Investigating the Time-Dependent Reversibility of DCM-Induced Epigenetic, Matrix, and Functional Remodeling

项目摘要 扩张型心肌病（DCM）是一种高度流行的遗传性心脏病，其特征在于收缩压升高， 功能障碍、离心性肥大和左心室扩张。虽然药物和机械治疗 已被证明部分改善心脏功能，这些结果往往是短暂的和高度可变的 从病人到病人。此外，最近的研究表明，表观遗传和基质失调， 即使在治疗后收缩功能改善的患者中，也会持续存在。鉴于异常染色质 重塑和细胞外基质（ECM）沉积已被确定为扩张重塑的驱动因素， 染色质和ECM重塑随着时间的推移会变得不可逆转-了解时间是至关重要的- DCM突变对逆转基因组、肌细胞和基质适应不良重构的依赖性影响 程度.该建议的中心假设是扩张型心肌病表型的可逆性是时间依赖性的 这是由于随着疾病的进展，永久性ECM的积累和染色质重塑。完成 在这个提议中，我将利用Davis实验室创建的DCM小鼠模型，该模型包含突变（I61 Q）， 心肌肌钙蛋白C（cTnC）直接降低肌丝的Ca 2+敏感性，导致偏心性 肥大、收缩功能障碍和左心室扩张。重要的是，该突变体的表达可以是 用强力霉素暂时控制，以专门测试我们的中心假设。在这篇文章中，我将（1） 确定I61 Q cTnC表达对肌细胞、基质和染色质可及性的时间依赖性影响， 2)检查DCM重塑在不同阶段的心肌细胞、基质和染色质可及性的可逆性 3）确定肌细胞是否保留了机械不平衡引起的表观遗传记忆 DCM的。提高我们对扩张型心肌病重塑的时间依赖性可逆性的理解， DCM患者的治疗目标和治疗窗口。此外，该项目的完成将 加强贝拉的训练，作为一个独立的科学家，并准备她有一天成为一名教授， 心血管工程学获得NIH F31博士前奖学金将促进重要的实验 和培训，这将有助于她追求这一职业目标。在这个项目中，贝拉将获得多组学的专业知识， 方法-如蛋白质组学，RNAseq和ATACseq-由于其 无偏见的筛选能力。华盛顿大学基因组学核心将帮助贝拉学习如何有效地使用这些 工具，这不仅有利于这个项目，但也将是一个令人难以置信的有用的技能贝拉的未来职业生涯。 鉴于该项目的临床相关性，我们聘请了代理医生Farid Moussavi-Harami，MD- 在华盛顿大学医学部从事心脏病学工作的科学家。穆萨维-哈拉米博士作为 临床医生将是至关重要的，以确保我们的研究问题和目标仍然与患者有关，他的 整个项目的指导将加强贝拉的培训和职业发展，因为她的目标是 最终经营一个实验室，重点是心血管疾病的转化疗法和技术。

Project Summary Dilated cardiomyopathy (DCM) is a highly prevalent inherited cardiac disease, characterized by systolic dysfunction, eccentric hypertrophy, and left ventricle dilation. While pharmacologic and mechanical treatments have been shown to partially improve cardiac function, these results are often short lived and highly variable from patient to patient. Moreover, recent studies have demonstrated that epigenetic and matrix dysregulation can persist, even in patients with improved systolic function after treatment. Given that aberrant chromatin remodeling and extracellular matrix (ECM) deposition have been identified as drivers of dilated remodeling—and that chromatin and ECM remodeling can become irreversible over time—it is crucial to understand the time- dependent effects of DCM mutations on reversing maladaptive remodeling at the genome, myocyte, and matrix levels. The central hypothesis of this proposal is that the reversibility of the DCM phenotype is time-dependent due to the accumulation of permanent ECM and chromatin remodeling as the disease progresses. To complete this proposal, I will utilize a DCM mouse model created by the Davis lab which contains a mutation (I61Q) in cardiac troponin C (cTnC) that directly decreases the myofilament’s Ca2+ sensitivity, leading to eccentric hypertrophy, systolic dysfunction, and left ventricle dilation. Importantly, expression of this mutant can be temporally controlled with doxycycline to specifically test our central hypothesis. In this proposal I will 1) Determine the time-dependent effects of I61Q cTnC expression on myocyte, matrix, and chromatin accessibility, 2) Examine the reversibility of DCM remodeling in myocyte, matrix, and chromatin accessibility at different stages of the disease, and 3) Determine if myocytes retain epigenetic memories of the mechanical disequilibrium caused by DCM. Improving our understanding of the time-dependent reversibility of DCM remodeling will better inform therapeutic targets and treatment windows for patients with DCM. Moreover, completion of this project will enhance Bella’s training as an independent scientist and prepare her to one day become a professor in cardiovascular engineering. Receiving the NIH F31 predoctoral fellowship will facilitate important experiments and training that will aid in her pursuit of this career goal. In this project, Bella will gain expertise in multi-omic approaches—such as proteomics, RNAseq, and ATACseq—which are growing in popularity due to their unbiased screening capabilities. The UW Genomics Core will assist Bella in learning how to effectively use these tools, which will not only benefit this project but will also be an incredibly useful skillset for Bella’s future career. Given the clinical relevance of this project, we have engaged Farid Moussavi-Harami, MD, an acting physician- scientist who practices cardiology within the UW Department of Medicine. Dr. Moussavi-Harami’s input as a clinician will be crucial for ensuring our research questions and aims remain relevant to patients, and his mentorship throughout this project will enhance Bella’s training and career development as she aims to eventually run a lab with an emphasis in translational therapeutics and technologies for cardiovascular diseases.