| Objectives:To establish a reliable animal model of rat orthotopic liver transplantation(OLT) with modified cuff technique.Silence the expression of heme oxygenase-1(HO-1) gene by delivery of synthetic HO-1 siRNA in rat liver graft,to investigate the impact of diazoxide preconditioning on the expression of HO-1 in liver cell,and explore the function and potential mechanism of HO-1 in rat liver grafts after ischemiareperfusion injury.Methods:1.Orthotopic liver transplantations in rats were performed by Kamada's doubl cuff techniques with some modification.2.OLT in rat were performed after HO-1 siRNA sequence was delivered in donor 48h.Rat were randomly divided into five groups namely:group A(I/RI group),group B(HO-1 siRNA group),group C(diazoxide group),group D(HO-1 siRNA+diazoxide group) and group E(control siRNA group).At 6h after liver transplantation,the pathological changes of liver tissue and the serum AST,ALT levels were observed in each group.The IL-6,TNF-αlevels of serum were detected in each group.The expression of HO-1 mRNA and protein levels were determined by RT-PCR and Western blot.Results:1.120 cases of rat OTL operations were performed in total.The operation time of donor was(34.2±2.9) min,the operation time of recipient was(43.6±2.9) min,the operation time of suprahepatic vena cava was(8.4±1.3) min,and the anhepatic phase was(14.5±1.1) min.We successfully established a stable rat liver transplantation model.2.The levels of HO-1 mRNA and protein in group C were remarkably higher than group A(p＜0.01),in group B and D were remarkably lower than group A(p＜0.01). There was no statistical difference of HO-1 mRNA and protein levels between group B and group D(p＞0.05).The pathological injure degree of liver tissue and the serum AST,ALT,IL-6,TNF-αlevels in group C were significantly lower than other groups; The pathological injure degree of liver tissue and the serum AST,ALT,IL-6,TNF-αlevels in group B and D were significantly higher than other groups.Conclusion:1.The modified Kamada's "doubl-cuff" technique is a stable,reliable,convenient and practicable model.2.HO-1 siRNA sequence inhibits HO-1 expression in vivo and enhances liver ischemia-reperfusion injure,whereas the nonspecific siRNA had no measurable effect on HO-1 expression.3.Diazoxide,which open the mitoKATP,through enhancing of HO-1 activity can relieve I/RI after liver transplantation,The potential mechanism is that HO-1 exerts cytoprotection against I/RI by reducing IL-6,TNF-αand inhibiting hepatocyte apoptosis. |
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