Engineering a Human Skeletal Muscle Tissue Model of LGMD2B

设计 LGMD2B 的人体骨骼肌组织模型

基本信息

批准号：
10719721
负责人：
Nenad Bursac
金额：
$ 52.59万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-15 至 2028-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10719721
关键词：
3-Dimensional Adipose tissue Age Animal Model Beds Biochemical Biopsy Calcium Cell Culture Techniques Cell Separation Cell membrane Cells Cholesterol Cholesterol Homeostasis Clinical Clinical Trials Coculture Techniques Complement Complex DYSF gene Disease Disease Progression Disease model Engineering Environment Esterification Exercise Exhibits Fatty acid glycerol esters Functional disorder Generations Genes Goals Histologic Homeostasis Human Impairment In Vitro Infiltration Inflammation Inflammatory Injury Intramuscular Investigation Limb structure Limb-Girdle Muscular Dystrophies Lipids Macrophage Mediating Membrane Membrane Proteins Metabolic Mitochondria Modeling Mus Muscle Muscle Cells Muscle Fibers Muscle Weakness Muscle satellite cell Muscular Atrophy Muscular Dystrophies Mutation Myocardium Myopathy Organelles Pathogenicity Patients Phagocytosis Pharmaceutical Preparations Pharmacology Study Phenotype Play Process Proteins Regulation Reproducibility Role Severity of illness Signal Pathway Skeletal Muscle Source Study models System Testing Time Tissue Engineering Tissue Model Tissues Toxin acetyl-LDL cell motility cholesterol biosynthesis cytokine disease phenotype drug discovery dysferlinopathies experience human model immune cell infiltrate in vitro Model in vivo induced pluripotent stem cell inflammatory milieu interstitial cell lipid metabolism muscle engineering new therapeutic target novel organ growth organ on a chip pharmacologic pre-clinical repaired response response to injury therapeutic target translational study uptake

项目摘要

Limb girdle muscular dystrophy 2B (LGMD2B) is a late-onset progressive muscular dystrophy resulting from mutations in the dysferlin gene. Dysferlin is a membrane-associated protein, highly expressed in skeletal and cardiac muscle fibers where it orchestrates membrane repair in response to various injuries. Currently, there are no ongoing clinical trials or therapies to slow disease progression or cure LGMD2B. While useful for in vivo mechanistic studies, dysferlin-deficient (BLAJ) mice, a model of LGDM2B, exhibit a mild disease phenotype compared to humans, limiting mouse utility for translational studies. Developing a high-fidelity in vitro model of human LGMD2B muscle would complement mouse studies and allow patient-specific disease modeling and drug discovery. Thus, the overarching goal of this project is to engineer a novel 3D human skeletal muscle tissue model (“myobundle”) that replicates the main structural, functional, and metabolic features of LGMD2B. Specifically, we will utilize human iPSC lines from three healthy and three LGMD2B donors to engineer LGMD2B myobundles that exhibit reproducible deficits in muscle contractile function, calcium homeostasis, and lipid handling, while showing drug responses consistent with studies in BLAJ mice and LGMD2B patients. Importantly, a defining feature of LGMD2B muscle is the ectopic fat formation suggested to occur due to adipogenic differentiation of muscle interstitial cells (MICs). We will thus develop a novel tissue-engineered model of intramuscular adipose tissue (IMAT) accumulation in LGMD2B muscle by co-culturing MICs isolated from LGMD2B human muscle biopsies and iPSC-derived muscle progenitor cells. In this novel co-culture system, we will identify pro-adipogenic factors secreted from LGMD2B muscle and study their ability to induce ectopic fat formation. Since immune cell infiltration and biased macrophage polarization are additional defining features of LGMD2B muscle, we will engineer co- and tri-cultured muscle-macrophage myobundles to further characterize roles of heterocellular interactions and inflammatory milieu in injury response and fat accumulation in LGMD2B. Finally, our preliminary studies suggest that the cholesterol metabolism in LGMD2B muscle is impaired and contributes to the disease, which we will further study pharmacologically, biochemically, and histologically in LGMD2B myobundles and BLAJ mice. Overall, we expect that the novel tissue-engineered model of human LGMD2B muscle developed in this project will enable new mechanistic and pharmacological studies, eventually leading to first clinical trials for LGMD2B.

肢带型肌营养不良症 2B (LGMD2B) 是一种迟发性进行性肌营养不良症，由 Dysferlin 基因突变是一种膜相关蛋白，在骨骼和细胞中高度表达。目前，心肌纤维负责协调膜修复以应对各种损伤。尚无正在进行的临床试验或疗法来减缓疾病进展或治愈 LGMD2B，但对体内有用。机制研究显示，dysferlin 缺陷 (BLAJ) 小鼠（LGDM2B 模型）表现出轻微的疾病表型与人类相比，限制了小鼠在转化研究中的应用。人类 LGMD2B 肌肉将补充小鼠研究并允许患者特定疾病建模和因此，该项目的首要目标是设计一种新型 3D 人体骨骼肌组织。模型（“myobundle”）复制了 LGMD2B 的主要结构、功能和代谢特征。具体来说，我们将利用来自三名健康人和三名 LGMD2B 供体的人类 iPSC 系来设计 LGMD2B 肌束在肌肉收缩功能、钙稳态和脂质方面表现出可重复的缺陷处理，同时显示与 BLAJ 小鼠和 LGMD2B 患者研究一致的药物反应。 LGMD2B 肌肉的一个决定性特征是由于脂肪形成而发生的异位脂肪形成因此，我们将开发一种新型的组织工程模型。通过共培养分离自 LGMD2B 的 MIC 来增加肌内脂肪组织 (IMAT) 的积累 LGMD2B 人类肌肉活检和 iPSC 衍生的肌肉祖细胞在这个新颖的共培养系统中。将鉴定 LGMD2B 肌肉分泌的促脂肪因子并研究它们诱导异位脂肪的能力由于免疫细胞浸润和偏向巨噬细胞极化是额外的定义特征。 LGMD2B 肌肉，我们将设计共培养和三培养的肌肉巨噬细胞肌束以进一步表征异细胞相互作用和炎症环境在 LGMD2B 损伤反应和脂肪积累中的作用。最后，我们的初步研究表明 LGMD2B 肌肉中的胆固醇代谢受损，并且导致该疾病，我们将在药理学、生化和组织学方面进一步研究总体而言，我们期望新型的人类组织工程模型。该项目中开发的 LGMD2B 肌肉最终将实现新的机制和药理学研究导致 LGMD2B 的首次临床试验。