Program Project: GENE MUTATION AND RESCUE IN HUMAN DIAPHRAGMATIC HERNIA

描述（由申请人提供）：为了揭示先天性膈疝（CDH）的病因，这是一种普遍的（1:2500活产）高影响（50%死亡率和高发病率）的先天缺陷，具有表型和遗传复杂性，我们将新颖和传统的调查策略结合起来，对人类CDH患者进行最大的队列之一。总的来说，我们正在使用分子遗传学、分子细胞遗传学、基因组学、发育学、生物信息学和干细胞策略以及多种模式生物的使用。我们采用的策略可以作为一个模板，用于识别不仅与CDH有关的基因通路，也与其他先天性异常有关的基因。我们工作的基础是为确定和招募几乎所有CDH患者而开发的基础设施，这些患者来自两个学术三级儿科外科中心，马萨诸塞州总医院（MGH）和波士顿儿童医院（CHB），每个中心都提供最先进的医学、外科和基因诊断和管理。研究人员对CDH患者和家庭进行了全面的访谈，并由资深遗传学家进行了仔细的表型分析；使用适用于每个先证者和尽可能多的家庭成员的婴儿和儿童的敏感方案获得生物样本，并将其储存起来用于拟议的迭代遗传研究。当多个查询线存在收敛时，选择候选基因进行测序[即，在阵列比较基因组杂交（aCGH）或单核苷酸多态性（SNP）阵列上检测到拷贝数变异（CNVs），在受影响的连锁分析中检测到属于多个CDH家族的共享间隔基因，在dna序列中表达的基因

DESCRIPTION (provided by applicant): In the quest to unravel the etiology of Congenital Diaphragmatic Hernia (CDH), a prevalent (1:2500 live births) high impact (50% mortality with high morbidity) birth defect with phenotypic and genetic complexity, we are combining novel and traditional investigative strategies to one of the largest cohort of human patients with CDH. Collectively, we are employing molecular genetic, molecular cytogenetic, genomic, developmental, bioinformatic, and stem cell strategies along with the use of multiple model organisms. The strategies that we employ can serve as a template for identifying genes in pathways responsible not only for CDH, but for additional congenital anomalies, as well. The foundation of our work is the Infrastructure developed for ascertaining and recruiting almost all patients with CDH from two academic tertiary pediatric surgical centers, Massachusetts General Hospital (MGH) and Children's Hospital Boston (CHB), each offering state of the art medical, surgical, and genetic diagnosis, and management. Patients and families with CDH are interviewed comprehensively by study staff and carefully phenotyped by a senior geneticist; biological samples are obtained using sensitive protocols adapted for infants and children from each proband and from as many family members as possible, and are banked for proposed iterative genetic studies. Gene candidates are selected for sequencing when there is convergence from multiple lines of inquiry [i.e., copy number variations (CNVs) detected on array Comparative Genomic Hybridization (aCGH) or Single Nucleotide Polymorphism (SNP) arrays, genes in shared intervals detected on linkage analysis in affected belonging to multiplex CDH families, expressed genes in the developing primordial diaphragm, and bioinformatic algorithms that prioritize multiple candidate variants detected by the aforementioned platforms and from annotation of animal models with diaphragm defects]. The functional significance of the most promising candidates is subsequently assessed in multiple animal models. The evolution of available sequencing capabilities to lower costs opens the door to examine the multiple genes or polygeneity that we now know will contribute to a birth defect phenotype such as CDH. Given the national and International outreach and reputation of our genetic partnerships and the strength of the voluntary support network provided by the exceptional parents of patients with CDH, the study has extended beyond our two institutions to ascertain a number of seminal multiplex kindreds, including both consanguineous and non-consanguineous families, from the U.S. and internationally. RELEVANCE: The overarching goal of this study is to converge the multiple genes detected by these complementary approaches into cogent molecular pathways that will reveal polygeneic defects in the pathways and point toward potential treatment paradigms to prevent or alleviate this mortal or morbid congenital anomaly. Further, the methods devised for this study of CDH may have broader implications across other congenital anomalies.