Understanding and regulating skeletal muscle progenitor and stem cell states through metabolic and epigenetic modulation

通过代谢和表观遗传调节了解和调节骨骼肌祖细胞和干细胞状态

• 批准号: 10654658
• 负责人: Peggie Jane Chien
• 金额: $ 4.4万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10654658
• 关键词: Adult; Affect; Binding; Biological Assay; Biology; Birth; California; Cell Maturation; Cell Therapy; Cells; Cellular Assay; Chromatin; Citric Acid Cycle; Development; Duchenne muscular dystrophy; Dystrophin; Embryo; Enzymes; Epigenetic Process; Fetal Skeleton; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Glycolysis; Homeostasis; Human; Human body; Isotope Labeling; Laboratory Finding; Link; Los Angeles; Measures; Mediating; Mentorship; Metabolic; Metabolic Pathway; Metabolism; Molecular; Muscle; Muscle Cells; Muscle Fibers; Muscle function; Muscle satellite cell; Muscular Atrophy; Myoblasts; Natural regeneration; Nucleic Acid Regulatory Sequences; Nutrient; Oxidative Phosphorylation; Pathway interactions; Patients; Play; Population; Process; Proteins; Regenerative Medicine; Research; Resources; Role; Scientist; Skeletal Muscle; Time; Tissues; Transplantation; Transposase; Universities; Wasting Syndrome; Work; cell replacement therapy; differentiation protocol; enzyme activity; epigenetic regulation; epigenome; fetal; gene regulatory network; human pluripotent stem cell; improved; loss of function mutation; male; metabolic profile; metabolomics; muscle degeneration; muscle regeneration; myogenesis; overexpression; postnatal; regeneration potential; regenerative; regenerative tissue; satellite cell; self-renewal; single-cell RNA sequencing; small molecule; stem cells; transcription factor; uptake

Project Summary Skeletal muscle is one of the most regenerative tissues in the human body. Skeletal muscle progenitor cells (SMPCs) contribute to developmental myogenesis, and skeletal muscle stem cells (satellite cells, SCs) contribute to postnatal muscle homeostasis and regeneration. In Duchenne Muscular Dystrophy (DMD), a prevalent but fatal X-linked muscle wasting disease that affects 1:5000 live male births, a loss-of-function mutation in the DMD gene results in the absence of functional dystrophin protein to stabilize muscle fibers. This leads to continuous damage from repeated muscle degeneration and regeneration and compromises the regenerative ability of SCs. There is currently no cure for DMD. Differentiating human pluripotent stem cells (hPSCs) into SCs is a valuable resource for developing cell replacement therapies for DMD. However, current hPSC directed myogenic differentiation protocols result in embryonic/fetal-like SMPCs that cannot be maintained over multiple passages (i.e. self-renew) in culture. SMPCs also do not engraft as efficiently as postnatal SCs, and it is not known how to mature SMPCs to SCs because it is unclear how they molecularly differ. Metabolism plays a key role in regulating cell state and function in stem cells across development. We performed single cell RNA sequencing analysis that demonstrated that expression of most genes in glycolysis, tricarboxylic cycle, and oxidative phosphorylation decrease across human myogenic development. However, how metabolic activity supports SMPCs or can be used to transition SMPCs to SCs in humans has not been investigated beyond the transcriptional level. By more closely evaluating the metabolic profiles of tissue-derived SCs and hPSC-derived myogenic subpopulations including hPSC-derived SMPCs (hPSC-SMPCs), specific metabolic enzymes and metabolites in the aforementioned pathways will be targeted as candidates to support hPSC-SMPC self-renewal or promote hPSC-SMPC maturation to SCs in culture. Tightly connected to metabolism is the epigenome which has a direct role in transitioning between cell states, particularly through modulating chromatin accessibility at cis-regulatory regions to mediate transcription factor (TF) binding and control gene expression. Evaluating differences in chromatin accessibility of TF binding motifs between the same myogenic populations identifies TFs that support and regulate SMPC and SC states. Several TF candidates have been identified as potential regulators of SMPC-to-SC maturation. Expression of TF candidates will be modulated to support hPSC-SMPCs or promote their maturation toward SC fate. This work will for the first time shed light on the metabolic and epigenetic roles in maintaining and transitioning between human muscle progenitor and stem cell states. These findings will enhance the regenerative potential of hPSC-derived muscle cells and enable the development of improved myogenic cell therapies for DMD. This study will be performed at the University of California, Los Angeles under the mentorship of Dr. April Pyle, an expert in hPSC and skeletal muscle biology, in my endeavor to becoming a research scientist in the field of regenerative medicine.

项目概要 骨骼肌是人体再生能力最强的组织之一。骨骼肌祖细胞 (SMPC) 有助于发育肌生成和骨骼肌干细胞（卫星细胞，SC） 有助于产后肌肉稳态和再生。在杜氏肌营养不良症 (DMD) 中， 流行但致命的 X 连锁肌肉萎缩疾病，影响 1:5000 活产男性，这是一种功能丧失 DMD 基因突变导致缺乏功能性肌营养不良蛋白来稳定肌纤维。这 导致反复的肌肉退化和再生造成持续损伤，并损害 SC 的再生能力。目前 DMD 尚无治愈方法。分化人类多能干细胞 (hPSC) 转化为 SC 是开发 DMD 细胞替代疗法的宝贵资源。然而，目前 hPSC 定向生肌分化方案导致胚胎/胎儿样 SMPC 无法维持 文化中的多个阶段（即自我更新）。 SMPC 的移植效率也不如出生后 SC， 目前尚不清楚如何将 SMPC 成熟为 SC，因为尚不清楚它们在分子上有何不同。代谢 在干细胞发育过程中调节细胞状态和功能方面发挥着关键作用。我们进行了单细胞 RNA 测序分析表明，大多数基因在糖酵解、三羧酸循环和 人类肌原性发育过程中氧化磷酸化减少。然而，代谢活动如何 在人类中支持 SMPC 或可用于将 SMPC 转变为 SC 尚未进行过研究 转录水平。通过更仔细地评估组织源性 SC 和 hPSC 源性的代谢特征 肌源性亚群，包括 hPSC 衍生的 SMPC (hPSC-SMPC)、特定代谢酶和 上述途径中的代谢物将作为支持 hPSC-SMPC 自我更新的候选物 或促进 hPSC-SMPC 在培养物中成熟为 SC。与新陈代谢密切相关的是表观基因组 在细胞状态之间的转变中具有直接作用，特别是通过调节染色质可及性 顺式调控区介导转录因子 (TF) 结合并控制基因表达。评估中 相同生肌群之间 TF 结合基序染色质可及性的差异可确定 支持和监管 SMPC 和 SC 状态的 TF。几位 TF 候选人已被确定为潜在候选人 SMPC 到 SC 成熟的调节因子。将调节 TF 候选者的表达以支持 hPSC-SMPC 或者促进他们走向 SC 命运的成熟。这项工作将首次揭示新陈代谢和 表观遗传在人类肌肉祖细胞和干细胞状态之间的维持和转变中发挥作用。这些 研究结果将增强 hPSC 来源的肌肉细胞的再生潜力，并促进 改进了 DMD 的生肌细胞疗法。这项研究将在加州大学洛杉矶分校进行 在 hPSC 和骨骼肌生物学专家 April Pyle 博士的指导下，Angeles 在我的努力下 成为再生医学领域的研究科学家。