c-Jun氨基末端激酶信号通路在不同剂量舒芬太尼预处理大鼠肝脏缺血-再灌注损伤中的作用

目的研究舒芬太尼预处理对大鼠肝脏缺血-再灌注损伤的作用及其与c-Jun氨基末端激酶(JNK)信号通路的关系。方法健康SD大鼠162只,雌雄不拘,体重250~300g,采用随机数字表法将其分为七组:假手术组(S组,n=30)、肝脏缺血-再灌注组(IR组,n=30)、1μg/kg舒芬太尼预处理组(SF1组,n=30)、5μg/kg舒芬太尼预处理组(SF5组,n=30)、10μg/kg舒芬太尼预处理组(SF10组,n=30)、阻断剂SP600125组(SP组,n=30)及二甲基亚砜组(DMSO组,n=6)。舒芬太尼预处理组分别在缺血前30min静脉输注不同剂量(1、5、10μg/kg)舒芬太尼,S组及IR组给予等体积生理盐水,SP组于缺血前30min腹腔注射15mg/kg JNK阻断剂SP600125,DMSO组于缺血前30min静脉注射与溶解SP600125等体积的DMSO;S、IR、SF1、SF5、SF10组于再灌注即刻(T1)、1h(T2)、2h(T3)、4h(T4)、6h(T5),SP、DMSO组于T3时取肝组织及腹主动脉血2ml;测定血清ALT和AST活性;观察肝组织MDA和SOD的变化;HE染色观察肝组织病理学改变;免疫组化法测定p-JNK;Western blot法测定肝组织p-JNK的蛋白表达水平。结果 T1~T5时IR、SF1、SF5、SF10组,T3时SP、DMSO组血清ALT、AST明显高于S组(P0.05);T1~T5时SF1、SF5、SF10组,T3时SP组血清ALT、AST明显低于IR组(P0.05);T1~T5时IR、SF1、SF5、SF10组,T3时SP、DMSO组肝组织MDA、SOD水平明显高于S组(P0.05);T1~T5时SF1、SF5、SF10组,T3时SP组肝组织MDA、SOD水平明显低于IR组(P0.05);T4时SF10组MDA、SOD水平明显低于SF1、SF5组(P0.05)。与T1时比较,T3时IR组p-JNK表达明显升高(P0.05);T3时IR、SF1、SF5、SF10、SP、DMSO组p-JNK表达明显高于S组(P0.05),SF1、SF5、SF10、SP组p-JNK表达明显低于IR组(P0.05),SF5、SF10组p-JNK表达明显低于SF1组(P0.05),SF10组p-JNK表达明显低于SF5组(P0.05)。结论舒芬太尼预处理可减轻大鼠肝脏缺血-再灌注损伤,且10μg/kg舒芬太尼保护作用最为显著,其机制可能是通过抑制JNK通路,降低p-JNK的表达,从而减轻大鼠肝脏缺血-再灌注损伤。

Role of c-Jun N-terminal kinase signal pathway in sufentanil preconditioning against hepatic ischemia-reperfusion inj ury in rats

Objective To detect the protective effect of sufentanil preconditioning on hepatic ischemia-reperfusion injury in rats and the role of c-Jun N-terminal kinase signal pathway in the mech-enism.Methods One hundred and sixty-two SD rats(in either gender,weighing 250-300 g)were ran-domly divided into seven groups:Sham-operated group (group S,n = 30 ),ischemia-reperfusion group (group IR,n =30),sufentanil preconditioning group (group SF1:1 μg/kg,n =30;group SF5:5 μg/kg,n =30;group SF10:10 μg/kg,n =30),SP600125 group (group SP,n =30),and dimethyl sulphoxide control group (group DMSO,n =6),different doses of sufentanil was administered 30 min before hepatic ischemia in group SF1,SF5 and SF10.Blood and liver samples were collected from each group at 0(T1 ),1 (T2 ),2 (T3 ),4 (T4 ),and 6 (T5 )hours after reperfusion.Serum alanine amin-otransferase (ALT)and aspartate aminotransferase (AST)were measured by an automatic biochemi-cal analyzer.Malondialdehyde (MDA)and superoxide dismutase (SOD)in liver tissue was measured. Liver sample was stained with HE to observe the hepatic pathological changes.Immunohistochemical method was used to determine the expression of JNK and western blotting was used to detect the ex-pression of P-JNK.Results Compared with group S,levels of AST,ALT increased significantly in group IR,SF1,SF5,SF10 at T1-T5 and in group SP,DMSO at T3 (P <0.05 ).Compared with group IR,levels of AST,ALT decreased significantly in group SF1,SF5,SF10 at T1-T5 and in group SP at T3 (P <0.05).Compared with group S,levels of MDA,SOD increased significantly in group IR,SF1, SF5,SF10 at T1-T5 and in group SP,DMSO at T3 (P < 0.05 ).Compared with group IR,levels of MDA,SOD decreased significantly in group SF1,SF5,SF10 at T1-T5 and in group SP at T3 (P <0.05).Compared with group SF1 and SF5,levels of MDA,SOD decreased significantly in SF10 at T4 . Compared with T1 ,the expression of p-JNK in group IR increased significantly at T3 (P < 0.05 ). Compared with group S,the expression of p-JNK in groups IR,SF1,SF5,SF10,SP,DMSO increased significantly at T3 (P < 0.05 ).Compared with group IR,the expression of p-JNK in groups SF1, SF5,SF10,SP decreased significantly and that in groups SF5,SF10 were less than that in group SF1 (P <0.05 ).The expression of p-JNK in group SF10 was less than that in group SF5 (P < 0.05 ). Conclusion Sufentanil preconditioning can reduce the hepatic ischemia-reperfusion injury and the dos-age of 10 μg/kg was the most effective.The protective mechanisms may inhibit JNK pathway and re-duce the expression of JNK.