5-氮杂-2’-脱氧胞苷抑制肾癌细胞增殖及逆转γ-catenin基因甲基化作用
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江苏大学临床医学科技发展基金(JLY20050015)


Effect of 5-Aza-2’-deoxycytidine on growth inhibition of renal carcinoma cell line OS-RC-2 and reversion of γ-catenin gene hypermethylation
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    摘要:

    目的:研究甲基化抑制剂5-氮杂-2’-脱氧胞苷(5-Aza-CdR)对人肾癌OS-RC-2细胞生长的抑制作用及对γ-catenin基因甲基化状态的影响,初步探讨肾癌发病机制及临床治疗的可行性-方法:不同浓度5-Aza-CdR处理肾癌细胞OS-RC-2后,采用倒置显微镜观察5-Aza-CdR处理OS-RC-2肾癌细胞株前后细胞形态学变化;四唑蓝(MTT)比色观察细胞经药物处理前后的生长活性;流式细胞仪检测5-Aza-CdR对OS-RC-2肾癌细胞株凋亡的影响;Western blot检测5-Aza-CdR处理OS-RC-2肾癌细胞株后γ-catenin蛋白表达的变化;甲基化特异性PCR(methylation-specific PCR,MSP)检测细胞处理前后γ-catenin 基因的甲基化状态-结果:形态学观察显示,用药后癌细胞体积缩小,核固缩,染色质凝聚成块;5-Aza-CdR明显抑制肾癌细胞OS-RC-2的生长;细胞凋亡率增高;药物处理后γ-catenin蛋白表达恢复;未经5-Aza-CdR处理的OS-RC-2细胞中γ-catenin基因启动子区域高甲基化,经10-5 mol/L 5-Aza-CdR处理72 h后,γ-catenin基因启动子区域高甲基化得到逆转-结论:5-Aza-CdR能有效逆转肾癌OS-RC-2细胞γ-catenin基因的异常甲基化,恢复γ-catenin蛋白表达,诱导肾癌细胞凋亡-

    Abstract:

    Objective:To investigate the effects of 5-Aza-2’-deoxycytidine(5-Aza-CdR) a methylation inhibitor on the growth of human renal carcinoma cell line OS-RC-2,and the DNA CpG island demethylation of γ-catenin gene so as to provide insights into origin of renal carcinoma and the possibility of its application in clinical treatment. Methods:MTT method and flow cytometry were used to detect the growth and appoptosis of OS-RC-2 after 5-Aza-CdR treatment respctirely. The western blot was used to detevt γ-catenin protein levels in the cell line before and after treatment with 5-Aza-CdR.The methylation and demethylation status of γ-catenin gene were volume detected by methylation special PCR(MSP). Results:The morphological pattern changes were observed invert-microscope. OS-RC-2 cells treated with 5-Aza-CdR displayed a slowed growth in comparision with the control cells. The apoptotic rate was also increased after 5-Aza-CdR treatment. Western blot indicated that expression of γ-catenin protein was recovered by 5-Aza-CdR treatment. The high methylation status of γ-catenin gene promoter region was reversed with 10-5 mol/L 5-Aza-CdR treated for 72 h. Conclusion:5-Aza-CdR effectively causes the demethylation of γ-catenin gene CpG-rich promoter regions, and recover the γ-catenin protein expression, subequenely induce the appoptosis of OS-RC-2 cells.

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陶美满,孙 浩,马克均,张顺兴,尤海燕.5-氮杂-2’-脱氧胞苷抑制肾癌细胞增殖及逆转γ-catenin基因甲基化作用[J].南京医科大学学报(自然科学版),2009,29(6):831-835

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  • 收稿日期:2009-01-15
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