三氧化二砷对骨髓瘤细胞系U266抑制增殖及诱导凋亡的机制研究

本研究探讨三氧化二砷(As2O3)在体外对骨髓瘤细胞系U266的生物学效应及其机制。用MTT法观察As2O3对U266细胞增殖的影响,用流式细胞术、DNA凝胶电泳法分析As2O3对U266细胞凋亡的影响,以RT-PCR法检测2μmol/LAs2O3不同处理时间对U266细胞人端粒酶逆转录酶蛋白催化亚单位(hTERT)mRNA的表达变化,用Western blot法检测As2O3不同处理时间对U266细胞procaspase-3、bcl-2及hTERT蛋白水平的表达变化。结果表明:As2O3能明显抑制U266细胞增殖,半数抑制浓度(IC50)为2μmol/L;As2O3能诱导U266细胞凋亡,呈现剂量和时间依赖性;2μmol/L As2O3不同时间处理U266细胞后procaspase-3蛋白及hTERT mRNA、蛋白表达呈时间依赖性降低,而bcl-2蛋白表达未发生变化。结论:As2O3通过改变线粒体跨膜电位,触发了U266细胞的线粒体凋亡途径,导致了caspase-3的活化,最终诱导U266细胞凋亡;hTERT表达下调也是As2O3诱导U266细胞凋亡的重要作用机制。

Mechanisms Underlying the Effect of Arsenic Trioxide on Proliferation Inhibition and Apoptosis Induction in Myeloma Cell Line U266

The aim of this study was to explore the mechanisms underlying effect of arsenic trioxide (As2O3) on myeloma cell line U266 in vitro. The viability and apoptosis of U266 cells were observed by MTT assay, flow cytometry and DNA agarose gel electrophoresis. The expression of hTERT mRNA was assessed by RT-PCR analysis. The variation of procaspase-3, bcl-2 and hTERT protein expression were detected by Western blot. The results indicated that the As2O3 could inhibit the growth of U266 cells significantly and the concentration of 50% growth inhibition (IC50) was 2 micromol/L. After treatment with 2.5 micromol/L As2O3 at 24, 48 and 72 hours, a dose- and time-dependent apoptosis of U266 cells could be observed. After treating U266 cells with 2 micromol/L As2O3 at different time points, a time-dependent reduction of procaspase-3, hTERT mRNA and protein was found without any change of bcl-2 expression. It is concluded that the As2O3 can change the mitochondrial transmembrane potential, initiating the mitochondial apoptosis pathway, leading in turn to caspase-3 activation, and inducing the apoptosis of U266 cells. These findings suggest that the reduction of hTERT plays a critical role in the apoptosis of U266 cells induced by As2O3.