千金藤素基于Akt/mTOR通路诱导宫颈癌细胞自噬  

Cepharanthine induces autophagy in cervical cancer cells based on Akt/mTOR pathway

作　　者：郭洁[1] 李晶[1] 汤丽丽 刘蓉[2] GUO Jie;LI Jing;TANG Lili;LIU Rong(The Third Department of Women of the First Hospital of Handan City,Handan 056002,China;Department of Anesthesiology,Textile Hospital,Handan First Hospital,Handan 056002,China)

机构地区：[1]邯郸市第一医院妇三科,邯郸056002 [2]邯郸市第一医院纺医院麻醉科,邯郸056002

出　　处：《西北药学杂志》2023年第4期53-58,共6页Northwest Pharmaceutical Journal

基　　金：2021年河北省医学科学研究课题计划项目(编号:20210990)。

摘　　要：目的探讨千金藤素基于蛋白激酶B(protein kinase B,Akt)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)通路诱导宫颈癌细胞自噬的作用机制。方法取人宫颈癌HeLa细胞株进行培养、传代,千金藤素低剂量、中剂量及高剂量组分别以15、30、60μmol·L^(-1)千金藤素处理,对照组以等量生理盐水处理。检测细胞增殖活性、细胞凋亡率及自噬溶酶体的数量;检测微管相关蛋白1轻链3-Ⅱ(microtubule-associated protein 1 light chain 3Ⅱ,LC3-Ⅱ)、Bcl-2同源结构域蛋白(Bcl-2 homologous domain protein,Beclin1)及p62 mRNA的表达;检测细胞磷酸化丙酮酸脱氢酶激酶同工酶1(phosphorylated pyruvate dehydrogenase kinase 1,p-PDK1)、磷酸化Akt(phosphorylated Akt,p-Akt)、磷酸化磷脂酰肌醇-3激酶(phosphorylated phosphatidylinositol 3-kinase,p-PI3K)及磷酸化mTOR(phosphorylated mTOR,p-mTOR)蛋白的相对表达量。结果与对照组比较,千金藤素3组细胞的增殖活性降低,细胞凋亡率升高,且比较细胞增殖活性,高剂量组<中剂量组<低剂量组,比较细胞凋亡率,低剂量组<中剂量组<高剂量组(P<0.05);AO染色结果显示,与对照组比较,千金藤素3组亮红色荧光比例升高,自噬溶酶体的数量增多,自噬程度增强,且低剂量组<中剂量组<高剂量组;与对照组比较,千金藤素3组LC3-Ⅱ/LC3-Ⅰ、Beclin1 mRNA的表达水平升高,p62 mRNA的表达水平及p-PDK1/PDK1、p-Akt/Akt、p-PI3K/PI3K和p-mTOR/mTOR蛋白的相对表达量降低,且比较LC3-Ⅱ/LC3-Ⅰ、Beclin1 mRNA表达量,低剂量组<中剂量组<高剂量组,比较p62 mRNA表达及p-PDK1/PDK1、p-Akt/Akt、p-PI3K/PI3K和p-mTOR/mTOR蛋白的相对表达量,高剂量组<中剂量组<低剂量组(P<0.05)。结论千金藤素可诱导宫颈癌细胞自噬,降低细胞的增殖能力,促进细胞凋亡,其作用机制可能与抑制PDK1介导的Akt/mTOR信号通路有关。Objective To investigate the role of cepharanthine in inducing autophagy of cervical cancer cells based on protein kinase B(Akt)/mammalian target of rapamycin(mTOR)pathway.Methods Human cervical cancer HeLa cell lines were cultured and sub-cultured.The low,medium and high dose cepharanthine groups were treated with 15,30 and 60μmol·L^(-1) cepharanthine,respectively,and the control group was treated with the same amount of normal saline.The cell proliferation activity,apoptosis rate and autophagy lysosome number were detected.The expression of microtubule-associated protein 1 light chain3-Ⅱ(LC3-Ⅱ),Bcl-2 homologous domain protein(Beclin1)and p62 mRNA were detected.The relative expressions of phosphorylated pyruvate dehydrogenase kinase 1(p-PDK1),phosphorylated Akt(p-Akt),phosphorylated phosphatidylinositol 3-kinase(p-PI3K)and phosphorylated mTOR(p-mTOR)were detected.Results Compared with the control group,the cell proliferation activity decreased and the apoptosis rate increased in cepharanthine 3 groups,the cell proliferation activity was high dose group<medium dose group<low dose group,the apoptosis rate was low dose group<medium dose group<high dose group(P<0.05).AO staining results showed that compared with the control group,the proportion of bright red fluorescence increased,the number of autophagic lysosomes increased,and the degree of autophagy increased in cepharanthine 3 groups,and low dose group<medium dose group<high dose group.Compared with the control group,the expression of LC3-Ⅱ/LC3-Ⅰand Beclin1 mRNA increased,p62 mRNA and the relative expression of p-PDK1/PDK1,p-Akt/Akt,p-PI3K/PI3K and p-mTOR/mTOR protein decreased in cepharanthine 3 groups,the expression of LC3-Ⅱ/LC3-Ⅰand Beclin1 mRNA were low dose group<medium dose group<high dose group,the p62 mRNA expression and relative expression of p-PDK1/PDK1,p-Akt/Akt,p-PI3K/PI3K and p-mTOR/mTOR proteins were high dose group<medium dose group<low dose group(P<0.05).Conclusion Cepharanthine can induce autophagy,reduce cell proliferation and pro

关 键 词：千金藤素 宫颈癌 细胞自噬 蛋白激酶B 哺乳动物雷帕霉素靶蛋白 

分 类 号：R965[医药卫生—药理学]