF growth factor and the CXCL10 ( = IP10) interferon inducible protein. Values of control samples were adjusted to the 1 value. Asterisks indicate a significant difference between mean protein amount of control samples and mean protein amount of UVA1 exposed samples (p<0.05, Student's t test). AU, arbitrary units.</p

Cellular effects in human reconstructed skin exposed to UVA1.

<p>Sham-exposed (control) and UV-exposed samples were taken for classical histology and for vimentin staining (vimentin: green labeling, nuclei counterstaining: red labeling) at 48 h post UVA1 exposure. Arrows indicate fibroblast disappearance in human dermal equivalent.</p

Distribution of UVA1 modulated genes in functional families.

<p>In order to perform an exhaustive bibliographic analysis including literature related to skin and dermatology, the list of UVA1 modulated genes was reduced by using a fold change threshold>2 or <0.5, and an Adjp<0.001. Under these criteria, 134 and 141 genes were found modulated in fibroblasts and keratinocytes respectively of UVA1 exposed reconstructed skins. In fibroblasts 24 genes out of 134 could not be classified because their functions were poorly described; 110 genes were distributed in functional families (<b>A</b>). In keratinocytes 30/141 genes could not be classified; 111 genes were distributed in functional families (<b>B</b>). Some genes could be classified in several functional families. Lists of gene names associated with gene bank accession number, fold change values and their distribution in functional families are given in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0105263#pone.0105263.s011" target="_blank">Tables S6</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0105263#pone.0105263.s012" target="_blank">S7</a>, for fibroblasts and keratinocytes respectively.</p