3D Bioprinted Human Model of Duchenne Muscular Dystrophy (DMD) Cardiomyopathy to Study Disease Progression with Imposed Force and Precise Gene Editing

杜氏肌营养不良症 (DMD) 心肌病的 3D 生物打印人体模型，通过施加力和精确的基因编辑来研究疾病进展

• 批准号: 10628962
• 负责人: Forum D Kamdar
• 金额: $ 53.93万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10628962
• 关键词: 3-Dimensional; 3D Print; Acceleration; Biological Models; CRISPR/Cas technology; Calcium; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cell Line; Cells; Complex; Cytoskeleton; Development; Disease; Disease Progression; Duchenne cardiomyopathy; Duchenne muscular dystrophy; Dystrophin; Early identification; Extracellular Matrix; Generations; Genes; Genetic Transcription; Glycoproteins; Goals; Health; Heart; Heart Diseases; Heart Injuries; Heart failure; Homeostasis; Human; Individual; Injury; Length; Link; Mechanical Stress; Mechanics; Mission; Modeling; Molecular; Muscle; Mutation; National Heart, Lung, and Blood Institute; Organ; Outcome; Patients; Phenotype; Physicians; Physiological; Pre-Clinical Model; Prevention; Proteins; Public Health; Pump; Research; Research Personnel; Research Project Grants; Role; Sarcolemma; Scientist; Signal Transduction; Skin; Technology; Testing; Tissues; adrenergic stress; base editing; bioink; bioprinting; cardiac tissue engineering; career; clinically significant; design; early satiety; effective therapy; exon skipping; functional restoration; heart cell; human disease; human model; induced pluripotent stem cell; induced pluripotent stem cell derived cardiomyocytes; innovation; mdx mouse; mechanical force; mechanical load; mechanotransduction; mouse model; muscular dystrophy mouse model; mutation correction; novel; novel therapeutics; pre-clinical; pressure; prevent; prime editing; protein expression; restoration; therapy development; transcriptome sequencing

Duchenne muscular dystrophy (DMD) cardiomyopathy is ubiquitous, deadly, and results from mutations in the dystrophin gene. Dystrophin is an essential component of cardiac mechanotransduction (MT) and distributes mechanical stress across the sarcolemma. In DMD, the absence of dystrophin results in cardiomyocytes that are vulnerable to contraction-induced damage, which accelerates disease progression. Our understanding of the early progression of DMD cardiomyopathy is limited due to models that do not fully recapitulate human disease, thus limiting development of effective therapies. Given the essential role of dystrophin in connecting the contractile apparatus to the extracellular matrix (ECM) for MT, it is critical to augment DMD cardiomyopathy models to include cell-ECM engagement with applied physiological force to recapitulate early changes at the organ level necessary to test novel therapies. The human chambered muscle pump (hChaMP) can do just that. The hChaMP is generated by 3D printing human induced pluripotent stem cells and bio-ink to yield a pump that can be pressurized to impose progressive strain. The long-term goal is to determine mechanisms by which DMD cardiomyopathy progresses to develop novel disease specific therapies to prevent cardiomyopathy. The overall objective is to assess the role of altered MT and ECM dynamics in the absence of dystrophin on DMD cardiomyopathy disease progression and to test dystrophin gene editing in a DMD hChaMP with volumetric loading. The central hypothesis is that the loss of dystrophin leads to increased vulnerability to mechanical stress resulting in early altered cardiac MT and ECM dynamics that promote disease progression and that early dystrophin replacement will limit contraction-induced injury by restoring MT and ECM homeostasis and thereby rescue DMD cardiomyopathy. Our central hypothesis will be tested in two specific aims: 1) To evaluate the impact of altered MT on DMD cardiomyopathy disease progression using the hChaMP model system with progressive volumetric loading; 2) To determine the impact of dystrophin restoration with DMD precise gene editing on cardiac remodeling mechanisms dictating disease progression in DMD cardiomyopathy. In aim 1, we will generate a DMD hChaMPs to assess the physiologic impact of increased volumetric pressure on the human DMD phenotype early and later. In aim 2, we will introduce DMD precise gene editing to restore dystrophin both early and late in DMD hChaMPs with volumetric loading and assess the physiologic and transcriptional changes. At the successful completion of the proposed research, the expected outcomes of our study is the characterization of early DMD cardiomyopathy progression with loading and the underlying molecular mechanisms. The proposed research is innovative as it combines enabling technologies to develop a 3D preclinical model of human DMD cardiomyopathy that mimics disease progression and correction with precise gene editing. These findings will have a significant impact on human health by increasing our understanding of disease progression and a strong basis for treating and preventing DMD cardiomyopathy.

杜氏肌营养不良症 (DMD) 心肌病普遍存在且致命，是由基因突变引起的 肌营养不良蛋白基因。抗肌营养不良蛋白是心脏机械转导 (MT) 的重要组成部分，并分布在 穿过肌膜的机械应力。在 DMD 中，肌营养不良蛋白的缺乏导致心肌细胞 容易受到收缩引起的损伤，从而加速疾病进展。我们的理解 由于模型不能完全再现人类，DMD 心肌病的早期进展受到限制 疾病，从而限制了有效疗法的开发。鉴于肌营养不良蛋白在连接 MT 的细胞外基质 (ECM) 收缩装置，对于增强 DMD 心肌病至关重要 模型包括细胞-ECM 与施加的生理力的相互作用，以重现细胞的早期变化 测试新疗法所需的器官水平。人体腔室肌肉泵 (hChaMP) 就能做到这一点。 hChaMP 通过 3D 打印人类诱导多能干细胞和生物墨水生成泵， 可以加压以施加渐进应变。长期目标是确定 DMD 的机制 心肌病不断发展，以开发新的疾病特异性疗法来预防心肌病。整体 目的是评估在 DMD 缺乏肌营养不良蛋白的情况下 MT 和 ECM 动力学改变的作用 心肌病疾病进展并在 DMD hChaMP 中测试肌营养不良蛋白基因编辑 加载中。核心假设是抗肌萎缩蛋白的丧失导致对机械应力的脆弱性增加 导致心脏 MT 和 ECM 动态的早期改变，从而促进疾病进展，并且早期 肌营养不良蛋白替代将通过恢复 MT 和 ECM 稳态来限制收缩引起的损伤，从而 拯救DMD心肌病。我们的中心假设将在两个具体目标上进行检验：1）评估 使用 hChaMP 模型系统改变 MT 对 DMD 心肌病疾病进展的影响 渐进式体积加载； 2) 确定DMD精确基因对肌营养不良蛋白恢复的影响 编辑心脏重塑机制决定 DMD 心肌病的疾病进展。在目标 1 中，我们 将生成 DMD hChaMP 来评估体积压力增加对人体的生理影响 DMD表型早期和晚期。在目标2中，我们将引入DMD精确基因编辑来恢复肌营养不良蛋白 在 DMD hChaMP 的早期和晚期进行体积负荷分析，并评估生理和转录变化。 在成功完成拟议的研究后，我们研究的预期结果是 早期 DMD 心肌病随负荷进展的特征和潜在的分子机制 机制。拟议的研究具有创新性，因为它结合了开发 3D 的支持技术 人类 DMD 心肌病的临床前模型，通过精确模拟疾病进展和纠正 基因编辑。这些发现将加深我们对人类健康的了解，从而对人类健康产生重大影响。 疾病进展以及治疗和预防 DMD 心肌病的坚实基础。