Elucidating Genotype-Phenotype Relationship of Polygenic Dilated Cardiomyopathies

项目总结 扩张型心肌病(DCM)是一种发病率较高的遗传性心脏病，发病率为1：250。 虽然通常被认为是一种单基因遗传病，但最近的遗传学研究表明 复杂的多基因原因。多基因心肌病的功能谱还不太清楚，这 阻碍了有效的治疗发展。干细胞技术的最新进展，如患者- 衍生诱导多能干细胞(IPSCs)和基因组编辑提供了前所未有的机遇 研究这种疾病的表型。心肌细胞将从患者的IPSCs中分化出来 两个DCM相关突变并测序以了解可能与DCM相关的转录组差异 疾病易感性。分析将通过使用CRISPR来标准化，以创建等基因对照， 插入与这些患者相关的健康个体的突变。然后我们将利用IPSC 与组织工程学相结合的平台构建心脏组织工程 细胞。我们将进行单细胞RNA测序，以测试细胞之间的相互作用和串扰 细胞。我们还将进行功能分析，如钙处理和收缩能力。CRISPR/dCas9系统 将使我们能够识别适合药物靶向的基因。母公司R01赠款的首要目标是 利用IPSC平台了解多基因扩张型心肌病的潜在机制。建议的多样性 补充剂通过研究不同种族的患者并考虑这种因素是如何在 疾病的严重性。非洲裔美国人和西班牙裔患者在健康研究中的代表性不足，而且 因此，更容易受到疾病管理不善的影响。卡洛斯·维拉博士将扩大我们的20人队列 来自这些少数群体的六个IPSC系的个体，并执行拟议的一套 实验。

PROJECT SUMMARY Dilated cardiomyopathy (DCM) is a genetic cardiac disorder with high morbidity and an incidence of 1:250. Although generally thought of as a disease with monogenic heritability, recent genetic studies suggest complex polygenic causes. Functional profiles of polygenic cardiomyopathies are poorly understood and this hampers effective therapeutics development. Recent advances in stem cell technology such as patient- derived induced pluripotent stem cells (iPSCs) and genome-editing, provides an unprecedented opportunity to study such disease phenotypes. Cardiomyocytes will be differentiated from iPSCs taken from patients with two DCM related mutations and sequenced to understand transcriptome differences that may correlate with disease susceptibility. The analysis will be normalized by using CRISPR to create isogenic controls and also insert the mutations in healthy individuals that are related to these patients. We will then utilize the iPSC platform coupled with tissue engineering to develop engineered heart tissues composed of cardiac specific cells. We will perform single cell RNA sequencing to test the cell-cell interactions and crosstalk between the cells. We will also perform functional assays like calcium-handling and contractility. CRISPR/dCas9 systems will allow us to identify genes suitable for drug targeting. The overarching goal of the parent R01 grant is to understand underlying mechanisms of polygenic DCM using the iPSC platform. The proposed diversity supplement extends this work by studying patients of different ethnicities and considering how this factors in disease severity. African-American and Hispanic patients are underrepresented in health studies, and therefore more vulnerable to poor disease management. Dr. Carlos Vera will expand our cohort of 20 individuals by six iPSC lines derived from these minority populations and perform the proposed set of experiments.