【摘要】 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。更多还原

【Abstract】 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.更多还原