高三尖杉酯碱通过mTOR通路诱导慢性髓系白血病K562细胞凋亡

Mechanism of mTOR Pathway in K562 cell Apoptosis Induced by Homoharringtonine

目的:观察高三尖杉酯碱(Homoharringtonine,HHT)对人CML细胞株K562的增殖及凋亡的影响,并从mTOR通路探索HHT抗CM L效应的作用机制。方法:采用不同浓度HHT或联合mTOR抑制剂雷帕霉素(RAPA)处理K562细胞,用CCK-8法检测细胞增殖抑制活力,流式细胞术检测细胞凋亡率,Western blot检测BCL-6、Caspase-3及mTOR通路相关蛋白的表达水平,RT-PCR法检测BCL-6 mRNA表达水平。结果:HHT可抑制K562细胞的增殖并促进其凋亡,且呈浓度依赖性(r=0.970)。随着HHT作用浓度的增加,mTOR通路相关蛋白表达水平升高(r=0.908),而BCL-6 mRNA及蛋白的表达水平下降(r_(mRNA)=-0.961,r_(protein)=-0.981)。与HHT单用组相比,联用RAPA可显著下调mTOR及Caspase-3的表达,并上调BCL-6表达。结论:HHT可通过激活mTOR通路抑制抗凋亡BCL-6蛋白的表达,进而诱导CML细胞凋亡。

Objective： To investigate the effect of homoharringtonine（ HHT） on proliferation and apoptosis of CML cell line K562 cells and to explore its possible mechanism through mTOR pathway. Methods： K562 cells were cultured with different concentrations of HHT or in its combination with mTOR inhibitor rapamycin（ RAPA） for 24 hours. The cell viability was analyzed by CCK-8 assay,the cell apoptosis was detected by flowcytometry,the expressions of BCL-6,Caspase-3 and mTOR signal pathway related proteins was assayed by Western blot,the expression of BCL-6 mRNA was determined by RT-PCR. Results： The HHT inhibited proliferation and induced apoptosis of K562 cells in a concentration-dependent manner（ r = 0. 970）. With the increasing of HHT concentration,the expression level mTOR signal pathway related proteins increased（ r = 0. 908）,while the mRNA and protein expression levels of BCL-6 decreased（ rmRNA=-0. 961,rprotein=-0. 981）,as compared with the HHT alone,the combination of HHT with RAPA could down-regulate the expression of mTOR signal pathway related protein and caspase-3,and up-regulated expression of BCL-6. Conclusion： HHT induces apoptosis of K562 cells by inhibiting BCL-6 expression through mTOR signal pathway.