咖啡因和解热镇痛抗炎类药物阿司匹林对HERV-W env基因表达的影响

摘要

目的 探讨精神活性类药物咖啡因和解热镇痛抗炎类药物阿司匹林对人内源性逆转录病毒W(HERV-W)env基因表达的影响.方法mTT法检测不同浓度的药物作用人神经母细胞瘤细胞系SH-SY5Y后,产生的毒性作用和对其增殖能力的影响.应用半定量RT-PCR检测不同浓度的药物作用SH-SY5Y后,对HERV-W env基因表达的影响.应用荧光定量PCR进一步检测不同浓度的药物作用SH-SY5Y后,对HERV-W env基因表达的影响.不同浓度的药物作用SH-SY5Y后,扩增并克隆HERV-W env基因,并进行测序和序列分析.将pGL3-env promoter重组质粒转染SH-SY5Y后,通过检测萤光素酶活性的变化,分析咖啡因和阿司匹林对env基因启动子活性的影响.结果咖啡因和阿司匹林对SH-SY5Y细胞系都有明显的增殖抑制作用以及毒性作用,并且这种增殖抑制作用和毒性作用呈时间和剂量依赖性.咖啡因和阿司匹林对env基因的表达都有上调作用,并且这种上调作用呈剂量依赖性.与未加药组相比,用低浓度的咖啡因(0.05mM/L)和高浓度的咖啡因(5mM/L)作用SH-SY5Y后,扩增获得的env基因序列中都检测到了有突变的序列;同样,用低浓度的阿司匹林(0.5mM/L)和高浓度的阿司匹林(5mM/L)作用SH-SY5Y后,扩增获得的env基因序列中也检测到了有突变的序列.咖啡因能增强env基因启动子的活性,而阿司匹林对env基因启动子的活性没有显著的影响.结论 环境因素能调节HERV-W家族env基因的表达,env基因的表达调控机制是一个非常复杂的体系.%Objective To explored the influence of caffeine,a kind of psychoactive drug,and aspirin,a sort of antipyretic analgesics,on transcription of env gene.Methods Different concentrations of drug mediated cytotoxicity were detected by MTT.The expression of HERV-W env gene was detected by semi-quantitative reverse-transcription polymerase chain reaction(RT-PCR)methods.The expression of HERV-W env gene was further detected by quantitative real-time PCR methods.The detected sequences were cloned,sequenced and analysised.The regu-lation of env promoter after drug treatment was detected by the Luciferase Assay System.Results Cell proliferation was inhibited after caffeine and aspirin treatment in a dose-dependent manner.The expression of HERV-W env gene was increased after caffeine treatment in SH-SY5Y cell-line.For aspirin treatment,the expression of env gene was increased,either.After caffeine treatment at different concentration,with low concen-tration(0.05 mM/L)and high concentration(5 mM/L),HERV-W env gene was cloned and sequenced.Mutation was found in HERV-W env gene.After aspirin treatment,mutation was found at the concentration of 0.5mM/L and 5 mM/L.Interestingly,no mutation was found in SH-SY5Y cell-line without any drug treatment.The activity of env promoter was increased after caffeine treatment,but remained the same after as-pirin treatment.Conclusion HERV-W env gene can be modulated by environmental influences.The mechanisms behind the regulation of ex-pression of HERV-W env gene are very complex.