Shear stress, endothelial miRNAs, and AV calcification

剪切应力、内皮 miRNA 和 AV 钙化

• 批准号: 10321908
• 负责人: Hanjoong Jo
• 金额: $ 53.94万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-12 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10321908
• 关键词: Antineoplastic Agents; Bicuspid; Biomechanics; Blood flow; Breast Microcalcification; Cancer Etiology; Cardiovascular system; Cells; Data; Development; Disease; Disease model; Drug Targeting; Endothelial Cells; Endothelium; Environment; Enzymes; Event; Exposure to; FDA approved; Family suidae; Fibrosis; Functional disorder; Funding; Gene Targeting; Genes; Genetic Transcription; Goals; HIF1A gene; Human; Hypertension; Hypoxia Inducible Factor; Hypoxia-Inducible Factor Pathway; In Vitro; Inflammation; Injections; Knowledge; Lead; Literature; Malignant Neoplasms; Mechanics; Mesenchymal; MicroRNAs; Molecular; Morbidity - disease rate; Mus; Pathogenesis; Pathogenicity; Pathologic; Pathway interactions; Pharmacology; Pharmacotherapy; Physiological; Play; Process; Risk Factors; Role; Sclerosis; Side; Small Interfering RNA; Stains; Stretching; Testing; Therapeutic; Therapeutic Studies; Transforming Growth Factor beta; Ubiquitin-Conjugating Enzymes; Ubiquitination; Vascular Endothelial Growth Factors; aging population; angiogenesis; aortic valve; aortic valve disorder; calcification; hypercholesterolemia; in vivo; inhibitor; insight; malignant breast neoplasm; mechanical force; mouse model; novel; novel therapeutics; overexpression; repaired; response; shear stress; valve replacement; vector

Calcific aortic valve disease (CAVD) is a significant cause of morbidity among the aging population and is a strong risk factor for additional cardiovascular events. Currently, there are no therapeutic options for CAVD other than valve replacement or repair due in part to the incomplete understanding of the underlying mechanisms. Interestingly, AV calcification develops in a side-specific manner, occurring preferentially on the fibrosa side exposed to d-flow while the ventricularis side exposed to stable flow is spared. Another mechanical force, elevated stretching commonly observed in bicuspid (BAV) and diseased valves and hypertension, also correlates well with CAVD. These suggest a potential role for d-flow and elevated stretch in the pathogenesis of CAVD, but the mechanism is unclear. Our goal is to define the role and mechanisms of the mechanical forces in CAVD pathogenesis and use the knowledge to develop novel anti-CAVD therapeutics. In the previous funding cycle, we have identified several flow- and side-dependent microRNAs (miRNAs) in human AV endothelial cells (HAVECs) and pig AVs (PAVs) and began determining their roles in CAVD. Recently, we have identified a novel flow- and stretch-sensitive miR-483-3p (miR-483), which has exciting potential as a critical regulator of CAVD pathogenesis. Our preliminary data show that miR-483 expression is decreased 1) by d-flow (OS) compared to stable flow (LS) in HAVECs, 2) in the fibrosa layer compared to the ventricularis in human and pig AVs, and 3) by pathological stretch conditions in PAVs ex vivo. Further data indicate that miR-483 inhibits EC inflammation and endothelial-to-mesenchymal transition (EndoMT), critical pathobiological events in CAVD, and that a key gene target of miR-483 is Ube2c (E2 ubiquitin-conjugating enzyme), which in turn may target the hypoxia- inducible factor (HIF1α) via controlling its upstream regulator pVHL. HIF1α's role in CAVD is unclear, but its well- known target genes include VEGF (angiogenesis and inflammation), TGFβ (fibrosis and calcification), Runx2 (calcification) and Twist1 (EndoMT), key CAVD pathogenic processes. Therefore, our overarching hypothesis is that miR-483 is an anti-CAVD miRNA, which is reduced under OS/pathological stretch conditions, leading to an increase in Ube2c, which in turn ubiquitinates pVHL for its degradation and increases the HIF1α level. HIFα, then, stimulates its target genes leading to inflammation, EndoMT, AV sclerosis and calcification. We will test this in 3 Aims. Aim 1 will determine the mechanisms by which miR-483 regulates shear-dependent responses of HAVECs and PAVs in a Ube2c- and the HIF1α-dependent manner in vitro and ex vivo. Aim 2 will determine the role of miR-483 in stretch-dependent calcification of HAVICs and PAVs ex vivo via Ube2c and HIF1α-dependent mechanisms. Aim 3 is an in vivo study where miR-483, Ube2c, and HIF1α will be modified genetically, molecularly or pharmacologically in a novel mouse model of CAVD that we just developed by treating GATA5-/- BAV mice with AAV-PCSK9 to induce hypercholesterolemia. Here, we will test their roles and their anti-CAVD therapeutic potential.

