p16^(INK4A)与HPV联合检测宫颈病变预后关系的实验研究

Experimental study of the relationship between jointly detecting p16^(INK4A) and HPV in the prognosis of cervical cancer

目的:探讨探讨p16基因启动子甲基化在宫颈癌发展中的作用及其与临床-病理因素及HPV感染的关系。方法:采用MSP的方法的p16甲基化测定和导流杂交基因芯片技术的HPV分型测定实验。结果:在正常宫颈组织、CIN和宫颈鳞癌中p16基因启动子甲基化的表达有统计学意义(P<0101);p16甲基化在低分化的病理类型G3(8/15)与G1+G2(8/45)比较有统计学意义(P=0.01)。HR-HPV感染随着病变级别增加阳性率增高,与正常宫颈比较有差异有显著性意义(P<0.05),HPV16/58的感染占所有HR-HPV的50%。p16在三组不同HPV亚型的甲基化阳性率无显著性差别(P>0.05)。结论:①p16基因启动子的甲基化发生在宫颈癌变的早期,随着宫颈癌变级别进展而增加,p16的异常甲基化有可能成为预测宫颈癌变级别进展的生物学指标之一,有助于辅助宫颈癌的早期诊断。②p16基因启动子甲基化与宫颈癌的临床病理特征有关联,甲基化更多发生在年龄小于40岁、病理低分化的宫颈鳞癌患者;p16基因甲基化的出现有可能促使宫颈癌恶性进展。③p16基因启动子甲基化在HR-HPV的发生率较LR或无HPV感染高,但无统计学意义。提示p16甲基化的发生是独立于HPV感染的事件,它们在宫颈癌的致病过程中有可能是选择性的作用于不同的时期。

Objective： To evaluate promoter hypermethylation of p16 in the preinvasive cervical epithelial lesions and invasive cervical carcinomas by MSP-PCR in order to determine whether hypermethylation of p16 might be associated with tumor progression.It is also focued on whether hypermethylation of p16 might be a clinically useful marker of lesions progress to invasive disease in which the correlation between HPV infection and hypermethylation of p16 gene is included in this study. Methods： MSP-PCR methods was used to examine promoter hypermethylation of p16 gene in cervical lensions-ICC,CINI,CINII-III,and normal cervix specimens and flow-through hybridization gene chip technology were used to examine HPV types in cervical lensions. Results： Promoter hypermethylation of p16 gene were absent in all normal cervical specimens in which there was a significant trend increasing hypermethylation from normal cervix or CINI to ICC（P0.05）. Also,hypermethylation of p16 gene were more frequent both in CINII-III and ICC than that in CINI（P0.05）. The difference between CINII-III and ICC was no significance. Promoter hypermethylation of p16 gene had significant differences in ages and histopathology which was higher in no more than 40 ages and in poorly differenciated grade（P0.05）. Moreover,hypermethylation were totally positive in cervical squamous carcinomas（SCC） and negative in adenocarcinomas（AC）. There were no significant differences in the frequencies of hypermethylation between clinical stage I and stage II,and so on lymph nodes metastases. HR-HPV infection had been detected negative in normal cervical samples,and HPV16/58 was the predominant types（50%） of HR-HPV infection in CIN and ICC. Hypermethylation of p16 gene were found in 25% of HPV16/18 positive cases,16.7% of the other HR-HPV cases,and 10.7% of LR-HPV or HPV negative cases. There were no significance in three groups（P0.05）. Conclusion： ①Promter hypermethylation of p16 gene was an early event in progression of cervical cancer which may be useful as cancer-specific marker. ②The inactivation of p16 hypermethylation may be responsible for cervical cancer malignant development. ③There was no association between hypermethylation of p16 gene and HPV infection.It suggests that they may be a selected event with time during multistage pathogenesis of cervical cancer.