Objective To investigate apoptosis inducted by levo-gossypol combined witn low concentration aoriamycm in HT1080 cells and its mechanisms.Methods The anti-proliferation of drugs on cells was detected by MTT assay.The changes of cells morphology were observed by Wright-Giemsa staining,Hoechst 33258 staining and transmission electron microscopy.The apoptosis rate and cell cycle were detected by flow cytometry.And the protein expression of Bcl-2 and Bax in HT1080 cell were detected by Western-Blot.The changes of caspase-3 and caspase-9 were detected by colorimetric assay.Results Levo-gossypol and low concentration adriamycin showed inhibition of HT1080 cells in a dose and time dependent.The combination had more obvious effect.Levo-gossypol and low concentration adriamycin induced apoptosis of HT1080 cell,as evidenced by typical morphological changes.Flow cytometry detected that the drugs induced apoptosis of HT1080 cell.The combination had more obvious apoptosis-inducing effect.Levo-gossypol could induce G1 arrest and adriamycin could induce S arrest.The combination had more obvious increase of S arrest.Levo-gossypol and low concentration adriamycin can result in decrease of Bcl-2 protein,increase of Bax protein,and activation of caspase-9 and caspase-3.The combination had more obvious changes.Conclusion Levo-gossypol and low concentration adriamycin have the effect of anti-proliferation and induction of apoptosis in HT1080 cell.The combination has more obvious effect.%目的 研究左旋棉酚联合低浓度阿霉素(ADM)诱导纤维肉瘤HT1080细胞凋亡及其作用机制.方法 采用噻唑蓝(MTT)检测药物对细胞的抗增殖作用,应用瑞氏-姬姆萨(WG)染色、赫斯特33258(hoechst 33258)染色、透射电镜观察细胞的形态学变化,流式细胞仪检测细胞的凋亡率和细胞周期.蛋白免疫印迹法(Western blotting)检测Bcl-2、Bax蛋白的表达,比色分析法检测细胞半胱氨酸天冬氨酸蛋白酶(caspase)-3、caspase-9的活性变化.结果 左旋棉酚与低浓度ADM对HT1080细胞具有抑制增殖作用,联合应用作用增强,有浓度与时间依赖性.药物作用后出现细胞凋亡的特征性改变,琼脂糖凝胶电泳观察到梯状带型,流式细胞仪检测发现两药能诱导肿瘤细胞凋亡,联合应用更强.左旋棉酚使细胞阻滞于G1期,ADM使细胞阻滞于S期,联合应用S期的细胞明显增加.细胞内Bcl-2蛋白表达减少,Bax蛋白表达增多,同时出现caspase-9、caspase-3的相继活化,联合应用变化更加明显.结论 左旋棉酚与低浓度ADM对纤维肉瘤HT1080细胞有明显抑制增殖及诱导凋亡的作用,联合应用作用加强.
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