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Role of proteoglycan sulfation during muscle regeneration in dystrophic animals

蛋白多糖硫酸化在营养不良动物肌肉再生过程中的作用

基本信息

批准号：
9066087
负责人：
Matthew Tierney
金额：
$ 0.95万
依托单位：
SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-06-01 至 2016-09-23
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9066087
关键词：
Adult Animals Behavior Behavioral Binding Biological Assay Biological Availability Cells Communication Complex Cues Development Disease Progression Donor person Duchenne muscular dystrophy Environment Event Exhibits FGF2 gene Fetal Tissues Fibroblast Growth Factor Fibroblast Growth Factor Receptors Fluorescence-Activated Cell Sorting Gene Expression Goals Growth Health Heparan Sulfate Proteoglycan Impairment In Vitro Intervention Ligands Measures Mediating Mediator of activation protein Membrane Mitogens Molecular Monitor Mus Muscle Muscle satellite cell Muscular Atrophy Natural regeneration Pathway interactions Patients Pattern Phosphorylation Play Post-Translational Protein Processing Pre-Clinical Model Proteoglycan Quality of life Regulation Resistance Role Signal Pathway Signal Transduction Signaling Molecule Skeletal Muscle Staging Stem cells Sulfatases Testing Therapeutic Effect Transplantation Wasting Syndrome Western Blotting Work beta catenin bioluminescence imaging cell behavior extracellular fetal functional disability human disease improved in vivo innovation insight mouse model muscle regeneration myogenesis neutralizing antibody new therapeutic target pre-clinical prospective receptor regenerative repaired response self-renewal sulfation syndecan therapy design tool

项目摘要

DESCRIPTION (provided by applicant): Muscle stem cells (MuSC) are required for the development and repair of skeletal muscle. During these dynamic periods of muscle growth and regeneration, MuSC must selectively interpret a host of signaling molecules to self-renew. This ability is progressively impaired in Duchenne muscular dystrophy (DMD), a devastating muscle wasting disease for which there is no cure. DMD is characterized by altered microenvironmental signaling that negatively influences MuSC-mediated regenerative potential. To better understand how MuSC communicate with their extracellular environment, I explored several membrane-bound mediators of signal transduction. Heparan sulfate proteoglycans (HSPG), in particular syndecans, are highly expressed in MuSC and can regulate ligand bioavailability and receptor formation. Heterogeneous sulfation patterns on HSPG via post-translational modifications increase behavioral complexity, leading to a differential activity of critical signalng pathways. Because regeneration in the adult recapitulates several aspects of myogenesis, I focused my preliminary studies on HSPG sulfation in fetal MuSC, the developmental precursors of adult MuSC. I have demonstrated that Sulf1, a specific regulator of HSPG 6-O-sulfation, is downregulated in fetal MuSC. I provide evidence that fetal MuSC are resistant to myogenic commitment in vitro and capable of rapid in vivo expansion and long-term self-renewal following transplantation. Furthermore, uncommitted fetal MuSC are preferentially sensitive to the potent mitogen FGF2 while less responsive to Wnt/β-catenin-mediated myogenic differentiation. Indeed, Sulf1 has been previously shown to simultaneously promote Wnt/β-catenin and inhibit FGF2 signaling. Sulf1 and HSPG expression are dysregulated in muscle wasting conditions. Therefore, the goal of this project is to examine the role of HSPG 6-O-sulfation on the regulation of dystrophic MuSC self-renewal and regenerative potential. To accomplish this, I propose (Aim 1a) to validate HSPG sulfatase and FGF and Wnt/β-catenin signaling component expression levels throughout myogenesis and in dystrophic animals. To determine MuSC responsiveness to FGF and Wnt/β-catenin pathway activation (Aim 1b), I will measure the expression and phosphorylation levels of downstream signaling pathway effectors. Inhibition of HSPG 6-O-sulfation will ascertain its role in FGF receptor complex formation and Wnt3a ligand bioavailability, defining the mechanisms underlying HSPG regulation of proper pathway activation. (Aim 1c). I will determine the impact of Sulf1 inhibition on dystrophic, myofiber-associated MuSC self-renewal (Aim 2a). Finally, I will evaluate the functional role of HSPG 6-O-sulfation in a preclinical model of DMD by inhibiting Sulf1 expression in (Aim 2b) transplanted, donor MuSC and (Aim 2c) native, dystrophic MuSC. Through these studies I will investigate a molecular mechanism, HSPG 6-O-sulfation, able to regulate FGF and Wnt/β-catenin signaling and MuSC regenerative potential in dystrophic animals. This work will potentially identify Sulf1 as a novel therapeutic target for the enhancement of MuSC self-renewal and the amelioration of DMD.

描述（由申请人提供）：骨骼肌的发育和修复需要肌肉干细胞（MuSC）。在肌肉生长和再生的这些动态时期，MuSC必须选择性地解释许多信号分子以自我更新。这种能力在杜氏肌营养不良症（DMD）中逐渐受损，这是一种无法治愈的破坏性肌肉萎缩疾病。DMD的特征在于改变的微环境信号传导，其对MuSC介导的再生潜力产生负面影响。为了更好地了解MuSC如何与其细胞外环境通信，我探索了几种膜结合的信号转导介质。硫酸乙酰肝素蛋白聚糖（HSPG），特别是多配体聚糖，在MuSC中高度表达，并且可以调节配体生物利用度和受体形成。通过翻译后修饰的HSPG上的异质硫酸化模式增加了行为复杂性，导致关键信号通路的差异活性。由于再生在成人重演肌发生的几个方面，我集中我的初步研究胎儿MuSC，成人MuSC的发育前体的HSPG硫酸化。我已经证明，Sulf 1，一个特定的调节HSPG 6-O-硫酸化，下调胎儿MuSC。我提供的证据表明，胎儿MuSC在体外对肌源性定型具有抗性，并且能够在移植后在体内快速扩增和长期自我更新。此外，未定型的胎儿MuSC优先对有效的有丝分裂原FGF 2敏感，而对Wnt/β-连环蛋白介导的肌源性分化反应较低。事实上，Sulf 1先前已被证明可以同时促进Wnt/β-catenin和抑制FGF 2信号传导。Sulf 1和HSPG表达在肌肉萎缩条件下失调。因此，本项目的目标是研究HSPG 6-O-硫酸化对营养不良MuSC自我更新和再生潜力的调节作用。为了实现这一点，我建议（目的1a），以验证HSPG硫酸酯酶和FGF和Wnt/β-catenin信号转导成分的表达水平在整个肌发生和营养不良的动物。为了确定MuSC对FGF和Wnt/β-catenin通路激活的反应性（目的1b），我将测量下游信号通路效应物的表达和磷酸化水平。HSPG 6-O-硫酸化的抑制将确定其在FGF受体复合物形成和Wnt 3a配体生物利用度中的作用，从而确定HSPG调节适当途径活化的潜在机制。(Aim 1 c）。我将确定Sulf 1抑制对营养不良性肌纤维相关MuSC自我更新的影响（Aim 2a）。最后，我将通过抑制（Aim 2b）移植的供体MuSC和（Aim 2c）天然的营养不良MuSC中Sulf 1的表达来评估HSPG 6-O-硫酸化在DMD临床前模型中的功能作用。通过这些研究，我将研究一种分子机制，HSPG 6-O-硫酸化，能够调节营养不良动物中FGF和Wnt/β-catenin信号传导以及MuSC再生潜力。这项工作将有可能确定Sulf 1作为一种新的治疗靶点，用于增强MuSC自我更新和改善DMD。