二甲双胍对人口腔癌KB细胞增殖及凋亡的影响  

Effects of metformin on human oral cancer KB cell proliferation and apoptosis in vitro

作　　者：王芳[1,2] 徐锦程[1,2] 夏飞[1,2] 刘哲[1,2] 赵素容 刘浩[1,2] 蒋志文 

机构地区：[1]蚌埠医学院第一附属医院 [2]蚌埠医学院药学系//安徽省生化药物工程技术研究中心,安徽蚌埠233030

出　　处：《南方医科大学学报》2014年第2期159-163,共5页Journal of Southern Medical University

基　　金：国家自然科学基金(81000992;81072207);安徽省自然科学基金(1208085MH136);安徽省高校省级自然科学研究重点(KJ2012A201)~~

摘　　要：目的研究二甲双胍(Metformin)对人口腔癌KB细胞增殖及凋亡的影响,以期为口腔癌的治疗开辟新的路径。方法 MTT法检测不同浓度(1.25、2.5、5、10、20 mmol/L)二甲双胍处理口腔癌KB细胞24、48、72 h后对细胞增殖的影响;分别用0.25、0.5、1 mmol/L的二甲双胍处理KB细胞,8 d后观察其对集落克隆形成的影响;5 mmol/L二甲双胍处理细胞24 h,线粒体膜电位检测试剂盒(JC-1)检测线粒体膜电位的变化;PI单染检测二甲双胍对KB细胞凋亡的影响;Western blot检测二甲双胍(5 mmol/L)处理KB细胞不同时间(0、6、16、24 h)GRP78以及caspase-3的表达。结果二甲双胍处理细胞后,对KB细胞的增殖有明显的抑制作用,并且呈药物浓度和时间依赖性。5 mmol/L的二甲双胍处理KB细胞24、48、72 h的存活率为68.0%、36.9%、14.5%,但24 h细胞凋亡率仅为11.99%,将浓度加大后凋亡率为24.11%,高于对照组;二甲双胍抑制KB细胞集落克隆的形成;JC-1荧光检测,红绿荧光的相对比例降低,提示膜电位下降;二甲双胍刺激口腔癌KB细胞GRP78的表达先上调后有减弱趋势,并且能诱导Caspase-3的激活。结论二甲双胍能显著抑制KB细胞的增殖并且诱导细胞凋亡,其机制与线粒体凋亡途径的激活以及过度的内质网应激有关。Objective To investigate the effects of metformin on the proliferation and apoptosis of human oral cancer cell line KB in vitro. Methods Human oral cancer cell line KB was exposed to different doses of metformin （0, 1.25, 2.5, 5, 10, and 20 mmol/L）, and the changes in cell viability were detected using MTT assay. Colony formation of the cells was observed following an 8-day metformin exposure. The changes in mitochondrial membrane potential were measured by JC-1 assay, and PI staining was used to observe the cell apoptosis. Western blotting was employed to detect the changes in the protein expressions of GRP78 and activated caspase-3. Results Metformin exposure caused time-and dose-dependent suppression of KB cell proliferation, and exposure to 5 mmol/L metformin for 24, 48 and 72 h resulted in cell survival rates of 68.0%, 36.9%, and 14.5%, respectively. Metformin significantly inhibited KB cell colony formation. Exposure of the cells to increased concentrations of metformin gradually increased the apoptotic rate and decreased mitochondrial membrane potential. Metformin caused an initial up-regulation followed by a down-regulation of GRP78 expression in KB cells and increased the expression of activated caspase-3. Conclusion Metformin can inhibit the proliferation and induce apoptosis of KB cells, the mechanism of which may involve the activation of the mitochondrial apoptotic pathway and endoplasmic reticulum stress.

关 键 词：口腔癌 二甲双胍 凋亡 葡萄糖调节蛋白78 CASPASE-3 glucose-regulated protein 78 CASPASE-3 

分 类 号：R739.85[医药卫生—肿瘤]