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华西医学  2012 

肿瘤坏死因子相关凋亡诱导配体对乙型肝炎病毒启动子的调控作用

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Keywords: 肿瘤坏死因子凋亡诱导配体,乙型肝炎病毒,自身启动子,相对荧光素酶活性

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Abstract:

目的 肿瘤坏死因子相关凋亡诱导配体(TRAIL)能够诱导乙型肝炎病毒(HBV)感染细胞发生凋亡,但抑制病毒复制的具体机制不清楚,研究通过非凋亡浓度TRAIL对4种HBV启动子调控作用的研究,探讨HBV复制的可能调控机制。方法 采用噻唑蓝法和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记荧光法,检测不同浓度TRAIL作用后人肝癌细胞株HepG2的存活率。使用HBV的4种启动子重组质粒,4种启动子分别控制乙肝表面抗原、X抗原、核心抗原、PS1抗原基因的转录与表达。将受HBV上述4种启动子调控的荧光素酶报道基因表达质粒(SpLUC、XpLUC、CpLUC、PS1pLUC)转染HepG2细胞,6h后按300、30、3ng/mL浓度梯度加入可溶性TRAIL,采用双荧光报道基因分析系统检测细胞化学发光值,计算相对荧光素酶活性。结果 300ng/mL是可溶性TRAIL诱导HepG2细胞凋亡的浓度阈值。采用远低于凋亡阈值浓度(30ng/mL)的TRAIL可明显上调对HBV的Sp启动子活性(P<0.001),另3种质粒的相对荧光素酶活性在加入TRAIL后改变不大。结论 TRAIL仅对HBV的Sp启动子活性具有上调作用,其生物学意义值得进一步研究。

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