微小染色体维持蛋白5在肝癌组织的表达及其对肝癌细胞增殖、迁移的调节作用  

Expression of minichromosome maintenance 5 in hepatocellular carcinoma and its regulation on proliferation,migration and apoptosis in hepatocellular cancer cell line

作　　者：刘健康[1] 田鹏[1] 付强 刘传江 张宏伟 Liu Jiankang;Tian Peng;Fu Qiang;Liu Chuanjiang;Zhang Hongwei(Department of General Surgery,Henan Provincial People’s Hospital,Central China Fuwai Hospital,Central China Fuwai Hospital of Zhengzhou University,Zhengzhou 450003,China;Department of Hepato-Biliary-Pancreatic Surgery,Henan Provincial People’s Hospital,People’s Hospital of Zhengzhou University,Zhengzhou 450003,China)

机构地区：[1]河南省人民医院普外科华中阜外医院普外科郑州大学华中阜外医院普外科,郑州450003 [2]河南省人民医院/郑州大学人民医院肝胆胰腺外科,郑州450003

出　　处：《中华实验外科杂志》2020年第3期454-456,共3页Chinese Journal of Experimental Surgery

摘　　要：目的探讨微小染色体维持蛋白5(MCM5)在肝细胞肝癌(HCC)组织中的表达和预后的关系,观察其对HCC细胞的增殖、迁移的调节作用。方法根据肿瘤基因组图谱(TCGA)和GEO数据库,分析HCC组织和癌旁组织中MCM5的差异表达,并绘制生存曲线。将MCM5低表达的HCC细胞株作为实验组(HepG2、HuH7),以转染空载体的HCC细胞株为对照组(shRNAHepG2、shRNAHuH7)。实时定量反转录聚合酶链反应(RT-qPCR)、蛋白质印迹法(Western blot)技术检测MCM5在人HCC细胞株和正常肝细胞株的表达量。短发卡RNA(shRNA)干扰MCM5表达后,细胞计数试剂盒(CCK-8)实验、细胞划痕修复实验检测HCC细胞的增殖和迁移能力;流式细胞术检测HCC细胞的凋亡率。多组资料间比较采用单因素方差分析和t检验,计数资料的比较采用χ2检验。结果MCM5在HCC组织较癌旁组织表达升高5.32倍(t=4.464,P<0.05),MCM5高表达患者5年生存率(38%)较低表达患者(51%)明显下降,差异有统计学意义(t=5.337,P<0.05)。MCM5在HCC细胞株(HepG2、HuH7)的表达量(3.95±0.37、3.09±0.27)较人正常肝细胞(HL7702)的表达量(0.94±0.13)明显增高,差异有统计学意义(t=14.532、16.339,P<0.01)。24、48、72、96 h细胞增殖能力检测显示,实验组HepG2细胞(0.51±0.04、0.94±0.06、1.25±0.15、0.94±0.06)较对照组的HepG2细胞(0.68±0.06、1.21±0.09、1.95±0.12、1.21±0.09)显著下降,差异有统计学意义(t=7.328、6.997、8.779、11.338、12.067,P<0.05);实验组HuH7细胞增殖能力(0.48±0.05、0.76±0.13、1.44±0.25、1.44±0.25)较对照组的HuH7细胞(0.58±0.07、0.98±0.13、1.86±0.14、1.98±0.13)显著下降,差异有统计学意义(t=4.787、5.062、5.997、8.932,P<0.05)。实验组的HepG2细胞、HuH7细胞划痕愈合百分比[(19.14±0.98)%、(19.57±1.01)%]较对照组划痕愈合百分比[(45.29±1.35)%、(40.22±1.66)%]明显减少,差异有统计学意义(t=25.369、22.891,P<0.01)。实验组HepG2细胞、HuH7细胞凋亡率[(14.68±3.21)%、Objective To analyze the expression of minichromosome maintenance 5(MCM5)in hepatocellular carcinoma(HCC)tissues and its effects on the prognosis of HCC,and detect the regulation of MCM5 on the proliferation,migration and apoptosis of HCC cell lines.Methods Construct short hairpin RNA(shRNA)to decrease the expression of MCM5 in HepG2 and HuH7 cells.The different expression genes of HCC and adjacent normal tissues were detected in The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)databases.The the expression levels of MCM5 were examined in HCC cell lines(HepG2,HuH7)and normal cells line(HL7702)by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR)and Western blotting.The short hairpin RNA(shRNA)to decrease the expression of MCM5 in HepG2 and HuH7 cells was constructed.The proliferation,migration and apoptosis of cells were observed by cell counting kit-8(CCK-8)assays,wound healing assays and flow cytometry(FC)respectively.Results The expression of MCM5 in HCC tissue was 5.32 times higher than adjacent normal tissues(t=4.464,P<0.05).The 5-year survival rate in high expression group was 38%,which obviously decreased in comparison with 51%in low expression group(t=5.337,P<0.05).The expression of MCM5 in HepG2 and HuH7 cells was respectively(3.95±0.37)and(3.09±0.27),which was significantly higher than(0.94±0.13)in HL7702 cells(t=14.532,16.339,P<0.01).CCK-8 assays showed that the proliferation was inhibited 24,48,72 and 96 hafter downregulation of MCM5 in HepG2 and HuH7 cells.A values were(0.51±0.04,0.94±0.06,1.25±0.15,0.94±0.06)in downregulated HepG2 versus(0.68±0.06,1.21±0.09,1.95±0.12,1.21±0.09)in control HepG2(t=7.328,6.997,8.779,11.338,12.067,P<0.05),and(0.48±0.05,0.76±0.13,1.44±0.25,1.44±0.25)in downregulated HuH7 versus(0.58±0.07,0.98±0.13,1.86±0.14,1.98±0.13)in control HuH7(t=4.787,5.062,5.997,8.932,P<0.05).Wound healing essay showed the recovery rate of downregulated HepG2 and HuH7 cells was(19.14±0.98)%and(19.57±1.01)%respectively,which decreased sig

关 键 词：癌 肝细胞 微小染色体维持蛋白5 增殖 迁移 

分 类 号：R735.7[医药卫生—肿瘤]