肝硬化大鼠肝缺血-再灌注损伤后Bsep、Mrp2表达在胆汁淤积中的作用研究

Expression of Bsep and Mrp2 in liver of cirrhotic rats with intrahepatic cholestasis after ischemia-reperfusion injury

目的研究肝硬化大鼠肝缺血-再灌注损伤(HIRI)后胆汁淤积的部分分子机制。方法将健康SD大鼠随机分成A、B、C 3组,每组15只,建立肝硬化、HIRI模型。A组为正常大鼠HIRI组,B组为肝硬化大鼠假手术组,C组为肝硬化大鼠HIRI组。每组大鼠肝门阻断时间为30min。分别检测各组大鼠术后第1、3、5天的胆汁DBil、TBil含量,血清AST、ALT、TBil、DBil含量;Western blot分析各组大鼠术后第1、3、5天胆盐输出泵(Bsep)、多耐药相关蛋白2(Mrp2)表达变化;免疫组织化学法分析Bsep蛋白的细胞亚定位。结果术后第1天,C组大鼠胆汁TBil、DBil含量均低于A组(均P<0.05);术后第3天,C组大鼠胆汁TBil、DBil含量均低于A组和B组(均P<0.05);术后第5天,3组大鼠胆汁TBil、DBil含量比较差异均无统计学意义(均P>0.05)。C组大鼠术后第1、3天血清AST、ALT水平均高于A组,术后第1、3、5天血清TBil、DBil含量均高于A、B组(均P<0.05)。术后第1、3、5天,C组大鼠Bsep蛋白表达量均低于A组,术后第3、5天均低于B组(均P<0.05);C组Mrp2表达均低于A、B组(均P<0.05)。肝细胞Bsep蛋白明显向细胞质移位。结论与正常肝脏相比,硬化肝对缺血耐受能力差,HIRI后硬化肝的胆汁淤积情况更加严重。Bsep、Mrp2表达降低以及Bsep蛋白由细胞膜向细胞质移位的改变在硬化肝HIRI后胆汁淤积中起了重要作用。

Objective To investigate the expression of Bsep and Mrp2 liver of cirrhotic rats with intrahepatic cholesta- sis after hepatic ischemia-reperfusion injury （HIRI）. Methods Forty five male sprague-dawley rats were randomly divided into 3 groups： normal rats with HIRI （group A）, liver cirrhosis rats with sham-operation （group B） and cirrhotic rats with HIRI （group C）. The ischemia time was maintained for 30 rain. The direct bilirubin （DBil） and total bilirubin （TBil） levels in bile and the DBil, TBil, AST, ALT level in serum were measured at dl, 3, 5 after operation. The expression of Bsep and Mrp2 proteins was detected by Western blotting and immunohistochemical method, Results The DBil, TBil levels in the bile of group C were significantly lower than those of group A at dl after operation（P〈0.05） and significantly lower than those of group A and B at d3（P〈0.05）. There was no significant difference among 3 groups at d5 after operation （P 〉0.05）. Serum ALT and AST levels in group C were significantly higher than those in group A and B at dl and d3 （P〈0.05）. Serum TBil and DBil levels in group C were significantly higher than those in group A and B at dl, d3 and d5 after surgery（P〈0.05）. The expression of Bsep in liver cells of group C were significantly lower than that in group A at dl, d3 and d5, and lower than that in group B at dl and d3. The expression of Mrp2 in group C was significantly lower than that in group A at dl, d3 and d5. Immunohistochemistry showed that Bsep protein was translocated to cy- toplasm of liver cells in group C. Conclusion Ischemic tolerance of cirrhotic liver is poor and intrahepatic cholestasis after HIRI in liver cirrhosis rats is more serious, which may be associated with the decreasied Bsep and Mrp2 protein expression and the ab- normal translocation of Bsep to cytoplasm of liver cells.