敲低核纤层蛋白B1通过抑制PI3K/AKT通路在肝癌中发挥抑癌作用  

Lamin B1 downregulation plays a tumor suppressive role on liver cancer by inhibiting the PI3K/Akt pathway

作　　者：展沙沙 范琪琪 李雪 Zhan Shasha;Fan Qiqi;Li Xue(Department of Geratric Medicine,The Sixth People’s Hospital of Qingdao,Qingdao 266011,Shandong Province,China;Department of Liver Disease,The Sixth People’s Hospital of Qingdao,Qingdao 266011,Shandong Province,China;Department of Medical Oncology,The Sixth People’s Hospital of Qingdao,Qingdao 266011,Shandong Province,China)

机构地区：[1]青岛市第六人民医院老年医学科,山东青岛266011 [2]青岛市第六人民医院肝病一科,山东青岛266011 [3]青岛市第六人民医院肿瘤内科,山东青岛266011

出　　处：《中国肝脏病杂志（电子版）》2023年第3期6-15,共10页Chinese Journal of Liver Diseases：Electronic Version

摘　　要：目的探讨核纤层蛋白B1(lamin B1,LMNB1)对肝癌进展的影响,发掘肝癌新的治疗靶点。方法利用工具网站基因表达水平值的交互式分析平台(Gene Expression Profiling Interactive Analysis,GEPIA)分析癌症基因组图谱数据库(The Cancer Genome Atlas,TCGA)中肝癌组织及癌旁组织LMNB1的表达水平,使用Kaplan-Meier分析LMNB1表达水平差异对肝癌患者预后的影响。采用实时定量聚合酶链式反应(polymerase chain reaction,PCR)检测正常肝细胞系及肝癌细胞系中LMNB1表达水平。选取LMNB1表达水平较高的两种人肝癌细胞株Hep3B、HepG2,转染小干扰RNA敲低LMNB1表达,在LMNB1表达水平较低的细胞SUN475中构建LMNB1稳定过表达细胞系并使用Western blot验证转染效率,采用CCK8法检测敲低或过表达LMNB1后肝癌细胞的增殖能力,采用平板克隆形成检测敲低或过表达LMNB1后细胞形成克隆的能力。采用Transwell检测敲低或过表达LMNB1后细胞侵袭及迁移能力的变化。采用Western blot实验检测磷酸化蛋白激酶B(phosphorylated protein kinase B,p-AKT)、蛋白激酶B(protein kinase B,AKT)、磷酸化细胞外调节蛋白激酶(phosphorylated extracellular regulated protein kinases,p-ERK)及细胞外调节蛋白激酶(extracellular regulated protein kinases,ERK)的变化。结果TCGA数据库分析表明肝癌组织中LMNB1表达水平较癌旁正常组织高(P<0.05),且LMNB1表达水平与肝癌患者预后呈负相关(P=0.003)。实时定量PCR显示相对正常人肝细胞系,肝癌细胞中LMNB1 mRNA表达量分别为MHCC97H:1.80±0.10(t=11.08,P<0.001);SUN475:1.29±0.03(t=5.88,P=0.004);HepG2:2.97±0.04(t=39.68,P<0.001);PLC/Prf/5:1.74±0.04(t=14.51,P<0.001);HuH7:1.70±0.02(t=15.22,P<0.001);SK-HEP-1:1.59±0.05(t=11.17,P<0.001);Hep3B:2.27±0.09(t=18.50,P<0.001);均显著高于正常人肝细胞系。Westernblot结果显示在HepG2及Hep3B细胞系中,与siScramble阴性对照组相比,转染siLMNB1实验组的LMNB1表达水平显著降低(HepG2:0.60±0.10 vs 1.6Objective To investigate the effect of LMNB1 on the progression of hepatocellular carcinoma(HCC),and to explore new therapeutic targets for HCC.Methods The expression level of LMNB1 in HCC tissues and normal tissues in The Cancer Genome Atlas(TCGA)database were analyzed by Gene Expression Profiling Interactive Analysis(GEPIA)website,and Kaplan-Meier curve was used to analyze the effect of LMNB1 expression on prognosis.Real-time quantitative polymerase chain reaction(PCR)was used to detect the expression level of LMNB1 in normal liver cell lines and HCC cell lines.Two human HCC cell lines Hep3B and HepG2 with higher expression levels of LMNB1 were selected and transfected with small interfering RNA to knockdown LMNB1 expression,LMNB1 stable overexpression cell lines were constructed in SUN475 cells with low level of LMNB1 expression and the transfection efficiency was verified by Western blot.The proliferation capacity of HCC cells after knockdown or overexpression LMNB1 was measured by CCK8 assay,and plate clone formation assay was used to detect the ability of clone formation of the cells.Western blot was used to detect the expression of phosphorylated protein kinase B(p-AKT),protein kinase B(AKT),phosphorylated extracellular regulated protein kinases(p-ERK)and extracellular regulated protein kinases(ERK).Results The results of TCGA database showed that the expression level of LMNB1 was higher in HCC tissues than that of normal tissues(P<0.05),and the expression level of LMNB1 was negatively correlated with the prognosis of patients with HCC(P=0.003).Real-time quantitative PCR showed that compared with the normal human liver cell lines,the mRNA expression level of LMNB1 in MHCC97H(1.80±0.10;t=11.08,P<0.001),SUN475(1.29±0.03;t=5.88,P=0.004),HepG2(2.97±0.04;t=39.68,P<0.001),PLC/Prf/5(1.74±0.04;t=14.51,P<0.001),HuH7(1.70±0.02;t=15.22,P<0.001),SK-HEP-1(1.59±0.05;t=11.17,P<0.001)and Hep3B(2.27±0.09;t=18.50,P<0.001)were significantly higher than those in normal human liver cell lines.Western blot showed that in

关 键 词：肝癌 核纤层蛋白B1 PI3K/AKT信号转导通路 

分 类 号：R735.7[医药卫生—肿瘤]