Measuring the effect of extracellular mechanics on smooth muscle contractility

描述（由申请人提供）：动脉存在于动态环境中，需要血管平滑肌细胞（VSMCs）不断适应，以维持机械稳态。在健康动脉中，血管张力的维持主要是通过VSMC的收缩和舒张来实现的，而收缩力的丧失表明VSMC功能障碍，这可能在动脉粥样硬化斑块形成和再狭窄的发病机制中起重要作用。由于动脉粥样硬化中的脂质池导致组织软化，或由于斑块钙化区域或植入支架导致组织硬化，机械环境可被斑块发展或支架植入显著扰乱。最近的研究结果发现，细胞周围环境的力学在其功能行为中起着重要作用，这表明由斑块或支架引起的非稳态组织力学也可能影响收缩功能。我们假设周围组织的力学直接影响血管平滑肌细胞的收缩功能。用目前的技术，很难评估组织刚度对VSMC功能的影响，因为现有的体外系统中力学是独立可控的，它们都局限于衬底刚度的狭窄范围，在这个范围内它们可以用来测量细胞或组织应力的产生。因此，为了验证这一假设，我们需要开发一种新的方法来测量组织应力，这种方法可以用于广泛的基质力学。我们之前已经开发了一种叫做血管肌肉薄膜（vMTFs）的方法，在这种方法中，我们将细胞微图纹成高度组织化的组织，并通过测量光束的曲率来确定细胞的应力。我们将采用这种方法来包括可变模量的基底，使我们能够测量模拟各种机械环境的vsmc的功能收缩性，例如软脂池和硬支架。然后，我们将把这种收缩性与VSMC表型标记表达联系起来，以确定与许多VSMC功能障碍有关的表型转换如何在VSMC底物调节的机械适应中发挥作用。这项工作的完成将为动脉病变中VSMCs不断演变的机械环境的功能含义提供更清晰的图像。

DESCRIPTION (provided by applicant): Arteries exist in a dynamic environment, necessitating constant adaptation by vascular smooth muscle cells (VSMCs), in order to maintain mechanical homeostasis. In a healthy artery, maintenance of vessel tension is primarily achieved by contraction and relaxation of VSMCs, and loss of contractility is indicative of VSMC dysfunction, which may play an important role in pathogenesis of atherosclerotic plaque formation and restenosis. The mechanical environment can be significantly perturbed by plaque development or stent implantation, either by softening of the tissue, due to lipid pools in atherosclerosis, or stiffening due to calcified regions of the plaque, or an implanted stent. Recent results have found that the mechanics of a cell's surroundings play a significant role in its functional behavior, suggesting that non-homeostatic tissue mechanics, due to plaques or stents, could also affect contractile function. We hypothesize that the mechanics of the surrounding tissue directly affects vascular smooth muscle cells' contractile functionality. With current techniques, it is difficult to assess the effects of tissue stiffness on VSMC function because existing in vitro systems in which mechanics are independently controllable are all limited to narrow ranges of substrate stiffness over which they can be used to measure cell or tissue stress generation. So, to test this hypothesis we will need to develop a new method for measuring tissue stress that can be employed with a wide range of substrate mechanics. We have previously developed a method called vascular muscular thin films (vMTFs), where we micropatterned cells into highly organized tissues on a thin rubber beam, and the cell stress was determined by measuring the curvature of the beam. We will adapt this method to include a variable modulus substrate, allowing us to measuring functional contractility of VSMCs mimicking a wide range of mechanical environments, such as soft lipid pools and stiff stents. We will then correlate this contractility with VSMC phenotype marker expression to determine how phenotype switching, which has been implicated in a number of VSMC dysfunctions, plays a role in VSMC substrate-modulated mechano-adaptation. Completion of this work will provide a sharper picture of the functional implications of the evolving mechanical environment of VSMCs in arterial pathologies.