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目的检测和分析人乳头瘤病毒(HPV)16型E6E7癌基因对结肠癌细胞增殖状况和细胞周期的影响。方法运用含有HPV16E6E7癌基因的重组腺伴随病毒(rAAVE6E7)感染HCT116和SW480结肠癌细胞株,应用流式细胞术检测感染效率,Westernblot法检测HPV16E6癌蛋白的表达。MTT法和流式细胞技术分别检测感染前后结肠癌细胞的增殖状况和细胞周期。结果rAAVE6E7感染结肠癌细胞后24h开始可以检测到HPV16E6癌蛋白的表达,未感染的细胞不表达HPV16E6蛋白。rAAVE6E7感染后,HCT116细胞群体倍增时间缩短为对照组的68.42%,OD540值升高47.48%,G0/G1期细胞比例下降,S期细胞比例增高。SW480细胞群体倍增时间缩短为对照组的77.06%、OD540值升高47.18%。结论高危型HPV的E6E7癌基因能够促进结肠细胞的增殖、干扰其细胞周期,有可能在结直肠癌的发生、发展过程中发挥作用。
Objective To detect and analyze the effect of human papillomavirus type 16 E6E7 on the proliferation and cell cycle of colon cancer cells. Methods HCT116 and SW480 colon cancer cell lines were infected with recombinant adeno-associated virus (HPV16E6E7) containing HPV16E6E7 oncogene. The efficiency of infection was detected by flow cytometry. The expression of HPV16E6 oncoprotein was detected by Western blot. MTT assay and flow cytometry were used to detect the proliferation and cell cycle of colon cancer cells before and after infection. Results The expression of HPV16E6 oncoprotein was detected at 24 h after rAAVE6E7 infection in colon cancer cells. The non-infected cells did not express HPV16 E6 protein. After rAAVE6E7 infection, the population doubling time of HCT116 cells was shortened to 68.42% of the control group, the OD540 value was increased by 47.48%, the proportion of cells in G0 / G1 phase decreased and the proportion of S phase cells increased. SW480 cell population doubling time was shortened to 77.06% of the control group, OD540 value increased 47.18%. Conclusion The high-risk HPV E6E7 oncogene can promote the proliferation of colon cells and interfere with the cell cycle, which may play a role in the occurrence and development of colorectal cancer.