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(申请人提供)：外周动脉疾病(PAD)导致相当大的发病率和死亡率，但几乎没有有效的治疗方法。促进血管生成治疗PAD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辅助作用的结果，例如局部分泌血管生长因子，尽管关于缺血肌肉中的驻留肌肉前体细胞是否能分化成新的血管以及如何实现这一点还知之甚少。我们实验室的初步研究表明，辅助伴侣蛋白Bag3在肌肉组织缺血保护中发挥着重要作用。众所周知，Bag3在肌肉中起着重要的作用，因为小鼠失去Bag3会导致暴发性肌病，而人类BAG3的突变与遗传性心肌病有关。我们研究了Bag3‘S在肢体缺血中的作用，因为它被确定为肢体存活(LSQ1)数量性状基因座的组成部分。除了对肌肉细胞的影响外，我们还意外地发现，外源性Bag3和腺相关病毒(AAV6)的肌肉特异性表达显著增加了非缺血和缺血肌肉的血管生成。此外，肌肉中Bag3的表达显著促进了血管的成熟，这一点从平滑肌肌动蛋白(SMA)阳性的血管中得到了证明。Bag3诱导Pax7+祖细胞增多，其SMA和CD31染色阳性，提示Bag3诱导血管祖细胞增多。肌肉中含有较多的Pax7+祖细胞，它们通过增殖而不是肥大在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系有关。基于这些观察结果，我们推测Bag3在肌肉中的表达促进了Pax7+细胞向血管细胞的分化，这可能为PAD的治疗提供了内源性血管干细胞的来源。为了验证这一假说，这一提议的具体目的是：1)确定外源Bag3在体内或体外表达Bag3后，Pax7+肌祖细胞是否能分化为内皮细胞或内皮周细胞；2)确定Bag3在体内表达后，Pax7+细胞是否需要Pax7+细胞来促进血管生成和血管成熟。这份探索性的R21提案是为了响应PA-09-248《心、肺和血液疾病的细胞治疗的定向干细胞分化》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的影响，作为PAD的潜在治疗方法。完成这一建议的具体目标将为骨骼肌组织中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，这些研究可能为PAD的治疗提供有效的治疗方法。