摘要
目的:研究粉防己碱通过干预人肝癌HepG_(2)细胞膜受体进而诱导HepG_(2)细胞凋亡的抗肿瘤机制。方法:CCK-8法检测HepG_(2)细胞的增殖情况;倒置显微镜、荧光显微镜观察HepG_(2)细胞形态;流式细胞仪检测HepG_(2)细胞凋亡率。试剂盒检测HepG_(2)细胞膜总蛋白、胆固醇、唾液酸含量;荧光染色技术检测HepG_(2)细胞膜流动性;试剂盒检测HepG_(2)细胞膜封闭度及Na^(+),K^(+)-ATPase、Ca^(2+),Mg^(2+)-ATPase活性变化;Western blot法检测HepG_(2)细胞膜葡萄糖转运蛋白(GLUT-1)和氨基酸转运蛋白(LAT-1)及HepG_(2)细胞凋亡相关蛋白(Bcl-2、Bax、Cyt-c)表达;试剂盒检测Caspase-3蛋白活性。结果:粉防己碱对HepG_(2)细胞具有增殖抑制作用;形态学观察显示给药组细胞排列变得稀疏,多数细胞由梭形和多角形变成了圆形;同时细胞间轮廓变模糊,出现细胞碎裂现象,细胞核固缩形成凋亡小体。流式细胞仪检测结果显示各给药组出现明显的凋亡峰。粉防己碱各给药组细胞膜总蛋白、胆固醇、唾液酸含量均显著降低,同时细胞膜的流动性和封闭度显著降低,其作用呈浓度依赖性。粉防己碱各给药组细胞膜Na^(+),K^(+)-ATPase和Ca^(2+),Mg^(2+)-ATPase活性显著降低,GLUT-1、LAT-1蛋白表达显著下调,细胞Cyt-c蛋白表达及Caspase-3活性显著升高,Bcl-2/Bax水平显著降低。结论:粉防己碱能通过干预肿瘤细胞膜受体诱导肝癌HepG_(2)细胞发生凋亡。
Objective:To study the anti-tumor mechanism of tetrandrine by interfering with human liver cancer HepG_(2)cell membrane receptors to induce HepG_(2)cell apoptosis.Methods:CCK-8 method was used to detect the proliferation of HepG_(2)cells;HepG_(2)cell morphology was observed under inverted microscope and fluorescence microscope,its apoptosis rate was detected by flow cytometry.HepG_(2)cell membrane total protein,cholesterol and sialic acid contents were detected by the kit.Fluorescence staining was used to detect cell membrane fluidity of HepG_(2).HepG_(2)cell membrane closure and Na^(+),K^(+)-ATPase,Ca^(2+),Mg^(2+)-ATPase activity changes were detected by kit.The expressions of HepG_(2)cell membrane glucose transporter(GLUT-1),amino acid transporter(LAT-1)and apoptosis-related proteins(Bcl-2,Bax,Cyt-c)in HepG_(2)cells were detected by Western blot;Caspase-3 protein activity was detected with the kit.Results:Tetrandrine could inhibit the proliferation of HepG_(2)cells.Morphological observation results showed that the cell arrangement became sparse in the administration group and most cells changed from spindle and polygon to round.At the same time,the contour between cells became blurred,the phenomenon of cell fragmentation appeared,and the cell nucleus constricted to form apoptotic bodies.Flow cytometry results showed that there were obvious peaks of apoptosis in each administration group.The contents of cell membrane total protein,cholesterol and sialic acid in each administration group of tetrandrine decreased significantly and the fluidity and closure of the cell membrane were significantly reduced in a concentration-dependent manner.The cell membrane Na^(+),K^(+)-ATPase and Ca^(2+),Mg^(2+)-ATPase activities of each administration group of tetrandrine were significantly decreased,and the expressions of GLUT-1 and LAT-1 were significantly down-regulated,Cyt-c protein expressions and Caspase-3 activity were significantly increased,Bcl-2/Bax level was significantly decreased.Conclusion:Tetrandrine can induce apoptosis of HepG_(2)cells by interfering with tumor cell membrane receptors.
作者
张晶
李吉业
于淼
刘斌
李钧
陈鹰翔
辛国松
ZHANG Jing;LI Ji-ye;YU Miao;LIU Bin;LI Jun;CHEN Ying-xiang;XIN Guo-song(Engineering Research Centre for Medicine,Harbin University of Commerce,Harbin 150076,China;Engineering Research Centre of Natural Anticancer Drugs,Ministry of Education,Harbin 150076,China)
出处
《中药材》
CAS
北大核心
2021年第7期1719-1725,共7页
Journal of Chinese Medicinal Materials
基金
中国博士后基金项目(2019M651296)
黑龙江省自然科学基金项目(LH2019H066)
黑龙江省教育厅高校创新人才项目(UNPYSCT-2017208)
哈尔滨商业大学创新人才项目(2019CX37,18XN024)
关键词
粉防己碱
细胞膜
膜受体蛋白
细胞凋亡
抗肿瘤
Tetrandrine
Cell membrane
Membrane receptor protein
Cell apoptosis
Anti-tumor
