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）导致大量发病率和死亡率，但几乎没有有效的治疗方法。用于治疗PAD的促进血管生成的策略几乎没有成功，这导致了对基于干细胞的干预的研究。基于细胞的方法的有益作用通常被认为是由辅助作用引起的，例如血管生长因子的局部分泌，尽管对缺血肌肉中的驻留肌肉祖细胞是否可以分化成新血管以及如何实现这一点知之甚少。我们实验室的初步研究表明，共伴侣蛋白Bag 3在保护肌肉组织免受缺血中起着重要作用。已知Bag 3在肌肉中起重要作用，因为小鼠中Bag 3的缺失导致暴发性肌病，并且人类Bag 3的突变与遗传性心肌病有关。我们研究了Bag 3的肢体缺血后，它被确定为肢体存活（Lsq 1）数量性状基因座（QTL）的组成部分的影响。除了对肌肉细胞的影响之外，我们意外地发现，用腺相关病毒（AAV 6）外源性肌肉特异性表达Bag 3诱导了非缺血和缺血肌肉中血管生成的显著增加。此外，Bag 3在肌肉中的表达显著增加血管成熟，如平滑肌肌动蛋白（SMA）阳性血管所证明的。Bag 3诱导Pax 7+祖细胞的增加，Pax 7+祖细胞对SMA和CD 31染色呈阳性，表明Bag 3诱导血管祖细胞的增加。肌肉含有相对大量的Pax 7+祖细胞，其通过增生而非肥大在出生后肌肉再生中发挥重要作用。有证据表明Pax 7+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Bag 3在肌肉中的表达驱动Pax 7+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。为了验证这一假设，本发明的具体目的是：1）确定在体内或体外骨骼肌细胞中外源性表达Bag 3后Pax 7+肌肉祖细胞是否可以分化为内皮细胞或内皮细胞;和2）确定在体内骨骼肌中表达Bag 3后Pax 7+细胞是否是血管生成和血管成熟所需的。该探索性R21提案是为了响应PA-09-248“用于心脏、肺和血液疾病的细胞基疗法的定向干细胞分化”而提交的，其目标是确定骨骼肌特异性Bag 3表达对Pax 7+祖细胞分化为血管细胞的影响，作为PAD的潜在治疗。实现这一建议的具体目标将为骨骼肌组织中的肌肉干细胞分化和血管与肌肉细胞之间的串扰机制提供重要的见解。此外，这些研究可能导致有效的治疗PAD的疗法。