心功能正常患者右室心尖起搏后外周血基因表达谱分析及基因标志物的筛选

发布时间：2018-04-03 07:17

本文选题：右室心尖起搏　切入点：基因芯片　出处：《南京医科大学》2016年博士论文

【摘要】：背景心脏起搏器植入是治疗缓慢性心律失常唯一有效的治疗措施。右室心尖(right ventricular apex,RVA)起搏是目前临床广泛应用的起搏方式。右室心尖起搏可导致心肌细胞电重构、组织学重构,房颤的发生率增加,最终可能导致心力衰竭。有研究报道心功能正常的患者RVA起搏对心室结构及功能的影响较小。RVA起搏术后在心室结构及功能均正常的阶段,分子生物学层面是否已经发生改变,目前研究较少。线粒体与心血管疾病的有关研究日益增多。线粒体融合蛋白视神经萎缩蛋白1(opticatrophy 1,OPA1)参与心力衰竭的发生发展。RVA起搏术后OPA1的外周血表达水平改变及其具体的生物学作用及相关调控机制,还需要进一步深入研究。目的对心功能正常患者RVA起搏术后外周血全基因进行检测,并筛选基因标志物,探讨其生物学作用。方法利用外周血表达谱基因芯片检测方法,测定10例患者RVA起搏术前和术后一周外周血全基因表达水平,另外有3例患者只测定了术前外周血全基因表达水平。Transcriptome Analysis Console 3.0 软件采用ANOVA 算法,计算P0.05 时的差异表达基因,通过DAVID(the Database for Annotation,Visualization and Integration Discovery)在线数据库对差异表达≥2倍的基因进行生物信息学分析。应用蒙特卡洛、逻辑回归进行分析运算,筛选出对起搏器植入前后样本有最佳甄别作用的基因组合。结果外周血表达谱基因芯片研究结果显示RVA起搏术后一周与术前比较存在大量差异表达基因,显著上调基因有228个,显著下调的基因有281个。差异表达的基因全部作用于细胞连接、细胞膜和糖蛋白,主要参与离子跨膜转运、细胞运动性、突触前膜聚集等生物过程。差异表达的基因参与钙离子信号通路、调节糖鞘脂的生物合成、神经活性受体与配体相互作用3条信号通路。其中钙离子调节信号通路发生差异表达的基因数目最多,包括CaMKⅡ2A、RYR2、IP3R。本研究基于外周血表达谱基因芯片研究的结果,根据逻辑回归分析方法,筛选出对起搏器植入前后样本有最佳甄别作用的基因组合(OPA1、NDUFA1、PRDX1、CTSA、STK10),敏感性91%,特异性92%,准确性91.3%。在起搏器植入术后患者与健康对照组的差异基因表达分析中,ROC曲线分析显示该基因组合的曲线下面积值(area under curve,AUC)0.90(95%CI 0.80-0.97,P0.001)。基因组合中除了 STK10,其余都与线粒体相关,其中OPA1与心血管疾病关系密切。结论心功能正常患者RVA起搏术后一周与术前术前比较存在大量差异表达基因,筛选出对起搏器植入前后有最佳甄别作用的基因组合,对差异表达基因进行生物信息学分析,为进一步的qRT-PCR验证奠定了一定基础。
[Abstract]:Background cardiac pacemaker implantation is the only effective treatment of slow arrhythmia treatment. Right ventricular apex (right ventricular, apex, RVA) is a widely used clinical pacing. The pacing right ventricular apical pacing can lead to myocardial cell electrical remodeling, tissue remodeling, increased incidence of atrial fibrillation, may ultimately lead to heart failure. Research reports normal cardiac function in patients with RVA pacing on ventricular structure and function after.RVA pacing in small ventricular structure and function were normal, whether the molecular level has changed, there is less research on the related research. More mitochondria and cardiovascular disease increased. Mitochondrial fusion protein of optic nerve atrophy protein 1 (opticatrophy 1 OPA1), changes in the expression of OPA1 in peripheral blood of the occurrence and development of.RVA pacing in heart failure and biological effects of specific and related The regulation mechanism, we still need further study. The purpose of normal cardiac function in patients with RVA pacing after peripheral blood gene detection and screening of genetic markers, study its biological effect. Methods the gene chip detection method using peripheral blood, 10 cases of patients with RVA pacing before and after surgery in a week the whole blood gene expression levels were measured in 3 cases only by preoperative peripheral blood gene expression level of.Transcriptome Analysis Console 3 software using ANOVA algorithm, calculate the differences between P0.05 gene expression by DAVID (the Database, for Annotation, Visualization and Integration Discovery) bioinformatics analysis of differentially expressed online database more than 2 times of the gene. The application of Monte Carlo, logistic regression analysis calculation, screened the best screening of samples before and after pacemaker implantation for combination. The results of peripheral blood gene expression microarray results showed that RVA pacing one week after operation compared with the preoperative, there are a large number of differentially expressed genes, 228 genes were up-regulated and 281 genes were down regulated significantly. The differentially expressed genes in the role of all cell junctions, cell membrane and egg white sugar, mainly involved in the transmembrane ion transport, cell motility, presynaptic membrane aggregation and other biological processes. The differential expression of genes involved in calcium signaling pathway, glycosphingolipid biosynthesis regulation, neural activity and receptor ligand interaction 3 pathways. The number of genes in which the calcium ion regulation signal pathway differentially expressed most, including CaMK II 2A, RYR2, IP3R. based on the study of peripheral blood gene expression microarray results, according to the logistic regression analysis method, screening the best combination of gene screening role of samples before and after pacemaker implantation (OPA1, NDUFA1, PR DX1, CTSA, STK10), the sensitivity was 91%, specificity 92%, accuracy analysis of differential gene expression in 91.3%. after pacemaker implantation in patients with healthy control group, ROC curve analysis showed that the gene combination area under the curve value (area under curve, AUC) 0.90 (95%CI 0.80-0.97, P0.001). In addition to the combination of genes STK10, the rest are related with mitochondria, the relationship between OPA1 and cardiovascular disease closely. Conclusion cardiac function in patients with normal RVA pacing a week after surgery and preoperative before the existence of a large number of differentially expressed genes, screening the best combination of gene screening role of pacemaker implantation after expression, bioinformatics analysis of gene differences. Lays the foundation for further verification of qRT-PCR.

【学位授予单位】：南京医科大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R541.7

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