EXPRESSION OF CYTOCHROME-P450 AND MICROSOMAL EPOXIDE HYDROLASE IN CERVICAL AND ORAL EPITHELIAL-CELLS IMMORTALIZED BY HUMAN PAPILLOMAVIRUS TYPE-16 E6/E7 GENES
EXPRESSION OF CYTOCHROME-P450 AND MICROSOMAL EPOXIDE HYDROLASE IN CERVICAL AND ORAL EPITHELIAL-CELLS IMMORTALIZED BY HUMAN PAPILLOMAVIRUS TYPE-16 E6/E7 GENES
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DOI：
10.1093/carcin/16.6.1391
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发表时间：
1995-06-01
期刊：
影响因子：
4.7
通讯作者：
OMIECINSKI, CJ
中科院分区：
文献类型：
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作者：
FARIN, FM;BIGLER, LG;OMIECINSKI, CJ
流行病学证据表明，人乳头瘤病毒(HPV)的存在，与吸烟行为相结合，极大地增加了患口腔癌、宫颈癌、外阴癌和/或肛门癌的风险。细胞色素P450(CYP)、微体环氧化物水解酶(MeH)等生物转化酶是许多环境化学物质生物活化和解毒的重要调节因子，包括烟草烟雾中的某些亚硝胺和多环芳烃(PAH)。由于口腔和宫颈上皮细胞特异性表达这些基因的信息很少，我们分析了已建立的原代和HPV永生化口腔和宫颈上皮细胞系的形态、特异性Cyps和meh的mRNA和蛋白表达模式。用HPV-16E6/E7基因逆转录病毒感染原代人口腔和宫颈上皮细胞，使其永生化。将HPV-18基因导入原代培养的人角质形成细胞，使其永生化，并用亚硝基甲基脲处理后致瘤。在PAH诱导剂存在或不存在的情况下，通过逆转录耦合聚合酶链式反应分析，在所有细胞系中均可检测到meh基因的表达，而在任何细胞系中均未检测到CYP2A6mRNA，无论培养条件如何，在口腔上皮细胞系中表达最高，而在除HPV-18永生化角质形成细胞系外的所有其他上皮细胞系中均可检测到。口腔上皮细胞系和HPV-16永生化宫颈上皮细胞系与其他细胞系相比，细胞色素P450 2D6基因表达水平均升高，所有细胞均可检测到细胞色素P1A1和细胞色素P1A2基因的表达，部分细胞可被多环芳烃诱导。为了证实RT/PCR数据，我们对选定的样本进行了免疫印迹实验，使用这些方法，在原代和HPV永生化的口腔和宫颈上皮细胞中检测到CYP1A1和CYP2E1蛋白，这些数据表明原代和HPV永生化细胞似乎都表达某些生物转化酶，这些酶是激活烟草特有的亚硝胺和多环芳烃所必需的。虽然HPV基因感染对这些系统表达的整体影响仍未完全阐明，但体外系统的特征将被证明是有用的，在鳞状细胞癌的病因学中，HPV与异源激活的相互作用机制被证明是有用的。
Epidemiological evidence suggests that the presence of human papillomaviruses (HPV), when combined with smoking behaviors, considerably enhances the risk of developing oral, cervical, vulvar, and/or anal carcinomas. It is well established that the cytochrome P450 (CYP), microsomal epoxide hydrolase (mEH), and other biotransformation enzymes are important modulators of the bioactivation and detoxification of many environmental chemicals, including constituents of tobacco smoke such as certain nitrosamines and polycyclic aromatic hydrocarbons (PAH). Since there is little information regarding oral and cervical epithelial-specific expression of these genes, established primary and HPV-immortalized oral and cervical epithelial cell lines were analyzed for morphology, mRNA and protein expression patterns of specific CYPs and mEH. Primary human oral and cervical epithelial cells were immortalized using retroviral infection with HPV-16 E6/E7 genes. Primary human keratinocyte cells were immortalized by transfection of HPV-18 and made tumorigenic with nitrosomethylurea treatment. Expression profiles for mEH, CYP1A1, CYP1A2, CYP2A6, CYP2B6, CYP2D6, CYP3A, and CYP2E1 were evaluated in these cultures in the presence or absence of a PAH inducer, using reverse transcriptase-coupled polymerase chain reaction analysis, mEH gene expression was evident in all cultures, while CYP2A6 mRNA was not detected in any of the cell lines, regardless of culture conditions, CYP2E1 mRNA expression was greatest in the oral epithelial cultures and detectable in all other epithelial cultures except for the HPV-18 immortalized keratinocyte cell line. Elevated levels of CYP2D6 mRNA existed in both oral epithelial cell lines and the HPV-16 immortalized cervical epithelial cells when compared to the other cell lines examined, CYP1A1 and CYP1A2 mRNAs were detected in all the cells and several cultures were inducible by PAH exposure. To corroborate the RT/PCR data, Western immunoblotting experiments were conducted on selected samples, Using these methods, CYP1A1 and CYP2E1 proteins were detected in primary and HPV-immortalized oral and cervical epithelial cultures, These data indicate that both primary and HPV immortalized cells appear to express certain biotransformation enzymes necessary for the activation of tobacco-specific nitrosamines and PAHs. Although the overall impact of HPV gene infection on expression of these systems remains to be fully elucidated, an in vitro system is characterized which should prove useful in examining interactive mechanisms of HPV with xenobiotic activation in the etiology of squamous cell carcinomas.