Single Cell RNA-seq to Identify Endocardial Ontogenic Factors for the Heart

翻译后摘要：本研究项目的目标是确定的签名基因，定义的个体发育生态位， 心脏内皮细胞，以产生关键的心脏结构，瓣膜和冠状动脉，使用小鼠 遗传学和单细胞转录组学。我们以前的研究表明，心脏内皮细胞是心脏的主要细胞， 心脏瓣膜和冠状动脉的祖细胞，来自不同的心脏区域和不同的 通过不同的机制。心脏流出道和房室管的内皮细胞 形成心脏瓣膜，而位于心室的则形成冠状动脉。这些观察结果 证明内皮细胞是具有不同发育命运的异质群体， 功能协调发展的然而，内在因素在时间上调节内源性个体发生生态位， 一些细胞与其他细胞的区别以及具体说明细胞命运和功能目前尚不清楚，至少是由于缺乏 实验模型和方法来分离和询问单个内皮细胞。这个研究项目 将使用小鼠遗传模型的协同方法填补这一严重缺失的知识空白， 发育心脏解剖学，先进的单细胞转录组技术，和系统生物学研究， 单个细胞的表达谱，以了解细胞异质性和揭示亚群。的 这项研究将导致确定细胞命运和启动冠状动脉粥样硬化的潜在调节剂， 血管生成，这可以进一步验证和功能确认。我们已经培育出了 这些模型使我们能够在不同的个体发育阶段分离心室和瓣膜内皮细胞。在这 我们将在这些模型上使用单细胞分选和RNA-seq来识别遗传因素 不同温度下瓣膜或冠状动脉形成区域的心内膜细胞亚群差异表达 阶段（目标1）。然后，我们将使用整合的形态学，细胞和分子分析来揭示其 在心脏瓣膜形成和冠状动脉发育中的潜在功能（目标2）。新信息将 揭示了心脏发育和疾病中的内皮细胞生物学。

Abstract: The goal of this research project is to identify the signature genes that define the ontogenic niche for cardiac endocardial cells to generate the critical heart structures, valves and coronary arteries, using mouse genetics and single cell transcriptomics. Our previous studies showed that cardiac endocardial cells are the progenitor cells for heart valves and coronary arteries, arising from different cardiac regions and at different stages through distinct mechanisms. Endocardial cells at the cardiac outflow tract and atrioventricular canal give rise to heart valves, whereas those at the ventricle generate coronary arteries. These observations document that the endocardial cells are a heterogeneous population with distinct developmental fates and functions. However, the intrinsic factors temporally regulating the endocardial ontogenic niche to distinguish some cells from the others and to specify cell fate and function are currently unknown, due at least to the lack of experiment models and methods to isolate and interrogate individual endocardial cells. This research program will fill this critically missing knowledge gap using a synergistic approach of mouse genetic models, developmental heart anatomy, advanced single cell transcriptome technology, and systems biology to study the expression profiles of individual cells in order to understand cell heterogeneity and uncover subpopulations. The study will lead to the identification of potential regulators for cell fate determination and for initiating coronary angiogenesis, which can be further validated and functionally confirmed. We have generated genetic mouse models that allow us to isolate ventricular versus valve endocardial cells at different ontogenic stages. In this research program we will use single cell sorting and RNA-seq on these models to identify the genetic factors differentially expressed by subpopulations of endocardial cells at valve- or coronary-forming regions at different stages (Aim 1). We will then use integrated morphologic, cellular, and molecular analysis to reveal their potential functions in heart valve formation and coronary artery development (Aim 2). The new information will shed light on endocardial biology in heart development and disease.