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目的:微小RNA(microRNA)异常在肿瘤发生发展中起重要作用,本研究探讨microRNA mir219在肝细胞癌中的异常甲基化和异常表达,可能参与肝细胞癌的发生发展过程。方法:采用pyrosequence、TA克隆测序等方法检测正常肝组织、肝细胞癌细胞株和肝癌标本中mir219上游预测启动子区域甲基化,Taqman实时定量RT-PCR测定肿瘤组织表达。结果:与正常肝脏组织相比,肝癌细胞株、肝癌标本中mir219上游预测启动子区域中茎环区和上游1kb区域的CpG岛存在高度甲基化。肝癌细胞株、肝癌标本中mir219表达显著下调,与其异常甲基化相关。结论:肝癌中微小RNA mir219茎环序列上游1094bp区域存在启动子区异常甲基化,表达下调,提示mir219异常可能在肝细胞癌发生发展中起重要作用,是潜在的抑癌基因。
OBJECTIVE: Abnormality of microRNA plays an important role in tumorigenesis. This study explored the abnormal methylation and abnormal expression of microRNA mir219 in hepatocellular carcinoma, which may be involved in the development of hepatocellular carcinoma. Methods: The methylation of the predicted upstream promoter region of mir219 in normal liver tissue, hepatocellular carcinoma cell lines and hepatocellular carcinoma specimens was detected by pyrosequence and TA cloning sequencing, and the expression of tumor tissue was detected by Taqman real-time RT-PCR. Results: Compared with the normal liver tissue, the CpG islands in the stem-loop region and upstream 1-kb region of the mir219 upstream predicted promoter region in the liver cancer cell line and the liver cancer specimen are highly methylated. The expression of mir219 in hepatocellular carcinoma cell lines and hepatocellular carcinoma was significantly down-regulated, which correlated with its abnormal methylation. CONCLUSION: Abnormal methylation of promoter region in 1094bp region upstream of the microRNA RNA219 stem-loop sequence in hepatocellular carcinoma is down-regulated, suggesting that aberrant mir219 may play an important role in the occurrence and development of hepatocellular carcinoma and is a potential tumor suppressor gene.