Muscle-Resident Stem Cells for Angiogenesis and Vascular Maturation in PAD

用于 PAD 血管生成和血管成熟的肌肉驻留干细胞

• 批准号: 8707552
• 负责人: Christopher D Kontos
• 金额: $ 18.75万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8707552
• 关键词: Arteries; Automobile Driving; BCL2 gene; Basal lamina; Blood Vessels; CSPG4 gene; Cardiomyopathies; Cell Therapy; Cells; Coronary heart disease; Dependovirus; Diphtheria Toxin; Electrons; Embryo; Endothelial Cells; Erythrocytes; Flow Cytometry; Genetic Variation; Goals; Growth; Growth Factor; Heart Diseases; Hematological Disease; Heterogeneity; Human; Hyperplasia; Hypertrophy; Immunofluorescence Microscopy; In Vitro; Inherited; Intervention; Investigation; Ischemia; Knockout Mice; Label; Lead; Limb structure; Link; Lower Extremity; Lung diseases; Mediating; Modality; Molecular Chaperones; Morbidity - disease rate; Mus; Muscle; Muscle Cells; Muscle Fibers; Muscle function; Muscle satellite cell; Mutation; Myoblasts; Myopathy; Myosin Heavy Chains; Obstruction; PECAM1 gene; Pericytes; Peripheral; Peripheral arterial disease; Play; Prevention; Production; Proteins; Quantitative Trait Loci; Role; Sarcolemma; Serotyping; Skeletal Muscle; Smooth Muscle; Smooth Muscle Actin Staining Method; Source; Staining method; Stains; Stem cells; Tamoxifen; Testing; angiogenesis; base; cell growth; control trial; density; effective therapy; in vivo; insight; interest; meetings; mortality; muscle necrosis; muscle regeneration; neovascularization; postnatal; promoter; public health relevance; response; satellite cell; stem; stem cell differentiation; success; tissue regeneration

DESCRIPTION (provided by applicant): Peripheral artery disease (PAD) causes substantial morbidity and mortality but has few effective treatments. Strategies to promote angiogenesis for the treatment of PAD have met with little success, which has led to the investigation of stem cell-based interventions. Beneficial effects of cell-based approaches are generally thought to result from accessory effects, such as the local secretion of vascular growth factors, although less is known about whether resident muscle progenitor cells in ischemic muscle can differentiate into new blood vessels and how this can be achieved. Preliminary studies in our lab have demonstrated that the co-chaperone protein Bag3 plays an important role in the protection of muscle tissue from ischemia. Bag3 is known to play an important role in muscle, as loss of Bag3 in mice results in a fulminant myopathy, and mutation in human BAG3 has been linked to inherited cardiomyopathies. We investigated Bag3's effects in limb ischemia after it was identified as a component of the limb survival (Lsq1) quantitative trait locus (QTL). In addition t effects on muscle cells, we unexpectedly found that exogenous muscle-specific expression of Bag3 with adeno-associated virus (AAV6) induced a dramatic increase in angiogenesis in non-ischemic and ischemic muscle. Moreover, Bag3 expression in muscle significantly increased vascular maturation, as evidenced by smooth muscle actin (SMA)-positive vessels. Bag3 induced an increase in Pax7+ progenitor cells, which stained positive for SMA and CD31, suggesting that Bag3 induced an increase in vascular progenitor cells. Muscle contains a relatively large number of Pax7+ progenitor cells, which play an important role in postnatal muscle regeneration through hyperplasia but not hypertrophy. Evidence has linked Pax7+ progenitor cells to both endothelial and pericyte lineages. Based on these observations, we hypothesize that expression of Bag3 in muscle drives differentiation of Pax7+ cells into vascular cells, which may provide a source of endogenous vascular stem cells for the treatment of PAD. To test this hypothesis, the Specific Aims of this proposal are to: 1) Determine whether Pax7+ muscle progenitor cells can differentiate into endothelial or peri- endothelial cells after exogenous expression of Bag3 in skeletal muscle cells in vivo or in vitro; and 2) Determine whether Pax7+ cells are required for angiogenesis and vascular maturation after Bag3 expression in skeletal muscle in vivo. This exploratory R21 proposal is submitted in response to PA-09-248, "Directed Stem Cell Differentiation for Cell-Based Therapies for Heart, Lung, and Blood Diseases", with the goal of determining the effects of skeletal muscle-specific Bag3 expression on the differentiation of Pax7+ progenitor cells into vascular cells as a potential treatment for PAD. Accomplishing the Specific Aims of this proposal will provide important insights into the mechanisms of muscle stem cell differentiation and crosstalk between blood vessels and muscle cells in skeletal muscle tissue. Furthermore, these studies may lead to effective therapies for the treatment of PAD.

描述（由申请人提供）：外周动脉疾病（PAD）导致大量的发病率和死亡率，但有效的治疗方法很少。用于治疗 PAD 的促进血管生成的策略收效甚微，这导致了基于干细胞的干预措施的研究。基于细胞的方法的有益效果通常被认为是由附加效应产生的，例如血管生长因子的局部分泌，尽管人们对缺血肌肉中的常驻肌肉祖细胞是否可以分化成新血管以及如何分化知之甚少。实现了。我们实验室的初步研究表明，辅助伴侣蛋白Bag3在保护肌肉组织免受缺血方面发挥着重要作用。 Bag3 已知在肌肉中发挥重要作用，因为小鼠中 Bag3 的缺失会导致暴发性肌病，而人类 BAG3 的突变与遗传性心肌病有关。 Bag3 被确定为肢体存活 (Lsq1) 数量性状基因座 (QTL) 的组成部分后，我们研究了 Bag3 对肢体缺血的影响。除了对肌肉细胞的影响外，我们意外地发现，腺相关病毒（AAV6）外源性肌肉特异性表达Bag3可诱导非缺血性和缺血性肌肉中血管生成的显着增加。此外，平滑肌肌动蛋白（SMA）阳性血管证明，肌肉中的 Bag3 表达显着增加血管成熟。 Bag3 诱导 Pax7+ 祖细胞增加，其 SMA 和 CD31 染色呈阳性，表明 Bag3 诱导血管祖细胞增加。肌肉中含有相对较多的Pax7+祖细胞，它们通过增生而不是肥大在出生后肌肉再生中发挥重要作用。有证据表明 Pax7+ 祖细胞与内皮细胞和周细胞谱系有关。基于这些观察，我们假设肌肉中 Bag3 的表达驱动 Pax7+ 细​​胞分化为血管细胞，这可能为 PAD 的治疗提供内源性血管干细胞的来源。为了检验这一假设，本提案的具体目标是： 1）确定Pax7+肌肉祖细胞在体内或体外在骨骼肌细胞中外源表达Bag3后是否可以分化为内皮细胞或内皮周细胞； 2)确定体内骨骼肌中Bag3表达后血管生成和血管成熟是否需要Pax7+细胞。此探索性 R21 提案是为了响应 PA-09-248“心脏、肺和血液疾病细胞疗法的定向干细胞分化”而提交的，目的是确定骨骼肌特异性 Bag3 表达对Pax7+祖细胞分化为血管细胞作为PAD的潜在治疗方法。实现该提案的具体目标将为了解肌肉干细胞分化以及骨骼肌组织中血管和肌肉细胞之间的串扰机制提供重要见解。此外，这些研究可能会带来治疗 PAD 的有效疗法。