Neural Crest and Cardiac Outflow Tract Formation

（改编自申请人的摘要）候选人是一名医学博士， 在儿科、围产期医学、 儿科心脏病学 他在分子和生物学领域获得了研究经验， 他目前正在寻求深入的基础研究 培训过渡到独立调查员，最终目标是 成为一名学术性的医生科学家。 颁奖时， 候选人将是费城儿童医院的教员 宾夕法尼亚大学儿科讲师 这项建议的重点是在胚胎心脏发生的关键时间， 流出道的分隔和形成发生， 包括神经嵴的贡献 发育缺陷发生在 这一时期占了大量的先天性心脏缺陷。 阐明心脏流出道形成的机制将 提高我们对这些疾病的认识。 初步实验 申请人进行的研究旨在鉴定已知的 当流出道分隔时， 开始（E11.5）。 其中一个（克隆35）编码一种功能未知的蛋白质 在进化过程中得以保存。 它具体表现在 流出道和神经嵴来源的间充质在时间的 间叶分隔 在此，提出了一个深入的分析， 新基因，特别关注其在神经嵴中的潜在作用 介导的心脏形态发生。 此外，一种新的转基因小鼠创造了 将用于分析神经嵴的命运 细胞在几个突变的小鼠品系与先天性心脏病。 具体 目的是：1）深入分析克隆35。 这将包括隔离 全长cDNA，广泛的表达分析，抗体的制备， 允许亚细胞定位，基因组染色体定位，并产生 有针对性地缺乏该基因的转基因小鼠。 2)细胞命运 先天性巨噬细胞瘤小鼠突变系中神经嵴细胞的测定 心脏病（斑点，补丁，NFL和崩溃3突变胚胎），通过使用 从近端Pax 3启动子表达Cre重组酶的转基因小鼠， 在神经嵴细胞中指导表达， 小鼠

(Adapted from applicant's abstract) The candidate is an MD with extensive medical training in the fields of pediatrics, neonatal-perinatal medicine and pediatric cardiology. He has acquired research experience in molecular and developmental biology and he is currently seeking intensive basic research training to transition to an independent investigator with the ultimate goal to become an academic physician scientist. At the time of the award the candidate will be a faculty member of the Children's Hospital of Philadelphia and Instructor in Pediatrics at the University of Pennsylvania. This proposal focuses on a critical time during embryonic cardiogenesis in which septation and formation of the outflow tract take place, a process involving neural crest contribution. Developmental defects occurring during this period account for a large number of congenital heart defects. Elucidation of the mechanisms involved in cardiac outflow tract formation will enhance our understanding of these diseases. Preliminary experiments performed by the applicant were aimed towards the identification of known and novel genes activated in the heart at the time when outflow tract septation begins (E11.5). One of them (Clone 35) encodes a protein of unknown function that is conserved through evolution. It is expressed specifically in the outflow tract and in neural crest derived mesenchyme at the time of aortopulmonary septation. Here, it is proposed an in depth analysis of this novel gene with particular attention to its potential role in neural crest mediated cardiac morphogenesis. In addition, a novel transgenic mouse created in the sponsor's laboratory will be used to analyze the fate of neural crest cells in several mutant mouse lines with congenital heart disease. Specific aims are: 1) In depth analysis of Clone 35. This will include isolation of full length cDNA, extensive expression analysis, preparation of antibodies to allow subcellular localization, genomic chromosomal mapping, and generation of transgenic mice with targeted deficiency of this gene. 2) Cell fate determination of neural crest cells in mouse mutant lines with congenital heart disease (Splotch, Patch, Nfl and collapsing 3 mutant embryos) by using transgenic mice expressing Cre-recombinase from the proximal Pax 3 promoter, that directs expression in neural crest cells, and appropriate Cre-reporter mice.