Study On The Effect Of Allogeneic Skin Transplantation And Its Anti-Rejection Reaction And Wound Healing In Small Pigs

The skin has the role of protection,excretion,regulation of body temperature and sensing external stimuli,and it is also one of the most important immune organs of the body.However,when a large skin defect is caused by burns or due to various traumatic injuries,the skin is damaged in a large area.The treatment is usually skin grafting surgery.Skin grafting is divided into autologous skin grafting and allogeneic skin grafting,and autologous skin grafting is preferred when grafting is needed because it has the advantages of low immunogenicity,fast establishment of blood supply,remarkable skin healing,and significant reduction of skin scars.However,when the autologous skin does not have enough graftable area to be transplanted into the wound,a suitable allogeneic skin can be selected for transplantation between homografts.For allogeneic skin grafts,whether mated or not,immune rejection will occur and the main solution for clinical treatment is the use of immunosuppressive drugs.Numerous studies in recent years have shown that intervention with mesenchymal stem cells(MSCs)can improve skin wound fibrosis,prevent lipid peroxidation,inhibit inflammation and promote skin regeneration,a finding that has led to an increasing number of researchers using MSCs as drugs for the treatment of skin-related diseases.In this experiment,12 small Bama pigs aged about 1 year old of 30-40 kg were taken to establish an allogeneic skin graft model.According to the different postoperative interventions on the allogeneic skin grafts,the small pigs were randomly divided into 4 groups: allogeneic skin graft group(Control),methylprednisolone group(MP),adipose mesenchymal stem cells group(ADSCs),and adipose mesenchymal stem cells combined with methylprednisolone(ADSCs+MP).Blood and tissue samples were collected preoperatively and at 7 d and 14 d postoperatively,and clinical changes of the grafted epidermis were photographed and recorded immediately,at 7 d,14 d and 21 d postoperatively.Through the superficial examination of the grafted skin pieces,pathological histological examination,routine blood(WBC,NEU,LYM)and serum biochemistry(GLB),Transplanted skin slice tissue oxidative stress(ELISA kits: SOD,GSH),inflammation(immunohistochemistry: CD4,ELISA kits: IL-2,IL-4,IL-10,IL-1β,IL-6,TNF-α,IFN-γ,Western blot with q PCR:PI3K,AKT,p-AKT,NF-κB,p-NF-κB),regeneration(ELISA kits: cortisol,CXCR4,SDF-1,Western blot with q PCR: 11β-HSD1,11β-HSD2,TGF-β,VEGF)were detected.The effects of the four groups of different interventions on the immune rejection and wound healing aspects arising from allogeneic skin grafts in minipigs were evaluated comprehensively.The results of experimental studies showed that it could be known from the apparent examination and pathological analysis of the skin sheet.At 7 days,the skin grafted in the control group turned black and hardened,and the epidermis fell off,and the epidermis layer was separated from the dermis layer.This indicates that the control group skin sheet necrosis at 7 d.The skin sheet in the MP group was flushed in color,thickened stratum corneum,and there were a small number of inflammatory cells at 7 days,and the survival time was about 7-14 days,and the ADSCs group and ADSCs+MP group showed that the skin sheet turned black and the texture was soft when 14 days,and a small part of the epidermal dermis turned black when the skin sheet was cut,and the epidermis and dermis layer were not separated,there were a small number of inflammatory cells,and its survival time was 14-21 days.The expression of GSH and SOD was significantly higher in the ADSCs+MP group at 7 d and 14 d.In terms of immune rejection,the number of CD4 T cells,the protein and gene expression of PI3 K,AKT,NF-κB,and the expression of IL-2,IFN-γ,TNF-α,IL-1β,and IL-6 pro-inflammatory factors were significantly lower in the ADSCs+MP group relative to the Control group.In terms of wound healing,the protein expressions of VEGF and TGF-β,and the expressions of SDF-1 and CXCR4 were significantly increased in the ADSCs+MP group relative to the Control group.And the protein and gene expression of11β-HSD1,11β-HSD2 were significantly decreased in the ADSCs+MP group relative to the Control group.In summary,ADSCs and ADSCs+MP can effectively prolong the survival time of skin slices in small pig allogeneic skin grafting,significantly inhibit the differentiation of CD4 T lymphocytes and the expression of inflammatory response factors,significantly reduce oxidative stress response,and promote vascular regeneration and migration of skin tissue.The results showed that ADSCs and ADSCs+MP had the functions of anti-oxidation,anti-immune rejection and tissue regeneration,and the effect of ADSCs 、 ADSCs+MP in inhibiting inflammation and promoting vascular regeneration was significantly better than that of ADSCs.This provides a theoretical basis for the clinical application of ADSCs、ADSCs+MP.