HCV感染通过Erk信号的激活上调PTTG1的表达

HCV Infection Up-regulates PTTG1 Expression via MAPK/Erk Pathway

为探讨丙型肝炎病毒(HCV)感染导致肝细胞癌发生的分子机制,利用前期研究建立的体外HCV细胞培养体系,将HCVJFH-1RNA转染CD81-Huh7细胞,确定能够获得感染性HCV后,收集HCV转染后10,50,100和150d的细胞,采用RealtimePCR及WesternBlot法检测不同时间点细胞内垂体瘤转化基因1(PTTG1)基因和蛋白水平的表达情况,同时检测总MAPK/Erk1/2,磷酸化Erk1/2(p-Erk1/2)蛋白的表达。随后,用干扰素抑制HCV的感染,再检测PTTG1及MAPK/Erk1/2的表达情况,以及用MAPK/Erk抑制剂(PD98059)阻断MAPK/Erk1/2的磷酸化后,检测PTTG1的表达情况。结果显示,HCV转染的细胞与未转染细胞比较,PTTG1的mRNA和蛋白表达均显著增加,p-Erk1/2蛋白显著升高(P<0.05);抑制HCV感染显著降低PTTG1的表达和MAPK/Erk1/2的磷酸化(P<0.05);MAPK/Erk1/2抑制剂显著降低了PTTG1基因和蛋白的表达(P<0.05)。体外HCV感染可以导致MAPK/Erk信号通路的磷酸化,进一步导致原癌基因PTTG1的表达增加,这可能是慢性HCV感染导致HCV相关性肝细胞癌发生的分子机制之一。

To investigate the possible molecular pathogenesis of hepatocellular carcinoma （HCC） induced by HCV infection, CD81-Huh7 cell was transfected by HCV-JFH1 RNA as described in our previous studies. Cells were collected 10, 50, 100 and 150 days after the HCV RNA transfection and were passaged every 3-4 days. PTTG1 mRNA and protein expressions were detected by Real time PCR and Western Blot. The phosphorylated Erk kinase （p-Erk） and the total Erk1/2 protein level were measured by Western blot. MAPK/Erk inhibitor （PD98059） was applied to block the phosphorylation of Erk1/2. HCV was inhibited by IFN. It is found that the levels of PTTG1 mRNA and protein are significantly increased at above time point in HCV transfected cells as compared to the untransfected cells. Parallel with PTTG1, the p-Erk is significantly increased in the transfected groups as compared with the untransfected groups. The inhibition of HCV has significantly decreased the expression of PTTG1 and the phosphorylation of MAPK/Erk1/2; MAPK/Erk1/2 inhibitor has significantly decreased the PTTG1 expression. The HCV infection activates MAPK/Erk1/2 pathway and leads to the increase of oncogenic PTTG1 over-expression, which may be one of the pathogenesis of the HCV related HCC development.