HPV E2基因在鼻咽上皮细胞中的表达以及对细胞增殖的影响  

The expression and the regulation of HPV E2 gene in nasopharyngeal mucosa epithelial cells

作　　者：王甜甜[1] 王军[1] 肖洋[1] 

机构地区：[1]首都医科大学附属北京同仁医院,北京100005

出　　处：《临床耳鼻咽喉头颈外科杂志》2017年第3期211-214,共4页Journal of Clinical Otorhinolaryngology Head And Neck Surgery

摘　　要：目的:克隆人乳头状瘤病毒(HPV)11型E2基因片段,构建真核生物表达载体,研究低危型HPV E2基因的表达对正常鼻咽部黏膜细胞(NP69-SV40T)增殖的影响作用。方法:提取HPV11型DNA,设计E2基因引物,PCR扩增E2基因全长序列,分子克隆至含有绿色荧光蛋白的pEGFP-C1表达载体,构建真核表达载体pEGFP-C1-E2,转染至NP69-SV40T细胞内,转染后行Western blot、流式细胞仪检测以及CCK8检测,观察对NP69-SV40T细胞增殖活性的影响。结果:成功克隆和构建了pEGFP-C1-E2表达载体,转入NP69-SV40T细胞后,证明了其在NP69-SV40T细胞的表达,CCK8和流式细胞仪检测细胞周期,实验结果表明低危型HPV E2转染NP69-SV40T细胞可以促进细胞增殖,同时Western blot检测细胞周期素CDK2和CDK4发现,在3组(转染组、对照组和空白组)中表达量无统计学差异。结论:表达E2的pEGFP-C1-E2真核表达质粒能有效转染NP69-SV40T,并且转染后E2基因的表达能促进细胞的增殖。这些结果为深入研究复发性呼吸道乳头状瘤病的发生和发展提供了新的思路和方法。Objective：Through cloning the E2 gene of Human papillomavirus and constructing the eukaryotic expression vector, we can research the effect of HPV E2 gene expression on the cell proliferation of NP69. Method：By extacting the DNA of HPVll, designing the primers and amplificating the sequences of E2 gene, building the eukaryotic expression vector, we transfected it into the cells of NP69. Cultivate the transfected cells, and observe the cell proliferation by the western blot, flow cytometry and CCK8. Result： We cloned and constructed pEG- FP-C1-E2 expression vector successfully. After transfected, we certificated the expression of the E2 gene. Through detected by CCK8 and flow cytometry, we found that the E2 gene could promote the cell proliferation. By western blot, we found that the expression of CDK2 and CDK4 had no significant difference statistically. Conclusion.. These results laid the foundation for the further studying of the occurrence and development of recurrent respiratory papillomatosis.

关 键 词：复发性呼吸道乳头状瘤病 人乳头状瘤病毒 E2基因 细胞增殖 

分 类 号：R734[医药卫生—肿瘤]