The Paracrine Effects Of Adipose-derived Stem Cells Promoted The Cutaneous Wound Healing In Streptozotocin-induced Diabetic Rat Model

ObjectiveThe purpose of this study was to explore the paracrine effects of adipose-derived stem cells(ASCs)on the proliferation,migration and differentiation of fibroblasts in vitro.In addition,full-thickness wound model was made to detect the effect of adipose-derived stem cells conditioned medium(ASCs-CM)on wound healing in diabetic rats.MethodsThe adipose tissue of healthy SD rats was harvested and digested by collagenase.CD44,the specific surface antibody of adipose stem cell,was identified by immunofluorescence staining.The conditioned medium of the 5th generation ASCs was collected and concentrated by ultrafiltration centrifuge tube.ASCs-CM.Fibroblasts were divided into two groups: treatment group(ASCs-CM)and control group(N-CM).The effects of ASCs-CM on the proliferation and migration of fibroblasts were observed by CCK-8 and scratch experiments.Western blot and qRT-PCR were performed to study the effects of ASCs-CM on the differentiation of fibroblasts into myofibroblasts.The diabetic SD rat model was induced by streptozotocin,and the full-thickness skin wound model of diabetic rats was prepared.The diabetic rats were randomly divided into ASCs-CM group and N-CM group,and 0.5 mL of the above solution was injected around the wound surface.The existing wound area was measured on the day 4,8,12 and 16 after the postoperative day,and some of the wound tissues were collected for immunohistochemical staining and qRT-PCR quantitative study.ResultsIn this experiment,adipose stem cells were successfully isolated and identified by immunofluorescence,which meet the characteristics of adipose stem cells.The results of CCK-8 assay and cell scratch test showed ASCs-CM could significantly promote the proliferation and migration of fibroblasts.At the same time,Western blot and qRT-PCR further confirmed that ASCs-CM could promote the transformation of fibroblasts into myofibroblasts.The healing rate of full-thickness skin wounds in diabetic rats was significantly higher in the ASCs-CM group than in the N-CM group on day 4,8,12 and 16 after postoperative days respectively.Immunohistochemical staining and qRT-PCR results showed that the expression of ?-SMA was higher than that of N-CM group on the day 4 after ASCs-CM treatment(P<0.05)and the expression of TGF-?1 was significantly higher than the control group on the 4th,8th and 12 th day after treatment,and the difference was statistically significant.ConclusionsThis experiment demonstrates that adipose-derived stem cell conditioned medium may accelerate wound healing in diabetic SD rats by promoting the secretion of TGF-?1 and promoting the proliferation,migration and differentiation of fibroblasts.