5-Aza-CdR对肺癌A549细胞DNA甲基转移酶活性的影响  

Effects of DNA methyltransferase activity in lung cancer cells treated with 5-Aza-CdR

作　　者：赵莹[1] 孙丽华[1] 杨振华[1] 王锋[2] 

机构地区：[1]南京医科大学附属南京市第一医院呼吸科,江苏南京210006 [2]南京医科大学附属南京市第一医院核医学中心,江苏南京210006

出　　处：《南京医科大学学报（自然科学版）》2009年第6期767-770,778,共5页Journal of Nanjing Medical University(Natural Sciences)

基　　金：国家自然科学基金资助项目(30700186);南京市医学科技发展项目(YKK07030)

摘　　要：目的:研究5-氮杂-2’-脱氧胞苷(5-Aza-2’-deoxycitydine,5-Aza-CdR)处理肺癌细胞株A549后,DNA甲基化转移酶(DNA methyltransferase,Dnmts)转录水平及其活性的变化,以及细胞生长状态的改变。方法:5-Aza-CdR处理A549细胞株,半定量RT-PCR检测癌细胞Dnmts的mRNA转录水平;酶活性连续循环比色法检测Dnmts的催化活性;流式细胞仪(FCM)及MTT法检测细胞的生长状态。结果:Dnmts的转录水平5-Aza-CdR处理前后水平分别为:Dnmt1(1.23±0.253,1.15±0.166)、Dnmt3b(0.760±0.164,0.649±0.181),两组无显著性差异(P>0.05)。两组Dnmts催化活性分别为:Dnmt1(0.195±0.030,0.153±0.041)、Dnmt3b(0.172±0.029,0.116±0.050),药物组催化活性下降(P<0.05)。FCM结果提示细胞周期阻滞于G1/G0期,两组凋亡率分别为0.62±0.56,7.60±1.92,凋亡率明显增加(P<0.01)。MTT结果示药物组细胞生长受抑制。结论:5-Aza-CdR抑制A549细胞株增殖,促进凋亡,对Dnmts的抑制作用是通过降低催化活性实现,不影响其转录水平。Objective： To study transcription level and catalytic activity of DNA methyltransferase （Dnmts） in lung cancer cell line A549 exposed in 5-Aza-CdR （5-Aza-2＇-deoxycitydine）,and the role of Dnmts in the process. Methods：A549 cells were exposed in the 5-Aza-CdR for demethylation. Dnmts mRNA levels were determined by semi-quantitative RT-PCR. The catalytic activity of Dnmts was detected by enzyme colorimetric determination kit. The apoposis and changes of cell cycles were observed by flow cytometry （FCM） ,and MTT assay was used to study the cell proliferation. Results：The transcription level of the drug group and control group were Dnmtl （1.23 ± 0.253;1.15 ± 0.166）, Dnmt3b （0.760 ±0.164;0.649 ± 0.181）,No significant difference in quantity of Dnmts mRNA can be oberserved between the two groups （P 〉 0.05）. Catalytic activity of the drug group and control group were Dnmt1 （0.195± 0.030;0.153±0.041）,Dnmt3b （0.172 ± 0.029;0.116 ± 0.050）. Catalytic activity decreased compared with the control group （P 〈 0.05）. FCM recommended 5-Aza-CdR arrested cell cycle at G1/G0,the rates of apoptosis of the two groups were 0.62 ±0.56;7.60 ± 1.92, and the rate of apoptosis of drug group was higher than the control group （P 〈 0.01 ）. Cell proliferation of drug group was inhibited according to the results of MTT. Conclusion： 5-Aza-CdR has cytostatic agent in A549 ceils cultured in vitro,and accelerates its apoptosis. The inhibition of Dnmts catalytic activity can account for demethylating agent of 5-Aza-CdR in A549 cells,not the Dnmts transcription level.

关 键 词：肺癌 5-氮杂-2’-脱氧胞苷 甲基化 DNA甲基化转移酶 

分 类 号：R734.2[医药卫生—肿瘤]