HBV通过下调PTEN的表达拮抗IFN-α抗病毒活性

HBV antagonizes the antiviral activity of IFN-αby down-regulating the expression of PTEN

目的 探讨乙型肝炎病毒(HBV)感染所致的第10号染色体缺失的磷酸酶及张力蛋白同源的基因(PTEN)下调与α-干扰素(IFN-α)抗病毒活性的相关机制。方法 在适宜条件下培养HepG2细胞和HepG2.2.15细胞,以空白载体(pcDNA3.1)和HBV1.3质粒分别转染HepG2细胞,Western blot法检测PTEN蛋白质的表达;将pcDNA3.1和PTEN过表达(PTEN-OE)质粒分别瞬时转染HepG2.2.15细胞,化学发光法分析细胞培养上清液中HBV相关抗原的表达,实时荧光定量PCR(qRT-PCR)技术分析HBV前基因组RNA(HBV pgRNA)表达;使用合成RNA双链体poly(I∶C)刺激PTEN-OE的细胞,qRT-PCR技术分析IFN-αmRNA的表达,Western blot法分析JAK/STAT信号通路中干扰素调节因子9(IRF9)、黏病毒抗性蛋白1(MxA)的表达。结果 瞬时表达HBV的HepG2细胞和稳定表达HBV的HepG2.2.15细胞中,PTEN蛋白的表达降低;PTEN-OE的HepG2.2.15细胞中,HBV相关抗原及HBV pgRNA的表达较对照组降低,经poly(I∶C)作用后,IFN-αmRNA水平较对照组显著升高,且JAK/STAT信号通路相关蛋白IRF9、MxA的表达增加。结论 HBV可能通过下调PTEN的表达发挥拮抗IFN-α抗病毒活性的作用。

Objective To investigate the mechanism of down-regulation of the phosphatase and tensin homolog deleted on chromosome ten(PTEN)caused by hepatitis B virus(HBV)infection and the antiviral activity of interferonα(IFN-α).Methods HepG2 cells and HepG2.2.15 cells were cultured under suitable conditions.HepG2 cells were transfected with empty vector(pcDNA3.1)and HBV1.3 plasmid respectively,and the protein expression of PTEN was detected by Western blot;pcDNA3.1 and PTEN over-expression(PTEN-OE)plasmid were transfected into HepG2.2.15 cells respectively.Chemiluminescence was used to analyze the expression of HBV-related antigens in the cell culture supernatant,and real-time quantitative PCR(qRT-PCR)was used to analyze the expression of HBV pregenomic RNA(HBV pgRNA);the synthetic RNA duplex[poly(I∶C)]was used to stimulate PTEN-OE cells,qRT-PCR was used to analyze IFN-αmRNA expression and Western blot was used to analyze the expression of interferon regulatory factor 9(IRF9)protein as well as myxovirus resistance protein 1(MxA)protein in the JAK/STAT signaling pathway.Results In HepG2 cells expressing HBV transiently and HepG2.2.15 cells stably expressing HBV,the expression of PTEN protein both decreased;the expression of HBV-related antigens and HBV pgRNA decreased in PTEN-OE HepG2.2.15 cells compared with the control group.After the treatment by poly(I∶C),the level of IFN-αmRNA was significantly higher than that of the control group,and the expression of IRF9 and MxA ptotein related to the JAK/STAT signaling pathway both increased.Conclusion HBV may play a role in antagonizing the antiviral activity of IFN-αby down-regulating the expression of PTEN.