摘要
本研究探讨5-氮杂-2'-脱氧胞苷(5-Aza-CdR)对人慢性髓系白血病(CML)急红变细胞系K562细胞生物学活性和DNA结合抑制因子4(ID4)基因表达的影响,探寻白血病基因治疗的新靶点。应用甲基化特异性PCR方法检测K562细胞中ID4基因甲基化情况;实时荧光定量PCR检测5-Aza-CdR处理K562细胞后ID4 mRNA的表达水平;流式细胞术分析5-Aza-CdR处理后细胞凋亡率和细胞周期的变化。结果表明,K562细胞中存在ID4基因的甲基化;5-Aza-CdR处理后K562细胞ID4 mRNA表达增加,并具有浓度依赖性,不同浓度药物处理组之间差异具有统计学意义(p<0.01)。5-Aza-CdR可使K562细胞凋亡率增加,并且作用呈时间剂量依赖性。且细胞凋亡率与ID4 mRNA的相对表达水平呈高度相关(r=0.95)。5-Aza-CdR处理K562细胞48小时后,随着药物浓度的增加,G0/G1期细胞逐渐增多,G2/M期细胞逐渐减少,细胞阻滞在G0/G1期。结论:甲基化转移酶抑制剂5-Aza-CdR能促使CML急红变细胞系K562细胞中沉默的ID4基因重新表达,可能进而参与K562细胞凋亡和细胞周期阻滞的调控。
This study was aimed to investigate the effect of 5-Aza-CdR on the biological activity of human erythroleukemia cell line K562 and the expression of inhibitor of DNA binding 4(ID4).ID4 methylation in K562 cell line was detected by methylation-specific PCR.RQ-PCR was used to analyze the expression levels of ID4 mRNA in K562 cell line treated by different concentrations of 5-Aza-CdR.Cell apoptosis rate and cell cycle were analyzed by flow cytometry.The result showed that ID4 gene methylation existed in K562 cells,ID4 mRNA expression in K562 cells treated with 5-Aza-CdR increased in a concentration-dependent manner,the difference between experimental groups was statistical significant(p〈0.01).The 5-Aza-CdR could enhance the apoptotic rate of K562 cells in time and dose-dependent manner,the apoptotic rate of K562 cells highly correlated to relative expression level of ID4 mRNA(r=0.95).After the K562 cells were treated by 5-Aza-CdR for 48 hours,cells in G0/G1 phase increased,cells in G2/M phase decreased along with enhancement of drug concentration.It is concluded that methyltransferase inhibitor 5-Aza-CdR can re-express the silent ID4 gene in K562 cells.The upregulation of ID4 may be a key factor to give rise to cell apoptosis,and the cell cycle of K562 cells can be arrested by 5-Aza-CdR.
出处
《中国实验血液学杂志》
CAS
CSCD
2011年第6期1388-1392,共5页
Journal of Experimental Hematology
基金
湖北省自然科学基金
编号2009CDB416