人肝癌细胞系HepG2和Huh7中的基因组拷贝数变异分析

Analysis of genomic copy number variations in human hepatocellular carcinoma cell lines HepG2 and Huh7

目的利用人肝细胞癌拷贝数变异和转录组实验,结合临床公共数据,探索肝细胞癌的拷贝数变异对肝癌发生发展的影响。方法用光学基因组图谱技术识别肝癌细胞系HepG2和Huh7中的拷贝数变异,随后分析两种细胞系拷贝数变异基因的功能,并根据富集通路绘制蛋白质相互作用网络。选取两个细胞系核心网络中的关键基因,分析肝细胞癌拷贝数变异与基因表达之间的关系。GEPIA数据库分析基因表达与患者临床生存的关系。用RNA-seq实验和公共数据验证基因的表达水平。结果HepG2细胞主要存在拷贝数增加,相关基因富集在雌激素信号通路、金黄色葡萄球菌感染等通路;Huh7细胞系中既有拷贝数增加又有减少,相关基因主要富集嗅觉传导、细胞因子-细胞因子受体相互作用等通路。关键基因SRC、MAPK3和MAP3K7拷贝数与基因表达成正比,并且高表达这些基因的患者生存期显著降低(P<0.05)。相比于HEK293T细胞系,SRC、MAP3K7基因在两个肝癌细胞系的表达显著升高(P<0.001),提示了肝细胞癌的特异性变异,MAPK3无差异。结论肝细胞癌拷贝数变异基因SRC、MAP3K7的表达与患者的预后显著相关,可能影响肝细胞癌的发生发展和异质性。

Objective To explore the effect of copy number variation on the occurrence and development of hepatocellular carcinoma by using copy number variation and transcriptome experiment of hepatocellular carcinoma combined with public clinical data.Methods The copy number variation found in hepatoma cell lines HepG2 and Huh7 was identified by optical genome mapping.The function of the copy number variant genes in the two cell lines was analyzed,and the protein interaction network was mapped according to the enrichment pathway.Key genes in the core network of two cell lines were selected to analyze the relationship between copy number variation and gene expression in hepatocellular carcinoma.The relationship between gene expression and clinical survival was analyzed by GEPIA database.RNA-seq assay and public data were used to verify gene expression levels.Results HepG2 cells mainly showed increased copy number,and related genes were enriched in estrogen signaling pathway and Staphylococcus aureus infection pathway.Huh7 cells showed both increased and decreased copy number,and related genes mainly concentrated in olfactory conduction and cytokine-cytokine receptor interaction pathways.The copy number of key genes SRC,MAPK3 and MAP3K7 was proportional to gene expression,and survival was significantly reduced in patients with high expression of these genes(P<0.05).Compared with HEK293T cell line,the expression of SRC and MAP3K7 genes in the two hepatocellular carcinoma lines was significantly increased(P<0.001),suggesting the specific variation of hepatocellular carcinoma.MAPK3 had no difference.Conclusions The expression of copy number variant genes SRC and MAP3K7 in hepatocellular carcinoma is significantly correlated with the prognosis of patients,and may significantly affect the development and heterogeneity of hepatocellular carcinoma.