【摘要】 目的:观察去甲基化药物对人肺癌A549和H1299细胞增殖、凋亡以及小分子RNA let-7a-2表达的影响。方法:以不同浓度的5-杂氮-2’-脱氧胞苷(DAC)对肺癌细胞进行处理后,CCK-8检测对细胞增殖的影响,AnnexinV-PI双染法检测对细胞凋亡的影响,qRT-PCR对let-7a-2的表达进行检测。结果:A549和H1299细胞经不同浓度DAC处理后细胞增殖受到明显抑制,以60μM浓度DAC处理后d5抑制率分别为(40.80±6.50)%和(47.24±4.95)%(均P<0.05),同时细胞呈现显著性早期凋亡,凋亡百分比分别达(64.58±4.21)和(59.61±5.69)(均P<0.05)。20μM、40μM、60μM DAC处理后let-7a-2的表达量在A549中分别为(10.86±0.30)、(5.02±2.83)、(17.79±1.95),比对照组(1.12±0.24)显著上调;在H1299中分别为(55.13±5.69)、(35.67±4.13)、(14.94±2.46),比对照组(1.31±0.56)显著上调。结论:DAC处理可抑制肺癌A549和H1299细胞增殖、促进细胞凋亡,上调let-7a-2的表达,let-7a-2基因调控区域超甲基化可能是其在肺癌细胞中表达异常的机制之一。更多还原

【Abstract】 Objective:To observe the effects of demethylating agent on cell proliferation,apoptosis and expression of miRNA let-7a-2 in human lung cancer cell lines A549 and H1299.Methods:A549 and H1299 cells were treated with different concentration of demethylating agent 5-aza-2’-deoxycytidine(DAC).Cell proliferation was measured with cell counting kit(CCK-8).Apoptotic rate was determined with AnnexinV-PI double staining on flow cytometry.The expression of let-7a-2 was measured with quantitative real-time PCR.Results:Cell proliferation of A549 and H1299 was inhibited by DAC treatment.The inhibition rates on the fifth day after60μM DAC treatment were(40.80±6.50)%(P<0.05)and(47.24±4.95)%(P<0.05).The apoptotic rates in A549 and H1299 were(64.58±4.21)and(59.61±5.69),respectively.They were significant increased by DAC treatment.The expression of let-7a-2 was upregulated after 20μM,40μM and 60μM DAC treatment,quantities reached to(10.86±0.30),(5.02±2.83),(17.79±1.95)vs control group(1.12±0.24)in A549 cells,and(55.13±5.69),(35.67±4.13),(14.94 ±2.46)vs control group(1.31 ±0.56)in H1299 cells.Conclusion:DAC treatment can inhibit the growth and induce apoptosis of human lung cancer A549 and H1299 cells,downregulate the expression of let-7a-2.The hypermethylation change on let-7a-2 gene regulating region may be one of the possible reasons of the disregulation effect of let-7a-2 in these lung cancer cells.更多还原