南方医科大学学报

南方医科大学学报 ›› 2023, Vol. 43 ›› Issue (8): 1345-1355.doi: 10.12122/j.issn.1673-4254.2023.08.11

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TMEM64在肝癌组织中高表达并促进肝癌细胞的增殖和侵袭

曹丹萍，蔡 娟，李艳娜，董润雨，王智雄，左学良   

  1. 皖南医学院第一附属医院//弋矶山医院胃肠外科，肿瘤内科，安徽 芜湖 241001
  • 出版日期:2023-08-20 发布日期:2023-09-11

TMEM64 is highly expressed in hepatocellular carcinoma and promotes tumor cell proliferation and invasion

CAO Danping, CAI Juan, LI Yanna, DONG Runyu, WANG Zhixiong, ZUO Xueliang   

  1. Department of Gastrointestinal Surgery, Department of Oncology, First Affiliated Hospital of Wannan Medical College (Yijishan Hospital), Wuhu 241001, China
  • Online:2023-08-20 Published:2023-09-11

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摘要： 目的 分析TMEM64在肝癌中的表达情况及对肝癌患者预后评估价值，探讨TMEM64对肝癌细胞增殖和侵袭能力的影响。方法 采用癌症基因组图谱（TCGA）数据库和基因型组织表达项目（GTEx）分析 TMEM64 在肝癌中表达水平。采用Wilcoxon秩和检验方法比较分析TMEM64基因在肝癌和癌旁组织中的表达差异。利用Kaplan-Meier曲线和Cox回归分析评估TMEM64 的表达水平对肝癌患者总生存期的影响及预后预测价值。通过单样本基因富集分析评估TMEM64的表达与免疫细胞浸润水平之间的相关性。依据 TMEM64 表达水平及相关临床变量构建基于总体生存率的nomogram图并进行验证。细胞实验中，在HCCLM3细胞中敲低TMEM64，分为si-NC（对照组）、si-TMEM64#1（敲低组1）、si-TMEM64#2（敲低组2）3个组；在Huh7细胞中过表达TMEM64，并分为vector（对照组）和TMEM64（过表达组），采用CCK-8、EdU和克隆形成实验检测TMEM64对肝癌细胞增殖的影响，使用Transwell和细胞划痕实验验证TMEM64对肝癌细胞迁移和侵袭能力的改变。结果 TMEM64在肝癌中的表达水平显著高于癌旁组织（P<0.001）。Kaplan-Meier生存分析显示，TMEM64高表达与肝癌患者不良总生存时间相关，差异有统计学意义（P<0.05）。多因素Cox回归分析显示，TMEM64高表达是肝癌患者OS的独立危险因素（P<0.05）。TMEM64的表达与NK细胞、CD8+T细胞和pDCs细胞的免疫细胞浸润水平呈负相关（P<0.05）。GO、KEGG和GSEA富集分析结果显示TMEM64与信号传递及肿瘤转移相关通路显著富集。列线图和校准曲线显示模型预测效能与实际结果相符。敲低TMEM64能抑制肝癌细胞的增殖、迁移和侵袭（P<0.01），过表达TMEM64能促进肝癌细胞的增殖、迁移和侵袭（P<0.01）。结论 TMEM64在肝癌中高表达，TMEM64高表达是肝癌患者不良预后的独立危险因素。TMEM64能促进肝癌细胞的增殖、迁移和侵袭。

关键词: TMEM64；肝细胞癌；预后；侵袭

Abstract: Objective To analyze the expression of TMEM64 in hepatocellular carcinoma (HCC) and investigate the effect of TMEM64 expression level on proliferation and invasion of HCC cells in vitro. Methods We analyzed the expression level of TMEM64 in HCC and adjacent tissues based on data from TCGA and GTEx databases. The prognostic value of TMEM64 for HCC patients was examined using Kaplan-Meier survival analysis and a Cox regression model, and a nomogram was constructed based on TMEM64 expression and clinical characteristics of the patients. Functional enrichment analysis was performed to explore the potential signaling pathways, and immune cell infiltration was assessed using single sample gene set enrichment analysis. We also performed cell experiment to observe the changes in proliferation, migration, and invasion in HCCLM3 cells with TMEM64 knockdown and in Huh7 cells with TMEM64 overexpression using CCK-8, EdU, colony formation, Transwell, and wound healing assays. Results The expression level of TMEM64 was significantly higher in HCC than in the adjacent tissues (P<0.05). Kaplan-Meier analysis suggested that a high expression of TMEM64 was associated with poor outcomes of the patients (P<0.05). Multivariate Cox regression analysis indicated that a high TMEM64 expression was an independent risk factor for overall survival of HCC patients (P<0.05). TMEM64 expression level was negatively correlated with the levels of immune cell infiltration by NK cells, CD8 + T cells, and plasma pDCs cells (P<0.05). GO, KEGG, and GSEA enrichment analyses showed that TMEM64 was significantly enriched with tumor invasion and metastasis pathways. The nomogram and calibration curves indicated a moderate prediction reliability of the model. In the cell experiment, TMEM64 knockdown obviously suppressed and TMEM64 overexpression markedly promoted the proliferation, migration, and invasion of HCC cells (P<0.01). Conclusion A high TMEM64 expression may serve as an independent risk factor for poor prognosis of HCC and promotes proliferation, migration, and invasion of HCC cells in vitro.

Key words: TMEM64; hepatocellular carcinoma; prognosis; invasion