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目的:观察叶下珠复方Ⅱ号抑制二乙基亚硝胺(DEN)诱导的大鼠肝癌形成效果,并探讨其作用机制。方法:将240只SPF级雄性Wistar大鼠随机分为正常组,模型组,叶下珠复方Ⅱ号高、中、低剂量(23.14,11.57 g·kg-1)组,喃氟啶组,采用DEN诱导肝癌大鼠模型,造模18周,同时给药进行干预,每6周各组处死大鼠8只,观察大鼠一般情况,肝功能指标变化和肝癌结节形成情况,镜下观察大鼠肝组织病理变化,放免法检测大鼠血清白细胞介素-6(IL-6)含量;采用实时定量PCR(qRTPCR)法、免疫印迹法(Western blot)分别检测肝组织IL-6信号通路及microRNA let-7a调控网络基因的mRNA和蛋白表达改变。结果:实验第18周末,叶下珠复方Ⅱ号高、低剂量组大鼠肝癌结节数较模型组明显减少(P<0.05);叶下珠复方Ⅱ号高、低剂量组大鼠血清IL-6,谷丙转氨酶(ALT),谷草转氨酶(AST)水平与模型组比较均明显降低(P<0.05);与模型组比较,叶下珠复方Ⅱ号高剂量组大鼠肝组织microRNA let-7a表达量明显上升,转录因子蛋白家族(NF-κB-p65),IL-6,信号转导与转录激活因子3(STAT3),Harvery鼠肉瘤病毒Ras基因(Ras)和原癌基因(C-myc)mRNA表达量均明显降低(P<0.05)。实验第18周,叶下珠复方Ⅱ号高、低剂量组STAT3和p-STAT3蛋白表达均明显低于模型组(P<0.05)。结论:叶下珠复方Ⅱ号对DEN诱导的大鼠肝癌形成具有明显的抑制作用,作用机制与上调microRNA let-7a表达和下调NF-κB-p65的表达,从而抑制IL-6的表达和IL-6/STAT3信号通路活化有关。
OBJECTIVE: To observe the inhibitory effect of Phyllanthus Compound Ⅱ on rat hepatocarcinoma induced by diethylnitrosamine (DEN) and its mechanism. Methods: 240 SPF male Wistar rats were randomly divided into normal group, model group, Phyllanthus Compound Ⅱ high, medium and low dose (23.14,11.57 g · kg-1) group, DEN induced hepatocellular carcinoma rat model, model 18 weeks, at the same time to interfere with the intervention, every 6 weeks, 8 rats were killed in each group, to observe the general situation of rats, liver function indicators and liver cancer nodules, microscopic observation The levels of IL-6 in serum of rats were detected by radioimmunoassay. The expression of IL-6 in liver was detected by qRTPCR and Western blotting respectively. MicroRNA let-7a regulates network mRNA and protein expression changes. Results: At the end of the 18th week, the number of HCC in the Phyllanthus Compound II high and low dose groups was significantly lower than that in the model group (P <0.05). The serum IL (P <0.05) .Compared with the model group, the expression of microRNA let-6 in the Yexiazhu Compound II high-dose group was significantly lower than that in the model group The expressions of NF-κB-p65, IL-6, STAT3, Ras and C- myc) mRNA were significantly decreased (P <0.05). At the 18th week of experiment, the expression of STAT3 and p-STAT3 protein in Phyllanthus Compound II high and low dose groups were significantly lower than those in model group (P <0.05). Conclusion: Ye Xia Zhu Fu Fang Ⅱ DEN-induced rat liver cancer significantly inhibited the mechanism of action and upregulation of microRNA let-7a expression and down-regulated the expression of NF-κB-p65, thereby inhibiting the expression of IL-6 and IL -6 / STAT3 signaling pathway activation.