DNA去甲基化对急性B淋巴细胞白血病细胞株中DNA甲基转移酶基因及微RNA作用的研究  

Study of DNA demethylation acting on DNA methyltransferase gene and microRNA in B-cell acute lymphocytic leukemia cell line

作　　者：石懿玉 周永安[2] 张万红[3] 马云霞[2] 曾红艳[4] 杨慧芳[4] 

机构地区：[1]山西省心血管病医院实验室,太原030024 [2]太原市中心医院中心实验室,030009 [3]山西省肿瘤医院消化微创外科,太原030013 [4]山西医科大学研究生院,太原030001

出　　处：《白血病．淋巴瘤》2016年第2期111-115,共5页Journal of Leukemia & Lymphoma

基　　金：山西省基础研究计划（2012011038-3）;山西省卫生厅科技攻关计划（2011085）

摘　　要：目的：探索甲基化转移酶抑制剂5-杂氮-2＆#39;-脱氧胞苷（5-Aza-CdR）对急性B淋巴细胞白血病（B-ALL）细胞株NALM-6的作用以及对细胞中微RNA（miRNA）表达水平的影响。方法用不同浓度5-Aza-CdR处理NALM-6细胞，采用四甲基偶氮唑盐（MTT）法检测细胞增殖情况，采用荧光定量反转录聚合酶链反应（RT-PCR）检测5-Aza-CdR处理后细胞DNA甲基转移酶（DNMT）基因mRNA表达水平的变化，采用miScript miRNA PCR Array芯片检测去甲基化后细胞中表达量发生改变的miRNA。结果 NALM-6细胞经不同浓度5-Aza-CdR处理不同时间后，细胞生长受抑，最高抑制率达（74.163±0.381）%。5-Aza-CdR作用浓度与DNMT基因mRNA表达水平呈反比，浓度为1000μmol／L的5-Aza-CdR作用细胞72 h后，DNMT-1的相对表达量降至0.453±0.021，DNMT-3L的相对表达量为0.003±0.001， DNMT-3B的相对表达量为0.395±0.019。miScript miRNA PCR Array筛选出3个miRNA（miR-184、miR-23a-3p、miR-34a-5p）与DNA甲基化相关。结论5-Aza-CdR可下调NALM-6细胞中DNMT基因的表达，并对细胞增殖有抑制作用。miR-184、miR-23a-3p和miR-34a-5p在B-ALL的发生、发展中与DNA甲基化相关。Objective To explore the function of 5-Aza-CdR in B-cell acute lymphocytic leukemia cell line NALM-6 and its influence on the expression of microRNA （miRNA） in the cells. Methods NALM-6 was treated with different concentrations of 5-Aza-CdR. Cell proliferation was detected by methyl thiazolyl tetrazolium （MTT） test, and DNA methyltransferase （DNMT） mRNA expression level was detected by reverse transcription PCR （RT-PCR）. The expression changes of miRNA were detected by miScript miRNA PCR Array chip in cells after methylation. Results NALM-6 cell growth was inhibited by different concentrations of 5-Aza-CdR processing time, reaching to the maximum inhibitory rate was （74.163 ±0.381） %. 5-Aza-CdR affected concentrations was inversely proportional with expression level of DNMT mRNA. After 1 000 μmol/L of 5-Aza-CdR was dealed with cell 72 h, the relative expression of DNMT-1 was reduced to 0.453 ±0.021, DNMT-3L was 0.003±0.001, DNMT-3B was 0.395±0.019. MiScript miRNA PCR array sieved out 3 miRNA （miR-184, miR-23a-3p, miR-34a-5p） associated with DNA methylation. Conclusions 5-Aza-CdR down regulates the expression of DNMT gene in NALM-6 cells, and inhibits the proliferation of cells. MiR-184, miR-23a-3p and miR-34a-5p are related to DNA methylation in the occurrence and development of B-cell acute lymphocytic leukemia.

关 键 词：白血病 B淋巴细胞 急性 DNA甲基化 微RNA 5-杂氮-2'-脱氧胞苷 

分 类 号：R733.71[医药卫生—肿瘤]