用RT-PCR和免疫印迹的方法检测稳定表达NS4B的HeLa细胞中的XBP1;通过RT-PCR的方法在表达NS4B的HeLa和Huh-7细胞中检测ATF6,Grp78和caspase-12的转录,并且通过报告基因的方法分析XBP1和Grp78启动子活性。实验结果表明:在表达NS4B的HeLa细胞中检测到XBP1的两种形式(剪接和未剪接),此外,在细胞中ATF6、Grp78的转录水平和XBP1、Grp78启动子的荧光素酶活性较没有表达NS4B的HeLa和Huh-7细胞中的量有所增加;通过染色质免疫沉淀实验(ChIP)分析,这些增加可能是由于XBP1结合到了这些基因的启动子上引起的。总之,实验结果可提示HCVNS4B通过ATF6或XBP1途径引起内质网压力,导致UPR反应。NS4B可能在HCV的致病性中起着重要的作用,特别是在慢性肝炎,甚至肝细胞癌中。

The unspliced and spliced forms of XBP1 stably expressing NS4B in HeLa cells, the transcriptional levels of ATF6, Grp78 and caspase-12, and the luciferase activity in XBP1 and Grp78 promoter reporter assays in HeLa and Huh-7 cells expressing NS4B were detected. The results showed that the two forms of XBP1 were detected NS4B in HeLa cells, moreover, ATF6 and Grp78 transcription levels and the luciferase activity in XBP1 and Grp78 promoter assays in cells expressing NS4B increased compared with that of cells without NS4B expression due to XBP1 binding to their promoter sites. Collectively, the results imply the possibility that NS4B induces UPR through ATF6 or IRE1-XBP1 pathways upon ER stress, and maybe play some roles in HCV pathogenesis, in particular, in chronic hepatitis, even hepatocellular carcinoma.