金丝桃苷对耐长春新碱结肠癌HCT8/VCR细胞耐药逆转作用  

Reversal Effect of Hyperin on Vincristine-Resistant Colon Cancer HCT8/VCR Cells

作　　者：王丽敏[1] 杨玉[1] 张明远[1] 卢春凤[1] 朱秋双[1] 

机构地区：[1]佳木斯大学基础医学院,黑龙江佳木斯154007

出　　处：《中国实验方剂学杂志》2016年第22期92-96,共5页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81301918);黑龙江省自然科学基金项目(H201356)

摘　　要：目的：探讨金丝桃苷对耐长春新碱结肠癌HCT8/VCR细胞的耐药逆转作用及其可能的作用机制。方法：分别以不同质量浓度（2.5-80 mg·L-1）的金丝桃苷及不同质量浓度（1.25-40 mg·L-1）的长春新碱作用于体外培养的HCT8和HCT8/VCR细胞48 h,采用噻唑蓝（MTT）比色法测定细胞生长抑制率,计算各组细胞金丝桃苷和长春新碱的半数抑制浓度（IC50）及金丝桃苷抑制率为5%的药物浓度;采用非毒性剂量金丝桃苷分别与不同质量浓度（1.25-40 mg·L-1）的长春新碱联合作用HCT8/VCR细胞,检测长春新碱的IC50,计算逆转倍数;采用流式细胞仪检测细胞凋亡率及线粒体膜电位变化,比色法检测含半胱氨酸的天冬氨酸蛋白水解酶（Caspase）-3,Caspase-9蛋白活性的变化。结果：金丝桃苷非细胞毒性剂量为0.82mg·L-1。0.82 mg·L-1金丝桃苷可使长春新碱对HCT8/VCR细胞的IC50从65.97 mg·L-1下降至9.94 mg·L-1,逆转倍数为6.63;与单独用长春新碱组比较,长春新碱与金丝桃苷联合用药组细胞凋亡率,Caspase-3,Caspase-9蛋白活性均显著提高（P〈0.01）。线粒体膜电位检测发现,与单独用长春新碱组及金丝桃苷组比较,长春新碱与金丝桃苷联合用药组低荧光强度相对细胞数所占比率显著提高（P〈0.01）。结论：长春新碱与金丝桃苷联合应用后可增强Caspase-3,Caspase-9蛋白活性,降低线粒体膜电位,提高HCT8/VCR细胞的凋亡率,从而产生耐药逆转作用。Objective： To investigate the reversal effect of hyperin on vincristine（VCR）-resistant colon cancer HCT8 / VCR cells,and its possible mechanism. Method： HCT8 cells and HCT8 / VCR cells were cultured with hyperin at the concentrations of 2. 5-80 mg·L- 1and vincristine at concentrations of 1. 25-40 mg·L- 1.The inhibitory rate of the cell growth was measured by Methylthiazolyldiphenyl-tetrazolium bromide（MTT） assay.The 50% inhibitory concentration（IC50） of hyperin and vincristine,as well as non-cytotoxic dose of hyperin were counted. HCT8 / VCR cells were treated with hyperin at non-cytotoxic dose with different concentrations of vincristine（VCR,1. 25-40 mg·L- 1） to determine IC50 and the reversal multiple in all groups. The apoptotic rate and the changes in mitochondrial membrane potential were evaluated by flow cytometry. The activities of Caspase-3and Caspase-9 were detected by colorimetric assays. Result： Hyperin at the concentration of 0. 82 mg·L- 1has no significant cytotoxicity to HCT8 / VCR cells. Hyperin at the concentration of 0. 82 mg·L- 1decreased the IC50 value of vincristine to HCT8 / VCR cells from 65. 97 mg·L- 1to 9. 94 mg·L- 1,with the reversal multiple of 6. 63. The apoptotic rate and Caspase-3 and Caspase-9 activities in combination group were significantly increased compared with vincristine group（P 〈0. 01）. The data of mitochondrial membrane potential showed that the percentage of cells in low fluorescence in combination group was increased compared with vincristine group（P〈 0. 01）.Conclusion： Vincristine combined with hyperin could increase Caspase-3 and Caspase-9 protein activities,decrease the mitochondrial membrane potential,and enhance apoptosis rate,so as to reverse the multidrug resistance of HCT8 / VCR cells.

关 键 词：金丝桃苷 长春新碱 HCT8/VCR细胞 多药耐药 含半胱氨酸的天冬氨酸蛋白水解酶-3 含半胱氨酸的天冬氨酸蛋白水解酶-9 半数抑制浓度 

分 类 号：R285.5[医药卫生—中药学]