Cyclic stretch of bicuspid aortic valves: elucidating its implications for cell signaling and tissue mechanics.

项目总结：虽然1.4%的人患有先天性二叶式主动脉瓣（BAV）缺陷， BAV患者占接受主动脉瓣置换术患者的50%。此外，患者 BAV患者更早发生主动脉瓣狭窄（AS），因此需要在比BAV患者更年轻的年龄进行置换术。 三尖瓣主动脉瓣（TAV）正常患者。鉴于替代品的耐久性有限，BAV 患者具有负面影响其寿命和生活质量的高手术负担。因此，我们认为， 药物治疗的发展将降低AS的发病率和死亡率。 我们的长期目标是在细胞水平上了解BAV疾病的过程，以开发有效的 缓解AS的治疗。由于没有药物治疗，我们的假设 存在于BAV中异常瓣膜间质细胞（维克）变形模式在 AS中的生化信号事件。由于AS与维克激活成肌纤维细胞有关 和转化生长因子-β（TGFB）信号，这项研究解决了之间的关系 机械调节的细胞形态和TGFB信号传导有两个具体目的： 1.确定不同主动脉瓣经历了哪些机械调节参数 解剖结构影响维克形态。肌成纤维细胞活化的水平将首先在 来自BAV和TAV的天然人瓣叶外植体。细胞形态从自然 将组织与在新的3D高通量中机械调节的细胞进行比较， 双轴振荡拉伸屏幕（3D HT-BOSS），以确定什么循环双轴拉伸，矩阵 刚度和维克基础收缩性是产生BAV和TAV中所见形态所必需的。 2.确定改变机械模式将如何改变维克对TGFB的反应。定量 蛋白质组学将被用于开发来自VIC的维克TGFB信号传导的稳态模型 新鲜分离自天然人瓣叶。然后将对3D HT-BOSS样品进行显微镜检查， 用于分析以下变化：1）α SMA蛋白的EC50和2）TGFB的核定位- 在外源性TGFB刺激下介导转录因子。 通过这项研究，AS的潜在驱动因素和药理学的新靶向途径 因此，治疗将被揭露。 此外，研究金申请人Toni West博士将接受的培训将使她能够 一跃成为独立调查员韦斯特博士将在 在她的赞助商Michal Sacks博士的实验室，以及她的合作者Aaron Baker博士的实验室， 德克萨斯大学作为培训的一部分，韦斯特博士将前往哥伦比亚大学，在那里她的同事- 赞助商乔瓦尼·法拉利博士经营着她将从中收集组织和细胞的生物银行。

PROJECT SUMMARY: While 1.4% of people have the congenital defect of bicuspid aortic valve (BAV), BAV patients make up 50% of the patients that receive aortic valve replacements. Moreover, patients with BAVs develop aortic stenosis (AS) earlier, and thus require replacements at younger ages than patients with normal tricuspid aortic valves (TAV). Given the limited durability of replacements, BAV patients have a high procedure burden that negatively affects their length and quality of life. Therefore, development of a pharmacological therapy will reduce morbidity and mortality from AS. Our long-term goal is to understand the BAV disease process at the cellular level to develop effective treatments that mitigate AS. Since no pharmacological treatment has been forthcoming, our hypothesis is that abnormal valve interstitial cell (VIC) deformation patterns present in BAVs have a crucial role in the biochemical signaling events in AS. Since AS is associated with VIC activation into myofibroblasts and transforming growth factor-beta (TGFB) signaling, this study addresses the relationship between mechanically-conditioned cellular morphology and TGFB signaling in two specific aims: 1. Determine what parameters of mechanical conditioning experienced in varying aortic valve anatomies affect VIC morphology. The level of myofibroblast activation will first be determined in native human valve leaflet explants from BAV and TAV. The morphologies of cells from native tissues will be compared to that of cells mechanically conditioned in a novel 3D high-throughput biaxial oscillatory stretch screen (3D HT-BOSS) to determine what cyclical biaxial stretch, matrix stiffness, and VIC basal contractility is required to produce morphologies seen in BAVs and TAVs. 2. Ascertain how altering mechanical pattern will modify VIC response to TGFB. Quantitative proteomics will be employed to develop steady-state models of VIC TGFB signaling from VICs freshly isolated from native human valve leaflets. Microscopy of 3D HT-BOSS samples will then be employed to analyze shifts in: 1) EC50 of αSMA protein and 2) nuclear localization of TGFB- mediated transcription factors upon exogenous TGFB stimulation. Through this investigation, underlying drivers of AS and novel target pathways for pharmacological treatment will therefore be uncovered. Furthermore, the training that the fellowship applicant, Dr. Toni West, will receive will enable her to make the leap to becoming an independent investigator. Dr. West will be conducting research in the lab of her sponsor, Dr. Michal Sacks, and in the lab of her collaborator, Dr. Aaron Baker, at the University of Texas. As part of her training, Dr. West will travel to Columbia University, where her co- sponsor Dr. Giovanni Ferrari runs the biobank she will be collecting tissues and cells from.