Exosome loaded genipin crosslinked hydrogel facilitates full thickness cutaneous wound healing in rat animal model

Abstract

Full thickness cutaneous wound therapy and regeneration remains a critical challenge in clinical therapeutics. Recent reports have suggested that mesenchymal stem cells exosomes therapy is a promising technology with great potential to efficiently promote tissue regeneration. Multifunctional hydrogel composed of both synthetic materials and natural materials is an effective carrier for exosomes loading. Herein, we constructed a biodegradable, dual-sensitive hydrogel encapsulated human umbilical cord-mesenchymal stem cells (hUCMSCs) derived exosomes to facilitate wound healing and skin regeneration process. The materials characterization, exosomes identification, and in vivo full-thickness cutaneous wound healing effect of the hydrogels were performed and evaluated. The in vivo results demonstrated the exosomes loaded hydrogel had significantly improved wound closure, re-epithelialization rates, collagen deposition in the wound sites. More skin appendages were observed in exosomes loaded hydrogel treated wound, indicating the potential to achieve complete skin regeneration. This study provides a new access for complete cutaneous wound regeneration via a genipin crosslinked dual-sensitive hydrogel loading hUCMSCs derived exosomes.

Keywords: genipin crosslinked hydrogel; hUCMSCs derived exosome; rat model; skin tissue regeneration; wound healing.

Figure 1.

Schematic of hydrogel synthesis, EXOs loading and treatment on rat model.

Figure 2.

Identification of isolated hUCMSCs derived EXOs. (A) TEM image of EXOs, scale bar: 200 μm; (B) particle size distribution of EXOs; (C) detection of EXOs markers (CD9, CD63, and Alix) expression by western blot analysis.

Figure 3.

Morphology of lyophilized hydrogel stent cross-section analyzed by SEM.

Figure 4.

In vitro degradation rate tests of genipin crosslinked hydrogel (n = 6).

Figure 5.

In vitro sustained release profile of EXOs loaded in genipin crosslinked hydrogel (n = 6).

Figure 6.

Hydrogel and EXOs facilitated wound closure rate in 14 days. (A) Representative images of healing process in wounds treated with saline, pure hydrogel and hydrogel and EXOs. (B) Wound closure rates of all three groups.

Figure 7.

Histological analysis of the wounds in different experimental conditions. (A) H&E staining microscopic images of healing wound sites. (B) MT staining of collagens in wound sites. Scale bar: 100 μm.

Figure 8.

Histochemical analysis of TNF-α and IL-1β expression in wound sites. (A, C) Immunohistochemistry staining images for TNF-α and IL-1β at 7 and 14 days post operation, respectively. Scale bar: 50 μm. (B, D) Quantitative analysis of relative density of TNF-α and IL-1β at 7 and 14 days after surgery, respectively.