SKP2基因表达水平对肝癌细胞放射敏感性的影响  

The expression level of SKP2 gene affects the radiosensitivity in hepatocellular carcinoma cells

作　　者：杨伟芳[1,2,3] 王学全 夏鑫杭 冉刚 王微[1,2] 周佳慧[1] 杨海华 皮文虎 Yang Weifang;Wang Xuequan;Xia Xinhang;Ran Gang;Wang Wei;Zhou Jiahui;Yang Haihua;Pi Wenhu(Department of Radiation Oncology,Taizhou Hospital of Zhejiang Province,Linhai 317000,China;Key Laboratory of Radiation Oncology of Taizhou,Linhai 317000,China;Key Laboratory of Minimally Invasive Techniques&Rapid Rehabilitation of Digestive System Tumor of Zhejiang Province,Linhai 317000,China)

机构地区：[1]浙江省台州医院放疗科,临海317000 [2]台州市放射肿瘤学重点实验室,临海317000 [3]浙江省消化系肿瘤微创诊治与快速康复研究重点实验室,临海317000

出　　处：《中华放射肿瘤学杂志》2022年第10期922-927,共6页Chinese Journal of Radiation Oncology

基　　金：浙江省医药卫生科技项目(2022KY1382);浙江省台州市科技计划项目(1901ky07)。

摘　　要：目的研究人肝癌组织中S期激酶相关蛋白2(SKP2)表达与预后的关系以及肝癌细胞中SKP2表达对人肝癌细胞放射敏感性的影响。方法使用TCGA数据库中肝癌组织和正常对照组织的测序结果,分析SKP2基因在肝癌组织中的表达情况及其与患者预后的关系。使用Western blot检测肝癌细胞放射照射前后SKP2蛋白水平表达变化。利用CRISPR/Cas9技术敲除肝癌细胞SKP2基因的启动子和第1外显子,建立SKP2基因敲除的PLC/PRF/5和Hep3B肝癌细胞系模型,并进一步进行放射照射。用克隆形成实验检测SKP2基因敲除细胞的放射敏感性变化情况。结果对TCGA数据库分析结果显示,与正常组织比较,SKP2基因在肝癌组织中表达升高。对SKP2高、低水平肝癌患者总体生存率进行K-M法分析,结果显示SKP2高表达肝癌患者预后相对较差,其5年生存率为34.6%,SKP2低表达肝癌患者则为50.6%(HR=2.18,95%CI为1.46~3.27,P<0.001)。体外细胞实验显示肝癌细胞受到放射照射后SKP2表达水平明显升高。在CRISPR/Cas9敲除SKP2^(-/-)肝癌细胞中,克隆形成实验结果显示SKP2^(-/-)肝癌细胞的放射敏感性较SKP2+/+明显增加。结论SKP2基因在肝癌组织中表达升高,且高表达者较低表达者预后差。放射可上调PLC肝癌细胞SKP2的表达水平,而敲除SKP2后其放射敏感性明显增加。Objective To investigate the effect of S-phase kinase-associated protein 2(SKP2)expression level on radiosensitivity of human hepatocellular carcinoma(HCC)cells and the correlation of SKP2 expression with clinical prognosis of patients with HCC.Methods The expression levels of SKP2 gene in liver cancer tissues and normal tissues were validated and its correlation with clinical prognosis of HCC patients was analyzed based on the TCGA database.Western blot was used to determine the SKP2 protein levels in HCC cell lines before and after radiation.CRISPR/Cas9 technology was employed to delete the promoter and first exon of SKP2 gene in PLC/PRF/5(PLC)and Hep3B HCC cells for generating the SKP2 knockout cell lines.The difference of radiosensitivity and cell survival rate between normal(SKP2+/+)and SKP2 knockout(SKP2^(-/-))HCC cells was determined by using cell clonogenic assay and CCK8 kit.Results Compared with normal tissues,the expression levels of SKP2 gene in HCC were increased based on the results of TCGA database analysis.K-M analysis showed that the HCC patients with high SKP2 expression had relatively poor prognosis.The 5-year overall survival(OS)was 34.6%in high SKP2 expression HCC patients and 50.6%in low SKP2 expression HCC patients,respectively(HR=2.18,95%CI=1.46-3.27,P<0.001).In vitro experiment showed that the expression levels of SKP2 were significantly increased after radiation in HCC cells.Simultaneously,deletion of SKP2 significantly increased the radiosensitivity of HCC cells.Conclusion The expression level of SKP2 gene is increased in HCC patients,and patients with high SKP2 expression have worse prognosis than those with low expression.Radiation can upregulate the SKP2 expression levels in HCC cells,while the radiosensitivity of the cells is significantly increased after SKP2 deletion.

关 键 词：S期激酶相关蛋白2 肝肿瘤 放射 放射敏感性 

分 类 号：R735.7[医药卫生—肿瘤]