摘要
目的:探讨胞质分裂蛋白调节因子1（PRC1）对肺腺癌细胞迁移、侵袭和增殖以及预后的影响。方法:从基因表达数据库（GEO）获得GSE7670、GSE32863和GSE68465非小细胞肺癌转录组表达队列，并对3个队列进行差异分析，然后对差异基因进行富集分析。从癌症基因组图谱（TCGA）获得差异基因的表达和临床数据，通过加权基因共表达网络分析（WGCNA）得到与肿瘤分期和淋巴结转移相关的模块，对模块基因进行蛋白质相关网络分析（STRING）得到核心基因，从mRNA和蛋白质两个层面验证PRC1在肺腺癌和正常组织中的差异表达。之后，在人肺腺癌细胞系H1299中敲除PRC1的表达，进行体外功能实验从而验证PRC1对肺腺癌细胞迁移、侵袭和增殖能力的影响。采用t检验比较两组间PRC1表达差异，二元logistic回归分析预测肿瘤的独立预后影响因子。最后，对PRC1在调控肺腺癌转移的下游机制进行相关研究。结果:肺腺癌中PRC1 mRNA和蛋白质的表达量均高于正常肺组织，差异均有统计学意义（均P<0.05）。Si-PRC1组细胞在迁移实验（P<0.001）、侵袭实验（P<0.001）和划痕实验（P<0.001）中均显著低于对照组；Si-PRC1组细胞的增殖能力在72和96 h处明显低于对照组。在肿瘤患者中，PRC1高表达的患者总生存期更短（P＝0.0014）。qPCR结果表明，在PRC1被敲低的细胞中E-cadherin表达显著上调（P<0.001），N-cadherin和Vimentin的表达均显著下调（P<0.01）。蛋白质印迹法结果显示，PRC1敲除的肿瘤细胞中激活的磷酸化MAPK信号减弱。结论:PRC1与患者不良预后有关，影响肺腺癌淋巴结的转移和肿瘤细胞增殖、迁移和侵袭能力，是肺腺癌潜在的生物标志物。
Abstract
Objective:To investigate the effect of protein regulator of cytokinesis 1 (PRC1) on lymph node metastasis, migration, proliferation and prognosis of lung adenocarcinoma.Methods:The non-small cell lung cancer expression matrix was obtained from Gene Expression Omnibus (GEO), including GSE7670, GSE32863 and GSE68465. Firstly, differential analysis was performed on the three datasets to obtain differentially expressed genes, which were then subjected to enrichment analysis. Secondly, the clinical and expression data of differentially expressed genes were obtained from The Cancer Genome Atlas (TCGA) database. The data was used for weighted gene co-expression network analysis (WGCNA) to obtain the module most relevant to clinical characteristics including tumor stage and lymph node metastasis. Thirdly, the hub gene was obtained through analysis of the module genes using the Search Tool for the Retrieval of Interaction Genes/Proteins (STRING) and binary logistic regression analysis. Subsequently, the expression level of PRC1 in both mRNA and protein levels between lung adenocarcinoma and normal tissues was validated. After knocking down the expression of PRC1 in human lung adenocarcinoma cell line H1299, in vitro functional experiments were conducted to validate the impact of PRC1 on the proliferation, migration, and invasion abilities of lung adenocarcinoma cells. T-test was used to compare the difference of PRC1 expression between normal and tumor tissues, and binary logistic regression analysis was performed to identify independent factors affecting tumor prognosis. Additionally, the downstream mechanism of PRC1 in regulating the metastasis of lung adenocarcinoma cells was investigated.Results:The expression level of PRC1 was higher in lung adenocarcinoma tissues compared to normal lung tissues, at both mRNA and protein levels, with statistical significance. The cells in the Si-PRC1 group were significantly lower than that in the control group in migration experiments (P<0.001), invasion experiments (P<0.001), and scratch experiments (P<0.001). The proliferation ability of Si-PRC1 group cells was significantly lower than that of the control group at 72 and 96 hours. Patients with high expression of PRC1 in lung adenocarcinoma had shorter overall survival (P=0.0014). The qPCR results showed that E-cadherin expression was significantly upregulated (P<0.001) in PRC1 knockdown cells, while N-cadherin and Vimentin expression were significantly downregulated (P<0.01). Western blot results showed that the activated phosphorylated MAPK signal was weakened in PRC1 knockout tumor cells.Conclusions:PRC1 is associated with poor prognosis of LUAD patients and affects lymph node metastasis and tumor cells proliferation, migration and invasion. PRC1 is a potential therapeutic target for lung adenocarcinoma.
基金项目
河南省科学技术厅科技攻关项目(222102310239)
北京医卫健康公益基金会医学科学研究基金资助项目(B20610AN)
万方数据