| Background and objectiveChronic wounds affect the life quality of patients and increase the social medical burden,which are a major livelihood issue that urgently needs to be resolved.Adipose-derived stem cells(ASCs)are abundant and readily available,and are the most attractive adult mesenchymal stem cells.Stromal vascular fraction(SVF)is a mixed cell population other than mature adipose cells within adipose tissue.Lots of published literatures have demonstrated the effectiveness of ASCs/SVF cell therapy in the field of wound healing.However,ASCs/SVF cell therapy need to introduce heterologous proteases,which is exogenous and non-clinically approved.In addition,conventional ASCs/SVF cell therapy is transplanted in the form of cell suspension,it not only suffers the risk of ectopic migration,but is vulnerable to phagocytosis from host immune cells,which reduces the safety and effectiveness of ASCs/SVF cell therapy.ECM/SVF gel is a gelatinous mixture enriched in biologically active ECM and SVF and result from adipose tissue treated with purely physical methods.In our previous study,ECM/SVF gel can promote wound healing in immunocompromised mice.However,it is unclear whether ECM/SVF gel can promote the chronic wounds with complicated pathogenesis.In addition,recent studies show that paracrine secretion of ASCs and/or adipose tissue is the main way to promote wound healing.Therefore,we speculate that the beneficial effects of ECM/SVF gel on wounds may also come from its paracrine.In summary,the purpose of this study is to investigate whether ECM/SVF gel promotes healing of diabetic rat wounds and clinical chronic wounds and whether paracrine is the main way to promote wound healing.This research is believed to be helpful to build a theoretical basis and provide stable experimental evidence for clinical application of ECM/SVF gel and paracrine components.Methods1.Basic research of accelerating skin wound healing in diabetic mice with allogeneic ECM/SVF gelThe full-thickness skin defect model from streptozotocin-induced diabetic rat was established.Skin wounds of diabetic rat was study after the ECM/SVF gel allograft.Observational indicators include wound healing rate,HE staining and Masson staining.2.Clinical research of promoting chronic wound healing with autologous ECM/SVF gelTwenty chronic wound patients were enrolled in this study.Ten patients underwent autologous ECM/SVF gel transplantation,and other ten patients underwent wound negative pressure closure therapy.Observational indicators include the healing rate of per week,HE staining,Masson staining,and CD31 immunohistochemistry.3.Preliminary mechanism study of accelerating wound healing with ECM/SVF gelThe ECM/SVF gel,SVF suspension and adipose tissue were cultured in vitro,and the conditioned media(Gel-CM,SVF-CM and Adi-CM)were obtained,respectively.Human dermal fibroblasts(HDFs)and immortal keratinocyte(HaCaT keratinocyte)cell lines were purchased.The effects of Gel-CM with different concentrations and three CMs on the proliferation and migration of HDFs and HaCaT cells cultured in vitro and secretory collagen from HDFs were studied.Three CMs were locally transplanted into full-thickness skin defect wounds of immunodeficient mice to observe wound healing.Observational indicators include wound healing rate and histology examination.4.Mechanism study of accelerating chronic wound healing with ECM/SVF gelChronic wound exudates and three CMs(Gel-CM,SVF-CM,and Adi-CM)were collected.The effects of three CMs on the proliferation and migration of HDFs and HaCaT cells and collagen secretion of HDFs after intervention with exudate were observed,Finally,ELISA was used to detect the expression of bFGF,EGF,VEGF and TGF-β1 in three CMs.Results1.Basic research of accelerating skin wound healing in diabetic mice with allogeneic ECM/SVF gelAfter ECM/SVF gel transplantation,inflammatory cell infiltration in the dermis reduced,adipose tissue regenerated significantly,and collagen deposition increased,resulting in earlier forming complete skin structure and improving wound healing rate in diabetic rats.2.Clinical research of promoting chronic wound healing with autologousECM/SVF gelThe healing rate of the ECM/SVF gel transplantation group was 34.55%±11.18%per week,while that of negative pressure treatment group was only 10.16%±2.67%.Lymphocytic infiltration of the deep dermis significantly decreased,collagen deposition increased,and a large number of neovascular and neo-adipogenesis were observed in the ECM/SVF gel transplantation group.3.Preliminary mechanism study of accelerating wound healing with ECM/SVF gelThree CMs could promote the proliferation and migration of HDFs and HaCaT cells,and affect the synthesis of type I collagen in HDFs,while the promotion effect of Gel-CM was more obvious and related to the dose.Gel-CM accelerated wound healing in nude mice and increased skin appendages regeneration and collagen synthesis.4.Mechanism study of accelerating chronic wound healing with ECM/SVF gelThree CMs could promote the proliferation and migration of HDFs and HaCaT cells after exudate intervention,and affect the synthesis of type I collagen by HDFs,while the promotion effect of Gel-CM was more obvious.Gel-CM contained high concentrations of bFGF,EGF,TGF-β1 and VEGF.Conclusions1.Allogeneic ECM/SVF-gel transplantation accelerates skin wound healing and improves healing quality of diabetic rats through promoting neo-adipogenesis and collagen deposition within dermis under the wound,and earlier forming complete skin structure.2.ECM/SVF-gel transplantation is a simple,safe and effective treatment for chronic wounds.3.ECM/SVF-gel transplantation promotes chronic wound healing by the paracrine mechanism.4.The mechanism of ECM/SVF-gel in chronic wounds is similar to that of ordinary wounds,which is the increasing of proliferation and migration of HDFs and HaCaT cells and collagen synthesis.5.Gel-CM has a wide application space in tissue regeneration and wound healing. |