钙化主动脉瓣疾病（CAVD）是衰老人群中发病率的重要原因，是一个 额外心血管事件的强大风险因素。目前，Cavd没有其他治疗选择 比更换阀门或维修的部分原因是对基本机制的不完全理解。 有趣的是，AV计算开发以特定于侧的方式，优先发生在纤维侧 暴露于D-flow时，却保留了暴露于稳定流量的室心侧。另一个机械力， 在双尖齿（BAV）和解散瓣膜和高血压中通常观察到的拉伸升高，也相关 和Cavd。这些表明在Cavd发病机理中d-flow和升高的潜在作用，但是 该机制尚不清楚。我们的目标是定义机械力在CAVD中的作用和机制 发病机理并利用知识开发新的抗CAVD疗法。在上一个资金周期中， 我们已经确定了人类AV内皮细胞中的几个流动和侧依赖的microRNA（miRNA） （Havecs）和Pig Avs（PAVS），并开始确定其在CAVD中的作用。最近，我们确定了一本小说 流动和拉伸敏感的miR-483-3p（miR-483），它具有令人兴奋的潜力，作为CAVD的关键调节剂 发病。我们的初步数据表明，与D-Flow（OS）相比，miR-483的表达降低了1） 与人和猪AV中的心室相比，HAVEC中的稳定流量（LS），2）在纤维层中，3） 通过病理拉伸条件，pavs ex vivo。进一步的数据表明miR-483抑制EC炎症 以及内皮到间质转变（内托），CAVD中的关键病理生物学事件以及该关键 miR-483的基因靶标是ube2c（E2泛素偶联酶），这又可能针对缺氧 - 通过控制其上游调节剂PVHL，诱导因子（HIF1α）。 HIF1α在CAVD中的作用尚不清楚，但其良好 已知的靶基因包括VEGF（血管生成和炎症），TGFβ（纤维化和钙化），Runx2 （钙化）和Twist1（endomt），关键CAVD致病过程。因此，我们的总体假设是 该miR-483是一种抗CAVD miRNA，在OS/病理拉伸条件下降低，导致 UBE2C的增加，进而泛素PVHL的降解并增加了HIF1α水平。 HIFα， 然后，刺激其靶基因，导致炎症，胚胎，AV硬化和钙化。我们将测试 这在3个目标中。 AIM 1将确定miR-483调节剪切依赖性响应的机制 ube2c-和hif1α依赖性方式的HAVEC和PAV在体外和离体中。 AIM 2将确定 miR-483在通过UBE2C和HIF1α依赖性的天堂和PAV的拉伸依赖计算中的作用 机制。 AIM 3是一项体内研究，其中miR-483，UBE2C和HIF1α将一般修饰， 我们刚刚通过治疗GATA5 - / - 开发的新型CAVD小鼠模型中的分子或药理学。 具有AAV-PCSK9的BAV小鼠诱导高胆固醇血症。在这里，我们将测试他们的角色及其反武器 治疗潜力。