ABHD5过表达对结肠癌细胞侵袭、迁移及AMPK/mTOR通路的影响

Effects of ABHD5 overexpression on invasion,migration and AMPK/mTOR pathway in colon cancer cells

目的探讨含自水解酶域蛋白5(ABHD5)过表达对结肠癌HCT116细胞侵袭、迁移以及单磷酸腺苷活化蛋白激酶(AMPK)/雷帕霉素靶体蛋白(mTOR)通路的影响。方法应用UALCAN数据库资料分析ABHD5 mRNA在结肠癌组织中的表达及其与结肠癌患者临床病理特征之间的关系。体外培养结肠癌HCT116细胞,转染ABHD5重组质粒并分为对照组、阴性转染组和ABHD5转染组,采用实时荧光定量PCR(qRT-PCR)方法检测各组HCT116细胞中ABHD5 mRNA的表达;CCK-8法检测各组HCT116细胞增殖情况;Transwell法检测细胞侵袭和迁移能力;蛋白免疫印迹法检测侵袭蛋白基质金属蛋白酶9(MMP-9)、E-钙黏附蛋白(E-cadherin)、Snail、AMPK/mTOR通路蛋白p-AMPK、AMPK、p-mTOR和mTOR的表达情况。结果UALCAN数据库资料分析结果表明,与正常结肠组织比较,ABHD5 mRNA在结肠癌组织中的相对表达量降低,且其在腺癌和N1分期中分别低于黏液腺癌(P<0.05)及N0分期(P<0.05)。与对照组和阴性转染组比较,ABHD5转染组的ABHD5 mRNA表达升高(P<0.05),HCT116细胞增殖抑制率升高(P<0.05),HCT116细胞迁移和侵袭细胞数目减少(P<0.05),蛋白MMP-9、Snail、p-mTOR和mTOR表达降低,蛋白E-cadherin、p-AMPK和AMPK表达升高(P<0.05)。结论ABHD5过表达能够抑制结肠癌细胞HCT116的侵袭和迁移,且激活AMPK,抑制mTOR的表达,ABHD5可能通过影响AMPK/mTOR通路在抑制结肠癌的过程中发挥作用。

Objective To investigate the effects of overexpression of alpha/beta hydrolase domain-containing protein 5(ABHD5)on the invasion and migration of human colon cancer cell line HCT116 and the pathway of adenosine monophosphate-activated protein kinase(AMPK)/mechanistic target of rapamycin(mTOR).Methods The expression of ABHD5 in colon cancer tissues and its relationship with clinicopathological features was analyzed by UALCAN database.HCT116 cells were cultured in vitro and transfected with ABHD5 recombinant plasmid,then they were divided into control group,negative transfection group and ABHD5 transfection group.Real time quantitative PCR(qRT-PCR)was used to detect the expression of ABHD5 mRNA in HCT116 cells.The proliferation of HCT116 cells was detected by CCK-8 method.Transwell assay was used to detect the invasion and migration of HCT116 cells.The expression of matrix metalloprotein 9(MMP-9),E-cadherin,Snail,and AMPK/mTOR pathway proteins p-AMPK,AMPK,p-mTOR and mTOR were detected by Western blot.Results The results of the UALCAN showed that compared with normal colon tissues,the expression of ABHD5 mRNA in colon cancer tissues was decreased(P<0.05),and which in the adenocarcinoma and the N1 stage was lower than that of the mucinous adenocarcinoma(P<0.05)and N0 stage(P<0.05),respectively.Compared with the control group and the negative transfection group,the expression of ABHD5 mRNA in the ABHD5 transfection group was increased(P<0.05),the proliferation inhibition rate of HCT116 cells in the ABHD5 transfection group was increased(P<0.05),the numbers of migration and invasion cells in the ABHD5 transfection group were decreased(P<0.05),the expressions of MMP-9,Snail,p-mTOR and mTOR were reduced,and the expressions of E-cadherin,p-AMPK and AMPK were increased(P<0.05).Conclusions The overexpression of ABHD5 can inhibit the invasion and migration of colon cancer HCT116 cells,activate AMPK,and inhibit the expression of mTOR.It suggests that ABHD5 may play a role in inhibiting colon cancer by affecting AMPK/mTOR pathway.