ABSTRACT
OBJECTIVES@#To explore the effect of hydrogel loaded with exosomes from Wharton's Jelly-derived mesenchymal stem cell (WJMSC) on wound healing.@*METHODS@#Exosomes were extracted from WJMSC, and the morphology and size of WJMSC-derived exosomes (WEX) were analyzed by transmission electron microscopy and nanoparticle size analyzer, respectively. The surface markers CD9, CD81, and Calnexin of WEX were detected by Western blotting. Exosome-loaded alginate hydrogel (WEX-gel) was prepared; its morphology was studied by scanning electron microscope, and its rheological behavior was examined by a rheometer. The in vitro drug release performance of WEX-gel was investigated by BCA method. RAW264.7 cells were treated with alginate hydrogel, WEX and WEX-gel, respectively; and the expression of CD86 and CD206 in macrophages was detected by flow cytometry. A full-thickness skin wound model was established in mice; the model mice were randomly divided into blank control group, WEX control group and WEX-gel group, and PBS, WEX and WEX-gel were applied to the wound area of mice, respectively. On day 3, the skin tissue of mice was excised, and the antibacterial effect of WEX hydrogel was evaluated by plate counting. On day 15, the mice were euthanized and the percentage of residual wounds was calculated. The histological changes of the skin wound were observed after hematoxylin and eosin (HE) and Masson stainings. The expression of CD86, CD206, CD31 and vascular endothelial growth factor (VEGF) in the skin wound tissue was detected by immunohistochemistry.@*RESULTS@#Exosomes were successfully extracted from WJMSC. WEX-gel presented a regular three-dimensional network structure, good rheology and controlled drug release performance. WEX-gel promoted the polarization of RAW264.7 cells from the M1 phenotype to M2 phenotype in vitro. The residual wound percentage in blank control group, WEX control group and WEX-gel group were (27.5±3.4)%, (15.3±1.2)% and (7.6±1.1)%, respectively (P<0.05). The antibacterial property of WEX-gel is better than that of WEX (P<0.05). The dermis thickness, the number of new hair follicles, and the rate of collagen deposition in the WEX-gel group were significantly higher than those in the other two groups (all P<0.05). The expression of CD206, CD31 and VEGF in skin wound tissue was higher and the expression of CD86 was lower in WEX-gel group than those in other two groups (all P<0.05).@*CONCLUSIONS@#WEX-gel can significantly promote wound healing in mice by regulating the polarization of macrophages.
Subject(s)
Mice , Animals , Vascular Endothelial Growth Factor A , Wharton Jelly , Exosomes , Hydrogels , Wound Healing/physiology , Mesenchymal Stem Cells , Anti-Bacterial Agents , AlginatesABSTRACT
Tolerogenic dendritic cells (tolDCs) facilitate the suppression of autoimmune responses by differentiating regulatory T cells (Treg). The dysfunction of immunotolerance results in the development of autoimmune diseases, such as rheumatoid arthritis (RA). As multipotent progenitor cells, mesenchymal stem cells (MSCs), can regulate dendritic cells (DCs) to restore their immunosuppressive function and prevent disease development. However, the underlying mechanisms of MSCs in regulating DCs still need to be better defined. Simultaneously, the delivery system for MSCs also influences their function. Herein, MSCs are encapsulated in alginate hydrogel to improve cell survival and retention in situ, maximizing efficacy in vivo. The three-dimensional co-culture of encapsulated MSCs with DCs demonstrates that MSCs can inhibit the maturation of DCs and the secretion of pro-inflammatory cytokines. In the collagen-induced arthritis (CIA) mice model, alginate hydrogel encapsulated MSCs induce a significantly higher expression of CD39+CD73+ on MSCs. These enzymes hydrolyze ATP to adenosine and activate A2A/2B receptors on immature DCs, further promoting the phenotypic transformation of DCs to tolDCs and regulating naïve T cells to Tregs. Therefore, encapsulated MSCs obviously alleviate the inflammatory response and prevent CIA progression. This finding clarifies the mechanism of MSCs-DCs crosstalk in eliciting the immunosuppression effect and provides insights into hydrogel-promoted stem cell therapy for autoimmune diseases.
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Objective To observe the effects of transplantation of autologous adipose-derived stem cells (ASCs) on osteoporosis (OP) in a rabbit ovariectomy (OVX) model.Methods A total of fifteen 6-month-old female New Zealand white rabbits were randomly divided into two groups:ovariectomy group (group A,n =12) and sham operation group (group B,n =3).All rabbits were subjected to bilateral ovariectomy in the group A.Six months later,bone mineral density (BMD) of group A and group B were measured by dual energy X-ray absorptiometry (DXA) to check the result of OVX-OP.ASCs harvested from adipose of OP rabbits were cultured to be expanded and differentiated in osteogenic medium in vitro.Osteogenesis was evaluated by alizarin red staining,alkaline phosphatase (ALP) staining and quantitative assays of osteocalcin (OCN).Autologous osteo-induced ASCs were mixed in calcium alginate hydrogel (CAH) and then transplanted in the left distal femurs,while CAH was transplanted in the right distal femurs of OP rabbits.At 12 weeks after implantation,BMD,micro-CT and histomorphological analyses were performed on these rabbits.Results The BMD of femurs in group A rabbits were obviously lower than that of group B rabbits (P ＜ 0.05) at 6 months after OVX.Compared with control group,ASCs cultured in osteo-induction medium had similar proliferation rate as the non-induced cells,but displayed positive ALP and alizarin red staining and OCN contents.At 12 weeks after implantation,the cell-treated femurs displayed higher BMD,bone trabecula number,trabecula thickness and separation than those of control group,while the structure model index and porosity were lower (P ＜ 0.05).Histological examination indicated that the trabecular thickness increased with complete CAH resorption in cell-treated group,while CAH remained in control group.Conclusions Transplantation of autologous ASCs can help strengthen osteoporotic bone in OVX-OP rabbits,providing a novel approach to OP treatment.