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目的 :探讨基因工程药物CNB通过线粒体途径引起肿瘤细胞凋亡的机制。方法 :本研究采用MTT比色法检测细胞存活和生长、流式检测细胞凋亡测定CNB对肿瘤细胞的杀伤效果;采用高内涵分析与成像仪检测细胞线粒体膜电势,以判断加入CNB后线粒体膜电势的变化情况等。结果 :在体外CNB能够有效的抑制Hep G2细胞的增殖。CNB对Hep G2细胞的抑制具有浓度依赖效应,浓度越高,抑制率相对越高。MTT实验证明,在体外CNB能够有效抑制肿瘤细胞的增殖;不同批次的CNB均具有同样的作用,能够抑制肿瘤细胞的增殖。但不同批次的CNB效果不同;CNB有浓度梯度效应,随着CNB浓度增加,能明显上调肿瘤细胞晚期凋亡的比例,当浓度达到700ug/ml时和抗肿瘤药品五氟尿嘧啶效果基本相同,说明CNB能促进肿瘤细胞凋亡;3种细胞加入CNB后均Caspase3表现为下调,说明Caspase3剪切体上调,Caspase3蛋白被激活,显示CNB能促进细胞凋亡;CNB能够定位到线粒体,导致线粒体膜通透性增加,从而引发线粒体介导的凋亡。结论 :CNB能够抑制肿瘤细胞增殖;CNB能够促进肿瘤细胞凋亡;CNB可促进线粒体途径的细胞凋亡。
Objective: To investigate the mechanism of gene engineering drug CNB through mitochondrial pathway causing tumor cell apoptosis. Methods: MTT assay was used to detect the survival and growth of cells. Flow cytometry was used to detect the killing effect of CNB on tumor cells. Mitochondrial membrane potential was measured by high-content analysis and imager to determine the mitochondrial membrane potential Potential changes and so on. Results: In vitro CNB can effectively inhibit the proliferation of Hep G2 cells. The inhibitory effect of CNB on Hep G2 cells in a concentration-dependent manner, the higher the concentration, the higher the inhibition rate. MTT experiments show that in vitro CNB can effectively inhibit tumor cell proliferation; different batches of CNB have the same effect, can inhibit the proliferation of tumor cells. CNB had different effects on concentration of CNB. Concentration gradient of CNB increased the proportion of advanced apoptosis cells with increasing concentration of CNB. When the concentration of CNB was 700ug / ml, the effect was similar to that of antitumor drug pentafluorouracil CNB could promote the apoptosis of tumor cells. Caspase3 expression was down-regulated after CNB treatment. Caspase3 was upregulated and Caspase3 protein was activated, which indicated that CNB could promote apoptosis. CNB could localize to mitochondria, Permeability increases, triggering mitochondria-mediated apoptosis. Conclusion: CNB can inhibit tumor cell proliferation; CNB can promote tumor cell apoptosis; CNB can promote mitochondrial pathway apoptosis.