miR-203过表达对结肠癌HT-29细胞紫杉醇化疗敏感性的影响及可能机制  

Influence of miR-203overexpression on paclitaxel chemotherapy sensitivity to colon cancer cells and its mechanism

作　　者：王康康 汪秀梅 张茂华 罗金键 吴慧丽 WANG Kang-kang;WANG Xiu-mei;ZHANG Mao-hua;LUO Jin-jian;WU Hui-li(Department of Gastroenterology,Zhengzhou Central Hospital of Zhengzhou University,Zhengzhou,Henan 450007,China)

机构地区：[1]郑州大学附属郑州中心医院消化内科,河南郑州450007

出　　处：《中华实用诊断与治疗杂志》2021年第3期220-225,共6页Journal of Chinese Practical Diagnosis and Therapy

基　　金：河南省高等学校重点科研项目计划(19A320079)。

摘　　要：目的采用脂质体转染法制备miR-203过表达结肠癌HT-29细胞,探讨miR-203过表达对结肠癌HT-29细胞紫杉醇化疗敏感性的影响及可能机制。方法取对数生长期结肠成纤维细胞CCD-18Co和结肠癌HT-29细胞,采用实时荧光定量PCR法检测miR-203相对表达量。取对数生长期结肠癌HT-29细胞,分为过表达组(转染miR-203-pcDNA 3.1质粒)、空载组(转染NC-pcDNA3.1质粒)和对照组(不作任何干预),采用实时荧光定量PCR法检测3组转染48h时miR-203相对表达量。取转染48h时过表达组、空载组、对照组细胞,用1、5、10、50、100μmol/L紫杉醇处理细胞48h,采用MTT法测定细胞增殖抑制率,并计算3组紫杉醇对细胞的半数抑制浓度(half inhibitory concentration,IC_(50))。转染48h,取对照组细胞分为对照1组和紫杉醇组,取过表达组细胞分为过表达1组和过表达+紫杉醇组,紫杉醇组和过表达+紫杉醇组应用对照组IC_(50)值(28.20μmol/L)浓度的紫杉醇处理,对照1组和过表达1组不进行处理,48h后采用流式细胞术检测4组细胞凋亡,采用Western blot法检测4组细胞磷脂酰肌醇-3激酶(phosphatidylinositol-3 kinase,PI3K)、蛋白激酶B(protein kinase B,Akt)、磷酸化Akt、caspase-3蛋白相对表达量。结果结肠癌HT-29细胞miR-203相对表达量(0.38±0.05)低于CCD-18Co细胞(1.01±0.09)(P<0.05)。转染48h,过表达组miR-203相对表达量(1.87±0.11)高于空载组(0.39±0.04)和对照组(0.40±0.05)(P<0.05),空载组与对照组比较差异无统计学意义(P>0.05)。1、5、10、50、100μmol/L紫杉醇处理48h,过表达组细胞增殖抑制率高于空载组和对照组(P<0.05),IC_(50)值低于空载组和对照组(P<0.05),空载组与对照组比较差异无统计学意义(P>0.05)。紫杉醇处理48h,过表达1组、紫杉醇组、过表达+紫杉醇组细胞凋亡率[(18.25±1.70)%、(19.11±1.83)%、(57.59±4.31)%]高于对照1组[(8.10±1.14)%](P<0.05),过表达+紫杉醇组高于过表达1组和紫杉醇组(P<0.05),过表�Objective To prepare the miR-203overexpressed colon cancer HT-29cells by liposome mediated transfection and to investigate the influence and mechanism of miR-203overexpression on paclitaxel chemotherapy sensitivity to colon cancer cells.Methods Real-time fluorescence quantitative PCR was used to detect the relative expression of miR-203in colon fibroblast cell lines and colon cancer HT-29cell lines in logarithmic growth phase.The HT-29cells in logarithmic growth phase was divided into overexpression group which was transfected with miR-203-pcDNA3.1plasmid,empty vector group which was transfected with NC-pcDNA3.1plasmid,and control group receiving no intervention.The relative expression of miR-203was detected after 48hof transfection in three groups by real-time fluorescence quantitative PCR.The cells in three groups were treated with 1,5,10,50and 100μmol/L of paclitaxel for 48h,the cell proliferation inhibition rate was measured by MTT method,and the half inhibitory concentration(IC_(50))of paclitaxel on the cells was calculated.Control group was divided into control group 1and paclitaxel group,and overexpression group was divided into overexpression group 1and the overexpression+paclitaxel group after 48hof transfection.Paclitaxel group and overexpression+paclitaxel group were treated with 28.20μmol/L of paclitaxel(control IC_(50)value).Control group 1and overexpression group 1received no treatment.The apoptosis was detected by flow cytometry in control group,overexpression group,paclitaxel group,overexpression+paclitaxel group after the treatment with paclitaxel for 48h,and the relative expressions of phosphatidylinositol-3kinase(PI3K),protein kinase B(Akt),phosphorylated Akt and caspase-3proteins were detected in four groups by Western blot.Results The relative expression of miR-203was lower in colon cancer HT-29cells(0.38±0.05)than that in CCD-18Co cells(1.01±0.09)(P<0.05).After 48hof transfection,the relative expression of miR-203was higher in overexpression group(1.87±0.11)than that in enmpty vector gr

关 键 词：结肠癌 HT-29细胞 miR-203 紫杉醇 磷脂酰肌醇-3激酶 蛋白激酶B caspase-3 

分 类 号：R735.35[医药卫生—肿瘤]