Experimental Study On Acute Rejection Of TGF-?1 Modified MSC In Beagles With Pancreas-kidney Transplantation

Objective:Establish a stable Beagles simultaneous pancreas-kidney transplants(SPK)model.Select appropriate experimental animals for this and subsequent related experiments,and explore and improve the surgical procedure for acute rejection after simultaneous pancreas-kidney transplants.To investigate the effect and molecular mechanism of TGF-?1 modified MSC on acute rejection after combined pancreas-kidney transplantation in dogsMethods:In this study,male beagle dogs weighing 11.5Kg-13.0Kg were selected as experimental subjects.They were randomly divided into donor groups and recipient groups,and each group received 25 pancreas-kid transplantation.Experimental group(5 recipients per group,5 donors):Group A:MSC intravenous infusion before recipient pancreas-kidney transplantation;Group B:TGF-?1 transfected MSC vein before recipient pancreas-kidney transplantation Infusion;Group C:Receptor pancreas-kidney combined with intravenous infusion of 5ml PBS and food for cyclosporine 150mg;Group D:recipient of pancreas-kidney transplantation before TGF-?1 transfection of MSC intravenous infusion,and food to the cyclosporine 150 mg;Group E:5 ml PBS was infused intravenously before donor pancreas-kidney transplantation.The cells in each group were intravenously administered 5 days before transplantation,and the number of cells was 1*107,which was resuspended in 5 ml of PBS.In the C and E groups,5 ml of PBS was intravenously administered five days before surgery.The donor graft is the whole pancreas and the duodenum in the 2nd and 3rd segments.The graft arterial blood supply preserves the abdominal aorta and the mesenteric artery segment.The venous end is a sufficiently long portal vein.Intestinal artery.The recipients were surgically resected to produce a type 1 diabetes model in the tail of the pancreas.The pancreaticoduodenal was transplanted into the right axillary fossa of the recipient,the kidney was transplanted into the left axillary fossa of the recipient,and the external drainage was performed with bladder drainage(BD).The inferior vena cava is recirculated(SV).The data of the operation time of the donor,the amount of perfusate,the time of the recipient operation,and the blood loss of the recipient during the operation were recorded.Before the pancreas transplantation,5 ml of the recipients were collected at 1,3,5,and 7 days after surgery.Blood,blood serum,C-peptide,IL-2,IL-4,IL-10 were detected by ELISA.Urine was collected at 8h,Id,3d,5d and 7d after operation to measure the amylase concentration.On the 7th day after transplantation,the recipient Beagle was sacrificed(if the recipient died within 7 days,the specimen was taken immediately),the transplanted pancreas and kidney were taken,and the pancreas and kidney were sliced and stained according to the standard procedure of HE staining.The score was based on the Nakhleh protocol,and the grade of acute rejection of transplanted kidneys was graded according to the Banff protocol.Results:1.25 groups of Beagle donors and recipients were successfully performed.The survival rate of the recipients was 96.0%after 24 hours(1 case of postoperative anatomy considering death due to hemorrhagic shock),donor surgery time:96.840±11.53min The amount of pancreatic kidney perfusate was 854.560±191.11ml,the operation time of the recipient was 137.464±1 8.28min,the blood loss of the recipient was 213.800±46.63ml,the thrombosis rate of the recipient was 8.00%,and the graft ischemia was 4.00%(1 Case of infarction).The Beagle pancreas-kidney transplantation model was successfully constructed.2.Comprehensive analysis of venous blood glucose,C-peptide value and urinary amylase after pancreas-kid transplantation in each experimental group:MSC group(group A),TGF-?1 modified MSC group(group B),cyclosporine group(C)There was no significant difference in the difference between TGF-?1 modified MSC combined with cyclosporine group(D group)(p>0.05).The 7d venous blood glucose level in the blank group(E)was significantly different from the TGF-?1 modified MSC combined with the cyclosporine group(D).The urinary amylase activity in the blank group(E)was lower than that in the other 4 groups,and the difference was statistically significant.Academic significance(p<0.05).3.Comprehensive analysis of peripheral blood immune related molecules IL-2,IL-4 and IL-10 in each group:Peripheral blood IL-2 levels in the blank group(E)after 5-7 days after transplantation were higher than those in the other 4 groups The difference was statistically significant(p<0.05),IL-4 level was lower than the other 4 groups,the difference was statistically significant(p<0.05),TGF-?1 modified MSC combined with cyclosporine group(D)5,7d peripheral The blood IL-2 water was relatively low,which was statistically different from the MSC group(A)(p<0.05),and the IL-4 level was higher than the other groups(p<0.05).4.Comprehensive pancreatic and renal pathological slice scores showed that MSC,TGF-?1 transfected MSC,cyclosporine can alleviate acute rejection after transplantation,and TGF-?1 transfected MSC combined with cyclosporine use than alone.MSC or TGF-?1 transfected MSCs were more effective in reducing immune rejection.Conclusion:This experiment established a model of pancreas-kidney transplantation for acute rejection after pancreas-kidney transplantation.MSC,TGF-?1 modified MSC,cyclosporine,TGF-?1 modified MSC combined with cyclosporine can alleviate beagle dogs.Acute rejection after pancreas-kidney transplantation,in which TGF-?1 modified MSC combined with cyclosporine was the best.TGF-?1-modified MSC combined with cyclosporine may attenuate IL-4 expression and inhibit IL-2 expression to alleviate acute rejection after beograft combined with pancreas-kidney.