【摘要】 目的探讨HPV16E6小干扰RNA(siRNA)与宫颈癌CaSki细胞中E6、p53、p21之间的关系。方法2004年9月至2005年3月于四川大学华西第二医院,应用化学合成针对HPV16E6的siRNA借脂质体转染CaSki细胞,实时荧光定量逆转录-聚合酶链反应(RT-PCR)和流式细胞术检测E6siRNA转染前后细胞中HPV16E6、p53、p21mRNA及其蛋白表达的变化。结果转染24h,E6mRNA的表达显著低于空白组(P<0.05)。各时间点p53、p21mRNA的表达差异无显著性意义(P>0.05)。转染48h,E6蛋白表达明显下调,p53、p21蛋白表达相应升高。结论HPV16E6siRNA能特异、高效地沉默宫颈癌细胞E6mRNA的表达,减少对野生型p53的降解,恢复p53蛋白的功能活性。RNA干扰(RNAi)技术可为HPV感染相关性疾病提供一种新的特异性基因治疗方法。更多还原

【Abstract】 Objective To investigate the relationship between HPV16 E6 small interfering RNA(siRNAs)and E6、p53、p21 in human cervical cancer cells line.Methods RNAi was induced using synthetic small interfering RNAs,which was transfected into CaSki cells line by lipofectamine.At various time points of post-transfection,the expression level of HPV16 E6,p53,p21 mRNA were detected by using real-time quantitative reverse transcription-polymerase chain reaction(real-time RT-PCR),and the level of these proteins were measured by FCM.Results At 24-hour post-transfection,the level of E6 mRNA was significantly decreased as compared with those in control(P<0.05).However,cellular p53 and p21 mRNA levels appeared unaffected at various time points of post-transfection(P>0.05).48 hours after transfection,the expression level of E6 protein was efficiently decreased,p53 and p21 protein level increased correspondingly.Conclusions It is identifided that HPV16 E6 siRNA could specially and efficiently silence E6 mRNA expression,decrease the degradation of wild type p53 protein,and then recover the function activity of p53 protein.RNA interference technology should provide a new rationale for specific gene-therapy approaches against HPV infection-related disease.更多还原