| Objective:To modify rat orthotopic liver transplantation method with two - cuff technique.To investigate the influence of pre-infusion of spleen cells from dexamethasone-treated rats in vivo on allograft survival of liver cirrhosis rats orthotopic liver transplantation, and develop new ideas and feasible ways for induction of clinic transplantation immune tolerance under no common immunosuppression.Methods:The animal model of liver cirrhosis was made with carbon tetrachloride, and the two-cuff orthotopic liver transplantation was performed. Wistar rats were used as donors, and SD rats as recipients. Some skills in Kamada's method were modified. Vein canulas were made of cotton - sticks according to vein size. Abdominal transverse incision was made both in donor and in recipient. Before the anhepatic period , tractive sutures were placed under the branch of portal vein(PV) . "Three poinT"anastomosis for suprahepatic vena cava(SVC) was used. PV and infrahepatic vena cava( IVC) anastomosesweremade using cuff technique, while the hooked injector was used to expel air. The SD rats were divided into normal liver function group, normal operation control group(N-Control), liver cirrhosis operation control group(C-Control), dexamethasone group(Dex), spleen cells group(SpC) and dexamethasone-treated spleen cells group(Dex-SpC). The N-Control and C-Control have nothing treatment; The Dex donor rats were treated with dexamethasone but the recipient rats did not infuse spleen cells; The SpC recipient rats were infused intravenously with donor spleen cells(5107) 7 days before transplantation but the donor rats were not treated with dexamethasone; The Dex-SpC recipient rats were infused intravenously with dexamethasone-treated donor spleen cells(5107) 7 days before liver transplantation. The survival time of each groups were observed. The blood alanine transaminase(ALT), total bilirubin(TBIL) and liver pathological changes of each groups were observed 1 week after the transplantation. Apoptotic percentage of spleen cells from dexamethasone-treated rats in vivo was measured by flow cytometry using Annexin V/PI double staining method.Results:(1) Orthotopic liver transplantations were performed in 26 rats, including 6 training operations and 20 experimental operations. With the modified method , The mean time for donor operation and liver preparation were 29 min and 10 min, and the recipient operative and anhepatic time were 51 min and 17 min respectively. The two - day and one - week survival rates were-8-90%(18/20) and 85%(17/ 20).(2) Dexamethasone-treated in vivo administrated induced donor spleen cells apoptosis (32.52?.24%) , while the non-treated groups did not(0.76?.11%).(3) The median survival time(MST) of N-Control, C-Control and Dex was 10, 17 and 16 days respectively, and had no difference each other(P0.05). The MST of group infused donor spleen cells was 28 days, and had significant difference to other groups (P0.01). The MST of group infused with donor in vivo dexamethasone-treated spleen cells was no less than 60 days, showed long-term survival, and had significant difference to other groups0.0l).(4) ALT, TBIL in C-Control, N-Control and Dex group were increased significantly 7 days after operation. ALT, TBIL were better in non-treated donor spleen cells group and the best in dexamethasone-treated donor spleen cells.(5) Rejection active index(RAI) of liver acute rejection pathology in C-Control, N-Control and Dex were between 6 and 9, and have no difference each other(P0.05). RAI in non-treated donor spleen cells group were between 4 and 6 and had significant difference to other groups (P0.01). RAI in dexamethasone-treated donor spleen cells were between 3 and 5 and had significant difference to other groups (P0.01).Conclusions: This modified two - cuff method in rat orthotopic liver transplantation can-9-simplify operation procedure , increase operative success rate. It can be performed safely , reliably and practicably by single one operator. All anastomosis must be kept in situ w...
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