Pathological arterial muscularization and the role of integrins

描述（由申请人提供）：心血管疾病对人类健康造成巨大负担，我们的长期目标是减轻血管平滑肌细胞（VSMC）的过度积累，这是许多这些疾病的核心。正常动脉的图尼卡中膜由VSMC和弹性膜交替的周向层组成。各种动脉疾病，包括动脉粥样硬化、再狭窄、肺动脉高压和瓣上主动脉瓣狭窄（SVAS）都受到弹性膜缺陷和肌化过度的困扰。SVAS是一种破坏性的人类疾病，其特征在于VSMC负荷增加，阻塞大动脉，由弹性蛋白基因ELN中的杂合无效突变引起。类似地，弹性蛋白突变的小鼠发展为大动脉（如主动脉）的肌肉过度化和狭窄。大血管手术是目前SVAS的唯一治疗方法，因为没有有效的药物选择。开发有效治疗血管疾病的主要障碍是对过量VSMC的细胞来源的理解不足;事实上，没有先前的研究追踪弹性蛋白突变体中任何细胞群体的谱系。此外，弹性蛋白突变体中主动脉肌化过度的分子和细胞机制尚未明确。我们的初步研究表明，整联蛋白β 3的表达和整联蛋白信号转导在弹性蛋白突变体主动脉中被强烈上调，并且β 3水平或活性的降低减弱了过度的肌肉化。此外，减少编码β 3的基因Itgb 3的剂量，延长了Eln（-/-）幼仔的生存力，这对于任何遗传或药理学干预都是前所未有的。减少SVAS中动脉过度肌化的一个潜在的有吸引力的策略是减弱增加的β 3表达。尽管对β 3表达的调控知之甚少，但生长停滞特异性同源盒（Gax）转录因子在VSMC中表达，Gax过表达降低β 3水平、增殖和迁移。我们假设在弹性蛋白突变体中，Gax下调诱导整合素β 3在预先存在的主动脉平滑肌中表达，导致VSMC异常定向、增殖和迁移，从而导致主动脉瓣狭窄。该建议利用转基因突变小鼠、从鼠主动脉分离的VSMC和人SVAS主动脉组织和细胞的研究来在三个特定目标中检验该假设：1）在弹性蛋白突变主动脉中，鉴定过量VSMC的细胞来源; 2）在弹性蛋白缺陷小鼠、鼠VSMC和人SVAS衍生的平滑肌细胞中，阐明整合素β 3诱导的过度主动脉肌化的分子和细胞机制; 3）阐明Gax在介导弹性蛋白突变体中整合素β 3表达增强和肌化过度中的作用。综上所述，我们提出的研究将阐明弹性蛋白缺乏、整合素β 3和主动脉肌肉过度化之间已确定的联系的机制，从而为开发SVAS和其他血管增生性疾病的新治疗策略提供具体步骤。

DESCRIPTION (provided by applicant): Cardiovascular diseases exert a massive burden on human health, and our long term objective is to attenuate the excessive accumulation of vascular smooth muscle cells (VSMCs) that is central to many of these diseases. The tunica media of the normal artery is composed of alternating circumferential layers of VSMCs and elastic lamellae. Diverse arterial disorders, including atherosclerosis, restenosis, pulmonary hypertension and supravalvular aortic stenosis (SVAS) are plagued by defective elastic lamellae as well as hypermuscularization. SVAS, a devastating human disease characterized by an increased VSMC burden that occludes large arteries, is caused by heterozygous null mutations in the elastin gene ELN. Similarly, elastin mutant mice develop hypermuscularization and stenosis of large arteries, such as the aorta. Major vascular surgery is the only current treatment for SVAS as no effective pharmacological options are available. A major obstacle to developing effective therapies for vascular disorders is the poor understanding of the cellular source(s) of excess VSMCs; indeed no prior studies have traced the lineage of any cell populations in elastin mutants. In addition, the molecular and cellular mechanisms underlying aortic hypermuscularization in elastin mutants are not well defined. Our initial studies indicate that integrin ß3 expression and integrin signaling is robustly upregulated in the elastin mutant aorta and that reduction of ß3 levels or activity attenuates the excessive muscularization. Furthermore, reduction of the dosage of the gene encoding ß3, Itgb3, extends the viability of Eln(-/-) pups which is unprecedented for any genetic or pharmacological intervention. A potentially attractive strategy for reducing arterial hypermuscularization in SVAS is attenuating the increased ß3 expression. Although little is known regarding regulation of ß3 expression, the growth arrest-specific homeobox (Gax) transcription factor is expressed in VSMCs and Gax overexpression reduces ß3 levels, proliferation and migration. We hypothesize that in elastin mutants, Gax downregulation induces integrin ß3 expression in pre-existing aortic smooth muscle resulting in aberrant VSMC orientation, proliferation and migration and thus aortic stenosis. This proposal utilizes studies of transgenic mutant mice, VSMCs isolated from the murine aorta and human SVAS aortic tissue and cells to test this hypothesis in three specific aims: 1) in the elastin mutant aorta, identify the cellular source(s) of excessive VSMCs; 2) in elastin deficient mice, murine VSMCs and human SVAS-derived smooth muscle cells, elucidate the molecular and cellular mechanisms underlying integrin ß3-induced excess aortic muscularization; and 3) elucidate the role of Gax in mediating enhanced integrin ß3 expression and hypermuscularization in elastin mutants. Taken together, our proposed studies will delineate mechanisms underlying the identified link between elastin deficiency, integrin ß3 and aortic hypermuscularization and thereby provide concrete steps towards developing novel therapeutic strategies for SVAS and other vasculoproliferative diseases.