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人乳头瘤病毒(human papillomavirus,HPV)是一种无包膜的环状闭合双链DNA病毒,具有严格的嗜人组织的特性.为探讨不同启动子驱动HPVE6/E7癌蛋白在裸鼠体内不同组织的表达效率,构建了带有角蛋白(K14)启动子和带有巨细胞病毒(CMV)启动子的E6/E7腺病毒载体(pAd-K14-E6/E7和pAd-CMV-E6/E7),pAd-K14-E6/E7和pAd-CMV-E6/E7、以及作为对照的重组腺病毒空载体pAdtrack-K14和pAd-CMV同源重组后,分别在293细胞中包装,收集重组病毒Ad-K14-E6/E7、Ad-CMV-E6/E7、Adtrack-K14和Ad-CMV,通过尾缘静脉注射到随机分组的裸鼠体内,并每d向裸鼠腹腔注射0.05 mg雌激素.采用RT-PCR和Western免疫印迹检测不同实验组E6/E7 mRNA和蛋白表达水平,免疫组化法检测P53和Bcl-2蛋白表达.结果显示,注射病毒Ad-K14-E6/E7(实验组1)裸鼠子宫体中E6/E7 mRNA、E6蛋白质、P53和Bcl-2蛋白高表达,而其它组织中低表达;注射病毒Ad-CMV-E6/E7(实验组2)裸鼠各组织E6/E7 mRNA、E6蛋白质、P53和Bcl-2蛋白均低表达.研究表明,在裸鼠体内角蛋白K14启动子可以调控E6/E7在子宫体中表达,CMV启动子未能诱导E6/E7在子宫体中表达.
Human papillomavirus (HPV) is a non-enveloped, circular, closed, double-stranded DNA virus that has stringent human-tissue properties.In order to investigate the effect of different promoters on HPVE6 / E7 oncoprotein in nude mice E6 / E7 and pAd-CMV-E6 / E7 were constructed with the keratin (K14) promoter and the E6 / E7 adenoviral vector carrying the cytomegalovirus (CMV) promoter ), pAd-K14-E6 / E7 and pAd-CMV-E6 / E7, and the control recombinant adenoviral empty vector pAdtrack-K14 and pAd-CMV were homologously recombined and then packaged in 293 cells respectively. Recombinant virus Ad K14-E6 / E7, Ad-CMV-E6 / E7, Adtrack-K14 and Ad-CMV were injected via tail vein into randomly assigned nude mice and peritoneal injections of 0.05 mg of estrogen every day The expression of E6 / E7 mRNA and protein in different experimental groups was detected by RT-PCR and Western blotting, and the expressions of P53 and Bcl-2 protein were detected by immunohistochemistry.The results showed that the injected virus Ad-K14-E6 / E7 (experimental group 1) E6 / E7 mRNA, E6 protein, P53 and Bcl-2 protein were highly expressed in the uterus of nude mice, but not in other tissues. The expression of E6 / E7 mRNA in E6 / E7 mRNA, E6 protein, P53 and Bcl-2 protein were low expression.Studies showed that keratin K14 promoter can regulate the expression of E6 / E7 in uterus in nude mice, and CMV promoter failed to induce E6 / E7 in uterine In the expression.