CDC20在肺腺癌组织中的表达及对肺腺癌细胞增殖和侵袭的影响研究

Expression of CDC20 in lung adenocarcinoma tissues and its effect on the proliferation and invasion of lung adenocarcinoma cells

背景与目的:肺腺癌具有早期发现难、肿瘤进展快及晚期手术切除率低等特点。尽管单药免疫治疗和免疫治疗联合化疗的相关研究在改善预后、克服耐药方面已初显成效,但是大部分肺腺癌患者从中获益仍有限。因此,迫切需要寻找具有相对较高灵敏度和特异度的新型生物标志物,以改善肺腺癌患者的预后。细胞分裂周期蛋白20(cell division cycle protein 20,CDC20)参与多种肿瘤的发生、发展,但在肺腺癌中的生物学作用及机制尚未明确。本研究旨在探究CDC20在肺腺癌中的表达情况及其对肺腺癌患者预后的预测价值,并分析CDC20对肺腺癌细胞增殖和侵袭能力的影响。方法:采用免疫组织化学(immunohistochemistry,IHC)检测CDC20在肺腺癌中的表达情况并结合生物信息学和临床病理学参数分析其与预后不良的相关性。采用Kaplan-Meier生存曲线描述CDC20对肺腺癌患者术后生存率的影响,采用COX多因素回归分析影响肺腺癌患者术后生存率的独立预后因素。通过受试者工作特征曲线分析CDC20表达在肺腺癌患者中的诊断价值。采用实时荧光定量聚合酶链反应(real-time fluorescence quantitative polymerase chain reaction,RTFQ-PCR)和蛋白质印迹法(Western blot)检测人正常肺上皮细胞系BEAS-2B、人肺腺癌细胞系A549和H1299中CDC20的表达水平。细胞实验中,通过敲低肺腺癌细胞中的CDC20,分为si-NC(对照组)、si-CDC20#1(敲低组1)和si-CDC20#2(敲低组2)3个组。采用细胞计数试剂盒-8(cell counting kit-8,CCK-8)、克隆形成、transwell和划痕实验检测细胞增殖、迁移和侵袭能力。通过基因本体论(Gene Ontology,GO)功能和京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)通路富集分析CDC20在肺腺癌中的生物学作用。通过基因集富集分析(gene set enrichment analysis,GSEA)CDC20在肺腺癌中可能的调控通路。本研究经河北省胸科医院伦理委员会批准(编号:2022051)。结果:生物信息学及IHC结果均显示,CDC20在肺腺癌组织中显著高表达(P<0.05)。生物信息学与临床参数分析结果均显示,CDC20高表达与患者预后不良相关。Kaplan-Meier生存分析和COX回归分析均显示,CDC20表达情况与患者术后生存率呈显著负相关(P<0.05)。敲低CDC20能抑制肺腺癌细胞增殖、迁移和侵袭(P<0.05)。GO功能、KEGG通路和GSEA结果均显示,CDC20与细胞周期相关。结论:CDC20在肺腺癌中高表达,CDC20高表达是肺腺癌患者不良预后的独立危险因素。CDC20能促进肺腺癌细胞增殖、迁移和侵袭。

Background and purpose:Lung adenocarcinoma has the characteristics of difficult early detection,rapid tumor progression and low surgical resection rate.Although studies on immunotherapy alone and immunotherapy combined with chemotherapy have shown initial success in improving prognosis and overcoming drug resistance,the majority of lung adenocarcinoma patients still receive limited benefits.Therefore,there is an urgent need to identify novel biomarkers with relatively high sensitivity and specificity to improve the prognosis of lung adenocarcinoma.Cell division cycle protein 20(CDC20) is involved in the occurrence and development of various tumors,but its biological role and mechanism in lung adenocarcinoma remain unclear.The aim of this study was to investigate the expression of CDC20 in lung adenocarcinoma and its predictive value for the prognosis of patients with lung adenocarcinoma,and to further explore the effects of CDC20 on the proliferation and invasion capabilities of lung adenocarcinoma cells.Methods:Utilizing immunohistochemistry(IHC) to detect the expression of CDC20 in lung adenocarcinoma tissues,we analyzed its correlation with poor prognosis in combination with bioinformatics and clinicopathological parameters.Kaplan-Meier survival curves were employed to illustrate the impact of CDC20 on postoperative survival rates in lung adenocarcinoma patients.COX multivariate regression analysis was conducted to identify independent prognostic factors influencing postoperative survival rates.Additionally,receiver operating characteristic(ROC) curves were applied to assess the diagnostic value of CDC20 expression in lung adenocarcinoma patients.Real-time fluorescence quantitative polymerase chain reaction(RTFQ-PCR)and Western blot were used to measure the expression levels of CDC20 in normal human lung epithelial cells(BEAS-2B) and human lung adenocarcinoma cells(A549 and H1299).In cellular experiments,CDC20 was knocked down in lung adenocarcinoma cells,which were divided into three groups:si-NC(control group),si-CDC20#1(knockdown group 1) and si-CDC20#2(knockdown group 2).Cell counting kit-8(CCK-8),colony formation,transwell and wound healing assays were conducted to assess cell proliferation,migration and invasion capabilities.Functional enrichment analysis using Geng Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathways was conducted to explore the biological roles of CDC20 in lung adenocarcinoma.Finally,gene set enrichment analysis(GSEA) was employed to investigate potential regulatory pathways of CDC20 in lung adenocarcinoma.This study was approved by the Ethics Committee of Hebei Chest Hospital(Number:2022051).Results:The results of both bioinformatics analysis and IHC demonstrated a significantly high expression of CDC20 in lung adenocarcinoma tissues(P<0.05).Both bioinformatics analysis and clinical parameter evaluation revealed a correlation between high CDC20 expression and poor patient prognosis.Kaplan-Meier survival analysis and COX regression analysis consistently indicated a significant negative correlation between CDC20 expression and postoperative survival rates in patients(P<0.05).Additionally,the expression levels of CDC20 were higher in human lung adenocarcinoma cell lines A549 and H1299 compared with BEAS-2B(P<0.05).Knockdown of CDC20 effectively inhibited the proliferation,migration and invasion of lung adenocarcinoma cells(P<0.05).The results of GO,KEGG pathways and GSEA consistently pointed to a relationship between CDC20 and cell cycle regulation.Conclusion:CDC20is highly expressed in lung adenocarcinoma.High expression of CDC20 is an independent risk factor for poor prognosis of lung adenocarcinoma patient.CDC20 can promote the proliferation,migration and invasion of lung adenocarcinoma cells.