The Effect Of Thalidomide On Acute And Chronic Rejection A Rat Kidney Transplantation Model And Its Immunoregulatory Mechanism

Inducing successful graft tolerance can make humans cross the natural barriers of the immune system and truly accomplish the ultimate goal of organ replacement.Successful immune tolerance can not only prolong graft survival,but also alleviate the huge burden of organ transplantation.Economic burden,physical and mental stress of patients and short-term medical conditions of organs.Although the current treatment plan can reduce the occurrence of acute and chronic rejection,it can not achieve tolerance.After 4 to 5 years of renal transplantation,about 30%of patients will show pathological manifestations of chronic rejection.Therefore,the primary goal of the current transplant community is to explore new treatment options.In particular,the development of new drugs has very obvious scientific value.The rat kidney transplantation model has the potential to study tolerance induction by means applicable to human conditions,and the researchers can understand the pathogenesis by manipulating rat kidney transplant rejection.Reconstruction of the urinary tract is one of the key techniques in the rat kidney transplantation model.Because the ureter has a very narrow caliber,it is easy to cause complications such as urinary leakage,ureteral necrosis and ureteral stricture,which may lead to deterioration of hydronephrosis or renal function.Urinary tract reconstruction can also lead to different outcomes of kidney transplantation.We established a rat kidney transplantation model using a donor vesicle-containing small bladder-receptor bladder double suture technique to reconstruct the urinary tract,and compared it with the donor bladder-receptor bladder anastomosis to improve the urinary tract.Reconstruction methods to reduce the incidence of complications.Our previous study found that thalidomide can significantly reduce atherosclerosis in rat vascular grafts and reduce graft vascular disease.The mechanism is to improve the inflammatory response,reduce lymphocyte infiltration,reduce the expression of VEGF,PDGF,ICAM,TNF-a and PCNA,delay the hardening process of the graft and reduce the intimal hyperplasia.In this experiment,we established a rat kidney transplantation model by establishing a modified donor small bladder-recipient bladder double-layer suture urinary tractreconstruction to observe the effect of thalidomide on acute and chronic renal allograft rejection in rats.Its immunological system.The effects of thalidomide on the proliferation and differentiation of alloreactive T cells in vitro were observed by mixed lymphocyte culture,and the regulation mechanism of reactive lymphocytes was elucidated.Part I:Application of modified small bladder patch-to-bladder double-layer sutures to establish a rat renal transplantation model.Objective:To establish a rat kidney transplantation model with modified donor small bladder-receptor bladder double-layer suture urinary tract reconstruction,and compare with donor bladder-receptor bladder anastomosis to identify improved rat kidney transplantation.The effect of urinary tract reconstruction.Methods:Rat kidney transplantation model was established by donor small bladder-recipient bladder double-layer suture urinary tract reconstruction and donor bladder-receptor bladder anastomosis urinary tract reconstruction:group A,donor small bladder valve-Receptor bladder mucosa muscle layer double suture(n=12);group B,donor bladder valve-receptor bladder monolayer suture(n=11).Compare urinary tract reconstruction time and complications.Results:The urinary tract reconstruction time was 14.12±1.73min in group A and 10.16±1.19min in group B.The difference between the two groups was statistically significant(P<0.05).The urinary tract complication rate in group A was 25%.The incidence of urinary tract complications in group B was 9.09%,and the difference between the two groups was statistically significant(P<0.05).Conclusion:The modified urinary tract reconstruction with small bladder-receptor bladder double suture can reduce urinary tract complications after renal transplantation in rats,although the urinary tract reconstruction time is longer than the traditional single-layer suture time.It can significantly reduce the possibility of urinary leakage,and is a highly reliable reconstruction procedure.Part Ⅱ:Effect of thalidomide on acute renal allograft rejection in ratsObjective:Objective:To elucidate the role of thalidomide in acute renal allograft rejection in rats.Methods:A Fischer-Lewis rat kidney transplant model was established.A homologous LEW-LEW graft was used as a control.Recipient rats were divided into a homologous transplantation group(Isograft,Iso),an allograft group(Allograft,Allo),a