Application of adipose mesenchymal stem cell-derived exosomes-loaded ß-chitin nanofiber hydrogel for wound healing.

INTRODUCTION: Clarifying the role and mechanism of exosome gel in wound repair can provide a new effective strategy for wound treatment. MATERIALS AND METHODS: The cellular responses of adipose mesenchymal stem cell-derived exosomes (AMSC-exos) and the wound healing ability of AMSC-exos-loaded ß-chitin nanofiber (ß-ChNF) hydrogel were studied in vitro in mouse fibroblasts cells (L929) and in vivo in rat skin injury model. The transcriptome and proteome of rat skin were studied with the use of sequenator and LC-MS/MS, respectively. RESULTS: 80 and 160 µg/mL AMSC-exos could promote the proliferation and migration of mouse fibroblastic cells. Furthermore, AMSC-exos-loaded ß-ChNF hydrogel resulted in a significant acceleration rate of wound closure, notably, acceleration of re-epithelialization, and increased collagen expression based on the rat full-thickness skin injury model. The transcriptomics and proteomics studies revealed the changes of the expression of 18 genes, 516 transcripts and 250 proteins. The metabolic pathways, tight junction, NF-κB signaling pathways were enriched in Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway. Complement factor D (CFD) and downstream Aldolase A (Aldoa) and Actn2 proteins in rats treated with AMSC-exos-loaded ß-ChNF hydrogel were noticed and further confirmed by ELISA and Western blot. CONCLUSION: These findings suggested that AMSC-exos-loaded ß-ChNF hydrogel could promote wound healing with the mechanism which is related to the effect of AMSC-exos on CFD and downstream proteins.

Adipose-derived mesenchymal stem cell-loaded ß-chitin nanofiber hydrogel promote wound healing in rats.

Because of stem cells are limited by the low efficiency of their cell homing and survival in vivo, cell delivery systems and scaffolds have attracted a great deal of attention for stem cells' successful clinical practice. ß-chitin nanofibers (ß-ChNF) were prepared from squid pens in this study. Fourier transform infrared spectroscopy, X-ray diffraction and scanning electron microscopy proved that ß-ChNFs with the diameter of 5 to 10 nm were prepared. ß-ChNF dispersion became gelled upon the addition of cell culture medium. Cell culture experiments showed that ß-ChNFs exhibited negligible cytotoxicity towards ADSCs and L929 cells, and it was found that more exosomes were secreted by the globular ADSCs grown in the ß-ChNF hydrogel. The vivo experiments of rats showed that the ADSCs-loaded ß-ChNF hydrogel could directly cover the wound surface and significantly accelerate the wound healing and promote the generation of epithelization, granulation tissue and collagen. In addition, the ADSCs-loaded ß-ChNF hydrogel clearly regulated the expressions of VEGFR, α-SMA, collagen I and collagen III. Finally, we showed that ADSCs-loaded ß-ChNF hydrogel activated the TGFß/smad signaling. The neutralization of TGFß markedly reduced Smad phosphorylation and the expressions of TIMP1, VEGFR and α-SMA. Taken together, these findings suggest that ADSCs-loaded ß-ChNF hydrogel promises for treating wounds that are challenge to heal via conventional methods. Graphical abstract.

Chitosan/Gelatin Composite Nonwoven Fabric Scaffold Seeding Minimal Function Unit of Skin for Functional Skin Regeneration.

The construction of intact functional skin is a challenging field in tissue engineering. Traditional skin tissue engineering, using "seed cells" as a bioactive source for scaffolding materials maybe not efficient enough. Here a new strategy is shown for constructing functional tissue-engineered skin with Minimal Functional Unit of Skin (MFUS) as the source of bioactivity. Chitosan/gelatin non-woven fabric is used as the scaffold. MFUS is derived from autologous skin with full-thickness skin microstructure and complete functional skin unit harvesting. A mathematical model is used to calculate the MFUS Minimal Harvest Diameter and Angle (MHDA). Chitosan/gelatin non-woven fabric (CS+GEL) is porous and absorbable, with an elastic modulus meeting the requirement of skin engineering. It supports layered and 3D growth of MFUS. The degradation rate of chitosan, including filament diameter and density is evaluated in vivo. MFUS-engineered skin could reduce the density of local nerve fibers in the early stage, potentially reducing pain during wound healing, as well as could limit excessive fibroblast cell migration in the later stage, potentially reducing scar formation. This study proposes a new strategy for the clinical treatment of large full-thickness skin defects by constructing intact functional at minimal cost.