宫颈病变程度与人乳头瘤病毒16型甲基化水平的关系研究

目的:探讨检测人乳头瘤病毒16型(HPV16)基因组甲基化水平判断宫颈病变阶段的可行性。方法:在行宫颈癌筛查的妇女中收集73例HPV16阳性标本,采用重硫酸盐-焦磷酸测序方法对HPV16型基因组L1基因开放阅读框(ORF)以及上游调控区(URR)的甲基化位点进行分析,甲基化结果与病理学检查结果进行比对分析,分析HPV16型甲基化与宫颈病变的关系。结果:HPV16基因的31,37,43,52和58位点在正常标本中存在中度甲基化,宫颈上皮内瘤变1度(CIN1)～CIN3标本存在低度甲基化,而宫颈癌(CA)标本表现为高度甲基化。正常标本、CIN标本以及CA标本各位点甲基化水平两两比较差异均有统计学意义(P均<0.05)。在CIN1,CIN2以及CIN3间甲基化水平差异无统计学意义;HPV16基因的3887,3927,3941和5602位点甲基化水平随着宫颈病变程度的加深而升高,并且正常标本、CIN、CA间两两比较均存在差异(P均<0.05);5608,5709,5611,5617,5762,6367,6389位点甲基化水平在正常标本、CIN1、CIN2和CIN3标本间无统计学差异。正常标本与CA标本、CIN标本与CA标本间比较,甲基化水平差异有统计学意义(P均<0.05)。结论:HPV16型基因组的甲基化与宫颈病变程度存在相关性,临床上通过检测HPV16型DNA的甲基化可对宫颈病变进行辅助诊断。

The relationship between grade of cervical lesions and HPV16 DNA methylation

Objective: To construct a new biomarker with which could evaluate the potential detection of HPV16 infection by explore the relationship between the grade of cervical lesions and HPV16 DNA methylation. Methods:Seventy-three HPV16 postive samples were collected from women undergoing routine cervical cancer screening. Complete methylation data were obtained for L1 gene ORF and URR CpGs in all samples by bisulfite-pyrosequencing. The methylation result was compared with that of pathological examination to analysis the relationship the grade of cervical lesions and HPV16 DNA methylation. Results:In sites of 31, 37, 43, 52 and 58, there was a moderate methylation level in no lesion samples, but with a low methylation level cervical intraepithelial neoplasia(CIN) samples and a high methylation level in Cervical cancer(CA) samples. The statistic analysis showed a significant difference among the normal and CIN and CA samples. There were significant differences among normal, CIN1, CIN2, CIN3 and CA samples in sites of 3887, 3927, 3941 and 5602. Besides, a significant difference existed between normal and CA sample or between CIN and CA samples in sites of 5608, 5709, 5611, 5617, 5762, 6367 and 6389. Conclusion: HPV16 DNA methylation level is associated with the grade of cervical lesions. And HPV16 DNA methylation level in clinical samples could be a biomarker of neoplastic cervical progression.