HCV感染人肝癌Huh7.5.1细胞后差异表达的lncRNA及其对细胞增殖的影响  

Differential Expression of Long Non-coding RNAs of Huh7.5.1 Cells After Infection with the Hepatitis C Virus and their Effects on Cell Proliferation

作　　者：付忠晓 李舒 向田[1] 谢焱 章晓联[1] 

机构地区：[1]武汉大学病毒学国家重点实验室/湖北省过敏及免疫相关疾病重点实验室/武汉大学医学研究院/武汉大学基础医学院免疫学系,武汉430071

出　　处：《病毒学报》2017年第2期192-199,共8页Chinese Journal of Virology

基　　金：国家自然科学基金(项目号:21572173);题目:以丙型肝炎病毒RNA-G4为新型靶标的配体分子的筛选及其抗病毒活性研究;国家自然科学基金(项目号:31370197);题目:基于适配子靶向病毒蛋白的策略研究HCV导致肝癌的机制与规律~~

摘　　要：丙型肝炎病毒(Hepatitis C virus,HCV)是导致人类肝脏疾病的重要病原体。长链非编码RNA(Long noncoding RNA,lncRNA)参与了许多疾病和生物过程的调控,但是lncRNA在HCV感染中的作用还了解很少。本研究旨在筛选HCV感染人肝癌细胞系Huh7.5.1后差异表达的长链非编码RNA(lncRNA),并研究相关lncRNA对细胞增殖及周期相关基因表达的影响。首先,分析HCV感染Huh7.5.1细胞72h前后lncRNA芯片表达谱;然后采用实时荧光定量PCR(Reverse tanscription-quantitative real time PCR,RT-qPCR)的方法对芯片中差异表达明显的17条lncRNA进行细胞水平验证;进一步采用CCK8以及ki67细胞增殖实验研究差异表达最明显的lncRNA对细胞增殖的影响;最后采用RT-qPCR探讨lncRNA对细胞周期蛋白基因cyclin B1/D1/E1的mRNA表达的影响。lncRNA芯片检测结果与RT-qPCR验证的相符的lncRNA中,发现HCV感染Huh7.5.1细胞后上调和下调最明显的两条lncRNA分别是RP11-288L9.1与ADAM20P1,而沉默RP11-288L9.1能抑制细胞周期蛋白基因cyclin B1/D1/E1的表达水平,并抑制Huh7.5.1细胞增殖。首次发现HCV感染后上调的RP11-288L9.1能促进细胞周期蛋白基因的表达水平和Huh7.5.1细胞增殖,另一种下调的ADAM20P1对细胞增殖影响不大。研究结果为HCV感染及肝癌细胞增殖提供了潜在的诊断标志物和治疗的新型靶点,具有一定的参考价值。We wished to study the differential expression of long non-coding RNAs （lncRNAs） in human Huh7.5.1 hepatoma cells after infection with the hepatitis C virus （HCV） and their effects on cell proliferation. Microarray analysis was used to analyze the differential expression of lncRNAs after Huh7.5.1 cells had been infected with HCV for 72 h. Reverse transcription-quantitative real-time PCR （RT-qPCR） was used to confirm the differential expression of IncRNAs. CCK8 and Ki67 assays were used to detect the effects of lneRNAs on cell proliferation, mRNA expression of the cell cycle-related genes cyclin B1/D1/E1 was analyzed by RT-qPCR. Microarray analysis and RT-qPCR confirmed that the mRNA expression of 17 lncRNAs in HCV-Huh7.5.1 cells was changed significantly. In particular, the transcription level of the lncRNA RP11-288L9.1 was increased the most, whereas that of ADAM20P1 was decreased the most, after HCV infection compared with those in untreated Huh7.5.1 cells. Silencing of RP11-288L9.1 could inhibit proliferation of Huh7.5.1 cells and expression of the cell cycle-related genes eyclin B1/D1/E1. Our results suggest that upregulation of expression of the lncRNA RP11-288L9.1 by HCV induces proliferation of Huh7.5.1 cells, whereas ADAM20P1 expression is downregulated by HCV and has no effect on cell proliferation. These findings show which lneRNAs profiles are associated with HCV infection and, as such, may offer new potent diagnostic/therapeutic targets for HCV infection and proliferation of hepatoma ceils.

关 键 词：丙型肝炎病毒(HCV) 长链非编码RNA 细胞增殖 细胞周期相关基因 

分 类 号：R373.2[医药卫生—病原生物学]