基于生物信息学分析CCN4在肝细胞癌中表达及其临床意义

doi:10.3877/cma.j.issn.2095-3232.2023.06.020

目的

基于生物信息学探讨细胞通信网络因子4（CCN4）基因在肝细胞癌（肝癌）中的表达及其临床意义。

方法

通过TCGA数据库分析肝癌组织中CCN4蛋白表达及其与临床病理参数的关系，利用肿瘤免疫评估资源（TIMER）数据库分析CCN4蛋白表达与肿瘤纯度和免疫细胞浸润的相关性。利用Kaplan-Meier Plotter数据库分析CCN4蛋白在肝癌中的表达与生存预后之间的关系。利用STRING数据库构建蛋白质相互作用（PPI）网络，并检验蛋白之间的相关性。CCN4表达与临床病理参数之间的关系分析采用χ²检验，相关性分析采用Spearman秩相关。

结果

提取TCGA肝癌数据集，共获得371例患者临床信息和相应RNA测序表达谱数据，数据分析显示，肝癌患者CCN4蛋白表达与种族、T分期、病理G分级有关（χ²=9.055，14.012，17.163；P<0.05）。Kaplan-Meier Plotter数据库生存分析显示，肝癌CCN4高表达患者总体生存期较短（HR=1.50，95%CI：1.05~2.14，P<0.05）。TIMER数据库分析显示，肝癌CCN4表达与肿瘤纯度呈负相关（r_s=-0.397，P<0.05），而与B细胞、CD8⁺ T细胞、CD4⁺ T细胞、巨噬细胞、中性粒细胞、树突细胞、M2巨噬细胞浸润水平呈正相关（r_s=0.306，0.279，0.460，0.490，0.465，0.404，0.532；P<0.05）。PPI网络分析显示，与CCN4主要相互作用的蛋白是核心蛋白聚糖（score=0.953）。

结论

CCN4蛋白表达与种族、T分期、病理G分级有关，可作为肝癌预后不良的生物标志物，机制可能与免疫细胞浸润、M2巨噬细胞极化及基质纤维形成相关。

关键词: 癌，肝细胞, CCN4基因, WISP1基因, 生物标志物, 生物信息学

Objective

To investigate the expression level and clinical significance of cellular communication network factor 4 (CCN4) gene in hepatocellular carcinoma (HCC) based on bioinformatics.

Methods

The expression level of CCN4 protein in HCC tissues and its relationship with clinicopathological parameters were analyzed by TCGA database. The correlation between the expression of CCN4 protein and tumor purity and immune cell infiltration was analyzed by TIMER database. Kaplan-Meier Plotter database was used to analyze the relationship between the expression of CCN4 protein and clinical prognosis in HCC. Protein-protein interaction (PPI) network was constructed by using STRING database, and the correlation between proteins was assessed. The relationship between CCN4 expression and clinicopathological parameters was analyzed by Chi-square test. Correlation analysis was conducted by Spearman's rank correlation.

Results

Clinical information and corresponding RNA sequencing data of 371 patients were obtained from TCGA HCC dataset. Data analysis showed that the expression of CCN4 protein in HCC patients was correlated with race, T stage and pathological G grade (χ²=9.055, 14.012, 17.163; P<0.05). Kaplan-Meier Plot demonstrated that HCC patients with high expression of CCN4 obtained shorter overall survival (HR=1.50, 95%CI: 1.05-2.14, P<0.05). TIMER database analysis showed that the expression level of CCN4 in HCC was negatively correlated with tumor purity (r_s=-0.397, P<0.05), whereas positively correlated with the infiltration levels of B cells, CD8⁺T cells, CD4⁺T cells, macrophages, neutrophils, dendritic cells and M2 macrophages (r_s=0.306, 0.279, 0.460, 0.490, 0.465, 0.404, 0.532; P<0.05). PPI network analysis indicated that decorin was the main protein interacting with CCN4 (score=0.953).

Conclusions

The expression level of CCN4 protein is associated with race, T stage and pathological G grade, which can be used as a biomarker for poor prognosis of HCC. The underlying mechanism may be related to the infiltration of immune cells, the polarization of M2 macrophages and the formation of stromal fibers.

Key words: Carcinoma, hepatocellular, CCN4 gene, WISP1 gene, Biomarker, Bioinformatics

表1 CCN4 mRNA表达与肝癌患者临床病理参数的关系（例）

参数	CCN4-H	CCN4-L	χ²值	P值
种族			9.055	0.029
黄种	69	85
黑种	7	10
白种	105	78
美国印第安人	5	12
T分期			14.012	0.003
T1	76	97
T2	38	47
T3	55	36
T4	17	5
G分级			17.163	0.001
G1	18	28
G2	61	86
G3	83	63
G4	24	8

表1 CCN4 mRNA表达与肝癌患者临床病理参数的关系（例）

参数	CCN4-H	CCN4-L	χ²值	P值
种族			9.055	0.029
黄种	69	85
黑种	7	10
白种	105	78
美国印第安人	5	12
T分期			14.012	0.003
T1	76	97
T2	38	47
T3	55	36
T4	17	5
G分级			17.163	0.001
G1	18	28
G2	61	86
G3	83	63
G4	24	8

图1 TCGA数据库371例肝癌组织和50例正常癌旁组织EPIC分析注：^*为P<0.05，^**为P<0.01，^***为P<0.001；TCGA数据为癌症基因组图谱，EPIC为免疫细胞和癌细胞的比例预估值，CCN4-H为CCN4基因高表达组，CCN为人类的细胞通信网络因子，CCN4-L组为CCN4基因低表达组，HCC为肝细胞癌

图2 CCN4基因表达与肝癌患者生存的Kaplan-Meier曲线注：CCN为人类的细胞通信网络因子

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Analysis of CCN4 expression and its clinical significance in hepatocellular carcinoma based on bioinformatics

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Analysis of CCN4 expression and its clinical significance in hepatocellular carcinoma based on bioinformatics