PON1基因低表达对肝癌细胞增殖和细胞克隆能力的影响  

Effects of PON1 gene low expression on proliferation and cell cloning capacity of hepatocellular carcinoma cells

作　　者：杨刚[1,2] 李永继 熊永福 唐涛[1] 李伟男[1] 李敬东[1] YANG Gang;LI Yongji;XIONG Yongfu;TANG Tao;LI Weinan;LI Jingdong(Department of Hepatobiliary Surgery,Affiliated Hospital of North Sichuan Medical College/Institute of Hepatobiliary Pancreas Intestines,Nanchong,Sichuan 637000,China;Department of Hepatobiliary Surgery,Affiliated Chengdu Xinhua Hospital,North Sichuan Medical College,Chengdu,Sichuan 610000,China)

机构地区：[1]川北医学院附属医院肝胆外科/川北医学院肝胆胰肠研究所,四川南充637000 [2]川北医学院附属成都新华医院肝胆外科,成都610000

出　　处：《重庆医学》2023年第22期3391-3396,共6页Chongqing medicine

基　　金：四川省医学会医学科研青年创新课题(Q21027)。

摘　　要：目的探讨对氧磷酶1(PON1)低表达对肝癌细胞增殖和细胞克隆能力的影响。方法提取肝癌组织及癌旁正常组织标本RNA测定PON1 mRNA的相对表达水平,ELISA法检测肝癌患者和健康人血清PON1蛋白水平。培养人肝癌细胞系PLC和Huh7,对PLC和Huh7细胞系进行转染,分为对照组和转染组(转染siPON1小干扰RNA进行基因沉默实验),使用实时荧光定量PCR及ELISA法验证转染效率,CCK-8实验检测转染后肝癌癌细胞增殖能力的变化,克隆形成实验检测细胞形成克隆的能力,流式细胞术检测细胞周期,Western blot实验检测AMP活化蛋白激酶(AMPK)信号通路相关蛋白变化。结果肝癌组织中PON1 mRNA相对表达水平低于癌旁正常组织(P<0.05),肝癌患者血清PON1蛋白水平低于健康人(P<0.05)。与对照组比较,转染组PON1 mRNA相对表达水平和PON1蛋白水平降低(P<0.05),细胞增殖能力明显增强(P<0.05),细胞克隆形成率明显增多(P<0.05),S期细胞比例明显增多(P<0.05),p-AMPK表达水平明显降低,p-mTOR表达水平明显升高。结论PON1表达调控了AMPK信号通路活化并抑制了肝癌细胞增殖。Objective To investigate the effect of PON1 low expression on the proliferation and cell cloning capacity of hepatocellular carcinoma cells.Methods RNA of hepatocellular carcinoma tissues and paracancerous normal tissues samples was extracted to detect the PON1 mRNA relative expression level.The serum PON1 protein level in the patients with hepatocellular carcinoma and healthy person was detected by ELISA.The human hepatocellular cancer cell line PLC and Huh7 were cultured.PLC and Huh7 cells conducted the transfection,then the cells were divided into the control group and transfection group(transfecting siPON1 small interfering RNA to conduct the gene silencing experiment).The transfection efficiency was verified by real time fluorescence quantification PCR and ELISA.The CCK-8 assay was used to detect the change of liver cancer cells proliferation ability after transfection,and the clone formation assay was used to detect the ability of cells forming clone.The cell cycle was detected by flow cytometry.The AMPK signaling pathway related protein change was detected by Western blot.Results The PON1 mRNA relative expression level in hepatocellular carcinoma tissues was lower than that in paracancerous tissues.The serum PON1 protein level in the patients with hepatocellular carcinoma was lower than that in healthy people(P<0.05).Compared with the control group,the PON1 relative expression level and PON1 protein level in the transfection group were decreased(P<0.05),the cellular proliferation ability was significantly enhanced(P<0.05),the cell clone forming rate was significantly increased(P<0.05),the proportion of S stage cells was significantly in-creased(P<0.05),the p-AMPK expression level was significantly decreased,and the p-mTOR expression level was significantly increased.Conclusion The PON1 expression regulates the activization of AMPK signaling pathway and inhibits the proliferation of hepatocellular carcinoma cells.

关 键 词：肝细胞癌 对氧磷酶1 细胞增殖 AMPK信号通路 生物标志物 

分 类 号：R735.7[医药卫生—肿瘤]