启膈散对食管癌EC9706细胞miR-133a/PI3K/Akt信号通路甲基化的影响  

Interventional effect of Qigesan on miR-133a/PI3K/Akt signaling pathway methylation in esophageal cancer EC9706 cells

作　　者：王蕊 高小玲[1] 李丹丹[1] 魏雨濛 李艳坤[1] 陈玉龙[1] WANG Rui;GAO Xiao-ling;LI Dan-dan;WEI Yu-meng;LI Yan-kun;CHEN Yu-long(Henan University of Chinese Medicine,Zhengzhou,Henan 450046 China)

机构地区：[1]河南中医药大学,河南郑州450046

出　　处：《时珍国医国药》2023年第3期516-520,共5页Lishizhen Medicine and Materia Medica Research

基　　金：国家自然科学基金(81803981);河南省科技研发计划联合基金(222301420073)。

摘　　要：目的观察启膈散乙酸乙酯提取物对食管癌EC9706细胞micoRNA-133a(miR-133a)/磷脂酰肌醇-3-激酶(PI3K)/蛋白激酶B(Akt)信号通路甲基化的干预作用。方法流式细胞术检测启膈散对EC9706细胞的凋亡的影响;蛋白印迹免疫法(Westernblotting)分别检测采用Akt抑制剂、转染miR-133a模拟物及DNMT1抑制剂后启膈散对DNMT1蛋白表达的影响;实时荧光定量聚合酶链式反应(Real-time PCR)检测启膈散对miR-133a及IGF-1R mRNA表达量的影响。结果与空白对照组比较,启膈散组可增加EC9706晚期凋亡率及总凋亡率(P<0.05),与DNMT1抑制剂组相比,启膈散组、联合用药低剂量组晚期凋亡率差异显著(P<0.05)。与空白对照组比较,启膈散及各联合用药组均能够抑制食管癌EC9706细胞DNMT1蛋白的表达(P<0.05);启膈散与PI3K/AKT信号通路抑制剂联合用药组、启膈散与转染miR-133a模拟物联合用药组均对DNMT1蛋白的抑制呈现一定程度的的协同增效作用。与空白对照组相比,启膈散组、DNMT1抑制剂组以及联合用药组均能够有效提高miR-133a的表达(P<0.05),其中联合用药低剂量组呈现了竞争抑制作用。与空白对照组相比,启膈散组及联合用药低剂量组能够促进IGF-1R mRNA的表达(P<0.05)。结论启膈散能通过调控食管癌EC9706细胞miR-133a/PI3K/Akt信号环路甲基化,促进miR-133a的表达,并抑制PI3K/Akt信号通路的激活,影响食管癌EC9706细胞的生长。Objective To observe the intervention effect of ethyl acetate extract of Qigesan on methylation of micoRNA-133a(miR-133a)/phosphatidylinositol-3-kinase(PI3K)/protein kinase B(Akt)signal pathway in esophageal cancer EC9706 cells.Methods Flow cytometry was used to detect the effect of Qigesan on the apoptosis of EC9706 cells;Western blot immunoassay(WB)was used to detect the effect of Qigesan on the expression of DNMT1 protein after Akt inhibitor,miR-133a mimic and DNMT1 inhibitor were transfected;and the effect of Qigesan on the expression of miR-133a and IGF-1RmRNA was detected by real-time fluorescence quantitative polymerase chain reaction(Real-time PCR).Results Compared with the blank control group,Qigesan group could increase the late apoptosis rate and total apoptosis rate of EC9706(P<0.05);Compared with DNMT1 inhibitor group,the late apoptosis rate in Qigesan group and combined low dose group was significantly different(P<0.05).Compared with the blank control group,Qigesan and each combination group could inhibit the expression of DNMT1 protein in esophageal cancer EC9706 cells(P<0.05);Qigesan combined with PI3K/Akt signal pathway inhibitor group,Qige San combined with transfected miR-133a mimic group showed synergistic effect on the inhibition of DNMT1 protein to a certain extent.Compared with the blank control group,Qigesan group,DNMT1 inhibitor group and combination group could effectively increase the expression of miR-133a(P<0.05),and the low dose group showed competitive inhibition.Compared with the blank control group,Qigesan group and combined low-dose group could promote the expression of IGF-1R mRNA(P<0.05).Conclusion Qigesan can regulate the methylation of miR-133a/PI3K/Akt signal loop in esophageal cancer EC9706 cells,promote the expression of miR-133a,inhibit the activation of PI3K/Akt signal pathway,and affect the growth of esophageal cancer EC9706 cells.

关 键 词：启膈散 食管癌 甲基化 miR-133a/PI3K/AKT信号通路 

分 类 号：R285.5[医药卫生—中药学]