Objective: To explore chromobox protein homolog 2 (CBX2) expressions in relation to clinical features of patients and elucidate its role in the progression of hepatocellular carcinoma. Methods: Using the Cancer Genome Atlas (TCGA) database, R language was used to analyze the distribution of differentially expressed mRNA in hepatocellular carcinoma. The different expression of CBX2 in HCC and adjacent tissues and its relationship with survival and clinical characteristics of patients were further analyzed. The expression of CBX2 in liver tissues, liver cancer tissue, and L02, HepG2 and SMMC-7721 cell lines was detected by real time-PCR and western blot. The expression of CBX2 was interfered by siRNA in hepatoma cell line. MTT, colony formation, transwell assays, and flow cytometry were used to identify the proliferation, apoptosis, invasion and clone-formation ability of HepG2 and SMMC-7721 cells after CBX2 down-regulation. According to the different data, t-test, ANOVA, chi-square test, and COX regression model were used for statistical analysis. Survival curve was plotted through Kaplan-Meier method. Results: TCGA public database analysis showed that the expression of CBX2 mRNA in hepatocellular carcinoma tissues (7.296 ± 1.6115) was significantly higher than normal liver tissues (4.706 ± 0.940) (P = 0.000). In addition, the overall survival time of patients with low CBX2 mRNA expression was significantly longer than that of patients with high CBX2 mRNA expression [(5.971 ± 0.411) years vs. (4.650 ± 0.503) years, P = 0.001]. The expression level of CBX2 mRNA was correlated with the pathological TNM stage (P = 0.025) and differentiation degree (P < 0.001) of liver cancer. COX regression analysis showed that CBX2 mRNA expression was an independent predictor of patient survival (P = 0.013). siRNA was transfected and compared with the blank control group. The transgenic ability of HepG2 and SMMC-77221 cells decreased significantly at 72h (P < 0.05) and 96h (P < 0.05), and the apoptosis rate (11.430% ± 0.215%) was higher than blank control group (6.6 00% ± 0.170%) (P = 0.003). The number of invasive cells ((both P < 0.05) and relative colony forming cells ((both P < 0.001) were significantly decreased. In 20 cases of tissue samples, the expression of CBX2 protein (relative expression level 3.020 ± 0.269) in liver cancer was higher than that in adjacent tissues (relative expression level 0.886±0.065) (P < 0.001). The overall survival time of patients with low CBX2 expression in liver cancer was longer than that of patients with high expression [(3.670 + 0.576) years vs. (0.834 + 0.153) years, P = 0.004]. Conclusion: An evident high expression of CBX2 is an independent poor prognostic factor in hepatoma. Down-regulation of CBX2 expression can inhibit the progression of liver cancer. Therefore, CBX2 may be a prognostic biomarker and a new target for HCC treatment.
目的： 探索色素框同源物2（CBX2）在肝细胞肝癌（HCC)中的表达，以及与患者临床特征的关系，阐述其在HCC进展中的作用。 方法： 利用癌症基因组图谱（TCGA）数据库，应用R语言分析肝癌差异表达mRNA的分布情况，CBX2 mRNA在肝癌组织和癌旁组织的表达差异及其与患者生存和临床特征的关系；应用实时定量反转录聚合酶链反应（Real Time-PCR）法和Western blot法检测CBX2在肝组织、肝癌组织、L02、HepG2、SMMC-7721细胞系中的表达情况；siRNA干扰肝癌细胞系CBX2表达，应用噻唑蓝（MTT）法、流式细胞技术、Transwell实验和克隆形成实验鉴定CBX2下调后HepG2、SMMC-7721细胞的增值、凋亡、侵袭和克隆形成能力。根据资料不同分别采用t检验分析、ANOVA分析、χ(2)检验、COX回归模型分析，并用Kaplan-Meier法绘制生存曲线。 结果： TCGA公共数据库分析表明CBX2 mRNA在肝癌组织（7.296±1.612）中表达明显高于正常肝组织（4.706±0.940） (P < 0.001)，且在肝癌患者中CBX2 mRNA低表达者总体生存时间明显长于高表达者[（5.971±0.411）年比(4.650±0.503）年，P = 0.001]；CBX2 mRNA的表达高低与肝癌的病理TNM分期（P = 0.025）和分化程度（P < 0.001）相关，COX回归分析表明CBX2 mRNA表达高低是患者生存的独立预测因素（P = 0.013）。转染siRNA后与空白组比，HepG2及SMMC-77221细胞的增值能力在72 h（P值均<0.05）和96 h（P值均<0.05）时明显下降，凋亡率（11.430%±0.215%）高于空白组（6.600%±0.170%）（P = 0.003）；侵袭细胞数减少（P值均<0.05）；相对克隆形成细胞数明显减少（P值均<0.001）。20例组织样本中，肝癌中CBX2蛋白（相对表达水平3.020±0.269）表达高于癌旁组织（相对表达水平0.886±0.065）（P < 0.001）；且肝癌中CBX2低表达患者的总体生存时间长于高表达者[（3.670±0.576）年比（0.834±0.153）年，P = 0.004]。 结论： 肝癌中CBX2显著高表达，与肝癌预后差明显正相关，下调CBX2表达可以抑制肝癌进展，因此CBX2不仅可以作为肝癌预后的生物标志物，也有可能是潜在的肝癌治疗靶点。.
Keywords: CBX2; Carcinoma, hepatocellular; Gene expression; Prognosis.