基础医学与临床 ›› 2024, Vol. 44 ›› Issue (2): 185-191.doi: 10.16352/j.issn.1001-6325.2024.02.0185

• 研究论文 • 上一篇    下一篇

下调去甲基化酶FTO抑制人肝癌细胞系HepG2增殖

卢慧莹, 王建国*   

  1. 新乡医学院第一附属医院 肝胆外科,河南 新乡 453002
  • 收稿日期:2023-08-25 修回日期:2023-11-03 发布日期:2024-02-05
  • 通讯作者: *WangJianGuo5777@126.com
  • 基金资助:
    河南省医学科技攻关计划项目(SBGJ202103035)

Downregulation of demethylase FTO inhibits proliferation of human liver cancer cell line HepG2

LU Huiying, WANG Jianguo*   

  1. Deparment of Hepatobiliary Surgery, the First Affiliated Hospital of Xinxiang Medical College, Xinxiang 453002, China
  • Received:2023-08-25 Revised:2023-11-03 Published:2024-02-05
  • Contact: *WangJianGuo5777@126.com

摘要: 目的 探究去甲基化酶脂肪质量和肥胖相关基因(FTO)抑制对人肝癌细胞系HepG2增殖的作用机制。方法 将肝癌细胞系HepG2分为对照组(转染空质粒)、FTO组(转染FTO过表达质粒)、si-FTO组(转染si-FTO)、si-FTO+si-FoxO1组(同时转染si-FTO和si-FoxO1)。RT-qPCR检测细胞中FTO相对表达量;用CCK-8、Transwell小室法和流式细胞仪分别检测细胞增殖、侵袭和凋亡;Dot blot检测m6A甲基化水平;蛋白质印迹检测细胞中FoxO1蛋白表达。结果 通过人类癌基因库(TCGA)分析肝癌患者总体生存期发现FTO高表达提示更短的生存期(P<0.05)。和正常肝细胞系HL7702相比,HepG2细胞中FTO表达明显升高(P<0.05)。和对照组相比,si-FTO组细胞中FTO相对表达量明显降低,FTO组细胞中FTO相对表达量明显升高(P<0.05);和si-FTO组相比,si-FTO+si-FoxO1组细胞中FTO相对表达量明显升高(P<0.05)。si-FTO组细胞活性、侵袭细胞数、m6A相对表达量均低于对照组,细胞凋亡率、FoxO1蛋白表达均高于对照组(P<0.05);FTO组细胞活性、侵袭细胞数、m6A相对表达量明显高于对照组,细胞凋亡率、FoxO1蛋白表达明显低于对照组(P<0.05);si-FTO+si-FoxO1组细胞活性、侵袭细胞数目、m6A相对表达量均高于si-FTO组,细胞凋亡率、FoxO1蛋白表达均低于si-FTO组(P<0.05)。结论 高表达FTO与临床预后较差相关,去甲基化酶FTO敲减可抑制肝细胞癌的增殖和侵袭,诱导肝细胞癌的凋亡,其作用机制可能和调控FoxO1表达有关。

关键词: 去甲基化酶, 脂肪质量和肥胖相关蛋白(FTO), 叉头盒蛋白O1(FoxO1), 肝细胞肝癌, 凋亡

Abstract: Objective To explore the mechanism of the demethylase fat mass and obesity associatal(FTO) gene on the proliferation of human liver cancer cell line HepG2. Methods HepG2 cells were divided into the control group, FTO group (transfected with FTO over-expression plasmid), si-FTO group (transfected with si-FTO)and si-FTO+si-FoxO1 group (simultaneously transfected with si-FTO and si-FoxO1). The expression of FTO in cells was detected by RT-qPCR. Cell proliferation, invasion and apoptosis were examined using CCK-8 assay, Transwell chamber assay and flow cytometry, respectively. Dot blot assay was performed to measure m6A methylation , and protein expression of FoxO1 in cells was detected by Western blot. Results Analysis of overall survival in liver cancer patients using The Cancer Genome Atlas (TCGA) showed higher expression of FTO associated with shorter survival (P<0.05). Compared with normal liver cells HL7702, FTO relative expression was significantly increased in human liver cancer cells HepG2 (P<0.05).The relative expression of FTO in si-FTO group cells was lower than that in the control group, while the relative expression of FTO in FTO group was higher than that in the control group (P<0.05). The relative expression of FTO in the si-FTO+si-FoxO1 group was higher than that in the si-FTO group(P<0.05). Compared with the Control group, cell viability, count of invasive cells, relative level of m6A were significantly decreased, while apoptosis rate and protein expression of FoxO1 were significantly increased in the si-FTO group(P<0.05). Cell viability, count of invasive cells, and relative expression level of m6A were significantly higher, while apoptosis rate and protein expression of FoxO1 were significantly lower in the FTO group compared to the Control group (P<0.05). Compared with the si-FTO group, cell viability, invasive cells and relative level of m6A were significantly increased, while apoptosis rate and protein expression of FoxO1 were significantly decreased in the si-FTO+si-FoxO1 group(P<0.05). Conclusion High expression of FTO is associated with poor clinical prognosis. Knockdown of the demethylase FTO gene inhibits proliferation and invasion of liver cancer cells and induces their apoptosis. The mechanism is potentially related to the regulation of FoxO1.

Key words: demethylase, fat mass and obesity associated(FTO) protein, forkhead box protein 1(FoxO1), hepatocellular carcinoma, apoptosis

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