[Experimental study on immune tolerance induced by human Ad-CTLA-4Ig in rats with pancreaticoduodenal transplantation].

OBJECTIVE: To study the immune tolerance induced by human Ad-CTLA-41g after pancreaticoduodenal transplantation of rats. METHODS: Wistar rats with diabetes were induced by caudal intravenous administration of streptozotocin (STZ) at a single dose of 60 mg/kg. Pancreaticoduodenal transplantations were performed with the SD rats as donors and the diabete Wistar rats as recipients. The SD pancreaticoduodenal grafts with human Ad-CTLA-41g infection via arterial perfusion in vitro were transplanted into Wistar rats with diabetes. The blood glucose of the rats was measured after transplantation. The graft functional survival time (GFST) was recorded. The concentration of IL-2 was measured by ELISA on day 3 and day 10 after operation. The human CTLA-41g was detected by Western blot after operation. RESULTS: The GFST of control group without gene transfer was (11+/- 1) d, while that of gene transfer group was (33 +/-4) d (P< 0. 01). The expression of serum human CTLA-41g was detected in rats of gene transfer group on the 10th day after transplantation, while no serum human CTLA-4Ig was detected in rats of control group without gene transfer. On the 3rd day after transplantation, the level of serum IL-2 was (33. 710+/-7. 803) pg/mL in rats of transfer group, and was (44. 772+/-9. 102) pg/mL in rats of control group without gene transfer. On the 10th day, the level of serum IL-2 was (47. 846+/-14. 050) pg/mL in rats of transfer group, and was (80. 806+/-18. 629) pg/mL in rats of control group without gene transfer. The levels of serum IL-2 in rats of transfer group was much lower than those of the control group without gene transfer (P

[The experimental research on the perforin and granzyme B expressions to the early diagnostic value of acute rejection response after pancreatic transplantation].

OBJECTIVE: To study the early diagnosis value of acute rejection response to perforin and granzyme B protein expressions around transplantation pancreas tissue after pancreatic transplantation. METHODS: Twenty-four hybrid landraces were randomly divided into three groups [control group ( I group), transplantation group ( II group), transplantation + immunosuppressive group (III group)]. The transplantation group was treated by pancreatoduodenal allotransplantation, and simultaneously the group of transplantation + immunosuppressants was intervened by cyclosporin A (CsA), mycophenolate mofetil (MMF) and metrisone. The peripheral venous blood was sampled via subclavian vein one day before operation and on the 1st, 3rd, 5th or 7th day after operation respectively, and then the glucose, insulin and glucagons in serum were measured. The grafted pancreatic tissues were collected for pathologic and immunohistochemical examinations at 1st, 3rd, 5th and 7th day after operation. RESULTS: (1) The levels of blood sugar, insulin and glucagons had no significant difference among groups of control, transplantation and transplantation + immunosuppressant group (P>0.05). (2) By comparing with two experimented groups, the expressions of perforin and granzyme B protein in transplantation pancreas tissue had a significant difference between transplantation group and transplantation + immunosuppressive group (P < 0.05). (3) The time of perforin and granzyme B protein expressions in transplantation pancreas tissue after transplantation was 2-4 days earlier than that of pathological changes in acute rejection. CONCLUSION: The expression alterations of perforin and granzyme B protein are earlier appearance than the pathological changes of acute rejection response in transplantation pancreas tissue, and by combining with routine pathological examination, the acute rejection response can be diagnosed on early stage.

[The experiment study on RNAi inhibiting the expression of survivin and inducing the apoptosis of pancreatic cancer cells].

OBJECTIVE: To investigate the inhibitory effects of RNAi expression vector suppressing the expression of survivin and inducing the apoptosis of pancreatic cancer cell PANC-1. METHODS: The expression of survivin was examined with RT-PCR and immunofluorescence protocols. The survivin gene was cloned into the T-vector and sequenced, at the same time, the RNAi expression vectors aimed survivin were successfully constructed, and then transfected into PANC-1 cells with liposomes. The expression of survivin mRNA was detected with RT-PCR and Western-blot techniques. The rate of cell apoptosis was measured with FACS. RESULTS: There was a high degree expression of survivin in PANC-1 cells. The DNA sequence was according with the survivin gene in GENE BANK. The survivin expression was inhibited about 70% by pTZU6 + 1-svv2 RNAi expression vectors, and the apoptosis cells increased. CONCLUSION: The RNAi expression vector can effectively inhibit the survivin expression and induce the apoptosis in PANC-1 cells.

Establishment of a pig model with enteric and portal venous drainage of pancreatoduodenal transplantation.

AIM: To establish the pig model of pancreatoduodenal transplantation with enteric drainage (ED) and portal venous drainage (PVD). METHODS: Forty-six hybrid Landrace pigs were divided into two groups (donors and recipients) randomly, and pancreatoduodenal allotransplantation was performed. Donors were perfused via abdominal aorta without clamping the portal venous outflow with UW solution at 80-100 cm H2O after heparinization. Whole pancreato-duodenal grafts were harvested with segments of abdominal aorta and portal vein, and shaped under 4 degrees UW solution. Then, end-to-end anastomosis was performed with the donor iliac artery bifurcation Y graft to the recipient superior mesenteric artery and celiac artery. Furthermore, type I diabetes model was made by removal of the recipient pancreas. The venous anastomosis was reconstructed between the donor portal vein and the recipient superior mesentery vein. Meanwhile, end-to-side anastomosis was performed with the donor common iliac artery bifurcation Y graft to the recipient abdominal aorta, and side-to-side intestinal anastomosis was performed between the donor duodenum and the recipient jejunum. External jugular vein was intubated for transfusion. Levels of plasma glucose, insulin and glucagon were measured during the operation and on the 1st, 3rd, 5th, and 7th d after operation. RESULTS: Pancreatoduodenal allotransplantation was performed on 23 pigs of which 1 died of complication of anesthesia. The success rate of operation was 95.6%. Complications of operation occurred in two cases in which one was phlebothrombosis with an incidence of 4.6%, and the other was duodenojejunal anastomotic leak with an incidence of 4.6%. The level of plasma glucose decreased within 30 min, after removal of pancreas and recovered on the 2nd d after operation. The level of plasma insulin and glucagon increased within 30 min after removal of pancreas and recovered on the 2nd d after operation. Rejection occurred on the 1st d and reached the worst level on the 7th d after transplantation, without change of plasma insulin and glucagon or clinical symptoms of rejection. CONCLUSION: Pancreatoduodenal transplantation in pigs can treat type I diabetes. ED and PVD can keep the function of endocrine in normal. The technique of pancreatoduodenal transplantation with ED and PVD may pave the way for the further application of pancreas transplantation in clinic.