摘要
目的探讨过表达Klotho基因对胃癌细胞SGC-7901细胞增殖、迁移及磷酸肌醇-3激酶/蛋白激酶B(PI3K/AKT)通路的影响。方法体外培养胃癌细胞SGC-7901、正常胃黏膜细胞CES-1,采用实时荧光定量PCR(qRT-PCR)法和Western blot检测细胞Klotho mRNA和蛋白相对表达水平;将SGC-7901细胞分为Klotho组、阴性对照组(NC组)、空白对照组(BC组),构建Klotho过表达载体,转染细胞后检测细胞Klotho mRNA和蛋白相对表达水平;利用CKK-8法检测细胞增殖情况,利用Transwell实验检测细胞迁移、侵袭情况,利用Western blot检测PI3K/AKT通路相关蛋白表达情况。结果胃癌细胞SGC-7901中Klotho mRNA及蛋白相对表达水平较正常胃黏膜细胞CES-1明显降低(P<0.05)。转染后,Klotho组、NC组检测到绿色荧光细胞比例达90%;以上Klotho组细胞中Klotho mRNA及蛋白相对表达水平较BC组与NC组明显升高(P<0.05);Klotho组细胞增殖率较BC组与NC组明显降低(P<0.05);Klotho组迁移、侵袭细胞数较BC组与NC组明显降低(P<0.05);Klotho组细胞PI3K、AKT蛋白相对表达水平与BC组、NC组比较无明显差异(P>0.05);Klotho组细胞磷酸化AKT(p-AKT)、磷酸化PI3K(p-PI3K)、CD44、基质金属蛋白酶9(MMP9)蛋白相对表达水平较BC组、NC组明显降低(P<0.05)。结论过表达Klotho可抑制胃癌细胞SGC-7901细胞增殖、迁移、侵袭,其机制可能与抑制PI3K/AKT信号通路有关。
Objective To explore the effects of overexpression of Klotho gene on proliferation,migration and phosphoinositol-3 kinase/protein kinase B(PI3K/AKT)pathway in gastric cancer SGC-7901 cells.Methods The gastric cancer cell line SGC-7901 and the normal gastric mucosal cell line CES-1 were cultured in vitro,and the relative expression levels of Klotho mRNA and protein were detected by real-time fluorescence quantitative PCR(qRT-PCR)and Western blot,respectively.The SGC-7901 cells were divided into the Klotho group,the negative control group(NC group)and the blank control group(BC group).The Klotho overexpression vector was constructed,and the relative expression levels of Klotho mRNA and protein were detected after transfection.The CKK-8 method was used to detect cell proliferation,the Transwell assay was used to detect cell migration,and the expression levels of PI3K/AKT pathway-related proteins were detected by Western blot.Results The relative expression levels of Klotho mRNA and protein in gastric cancer cell SGC-7901 were significantly lower than those in normal gastric mucosal cell CES-1(P<0.05).After transfection,the proportion of green fluorescent cells in the Klotho group and the NC group was above 90%.The relative expression levels of Klotho mRNA and protein in the Klotho group were significantly higher than those in the BC group and the NC group(P<0.05);the cell proliferation rate of the Klotho group was significantly lower than that of the BC group and the NC group(P<0.05);the number of migrating and invasivecells in the Klotho group was significantly lower than that in the BC group and the NC group(P<0.05);the relative expression levels of PI3K and AKT proteins in the Klotho group were not significantly different from those in the BC group and the NC group(P>0.05),while the relative expression levels of phosphorylated AKT(p-AKT),phosphorylated PI3K(p-PI3K),CD44 and matrix metalloproteinase 9(MMP9)proteins in the Klotho group were significantly lower than those in the BC group and the NC group(P<0.05).Con
作者
蒋茂芬
李君强
刘春姣
王敏
JIANG Maofen;LI Junqiang;LIU Chunjiao;WANG Min(Department of Pathology,People′s Hospital of Beilun District,Ningbo,Zhejiang 315800,China;Wenzhou Medical University,Wenzhou,Zhejiang 325035,China)
出处
《重庆医学》
CAS
2020年第13期2065-2070,共6页
Chongqing medicine
