Physical Regulation of Muscle Stem Cells in Bioengineered 3D Models

生物工程 3D 模型中肌肉干细胞的物理调节

• 批准号: 8166019
• 负责人: Penney Marie Gilbert
• 金额: $ 8.63万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8166019
• 关键词: Accidents; Adult; Advisory Committees; Aging; Architecture; Award; Bed rest; Behavior; Biocompatible Materials; Bioinformatics; Biological Assay; Biology; Biomechanics; Biomedical Engineering; Cachexia; Cell Culture Techniques; Cell Fate Control; Cell Line; Cell Shape; Cell Survival; Cell Therapy; Cells; Clinical; Critiques; Cues; Cytoskeleton; Data; Defect; Development; Disease; Elasticity; Encapsulated; Engineering; Ensure; Environment; Extracellular Matrix; Fiber; Focal Adhesions; Force of Gravity; Foundations; Funding; Future; Gene Expression; Genetic; Goals; HIV Infections; Healthcare Systems; Hereditary Disease; Hydrogels; K-Series Research Career Programs; Knowledge; Laboratories; Learning; Malignant Neoplasms; Mechanics; Mediating; Mentors; Methods; Microfabrication; Microscopy; Modeling; Molecular; Movement; Mus; Muscle; Muscle Fibers; Muscle satellite cell; Muscular Atrophy; Muscular Dystrophies; Natural regeneration; Nature; Pathway interactions; Patients; Pattern; Physical environment; Postdoctoral Fellow; Property; Public Health; Quality of life; RNA; RNA Sequences; Regulation; Research; Research Personnel; Rest; Risk; Role; Skeletal Muscle; Skeletal muscle injury; Stem cells; System; Testing; Therapeutic; Time; Time Management; TimeLine; Tissue Engineering; Tissues; Training; Universities; Work; Writing; adult stem cell; aged; base; career; career development; cell assembly; cell behavior; design; in vivo; injured; insight; interdisciplinary approach; multidisciplinary; muscle form; muscle regeneration; new therapeutic target; novel strategies; physical property; prospective; regenerative; repaired; sarcopenia; satellite cell; scaffold; self-renewal; skeletal; skeletal muscle wasting; skills; stem cell biology; stem cell fate; stem cell niche; stemness; success; therapeutic development; three-dimensional modeling; wasting

DESCRIPTION (provided by applicant): Skeletal muscle wasting that occurs during aging and numerous other pathological conditions including HIV infection and cancer is a serious public health concern and was a financial burden of more than $18.5 billion to the U.S. healthcare system in 20001. Currently there are no effective strategies to promote muscle regeneration in aged, injured, diseased or physical inactive skeletal muscle. The inherent capacity of muscle tissue to regenerate itself is mediated by resident skeletal muscle stem cells (MuSCs)2,3,4. Recent advances by our laboratory and others have identified strategies to prospectively isolate MuSCs from murine skeletal muscle5,6,7,8,9,10,11. Due to the relatively nascent nature of the MuSC field, we are only just beginning to understand the mechanisms underlying MuSC regulation. Elucidating MuSC regulation will be critical to harness the potential of this adult, tissue-specific stem cell and lend to the development of therapeutics to overcome muscle atrophy. In recent studies we demonstrated that substrate elasticity, a physical property of the MuSC niche, is a potent regulator of MuSC viability and self-renewal12,13. Using a biomaterials approach in conjunction with in vivo functional assays, we showed for the first time that 'stemness' is maintained by culturing MuSCs on substrates that match the softness of muscle tissue. In this Career Development Proposal, we aim to extend this finding to gain further insight into MuSC regulation by physical features of the in vivo niche. Specifically, we will use a multidisciplinary approach to explore MuSC fate modulation by substrate rigidity and niche architecture and will focus on revealing mechanistic insights. In Aim 1, we will elucidate the mechanisms of MuSC regulation imposed by 2D substrate elasticity. This aim will test the hypothesis that MuSCs possess 'rigidity sensing' systems and that these systems transduce information about the physical environment to modify gene expression and ultimately, cell fate. The goal of Aim 2 is to engineer a skeletal muscle tissue mimic using engineered extra-cellular matrix (eECM) to interrogate MuSC physical regulation in the context of a 3D environment. We hypothesize that MuSC regulation by substrate rigidity will be fundamentally different in a 3D setting as compared to our 2D observations and aim to decouple the effects of 'dimensionality' on MuSC viability, proliferation and fate (quiescence, activation, self-renewal, differentiation). Finally, in Aim 3 we will investigate the role of 3D niche architecture on MuSC fate. We will use microfabricated models to test the hypothesis that MuSC quiescence and activation are determined by 3D niche architecture. The knowledge gained from these Aims will lend to our fundamental understanding of niche composition and MuSC regulation. The ultimate goal of this work is to potentiate development of cell-based or systemically delivered therapeutics designed to promote skeletal muscle regeneration in specific contexts of muscle wasting. If funded, this Career Development Award (CDA) would afford me the opportunity to expand my background in skeletal muscle biology, gain necessary knowledge of biomaterials approaches, achieve the capacity to interpret bioinformatics data and acquire expertise in microscopy; skills critical to my future success as an independent researcher (see timeline). Formal and informal interactions with my Advisory Committee will assess my progress on the proposed Aims and provide critique and advice. Finally, professional skills (e.g. mentoring, research presentation, time management, writing, etc) vital to my long term success as an academic will be gained during the K99 mentored period through Stanford courses designed to prepare postdocs for the career transition and through career development advice acquired by my advisory committee and my sponsor, Dr. Helen Blau. Together, the proposed studies and career development training will ensure I achieve my long term career goal; to establish a successful and independently-funded laboratory at a major University and use bioengineering approaches to study the molecular mechanisms that control muscle stem cells in the resting tissue and during regeneration specifically in the context of a 3D tissue. PUBLIC HEALTH RELEVANCE: Skeletal muscle is critical to our day to day movement and loss of muscle mass and/or function due to genetic disorders, aging, cancer or physical inactivity impairs quality of life and increases the risk of mobility related accidents. Adult skeletal muscle contains stem cells that are responsible for the day to day repair of skeletal muscle injury. The research in this proposal used an interdisciplinary approach and is focused on identifying the mechanisms regulating muscle stem cells in the tissue and aims to harness this knowledge to develop novel strategies for promote skeletal muscle repair in patients.

