肝复康经JNK信号通路在肝星状细胞凋亡中的调节机制

摘要

Objective To explore the regulation mechanisms of Ganfukang (GFK) in hepatic stellate cells (HSC) apoptosis through JNK signaling pathways.Methods All the SD rats were randomly divided into 5 groups:normal control group (group C),model group (group M) and high,middle and low dose treatment groups (group HT,group MT and group LT).HSC-T6 cell line was randomly divided into three groups:control group,acetaldehyde group and GFK group.The pathological changes of hepatic tissues were observed under light microscope by HE staining.The mRNA expression of MKK4,MKK7,JNK1,JNK2,COX-2,Survivin,Caspase-3,α-SMA,typeⅠcollagen and typeⅢcollagen was measured by RT-PCR,and the hepatic expression of COX-2 was detected by immunohistochemistry and western blot.Results The COX-2 was rarely expressed in group C.The expression of COX-2 protein in group M increased more significantly than that of group C (P<0.05).Compared with group M,the expression of COX-2 protein decreased in group MT (P<0.05).The expression levels of main kinases in JNK pathway (MKK4,MKK7,JNK1 and JNK2) and Survivin in group M were higher than those of group C (P<0.05),and they were lower in GFK treatment groups than those of group M (P<0.05),while the tendency of expression levels in Caspase-3 mRNA was contrary.The expression of COX-2 was significantly positively correlated with Survivin,and negatively correlated with Caspase-3.The expression of colleganⅠ,collagenⅢ and α-SMA mRNA in group MT was lower than that of group M (P<0.05).Conclusion GFK has therapeutic effect on liver fibrosis by inducing the apoptosis of HSC through inhibiting the transduction of JNK signal pathway in liver fibrosis.%目的 探讨中药肝复康经JNK(c-Jun N-terminal kinase,JNK)信号通路在肝星状细胞凋亡中的调节机制.方法 将SD大鼠随机分为5组:正常对照组(C),模型组(M),肝复康高(HT)、中(MT)、低(LT)剂量治疗组,每组给予相应的处理;HSC-T6细胞株随机分为3组:正常对照组、乙醛组及肝复康治疗组.HE染色法观察肝组织形态结构变化;RT-PCR法测定MKK4、MKK7、JNK1、JNK2、COX-2、Survivin、Caspase-3、α-SMA、collagenⅠ和collagenⅢ的基因表达水平;免疫组织化学法和免疫蛋白印记法观察COX-2在肝组织和肝星状细胞中的表达.结果 正常组中COX-2几乎不表达.与正常组相比,模型组中COX-2表达显著增加(P<0.05).与模型组相比,肝复康治疗组各组中COX-2表达均明显降低,其中,中剂量组最明显(P<0.05).与正常组相比,模型组中MKK4、MKK7、JNK1、JNK2、COX-2、Survivin的基因表达水平均显著上调(P<0.05).与模型组相比,肝复康治疗组各基因表达水平均明显减弱,中剂量组最明显(P<0.05),其中Caspase-3基因表达呈相反趋势.COX-2和Survivin的表达呈正相关,和Caspase-3的表达呈负相关.检测α-SMA、collagenⅠ与collagenⅢ基因表达,与模型组相比,中剂量治疗组表达水平明显下调(P<0.05).结论 肝复康可通过抑制JNK信号通路的传导诱导肝星状细胞凋亡,发挥抗纤维化作用.