克山病患者差异基因表达及细胞凋亡通路研究

摘要

目的 观察克山病基因表达谱的变化,从基因水平探讨克山病细胞凋亡发生机制,寻找克山病诊断标志基因,建立克山病诊断质心图.方法 选择来自克山病病区符合克山病诊断指标的10例慢型克山病患者作为克山病组,10例健康居民作为对照组.取受检者空腹静脉血,提取全血RNA,采用Agilent表达谱芯片[Agilent 4×44K全基因组Oligo芯片(单标)]测试全基因组表达谱.采用Agilent Feature Extraction Software 提取数据,克山病组与对照组间差异基因分析采用t检验筛选,经基因关系网络分析(Pathway studio analysis)软件分析基因富集通路,并经分类预测分析(prediction analysis for microarray,PAM)筛选克山病诊断标志基因,建立诊断质心图.结果 与对照组比较,克山病组筛选出3 068个差异基因,其中克山病患者上调1 570个,下调1 498个.富集信息通路38条.其中凋亡通路居于首位,其关系网络图显示6个基因在克山病组表达上调,包括共济失调突变基因(ATM)、cAMP依赖蛋白激酶A(PKA)、杆状病毒IAP repeat-containing 2 (BIRC2)、NLR家族凋亡抑制蛋白(NAIP)、BCL2样11(Bim)、BCL2相关蛋白A1 (BCL2A1);7个基因表达下调,包括细胞凋亡相关蛋白酶8(CASP8)、BCL2结合成分3(BBC3)、BCL2相关永生基因(BAG1)、BCL2-相关X蛋白(BAX)、BCL2样1 (BCL2L1)、BCL2相关卵巢杀手(BOK)、凋亡相关蛋白酶6(CASP6).共筛选出42个诊断标志基因.结论 凋亡相关差异表达基因及信息通路在克山病的发病机制中具有重要作用,筛选的42个诊断标志基因可为克山病的诊断提供基础依据.%Objective To investigate the differences in gene expression profiles of peripheral blood from patients with Keshan disease (KD) and the apoptosis mechanism in KD,to obtain diagnostic markers and establish diagnostic centroids plot for KD.Methods RNA was isolated from ten patients with KD diagnosed according to the clinical criteria for KD in China and ten health controls.The expression profiles were evaluated by Agilent 4 ×44K Whole Human Genome density oligonucleotide microarray analysis.The data were extracted by Agilent Feature Extraction Software t test,Pathway studio analysis and prediction analysis for microarray (PAM) were used to identify differently expressed genes,gene pathways,diagnostic markers and establish diagnostic centroids plot.Results Totally 1 570 up-regulated genes and 1 498 down-regulated genes were identified.Thirty-eight enrichment pathways were also identified,and the highest ranked by Pathway studio analysis was related to apoptosis.Six genes involved in apoptosis pathway were up-regulated in KD included ataxia telangiectasia mutated (ATM),cAMP-dependent protein kinase,protein kinase A (PKA),baculoviral IAP repeat-containing 2 (BIRC2),NLR family,apoptosis inhibitory protein (NAIP),BCL2-1ike 11 (Bim),BCL2-related protein A1 (BCL2A1) and down-regulated were 7 which included caspase 8 (CASP8),BCL2 binding component 3 (BBC3),BCL2--associated athanogene (BAG1),BCL2-associated X protein (BAX),BCL2-1ike 1 (BCL2L1),BCL2-related ovarian killer (BOK),and caspase 6 (CASP6).Forty-two diagnostic markers were obtained through PAM analysis.Conclusions Apoptosis related to genes and pathways might play an important role in the pathogenesis of KD.Forty-two markers could be used as molecular markers for the diagnosis of KD,which is important to the diagnosis of KD.