盐酸青藤碱通过线粒体凋亡途径诱导胃癌细胞死亡

Sinomenine hydrochloride induces gastric cancer cell death through mitochondrial apoptotic pathway

目的探讨盐酸青藤碱诱导人胃癌BGC-823细胞死亡的作用及其可能机制。方法将胃癌BGC-823细胞分为对照组(未给药)和4个盐酸青藤碱(0.5、1、2、4 mmol/L)处理组,MTT法检测细胞活力;Annexin V-FITC/PI双标记后采用流式细胞仪定量检测凋亡细胞的比例;分光光度法检测半胱天冬酶-3(caspase-3)凋亡蛋白酶活力;JC-1染色标记后采用流式细胞仪检测线粒体膜电位水平;Western blotting检测细胞质中细胞色素C蛋白的表达。结果MTT结果显示,盐酸青藤碱可浓度和时间依赖性抑制人胃癌BGC-823细胞活力;流式细胞术检测结果显示,盐酸青藤碱浓度依赖性促进BGC-823细胞凋亡;盐酸青藤碱可激活caspase-3,促进细胞色素C从线粒体向细胞质中释放,导致细胞线粒体膜电位崩溃;总caspase抑制剂Z-VAD-FMK预处理能有效抑制盐酸青藤碱诱导的caspase-3活化和细胞凋亡。结论盐酸青藤碱可通过激活线粒体凋亡通路显著诱导人胃癌BGC-823细胞凋亡。

Objective To investigate the cell death-inducing effect of sinomenine hydrochloride on human gastric cancer BGC-823 cells and its potential mechanism.Methods Gastric cancer BGC-823 cells were divided into 5 groups:control group,sinomenine hydrochloride non-treatment group,and treatment groups that were respectively treated with various doses of sinomenine hydrochloride(0.5,1,2 mmol/L).MTT assay was used to evaluate the cell viability.Flow cytometer was employed to quantitatively detect the percentage of apoptotic cells after Annexin V-FITC/PI staining.Spectrophotometry was used to detect caspase-3 activity.Mitochondrial membrane potential was assessed using flow cytometry after the cells were dyed by JC-1.The expression of cytosolic cytochrome C protein was determined by Western blotting analysis.Results MTT assay showed that sinomenine hydrochloride inhibited the viability of human gastric cancer BGC-823 cells in concentration-and time-dependent manners.Sinomenine hydrochloride induced the apoptosis of BGC-823 cells in a concentration-dependent manner indicated by flow cytometry.Sinomenine hydrochloride could also activate caspase-3,induce release of cytochrome C from mitochondria to cytoplasm and promote the disruption of mitochondrial membrane potential.Pretreatment with Z-VAD-FMK,a pan-caspase inhibitor,could effectively revert caspase-3 activation as well as apoptosis induced by sinomenine hydrochloride.Conclusion Sinomenine hydrochloride can significantly promote the apoptosis via the activation of mitochondrial apoptotic pathway in human gastric cancer bgc-823 cells.