Developmental regulation of cranial tendon fibroblast diversity and ECM interactions

颅腱成纤维细胞多样性和 ECM 相互作用的发育调节

• 批准号: 10446059
• 负责人: Thomas F Schilling
• 金额: $ 37.77万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10446059
• 关键词: Address; Aging; Atrophic; Binding Proteins; Cells; Cephalic; Contracts; Data; Development; Disease; Embryo; Extracellular Matrix; Feedback; Fibroblasts; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Genetic; Genetic Diseases; Genetic Transcription; Genetic study; Goals; Head; Heterogeneity; Human; Image; In Situ; Knowledge; Ligaments; Limb structure; Location; Maintenance; Mesoderm; Methods; Molecular; Morphogenesis; Mus; Muscle; Muscle Contraction; Musculoskeletal Development; Musculoskeletal System; Neural Crest Cell; Paralysed; Pharmacological Treatment; Pharmacology; Physiological; Play; Population; Production; Property; Regulation; Research; Role; Scaffolding Protein; Shock; Signal Transduction; Site; Specific qualifier value; Tendon Injuries; Tendon structure; Testing; Thrombospondins; Tissues; Transforming Growth Factor beta; Tretinoin; Ursidae Family; Variant; Weight-Bearing state; Zebrafish; absorption; base; bone; cell type; craniofacial; developmental genetics; effective therapy; experimental study; genetic manipulation; in vivo; in vivo imaging; insight; ligament injury; mechanical force; mechanical load; mutant; novel; progenitor; repaired; response; scaffold; scleraxis; single-cell RNA sequencing; skeletal; stem; stem cells; tendon development; thrombospondin 4; transcription factor; transcriptomics

Project Summary/Abstract Tendons and ligaments are fundamental components of a functional musculoskeletal system. Tendons attach muscles to bone and interact with these tissues at distinct attachment sites called entheses (bone) and myotendinous junctions (MTJs, muscle). These interactions cause unique changes in gene expression and extracellular matrix (ECM) production that allow tendons to bear the forces exerted by muscle contraction. Transcription factors such as Scleraxis (Scx) specify early tendon progenitor cells (TPCs) and regulate ECM production. We previously discovered a critical ECM scaffolding protein called Thrombospondin-4b (Tsp4b) in zebrafish that is regulated by Scx, required for tendon maintenance and conserved in human tendons. How different types of tendon fibroblasts (tenocytes) are specified and influence ECM assembly at entheses or MTJs remains unclear. The current proposal addresses these issues using the advantages of the zebrafish for single cell RNA sequencing, in vivo imaging and genetic manipulation. The long-term goal of the proposed research is to understand the spatial dynamics of gene regulation and ECM assembly in tendons/ligaments and their regulation by mechanical force. Three primary hypotheses guide the research: 1) distinct subtypes of TPCs develop at entheses and MTJs in response to force, 2) ECM secreted by tenocytes regulates force-dependent signals that alter these distinct modes of gene expression in tenocytes and 3) retinoic acid is a novel force-dependent signal controlling tendon development. Aim 1 will perform scRNA-seq in tenocytes and see how force alters gene expression profiles. Aim 2 will study roles for Tsp4b, ECM, and TGF-beta signaling in force-dependent gene expression in tendons. Aim 3 will study the roles of RA signaling in tendons, and its responses to mechanical load. Each aim combines novel single cell approaches, genetic manipulation, live imaging and quantitative methods for physiological stimulation of muscles to get at mechanisms of tendon cell specification and ECM assembly in response to force.

项目摘要/摘要 肌腱和韧带是功能肌肉骨骼系统的基本组成部分。肌腱 将肌肉连接到骨骼上，并在不同的连接位置与这些组织相互作用，称为凹陷 (骨)和肌腱连接(MTJ、肌肉)。这些相互作用导致基因的独特变化 表达和产生细胞外基质(ECM)使肌腱承受由 肌肉收缩。转录因子如硬化轴(SCX)指定早期肌腱祖细胞 (TPC)和监管ECM生产。我们之前发现了一种关键的细胞外基质支架蛋白 在斑马鱼中被称为血栓反应蛋白-4b(Tsp4b)，受SCX调节，是肌腱所必需的 维持和保存在人体肌腱中。不同类型的肌腱成纤维细胞(腱细胞) 以及对端部或MTJ的ECM组装的影响仍不清楚。目前的提案 利用斑马鱼在体内进行单细胞RNA测序的优势解决这些问题 成像和基因操作。拟议研究的长期目标是了解 肌腱/韧带中基因调控和细胞外基质组装的空间动力学及其调控 机械力。指导这项研究的三个主要假设：1)不同亚型的TPC发生 2)肌腱细胞分泌的细胞外基质调节力依赖性 在腱细胞中改变这些不同基因表达模式的信号和维甲酸是一种新的 肌腱发育依赖于力的信号。目标1将在腱细胞中执行scRNA-seq和 看看力是如何改变基因表达谱的。AIM 2将研究Tsp4b、ECM和转化生长因子-β的作用 肌腱中力依赖基因表达的信号转导。目标3将研究RA信号在 肌腱及其对机械载荷的反应。每个目标都结合了新的单细胞方法， 基因操作、活体成像和肌肉生理刺激的定量方法 获得肌腱细胞规格和ECM组装对力的响应机制。