二甲双胍通过激活腺苷酸活化蛋白激酶/p53轴介导的线粒体功能诱导肿瘤细胞凋亡  

Metformin induces apoptosis in tumor cell through activation of adenosine monophosphate-actived protein kinase/p53 axis-mediated mitochondrial function

作　　者：胡晓乐 束媛媛 付瑛[1] Hu Xiaole;Shu Yuanyuan;Fu Ying(Department of Pharmacy,Qingdao Hospital of Rehabilitation University(Qingdao Municipal Hospital),Qingdao 266000,China)

机构地区：[1]康复大学青岛医院(青岛市市立医院)药剂科,青岛266000

出　　处：《中华实验外科杂志》2023年第11期2287-2291,共5页Chinese Journal of Experimental Surgery

基　　金：浙江省医药卫生科技计划项目(2021KY1118)。

摘　　要：目的:探讨二甲双胍(2型糖尿病的一线治疗药物)干预宫颈癌的疗效。方法:体外培养Caski细胞,不同浓度二甲双胍(2.5、5.0、10.0、20.0 nmol/L)干预Caski细胞,设立对照(CON)组,二甲双胍(2.5、5.0、10.0、20.0 nmol/L)干预组。分别采用细胞计数试剂盒(CCK-8)法、克隆形成实验、流式细胞术、DCFH-DA探针检测二甲双胍对Caski细胞增殖、凋亡、ROS产生的影响。应用蛋白质印迹法(Western blot)检测线粒体功能障碍的特异性蛋白以及腺苷酸活化蛋白激酶(AMPK)/p53途径表达。结果:细胞学实验结果显示,CON组和二甲双胍(2.5、5.0、10.0、20.0 nmol/L)干预组的A450值分别为0.98±0.10、0.85±0.10、0.58±0.08、0.46±0.04、0.38±0.03,组间差异有统计学意义(F=42.36,P<0.01);CON组和二甲双胍(2.5、5.0、10.0、20.0 nmol/L)干预组的细胞凋亡率依次为(3.85±0.35)%、(8.55±0.62)%、(12.50±1.02)%、(20.56±2.36)%、(25.76±2.45)%,组间差异有统计学意义(F=56.74,P<0.01);CON组B细胞淋巴瘤/白血病-2相关X蛋白(bax)/B细胞淋巴瘤/白血病-2(bcl-2)(1.00±0.05),细胞色素C(Cyt C)(1.00±0.04),裂解的半胱氨酰天冬氨酸特异性蛋白酶-3(cleaved Caspase-3)(1.00±0.03),Cyto Cyt C(1.00±0.03)和Mito Cyt C(1.00±0.05)浓度,与二甲双胍(2.5、5.0、10.0、20.0 nmol/L)干预组比较(2.5 nmol/L组:bax/bcl-2:1.95±0.09,Cyt C:1.45±0.06,cleaved Caspase-3:1.65±0.05,Cyto Cyt C:1.46±0.06,Mito Cyt C:0.72±0.04;5.0 nmol/L组:bax/bcl-2:3.26±0.25,Cyt C:1.86±0.18,cleaved Caspase-3:1.92±0.24,Cyto Cyt C:1.84±0.21,Mito Cyt C:0.65±0.03;10.0 nmol/L组:bax/bcl-2:4.14±0.28,Cyt C:2.25±0.25,cleaved Caspase-3:2.38±0.28,Cyto Cyt C:2.26±0.25,Mito Cyt C:0.53±0.03;20.0 nmol/L组:bax/bcl-2:6.18±0.36,Cyt C:3.65±0.42,cleaved Caspase-3:3.42±0.36,Cyto Cyt C:2.98±0.32,Mito Cyt C:0.42±0.02),组间差异有统计学意义(P<0.05);与CON组比较[磷酸化AMPK(p-AMPK)/AMPK:1.00±0.06,p-p53/p53:1.00±0.05],二甲双胍治疗后p-AMPK/AMPK(2.5 nmol/L组:1.65±0.12;5.0 nmol/L组Objective It aims to assess the efficacy of metformin(a first-line therapeutic agent for type 2 diabetes mellitus)in intervening in cervical cancer and to provide a research basis for the development of effective new chemotherapeutic agents.Methods Caski cells were cultured in vitro and different concentrations of metformin(2.5,5.0,10.0,20.0 nmol/L)intervened in Caski cells,setting up a control(CON)group and metformin(2.5,5.0,10.0,20.0 nmol/L)intervention groups.The effects of metformin on proliferation,apoptosis,and ROS production in Caski cells were detected by cell counting kit-8(CCK-8)assay,clone formation assay,flow cytometry,and DCFH-DA probe,respectively.Western blotting was applied to detect proteins specific for mitochondrial dysfunction and adenosine monophosphate-actived protein kinase(AMPK)/p53 pathway expression.Results The results of cytological experiments showed that the A450 values were 0.98±0.10,0.85±0.10,0.58±0.08,0.46±0.04,0.38±0.03 in the CON group and the metformin(2.5,5.0,10.0,20.0 nmol/L)intervention groups,respectively,and that the differences between the groups were statistically significant(F=42.36,P<0.01);The cell apoptosis rates in the CON group and metformin(2.5,5.0,10.0,20.0 nmol/L)intervention groups were(3.85±0.35)%,(8.55±0.62)%,(12.50±1.02)%,(20.56±2.36)%,and(25.76±2.45)%,in that order,with statistically significant differences between groups(F=56.74,P<0.01);B cell lymphoma/leukemia-2 associated X protein(bax)/B cell lymphoma/leukemia-2(bcl-2)(1.00±0.05),Cyt C(1.00±0.04),cleaved cysteinyl aspartate-specific protease-3(cleaved Caspase-3)(1.00±0.03),Cyto Cyt C(1.00±0.03),and Mito Cyt C(1.00±0.05)concentrations in the CON group compared to metformin(2.5,5.0,10.0,20.0 nmol/L)intervention groups(2.5 nmol/L group:bax/bcl-2:1.95±0.09,Cyt C:1.45±0.06,cleaved Caspase-3:1.65±0.05,Cyto Cyt C:1.46±0.06,Mito Cyt C:0.72±0.04;5.0 nmol/L group:bax/bcl-2:3.26±0.25,Cyt C:1.86±0.18,cleaved Caspase-3:1.92±0.24,Cyto Cyt C:1.84±0.21,Mito Cyt C:0.65±0.03;10.0 nmol/L group:bax/b

关 键 词：宫颈癌 CASKI细胞 线粒体功能障碍 腺苷酸活化蛋白激酶/p53 

分 类 号：R96[医药卫生—药理学]