加权基因共表达网络分析黏附基因在腹主动脉瘤中相关机制

Weighted gene co-expression network analysis identified adhesion genes as key mechanism in abdominal aortic aneurysms

目的通过加权基因共表达网络分析揭示腹主动脉瘤(AAA)发生的潜在机制。方法对数据库编号GSE47472和数据库编号GSE57691两个AAA转录组测序数据合并,进行基因差异表达分析得到差异基因,加权基因共表达网络分析(WGCNA)得到中心基因,两者取交集得到差异中心基因,并对其进行功能富集分析。建立小鼠AAA模型进行定量聚合酶链反应(qPCR)验证差异中心基因表达。使用GraphPad Prism 8进行统计分析。Kolmogorov-Smirnov检验数据正态性,采用独立样本t检验分析基因差异表达。结果共筛选出745个差异表达基因,建立16个基因共表达模块,其中中心模块包含119个基因,取交集后共得到60差异中心基因。对差异中心基因进行功能富集分析结果示AAA中平滑肌增殖分化功能和细胞黏附功能相关基因低表达。低表达的细胞黏附相关基因为钙黏着蛋白-2(CDH2),(钙黏着蛋白-13)CDH13,整合素相互作用蛋白-2(FERMT2),Rho关联含卷曲螺旋结合蛋白激酶1(ROCK1),层黏蛋白α5(LAMA5),进行qPCR验证其表达差异显著性。相对表达倍数分别为FERMT2=0.31 (t=2.454,P<0.05),ROCK1=0.22(t=3.686,P<0.05),LAMA5=0.45(t=3.168,P<0.05),CDH13=1.36(t=0.103,P>0.05),CDH2=1.71(t=0.702,P>0.05)。结论 FERMT2、ROCK1、LAMA5低表达可能通过介导血管平滑肌细胞与细胞外基质间黏附作用异常参与AAA形成。

Objective To explore the potential mechanism of abdominal aortic aneurysm(AAA)through weighted gene co-expression network analysis.Methods The transcriptome sequencing data of GSE47472 and GSE57691 were combined.Differential expression genes(DEGs)were obtained by differential gene expression analysis,and hub genes were obtained by weighted gene coexpression network analysis(WGCNA).The intersection of the two genes was conducted to obtain hub DEGs,and the Gene Set Enrichment Analysis was performed.Subsequently,a mouse model of AAA and a sham surgery group were established to verify hub DEGs by quantitative polymerase chain reaction(qPCR).Statistical analysis was performed using GraphPad Prism 8.Kolmogorov-Smirnov test data for normality,independent-sample student′s t test for significance of gene differential expression.Results 745 DEGs were screened out,and 16 gene co-expression modules were established.Hub module includes 119 genes.60 hub DEGs were obtained after intersection.Functional enrichment analysis of hub DEGs showed low expression of genes related to smooth muscle proliferation,differentiation,and adhesion function.The adhesion related genes were N-cadherin 2(CDH2),N-cadherin 13(CDH13),FERM domain containing kindlin 2(FERMT2),Rho associated coiled-coil containing protein kinase 1(ROCK1),and laminin subunitα5(LAMA5),and the significance of their expression differences was verified by qPCR.The relative expression times were FERMT2=0.31(t=2.454,P<0.05),ROCK1=0.22(t=3.686,P<0.05),LAMA5=0.45(t=3.168,P<0.05),CDH13=1.36(t=0.103,P>0.05),CDH2=1.71(t=0.702,P>0.05).Conclusion Low expression of FERMT2,ROCK1 and LAMA5 were involved in the formation of AAA by mediating the focal adhesion between vascular smooth muscle cells and extracellular matrix.