No abstract

Acute and chronic wounds contribute to increased morbidity and mortality in affected people and impose significant financial burdens on healthcare systems. For these challenging wounds, acellular dermal matrices (ADMs) have been used as a biological wound coverage. Unlike engineered dermal matrices, ADMs are prepared through the removal of cells from skin, while preserving the extracellular matrix structure and function. In this study, our primary objective was to develop a detergent-free method for decellularization of the skin to mitigate chemical stress on matrix molecules. Then, we performed a set of in vitro and in vivo experiments to compare this method with nonionic and anionic detergent methods. All decellularization methods satisfactorily removed cells and supported fibroblast growth and migration in vitro. Sulfated glycosaminoglycan content was reduced significantly (p < 0.05) only in the ionic detergent treatment group. In contrast to the detergent-free method, all detergent-based methods significantly reduced scaffold mechanical strength and elastin content (p < 0.05). Three weeks after transplantation, the results showed reepithelialization, angiogenesis, and migration of host cell into scaffolds with no induction of immunogenic reaction in all ADM groups tested. In our study, the detergent-free method showed better preservation of matrix composition and biomechanical properties, but after transplantation, all methods of ADM preparation resulted in equally biofunctional matrices as wound coverage.

Dermal fibrosis, characterized by excessive extracellular matrix (ECM), is a pathological condition with limited effective therapeutic modalities. Lack of an antiscarring dressing further impedes the preventive measures for this condition. Here, we develop a new antiscarring dressing and investigate its potential as a slow-releasing vehicle for kynurenic acid (KynA), an antifibrotic agent. KynA was incorporated into polymethyl methacrylate (PMMA) nanofibers, containing increasing concentration of polyethylene glycol (PEG). Fibre morphology, water absorption capacity, surface hydrophilicity, in vitro drug release profile, and in vivo antifibrotic effects were investigated. Increasing concentrations of PEG (1-20%) significantly increased surface hydrophilicity, water absorption capacity, and drug release. Based on the obtained release profiles, PMMA + 10% PEG was the preferred formulation for sustained KynA release up to 120 hours. In vitro studies confirmed the preservation of KynA antifibrotic properties during electrospinning, indicated by fibroblasts proliferation suppression and ECM expression modulation. In vivo application of KynA-incorporated films significantly inhibited collagen (23.89 ± 4.79 vs. 6.99 ± 0.41, collagen-I/β-actin mRNA expression, control vs. treated) and fibronectin expression (7.18 ± 1.09 vs. 2.31 ± 0.05, fibronectin/β-actin mRNA expression, control vs. treated) and enhanced the production of an ECM-degrading enzyme (2.03 ± 0.88 vs. 11.88 ± 1.16 MMP-1/β-actin mRNA expression, control vs. treated). The fabricated KynA-incorporated films can be exploited as antifibrotic wound dressings. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2334-2344, 2016.

Skin transplantation provides an excellent potential model to investigate the immunology of allograft rejection and tolerance induction. Despite the theoretical ease of performing skin transplantation, as well as the potential of directly observing the reaction to the transplanted tissue, the poor reliability of skin transplantation in the mouse has largely precluded the use of this model. Furthermore, there is controversy regarding the most appropriate skin graft donor site due to poor success of back skin transplantation, as compared with the thinner ear or tail skin. This study demonstrates a reliable method to successfully perform skin grafts in a mouse model, as well as the clinical and histologic outcome of syngeneic grafts. A total of 287 grafts were performed (in 126 mice) utilizing donor skin from the ear, tail or back. No graft failure or postoperative mortality was observed. Comparison of this technique with two previously established protocols of skin transplantation (5.0 absorbable Suture + tissue glue technique and no-suture technique) demonstrates the significant improvement in the engraftment success of the new technique. In summary, a new technique for murine skin grafting demonstrates improved reliability across donor site locations and strains, increasing the potential for investigating interventions to alter the rejection process.

SummaryThere is controversy about the immunomodulatory effect of fibroblasts on dendritic cells (DCs). To clarify this issue, in this study, we have evaluated different features of fibroblast-primed DCs including their ability to express co-inhibitory and co-stimulatory molecules, pro-inflammatory and anti-inflammatory cytokines and their ability to induce T-cell proliferation. We also examined migratory capacity of DCs to lymphatic tissues and present fibroblast-derived antigens after encountering fibroblasts. The results of our in vitro study showed that both co-inhibitory (programmed death ligand 1 and ligand 2 and B7H4) and co-stimulatory (CD86) molecules were up-regulated when DCs were co-cultured with fibroblasts. In an animal model, we showed that intra-peritoneal injection (IP) of both syngeneic and allogeneic fibroblasts significantly increased both total DC count and expression level of co-inhibitory and co-stimulatory molecules on DCs. Priming of DCs with syngeneic and allogeneic fibroblasts reduced the proliferation of CD4 + and CD8 + T cells. Even activation of fibroblastprimed DCs failed to restore their ability to induce T-cell proliferation. Likewise, priming of DCs with fibroblasts blocked the ability of ovalbumin-pulsed DCs to induce proliferation of ovalbumin-specific CD4 + T cells. Compared with non-activated DCs, fibroblast-primed DCs had significantly higher expression levels of interleukin-10 and indoleamine 2, 3 dioxygenase. Fibroblast-primed DCs had a significantly reduced interleukin-12 expression level compared with that of activated DCs. After priming with fibroblasts, DCs were able to migrate to lymphatic tissues and present fibroblast-derived antigens (ovalbumin). In conclusion, after priming with fibroblasts, DCs gain tolerogenic features. This finding suggests the potential role of fibroblasts in the maintenance of immune tolerance.