5-Aza-2'-dC通过上调DR4与DR5表达增加TRAIL诱导乳腺癌细胞凋亡的敏感性

摘要

目的 探讨5-Aza-2'-dC增加肿瘤坏死因子相关凋亡诱导配体(TRAIL)诱导乳腺癌细胞凋亡敏感性的作用及其可能机制.方法 5 μmol/L 5-Aza-2'-dC顸处理细胞96 h,TRAIL(0,50,100,200 ng/mL)处理细胞12h,MTT检测细胞增殖抑制率,Annexin V-FITC/PI法检测细胞凋亡,流式细胞技术检测TRAIL受体DR4及DR5的表达,Western blot检测caspase-8蛋白表达水平,Real-time PCR检测细胞DR4、DR5和caspase-8 mRNA表达.结果 5-Aza-2'-dC预处理组与对照组相比,TRAIL对细胞增殖抑制率和诱导凋亡率明显高于对照组(P＜0.05).5-Aza-2'-dC能增加细胞表面DR4和DR5表达,促进凋亡通路关键蛋白caspase-8活化.5-Aza-2'-dC在mRNA水平增加了DR4、DR5和caspase-8的表达.结论 5-Aza-2'-dC增加了TRAIL诱导乳腺癌细胞凋亡的敏感性,其可能机制是通过上调DR4、DR5和caspase-8表达实现.%Objective To observe the effect of 5-Aza-2'-dC on TRAIL-induced apoptosis of breast cancer cells and explore the potential mechanism. Methods Human breast cancer cells MDA-MB-231 were pretreated for 96 h with/without 5-Aza-2'-dC (5 μmol/L), and se-quentially treated with TRAIL(0,50,100,200 ng/mL) for 12 h. MTT assay was used to detect the cell proliferation and Annexin V-FTTC/PI was applied to analyze cell apoptosis. The expression of DR4 and DR5 was analyzed by flow-cytometry and the expression of caspase-8 was determined by Western blot. The mRNA expression of DR4, DR5 and caspase-8 was analyzed by Real-time PCR. Results The sequential treatment with 5-Aza-2'-dC and TRAIL resulted in significant inhibition of the cell proliferation and apoptosis. Moreover,pretreatment of breast cancer cells with 5-Aza-2'-dC caused enhancement of the expression of DR4,DR5 and caspase-8 both in protein and mRNA levels. Conclusion These findings suggest that pretreatment with 5-Aza-2'-dC enhances TRAIL-induced apoptosis in breast cancer cells. 5-Aza-2' -dC mediated up-regulation of DR4, DR5 and caspase-8 expression may the the potential mechanism.