Vascular Reactivity in Heart Failure

描述（由申请方提供）：在充血性心力衰竭（CHF）临床综合征期间，血管系统的特征为静息血管收缩、对应激的异常反应和对一氧化氮（NO）介导的血管舒张的抵抗。导致血管系统发生这些变化的机制尚不清楚。以前的研究还没有解决是否血管平滑肌收缩表型的变化导致这些异常。该应用程序的目标是确定与CHF相关的血管异常的机制。平滑肌激活依赖于20 kDa肌球蛋白轻链（MLC 20）磷酸化水平，这取决于MLC激酶和MLC磷酸酶的相对活性。MLC磷酸酶是由三个亚基组成的全酶;约-20 kDa亚基、约-38 kDa催化亚基和肌球蛋白结合亚基（MBS）。MBS的四种不同亚型的特征在于存在（133 kDa）或不存在（130 kDa）中心插入物和亮氨酸拉链。初步数据表明，与133 kDa同种型相比，130 kDa同种型具有增加的活性，并且中心插入物决定了平滑肌对激动剂诱导的力增强的敏感性。此外，我们的数据表明，亮氨酸拉链的剪接变体表达决定了对NO介导的血管舒张的敏感性。本申请基于这样的假设，即与CHF相关的血管异常是由于MBS的剪接变体表达的变化。我们认为，静息血管收缩是由于中央插入的剪接变体表达的变化，而对NO介导的血管舒张的阻力是由于亮氨酸拉链的剪接变体表达的变化。为了验证这一假设，我们将确定（1）CHF发生前后平滑肌对激动剂诱导的力增强、NO介导的血管舒张和MBS剪接变体亚型表达的敏感性;（2）MBS剪接变体亚型的强制表达是否改变了对激动剂诱导的力增强和NO介导的平滑肌舒张的敏感性。我们将使用几种不同的血管平滑肌的控制和大鼠心肌梗死模型的CHF，以及培养的平滑肌细胞来测试我们的假设。这些研究将阐明对激动剂诱导的力增强和NO介导的平滑肌松弛的敏感性的组织特异性差异的分子机制。

DESCRIPTION (provided by applicant): During the clinical syndrome of congestive heart failure (CHF), the vasculature is characterized by resting vasoconstriction, abnormal responses to stress, and resistance to nitric oxide (NO) mediated vasodilatation. The mechanism that leads to these changes in the vasculature is unknown. Previous studies have not addressed whether changes in the contractile phenotype of vascular smooth muscle cause these abnormalities. The goal of this application is to determine the mechanism for the vascular abnormalities associated with CHF. Smooth muscle activation is dependent on 20 kDa myosin light chain (MLC20) phosphorylation levels, which depend on the relative activities of MLC kinase and MLC phosphatase. The MLC phosphatase is a holoenzyme consisting of three subunits; a approximately -20 kDa subunit, a approximately -38 kDa catalytic subunit and a myosin binding subunit (MBS). Four distinct isoforms of the MBS are characterized by the presence (133 kDa) or absence (130 kDa) of a central insert and a leucine zipper. Preliminary data demonstrate that the 130 kDa isoform has an increased activity compared to the 133 kDa isoform, and that the central insert determines the sensitivity of smooth muscle to agonist induced force enhancement. In addition, our data suggest that splice variant expression of the leucine zipper determines the sensitivity to NO mediated vasodilatation. This application is based on the hypothesis that vascular abnormalities associated with CHF are due to changes in splice variant expression of the MBS. We suggest that the resting vasoconstriction is due to changes in splice variant expression of the central insert, while the resistance to NO mediated vasodilatation is due to changes in splice variant expression of the leucine zipper. To test this hypothesis, we will determine (1) the sensitivity of smooth muscle to agonist induced force enhancement, NO mediated vasodilatation, and the expression of splice variant isoforms of the MBS prior to and after the development of CHF; and (2) if forced expression of the splice variant isoforms of the MBS alters the sensitivity to agonist induced force enhancement mad NO mediated smooth muscle relaxation. We will use several different vascular smooth muscles of control and a rat infarct model of CHF, as well as cultured smooth muscle cells to test our hypothesis. These studies will elucidate the molecular mechanism underlying the tissue specific differences in the sensitivity to agonist induced force enhancement and NO mediated smooth muscle relaxation.