cyclosporine group(CsA),and a thalidomide group(Thalidomide,Tha).Dynamic monitoring of postoperative creatinine(Scr)changes,observed to 5 days after surgery,blood samples were taken for the detection of interleukin(IL)-2,IL-6,IL-17 and tumor necrosis factor-a(TNF-a)The concentration of the transplanted kidney was pathologically examined.Results:The creatinine of the Allo group and the Tha group increased progressively in the acute rejection model.There was no significant difference in the creatinine level between the Tha group and the Allo group,which was significantly higher than that in the Iso group and the CsA group.The pathological changes of renal transplantation in the Tha group were similar to those in the Allo group.All of them showed acute tubular injury,diffuse severe inflammatory infiltration with severe glomerulopathy,endometrial arteritis and hyaline degeneration of arterioles.The concentrations of IL-2,IL-6,IL-17 and TNF-a in the Allo group were significantly increased.The concentrations of IL-6,IL-17 and TNF-a in the serum of the Tha group were significantly lower than those in the Allo group,but the concentration of IL-2 was significantly higher than the Allo group.Conclusion:These results indicate that thalidomide has a weak immunosuppressive effect and is less effective in acute rejection.Part Ⅲ:The effect of thalidomide on chronic renal allograft rejection in rats and preliminary study on its immunoregulatory mechanismObjective:To observe the effect of thalidomide on chronic rejection in rat kidney transplantation model and to clarify its exact mechanism.Methods:A Fischer-Lewis rat kidney transplant model was established.A homologous LEW-LEW graft was used as a control.Recipient rats were divided into a homologous transplantation group(Isograft,Iso),an allograft group(Allograft,Allo),and a thalidomide group(Thalidomide,Tha).Serum levels of creatinine,IL-2,IL-6,IL-17 and TNF-α were measured at 8 weeks postoperatively.The distribution of T cell subsets in peripheral blood was detected by flow cytometry.Pathological examination of transplanted kidney was performed;immunofluorescence and Western blotting were used to detect the expression of transforming growth factor β1(TGF-β1),smooth muscle actin alpha(a-SMA)and vascular endothelial growth factor(VEGF)protein in renal tissues of each group..Results:At 8 weeks after kidney transplantation,pathological changes such as mononuclear cell infiltration,glomerular sclerosis,tubular atrophy,interstitial fibrosis,and small intima thickness were observed in the transplanted kidney.The pathological changes of the transplanted kidney in the Tha group were significantly lighter than those in the Allo group.The levels of serum creatinine and serum inflammatory cytokines in the Tha group were significantly lower than those in the Allo group.The proportion of CD4+CD25+,CD4+CD25+FoxP3+T cells in the peripheral blood of the Tha group was significantly increased,and the proportion of CD4+Th17+ T cells was significantly increased.Significantly decreased,the expression of TGF-β1,α-SMA and VEGF protein was significantly reduced.Conclusion:Thalidomide can significantly alleviate chronic rejection.The mechanism may be that thalidomide reduces the secretion of inflammatory factors,changes the distribution of T cell subsets,and reduces the expression of related fibrotic proteins.Part IV:Regulation of thalidomide on alloreactive lymphocytesObjective:To observe the effect of thalidomide on body-reactive lymphocytes in vitro and to elucidate its immunoregulatory effect on reactive lymphocytes.Methods:The Fischer-Lewis mixed lymphocyte culture(MLC)system was established and divided into control group(Control,Con)and thalidomide group(Thalidomide,Tha).Flow cytometry was used to detect the distribution of T cell subsets and apoptotic rate,the concentration of IL-2,IL-6,IL-17 and TNF-a in the culture medium was determined by ELISA.Results:The proportion of Th17 cells in the Tha group was significantly lower than that in the control group at the same time.The proportion of Treg cells was significantly higher than that in the Con group at the same time point(P<0.05).The IL in the culture medium of the Tha group The concentration of IL-2 was significantly higher than that of the Con group at the same time point(P<0.05).The apoptosis rate of T cells in the Tha group was significantly higher than that in the Con group(P<0.05).IL-6,IL-17 and TNF-a concentrations were significantly lower than the Con group at the same time point(P<0.05).Conclusion:Thalidomide can promote the proliferation of Treg cells in allergic T cells,inhibit the proliferation of Th17 cells,change the distribution of T cell subsets,increase the apoptosis of allogeneic T cells,and reduce the concentration of IL-6,IL-17 and the TNF-α.