Biomechanical regulation of valvulogenesis

描述（由申请人提供）：瓣膜相关的先天性心脏缺陷（CHD）是早产和婴儿死亡的主要原因。尽管有相当大的研究进展揭示了心脏形态发生的重要遗传调节因子，但由于缺乏对这些途径如何协调驱动瓣膜形成和重塑的理解，将其转化为临床效益受到阻碍。随着胚胎瓣膜的生长和成熟，它们暴露在越来越苛刻的机械环境中，但这些力的功能后果却鲜为人知。瓣膜发生可以被看作是一个工程过程，而不是一个纯粹的遗传程序。因此，揭示这些新的控制关系是必不可少的，但由于缺乏分析工具和实验方法来分离和量化活胚胎瓣膜的机械效应，因此一直具有挑战性。首先，我们开发了独特的生物力学测试设备，可以量化胚胎垫和瓣膜的生物力学。其次，我们创造了一种新的斑点跟踪算法，结合高频超声，可以非侵入性地量化胚胎瓣膜内的动态组织应变。第三，我们开发了一种独特的有限元模拟策略，该策略与体内测量迭代，以绘制解剖精确几何形状的局部生物力学参数。第四，我们创造了第一个实验策略，通过飞秒激光消融（FLP）在活禽胚胎中无创地制造局部孤立的心脏内缺陷。利用这些使能技术，我们的目标是确定和描述指导胚胎瓣膜形成的机械-遗传关系。我们的总体假设是，机械信号通过同时调节多种瓣膜形成信号通路来协调胚胎瓣膜的雕刻和加强。我们将首先量化瓣膜周围的体内机械环境变化及其生物力学适应性（目的1）。接下来，我们将确定体外机械刺激如何调节已知的瓣膜发生分子调节网络（目的2）。然后，我们通过局部控制的鸟类胚胎瓣膜光消融，在体内测试了先前实验提出的机械-遗传机制（Aim 3）。这一建议将提供胚胎瓣膜形成过程中体内生物力学环境的重要定量细节，以及这种环境如何调节局部基因表达以促进瓣膜形成和成熟。这一新信息将补充来自基因操作研究的现有数据集，并指导对这些结果的新解释。更好地了解指导瓣膜形成和重塑的机械生物学关系，将大大拓宽解释冠心病发病机制的范围，并为预防或修复冠心病提供新的临床策略。

DESCRIPTION (provided by applicant): Valve related congenital heart defects (CHD) are a major cause of preterm and infant death. Despite considerable research progress uncovering important genetic regulators of cardiac morphogenesis, translation to clinical benefit has been hampered by a lack of understanding of how these pathways coordinate to drive valve formation and remodeling. The embryonic valves are exposed to an increasingly demanding mechanical environment as they grow and mature, but the functional consequences of these forces are far less understood. Valvulogenesis can alternatively be considered an engineering process rather than a purely genetic program. Uncovering these new governing relationships is therefore essential but has been challenging because of the lack of analytical tools and experimental approaches that can isolate and quantify mechanical effects in live embryonic valves. First, we have developed unique biomechanical testing devices that can quantify embryonic cushion and valve biomechanics. Second, we have created a novel speckle tracking algorithm in conjunction with high frequency ultrasound that can non-invasively quantify dynamic tissue strains within the embryonic valves. Third, we developed a unique finite element simulation strategy that iterates with in vivo measurements to map local biomechanical parameters in anatomically precise geometries. Fourth, we have created the first experimental strategy to create locally isolated intracardiac defects in live avian embryos non-invasively through femtosecond laser photoablation (FLP). Using these enabling technologies, our objective for this proposal is to identify and characterize mechano-genetic relationships that guide the formation of the embryonic valves. Our overall hypothesis is that mechanical signaling orchestrates the sculpting and strengthening of the embryonic valves through simultaneous regulation of multiple valvulogenic signaling pathways. We will first quantify the changing in vivo mechanical environments surrounding the valves and their biomechanical adaptation (Aim 1). We will next determine how the network of known molecular regulators of valvulogenesis is modulated by mechanical stimulation in vitro (Aim 2). Then we test the mechano-genetic mechanisms suggested by the previous experiments in vivo using locally controlled photoablations to avian embryonic valves (Aim 3). This proposal will generate significant quantitative detail of the in vivo biomechanical environment during embryonic valvulogenesis and how this regulates local gene expression to promote valve formation and maturation. This novel information will complement existing datasets from genetic manipulation studies and guide new interpretations of these results. A better understanding of the mechanobiological relationships guiding valve formation and remodeling significantly broadens the array of mechanisms to explain the pathogenesis of CHD and tools to enable new clinical strategies to prevent or repair CHD.