ESTROGEN REGULATION OF SMOOTH MUSCLE BKCA CHANNELS

雌激素对平滑肌 BKCA 通道的调节

• 批准号: 6719855
• 负责人: Kathleen Dunlap
• 金额: $ 25.07万
• 依托单位: TUFTS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-02-01 至 2005-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6719855
• 关键词: Xenopus oocyte; biological signal transduction; cGMP dependent protein kinase; calcium ion; electrophysiology; enzyme activity; estrogen receptors; estrogens; gene expression; gene targeting; genetically modified animals; guanylate cyclase; hormone regulation /control mechanism; ion channel blocker; laboratory mouse; myocardial ischemia /hypoxia; nitric oxide synthase; phosphorylation; potassium channel; protein structure function; tissue /cell culture; transcription factor; vascular endothelium; vascular smooth muscle; vasodilatation

Estrogen protects the cardiovascular system from injury by complex mechanisms that have both short- and long-term components. Longer- term estrogen actions occurs following hours of estrogen exposure and is mediated by estrogen receptors acting in the nucleus as transcription factors for growth-related genes. The rapid effects of estrogen, by contrast, occurs in a seconds-to-minutes time frame and do not require gene transcription. Project 5 of the SCOR application addresses the molecular mechanisms underlying one of the most important of these rapid effects-vasodilation. The signaling pathways underlying this non- genomic action of estrogen are incompletely understood but new data support they involve the action of nitric oxide synthase in vascular cells, elevation of nitric oxide, stimulation of vascular smooth muscle cell soluble guanylyl cyclase, and substrate phosphorylation by cGMP- dependent protein kinase (PKG). Although the key substrate(s) for PKG is unknown, new data support one likely possibility is the smooth muscle large conductance Ca2+-activated K+ (BKCa) channel. Blockade of BKCa channels in vascular rings significantly reduces estrogen-induced vasodilation, demonstrating the central role that BKCa channels in vascular rings significantly reduces estrogen-induced vasodilation, demonstrating the central role that BKCa channels play in the acute physiological response to estrogen. Experiments in Aim 1 will use BKCa channels in vascular rings significantly reduces estrogen-induced vasodilation, demonstrating the central role that BKCa channels play in the acute physiological response to estrogen. Experiments in Aim 1 will use BKCa channel activity recorded from single smooth muscle cells dissociated from mouse aorta as an assay for characterizing upstream elements in the estrogen signaling pathway that targets BKCa channels. Experiments will assess the relative contributions of endothelial and smooth muscle cells, the types of estrogen receptor involved, and the requirement for nitric oxide synthase activation. Experiments in Aim 2 will focus on the molecular mechanism by which BKCa channel activity is regulated by estrogen. We will identify the molecular variants of BKCa channel activity is regulated by estrogen. We will identify the molecular variants. variants of BKCa channels that are expressed in smooth muscles cells from mouse aorta and study the recombinant channels expressed heterologously in order to test the hypothesis that estrogen brings about BKCa channel activation through a direct PKG-dependent phosphorylation of the channel protein. Such information is essential to further understand the protective effects of estrogen on vascular tissue and may form the basis for development of new therapies for cardiovascular disease.

雌激素通过复杂的机制保护心血管系统免受短期和长期损伤。长期雌激素作用发生在雌激素暴露数小时后，由雌激素受体介导，在细胞核中作为生长相关基因的转录因子。相比之下，雌激素的快速作用在几秒到几分钟的时间内发生，不需要基因转录。SCOR应用的项目5解决了这些最重要的快速效应之一-血管舒张的分子机制。雌激素的这种非基因组作用的信号通路尚不完全清楚，但新的数据支持它们涉及血管细胞中一氧化氮合酶的作用，一氧化氮的升高，血管平滑肌细胞可溶性鸟酰环化酶的刺激，以及cGMP依赖性蛋白激酶（PKG）的底物磷酸化。虽然PKG的关键底物尚不清楚，但新的数据支持一种可能的可能性是平滑肌大电导Ca2+激活K+ (BKCa)通道。阻断血管环中的BKCa通道可显著降低雌激素诱导的血管舒张，表明血管环中的BKCa通道可显著降低雌激素诱导的血管舒张，表明BKCa通道在雌激素的急性生理反应中发挥核心作用。Aim 1的实验将在血管环中使用BKCa通道显著降低雌激素诱导的血管舒张，证明BKCa通道在雌激素的急性生理反应中发挥核心作用。Aim 1的实验将使用从小鼠主动脉分离的单个平滑肌细胞中记录的BKCa通道活性，作为表征雌激素信号通路中靶向BKCa通道的上游元素的实验。实验将评估内皮细胞和平滑肌细胞的相对贡献，所涉及的雌激素受体类型，以及一氧化氮合酶激活的需求。Aim 2的实验将重点关注雌激素调节BKCa通道活性的分子机制。我们将确定BKCa通道活性受雌激素调节的分子变异。我们将识别分子变异。研究小鼠主动脉平滑肌细胞中表达的BKCa通道的变异体，并研究重组通道的异种表达，以验证雌激素通过直接依赖pkg的通道蛋白磷酸化激活BKCa通道的假设。这些信息对于进一步了解雌激素对血管组织的保护作用至关重要，并可能成为开发心血管疾病新疗法的基础。