DNA甲基化异常对mir-615-5p在胰腺癌细胞株PANC-1中表达影响
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江苏省自然科学基金(BK2006241);国家自然科学基金(30500492)


Abnormal DNA methylation in pancreatic cancer cell line PANC-1 affects mir-615-5p expression
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    摘要:

    目的:描述胰腺癌细胞株中DNA异常高甲基化对miRNA表达抑制所起的作用-观察DNA甲基转移酶抑制剂对miR-615-5p在胰腺癌细胞株PANC-1中甲基化状态和表达的影响-方法:甲基化芯片筛选出在胰腺癌细胞株PANC-1中存在异常甲基化的miRNA-采用去甲基化药物5-氮杂胞苷(5-aza-2-adeoxycytidine,5-aza-dC)处理体外培养的PANC-1细胞,甲基化特异性聚合酶链反应(methylation-specific PCR,MSP)结合亚硫酸氢盐修饰结合测序法(bisulfite-sequencing PCR,BSP)检测用药前后miR-615-5p的甲基化状态的改变-采用RT-PCR定量检测用药前后miR-615-5p表达量的差异-结果:通过MSP和BSP发现在胰腺癌细胞株PANC-1中甲基化率明显高于正常组织-对胰腺癌细胞株用去甲基化剂5-aza-dC的作用,发现miR-615-5p甲基化率减少并伴随着表达的重新激活-结论:胰腺癌细胞株PANC-1中的miR-615-5p表达与启动子区甲基化状态有关,启动子区的高甲基化导致PANC-1中miR-615-5p的表达沉默-

    Abstract:

    Objective: To observe the abnormal DNA methylation inhibits the miRNA expression in pancreatic cancer cell line, and DNA methyltransferase inhibitor on the miR-615-5p of the methylation status and expression in the pancreatic cancer cell line PANC-1. Methods: Methylation microarray was used to screen in pancreatic cancer cell line PANC-1 in the abnormal methylation of miRNA. PANC-1 cells were treated with demethylating agent 5-aza-2-adeoxycytidine (5-aza-dC) in vitro, Methylation specific polymerase chain reaction (methyla-tion- specific PCR, MSP) combination of bisulfite modified DNA sequencing method (bisulfite-sequencing PCR, BSP) were used to detect miR-615-5p methylation changes before and after treatment. The changes of miR-615-5p expression was detected by quantitative RT-PCR before and after treatment. Results: Methylation was significantly higher in pancreatic cancer cell line PANC-1 than in normal tissue by MSP and BSP methods. miR-615-5p reduction in methylation was associated with the expression reactivation in the pancreatic cancer cells PANC-1 treated with demethylation agent 5-aza-dC. Conclusion: This study demonstrates that aberrant DNA methylation of the miR-615-5p CpG island is closely linked to their inappropriate silencing in pancreatic cancer cell line PANC-1.

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朱 墨,高文涛,钱祝银,苗 毅. DNA甲基化异常对mir-615-5p在胰腺癌细胞株PANC-1中表达影响[J].南京医科大学学报(自然科学版),2011,(1):26-30

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  • 收稿日期:2010-07-30
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