染色体芯片分析技术应用于先天性心脏病分子诊断及发病机制研究

发布时间：2018-03-28 08:46

  本文选题：先天性心脏病　切入点：染色体芯片分析　出处：《上海交通大学》2014年博士论文

【摘要】：目的:本研究旨在评价染色体芯片分析(CMA)对先天性心脏病(CHD)的分子诊断率,比较综合征型CHD与单纯型CHD之间拷贝数变异(CNV)检出率的差异;分析CHD临床表型与CNV的相关性;并利用正常对照数据及基因优选分析鉴定新的CHD候选相关位点及基因。方法:本研究系统回顾了在2006年12月至2013年4月期间就诊于波士顿儿童医院(BCH)心脏科的CHD患儿的详细临床资料及临床CMA结果。所有纳入研究的BCH CHD病例被分为单纯型CHD和综合征型CHD两类,并依据患儿的表型进一步将该队列的CHD病例划分成9个亚型。本研究同时还将在2011年9月至2012年3月期间就诊于上海儿童医学中心(SCMC)心脏外科的圆锥动脉干畸形患儿作为研究对象。BCH及SCMC CHD队列的CMA分析分别基于Agilent 4×180K SNP+CGH芯片和Affymetrix Cytoscan HD芯片平台。所有经确定的非多态性CNV依据美国医学遗传协会(ACMG)指南进行临床致病性的评估。统计BCH队列中CMA分析对单纯型CHD和综合征型CHD的分子诊断率,计算并比较该队列中各个CHD亚型间CNV频率的差异。综合两个队列的CMA分析结果,通过与正常对照组的比对,筛选新的CHD候选相关CNV。最后,结合在线分析工具、基因表达数据库及通路分析,设计了一套基因优选分析流程以鉴定新的CHD候选相关基因。结果:纳入本研究的CHD病例共包括514例CHD患儿,其中422例来自BCH队列,92例来自SCMC队列。对于BCH队列,临床CMA检测对CHD的整体分子诊断率为12.8-18.48%。排除12例携带染色体非整倍体的病例,CMA对综合征型CHD的分子诊断率为14.11-20.56%,而对单纯型CHD的诊断率为4.32-9.26%。在所有CHD亚型中,致病性CNV在圆锥动脉干畸形及心脏间隔缺损病例中的频率最高。SCMC队列的分子诊断率略低(8.7-12%)。此外,本研究筛选出4个重现次数≥3且在CHD队列中显著富集的染色体区域(4q末端区域、15q11.2、16p12.2及Yp11.2)作为新的CHD候选相关性位点。最后通过基因优选流程,鉴定出20个新的CHD候选相关基因。结论:本研究的CNV分析结果为临床CMA平台作为诊断CHD的一线实验检测方法提供了重要证据。尽管综合征型CHD的致病性CNV检出率更高,但CMA对单纯型CHD仍具有一定的诊断价值,尤其对于患圆锥动脉干畸形及心脏间隔缺损的病例CNV的检出率更高。致病性CNV的分析及筛选为候选基因的鉴定提供了丰富的资源。
[Abstract]:Objective: to evaluate the molecular diagnostic rate of chromosomal microarray analysis (CMA) for congenital heart disease (CHD), to compare the difference in the detection rate of CHD between syndromic CHD and simplex CHD, and to analyze the correlation between CHD clinical phenotype and CNV. The new candidate loci and genes of CHD were identified by normal control data and gene analysis. Methods: the CHD from December 2006 to April 2013 in cardiac department of Boston Children's Hospital was systematically reviewed. Detailed clinical data and clinical CMA results of children. All BCH CHD cases included in the study were divided into two groups: simple CHD and syndromic CHD. The cohort of CHD cases were further divided into 9 subtypes according to their phenotypes. From September 2011 to March 2012, we will also visit Shanghai Children's Medical Center (SCC) from September 2011 to March 2012 to study conical artery trunk malformation in cardiac surgery. The CMA analysis of SCMC CHD cohort and. BCH cohort were based on Agilent 4 脳 180K SNP CGH and Affymetrix Cytoscan HD chip, respectively. All identified non-polymorphic CNV were evaluated for clinical pathogenicity according to the American Medical genetic Association (ACMG) guidelines. To estimate the molecular diagnostic rate of CMA analysis in BCH cohort for simplex CHD and syndromic CHD, The differences of CNV frequency among different CHD subtypes in the cohort were calculated and compared. The results of CMA analysis of the two cohorts were synthesised and compared with those of the normal control group. Finally, a new candidate for CHD was selected. Finally, an online analysis tool was used. Gene expression database and pathway analysis were used to identify new candidate genes for CHD. Results: 514 cases of CHD were included in this study. Of these, 422 were from BCH cohorts and 92 from SCMC queues. For BCH queues, The overall molecular diagnostic rate of CHD by clinical CMA was 12.8-18.48.The molecular diagnostic rate of CHD was 14.11-20.56 excluding 12 cases with chromosomal aneuploidy, and the diagnostic rate of CHD was 4.32-9.26 in all CHD subtypes. The frequency of pathogenetic CNV in the conical trunk malformation and cardiac septal defect was the highest. The molecular diagnostic rate of the cohort was slightly lower than 8.7-12. In addition, In this study, four chromosomal regions with recurrence times 鈮，

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