乳腺癌发展过程中的bcl-2甲基化及其与蛋白表达的关系

目的建立bcl-2基因甲基化特异的PCR（MSP）检测方法，检测乳腺癌发展过程中的bcl-2甲基化．并探讨bcl-2甲基化与蛋白表达的关系。方法设计bcl-2基因MSP引物，采用MSP方法检测30例正常、25例不典型增生（癌前病变）、40例腋窝淋巴结阴性（癌症初期）和45例腋窝淋巴结阳性（癌症后期）乳腺组织的bcl-2基因5’端启动子CpG岛甲基化状态。另外采用免疫组织化学S-P法检测bcl-2的蛋白表达。结果正常、不典型增生、淋巴结阴性和淋巴结阳性乳腺组织的bcl-2甲基化率分别为10．0％、32．0％、40．0％和53．3％。乳腺癌发展过程中的bcl-2甲基化率渐增（P〈0．01），bcl-2蛋白表达率渐减（P〈0．01）。不典型增生较正常乳腺组织的bcl-2甲基化水平显著提高（P〈0．05）。在乳腺癌发展过程中的4个阶段，bcl-2甲基化与蛋白表达两者之间均呈显著负相关性（P〈0．01）。结论该研究建立了bcl-2基因MSP检测方法，bcl-2基因MSP引物的设计是合理的。bcl-2启动子甲基化可能成为乳腺癌前病变的分子指标之一。在乳腺癌发展过程中，bcl-25’端调控区CpG岛甲基化可能是下调bcl-2表达的因素。

BCL-2 METHYLATION IN THE DEVELOPMENT OF BREAST CANCER AND THE RELATIONSHIP BETWEEN BCL-2 METHYLATION AND EXPRESSION

Objective To establish MSP (methylation-specific PCR) method for detecting bcl-2 gene, to study bcl-2 methylation in the development of breast cancer and the relationship between bcl-2 methylation and expression. Methods Designing the MSP primer of bcl-2 gene, the methylations in CpG island of bcl-2 gene 5'-regulatory region in 30 cases of normal, 25 cases of atypical hyperplasia(precancerous condition), 40 cases of lymphnode(-)(low-stage) and 45 cases of lymphnode( )(high-stage) breast tissue were detected by using MSP. The expressions of bcl-2 were detected by using SP immunohistochemical technique. Results The positive rates of bcl-2 methylation in normal, atypical hyperplasia, lymphnode(-) and lymphnode( ) breast tissue were 10.0%,32.0%,40.0% and 53.3% respectively. The methylation of bcl-2 was increased and the expression of bcl-2 was decreased in the development of breast cancer(P<0.01). The positive rate of bcl-2 methylation in atypical hyperplasia was significantly higher than that in norrnal(P<0.05). There was always a significant negative correlation between the bcl-2 methylation and the bcl-2 expression in the above four developing stages of breast cancer(P<0.01). Conclusion This study established MSP method for detecting bcl-2 gene. The design of the MSP primer of bcl-2 gene in this study was successful. The positive rate of methylation of bcl-2 promoter would become a molecular indicator of breast carcinogenesis. In the development of breast cancer, the methylation in CpG island of bcl-2 gene 5'-regulatory region could be a down-regulation factor of the expression.