NCAPH基因敲除对胰腺癌细胞增殖迁移及侵袭能力的影响  

Effect of NCAPH knockout on proliferation and invasion of pancreatic cancer cells

作　　者：李军强 杨志良 王彦 凡振玉[1] 关晓英[1] 王天成[1] Li Jun-Qiang;Yang Zhi-Liang;Wang Yan;Fan Zhen-Yu;Guan Xiao-Ying;Wang Tian-Cheng(Department of Neurology,Lanzhou University Second Hospital,Lanzhou 730000,China)

机构地区：[1]兰州大学第二医院神经内科,兰州730000

出　　处：《解放军医学杂志》2022年第6期555-560,共6页Medical Journal of Chinese People's Liberation Army

基　　金：甘肃省青年科技基金(20JR5RA323)。

摘　　要：目的探究NCAPH基因敲除对胰腺癌PANC细胞增殖、迁移及侵袭能力的影响。方法利用Oncomine数据库分析NCAPH在胰腺癌及其他癌组织中的表达水平;Kaplan-Meier plotter数据库分析NCAPH高低表达与胰腺癌患者预后的相关性;使用CRISPR/Cas9技术构建NCAPH基因敲除慢病毒建立NCAPH基因敲除细胞株;通过CCK-8实验、克隆形成实验、Transwell实验及划痕实验检测敲除NCAPH基因对细胞增殖、迁移及侵袭能力的影响,Western blotting检测MEK及ERK等蛋白的表达。结果NCAPH在胰腺癌、胃癌、结直肠癌及结肠癌组织中的表达均显著上调。Kaplan-Meierplotter数据库分析显示,NCAPH高表达组胰腺癌患者的预后不良(P<0.05);Western blotting检测结果显示,CRISPR/Cas9基因编辑技术能有效敲除胰腺癌PANC细胞系的NCAPH基因,从而抑制NCAPH蛋白的表达。CCK-8实验及克隆形成实验结果显示,与对照组相比,NCAPH敲除明显抑制了PANC细胞在48、72、96及120 h的增殖能力(0.488±0.007 vs.0.411±0.004、0.689±0.004 vs.0.497±0.010、1.071±0.034 vs.0.689±0.020、1.441±0.038 vs.0.855±0.025)及克隆形成能力(210.0±2.9 vs.144.0±16.4),差异有统计学意义(P<0.05);划痕实验及Transwell实验结果显示,与对照组相比,NCAPH敲除明显抑制了PANC细胞的迁移能力(34.9%±1.7%vs.15.1%±2.1%)及侵袭能力[(351±23.64)个vs.(194±13.0)个],差异有统计学意义(P<0.05)。与PANC正常细胞株相比,PANC敲除细胞株的p-ERK及p-MEK蛋白表达量明显降低,差异有统计学意义(P<0.05)。结论NCAPH敲除可明显抑制胰腺癌细胞的增殖、迁移及侵袭能力,其机制可能与MAPK/ERK信号通路有关。Objective The study explored the effect of the knockout of the NCAPH gene on the proliferation,migration and invasion of pancreatic cancer cells,using NCAPH knockout pancreatic cancer cells generated by CRISPR/Cas9 technology.Methods The expression levels of NCAPH were analyzed in multiple cancer tissues and normal tissues by the Oncomine database.Kaplan-Meier analysis was used to investigate the correlation between the expression of NCAPH and the survival of pancreatic cancer patients.NCAPH-knockout cells using lentivirus were produced using the CRISPR/Cas9 technique.The proliferationrelated molecules MEK and ERK were determined by Western blotting.CCK-8,colony formation,wound healing and Transwell assay were adopted to detect cell proliferation,migration and invasion of PANC cells.Western blotting detects the expression of MEK,p-MEK,ERK and p-ERK proteins.Results The results showed that NCAPH was significantly up regulated compared with paracancerous tissues in multiple cancer.Kaplan-Meier survival analysis revealed that patients with high expression of NCAPH were associated with a worse prognosis.The Western blotting showed that CRISPR/Cas9 technology efficiently disrupted the NCAPH gene and inhibited its expression in PANC cells.CCK-8 assay and colony formation assay showed that,compared with the control group,inhibiting the expression of NCAPH can inhibit the cell proliferation at 48,72,96 and 120 h(0.488±0.007 vs.0.411±0.004,0.689±0.004 vs.0.497±0.010,1.071±0.034 vs.0.689±0.020,1.441±0.038 vs.0.855±0.025)and the colony formation ability(210.0±2.9 vs.144.0±16.4),the difference was statistically significant(P<0.05).Cell scratch and Transwell invasion assay showed that,compared with the control group,NCAPH knockout significantly suppressed cell migration(34.9%±1.7%vs.15.1%±2.1%)and invasion[(351±23.64)cells vs.(194±13.0)cells],the difference was statistically significant(P<0.05).Then,we investigated the molecular mechanisms of this change,compared with the control group,NCAPH kno ckout significan

关 键 词：胰腺癌 NCAPH 细胞增殖 细胞迁移 

分 类 号：R735.9[医药卫生—肿瘤]