16型人乳头瘤病毒E5基因对永生化口腔上皮细胞EGFR、p21、p53和Rb基因表达的影响

Effects of human papillomavirus E5 gene on EGFR,p21,p53 and Rb gene expression in immortalized oral epithelial cells

目的分析16型人乳头瘤病毒E5基因(HPV16 E5)对永生化口腔上皮细胞(HIOEC)肿瘤相关基因表皮生长因子受体(EGFR)、p21、p53和Rb表达的影响,探究HPV16 E5在口腔鳞状细胞癌发生发展过程中的潜在作用机制。方法将含有目的基因HPV16 E5的慢病毒载体p LOV-E5及对照病毒载体p LOV-N分别转染HIOEC细胞,并筛选出各自稳定转染的细胞株。RT-q PCR和Western blot法分别检测EGFR、p21、p53和Rb基因mRNA和蛋白表达水平的变化。MTT法分析HPV16 E5对HIOEC细胞增殖活性的影响。结果p LOV-E5转染的HIOEC细胞中,p21和p53的mRNA转录水平较对照组显著提高(P=0.00、0.02),但EGFR的mRNA转录水平较对照组无明显变化;EGFR、p21和p53的蛋白表达水平较对照组均显著提高,Rb基因的mRNA和蛋白表达水平较对照组均无明显变化。MTT法检测结果表明,p LOV-E5转染HIOEC的增殖活力较对照组显著提高(P=0.00)。结论 HPV16 E5可以促进HIOEC的增殖,同时上调EGFR、p21和p53在HIOEC中的表达水平。

Objective To investigate the effects of HPV16 E5 on the expression of EGFR,p21 ,p53 and Rb in thehuman immortalized oral epithelial cell（ HIOEC）, and to discuss the potential mechanism of HPV16 E5 in the oc- currence and development of oral squamous cell carcinoma. Methods Open reading frame of HPV16 E5 was cloned into the lentiviral vector pLOV-N to prepare recombinant plasmid pLOV-E5, which was stably transfected in- to HIOEC. EGFR,p21 ,p53 and Rb were detected by RT-qPCR and Western blot on the mRNA level and the pro- tein level, respectively. Cell proliferation after stable trasfection was evaluated by MTT assay. Results After stable trasfection of HPV16 E5 ,the protein levels of EGFR,p21 and p53 in HIOEC were markedly overcxpressed, as well as the mRNA levels of p21 and p53 （P = 0. 00,0. 02）. However, the mRNA levels of EGFR had no obvious change after trasfection. There were also no significant changes of Rb on the mRNA and protein levels. Compared with the control cells, the proliferation activity of HIOEC/pLOV-E5 cells was significantly increased （ P = 0. 00 ）. Conclu- sion HPV16 E5 can stimulate the expression of EGFR,p21 and p53 as well as the proliferation of HIOEC.