DESCRIPTION (provided by applicant): Skeletal muscle wasting that occurs during aging and numerous other pathological conditions including HIV infection and cancer is a serious public health concern and was a financial burden of more than $18.5 billion to the U.S. healthcare system in 20001. Currently there are no effective strategies to promote muscle regeneration in aged, injured, diseased or physical inactive skeletal muscle.肌肉组织重生本身的固有能力是由常驻骨骼肌干细胞（MUSC）介导的2,3,4。我们的实验室和其他人的最新进展已经确定了将MUSC与鼠骨骼肌分离的策略5,6,7,8,9,10,11。由于MUSC领域的相对较新的性质，我们才刚刚开始了解MUSC调节的基础机制。阐明MUSC调节对于利用该成年，组织特异性干细胞的潜力至关重要，并为克服肌肉萎缩的治疗剂发展。 在最近的研究中，我们证明了MUSC利基市场的物理特性底物弹性是MUSC生存能力和自我续订12,13的有效调节剂。我们使用生物材料方法与体内功能测定法结合使用，我们首次通过对与肌肉组织柔软度相匹配的底物上培养MUSC来保持“茎”。在这项职业发展计划中，我们旨在扩展这一发现，以进一步了解MUSC调节的体内特征。具体而言，我们将使用多学科的方法来探索底物刚性和利基架构的MUSC命运调制，并将专注于揭示机械洞察力。在AIM 1中，我们将阐明由2D底物弹性施加的MUSC调节机制。这个目标将检验MUSC具有“刚度传感”系统的假设，并且这些系统会传达有关物理环境的信息以修改基因表达，并最终导致细胞命运。目标2的目的是使用工程的细胞外基质（EECM）来设计模拟骨骼肌组织，以在3D环境的背景下询问MUSC物理调节。我们假设与我们的2D观测值相比，在3D设置中，底物刚性的MUSC调节将在根本上有所不同，并旨在使“维度”对MUSC生存能力，增殖和命运的影响（静止，激活，激活，自我更新，差异，差异化）。最后，在AIM 3中，我们将研究3D利基结构在MUSC命运中的作用。我们将使用微型制造模型来测试MUSC静止和激活的假设，该假设是由3D小裂结构确定的。从这些目标中获得的知识将使我们对利基组成和MUSC监管的基本理解。这项工作的最终目的是增强基于细胞或系统地交付的治疗剂的开发，旨在在肌肉浪费的特定情况下促进骨骼肌再生。 如果获得资助，该职业发展奖（CDA）将使我有机会扩大自己的骨骼肌肉生物学背景，获得对生物材料方法的必要知识，实现解释生物信息学数据并获得显微镜方面的专业知识的能力；对于我作为独立研究人员的未来成功至关重要的技能（请参阅时间表）。与我的咨询委员会的正式和非正式互动将评估我在拟议的目标上的进步，并提供批评和建议。最后，专业技能（例如，指导，研究演示，时间管理，写作等）对于我的长期成功至关重要的是，在K99指导期间通过斯坦福大学的课程将获得旨在为职业过渡和我的顾问委员会获得的职业发展咨询准备的斯坦福课程，并获得我的顾问委员会和我的赞助人Helen Blau博士。拟议的研究和职业发展培训将共同确保我实现长期职业目标；为了在一所主要大学建立成功且独立的实验室，并使用生物工程方法研究分子机制，这些分子机制可以控制静息组织中的肌肉干细胞，并在再生过程中特别是在3D组织中。 公共卫生相关性：由于遗传疾病，衰老，癌症或身体不活动而引起的肌肉运动和肌肉质量和/或功能的损失至关重要，骨骼肌肉至关重要。成年骨骼肌含有负责骨骼肌损伤日常修复的干细胞。该提案中的研究采用了跨学科方法，并致力于确定调节组织中肌肉干细胞的机制，并旨在利用这种知识来制定新的策略来促进患者的骨骼肌修复。