The Role of Trp Proteins in Myometrial Calcium Dynamics

Trp 蛋白在子宫肌层钙动态中的作用

• 批准号: 6921016
• 负责人: BARBARA M SANBORN
• 金额: $ 18.96万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-01 至 2009-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6921016
• 关键词: calcium channel; calcium flux; calcium indicator; calcium transporting ATPase; clinical research; confocal scanning microscopy; estrogens; female; hormone regulation /control mechanism; human subject; laboratory rat; muscle contraction; myometrium; polymerase chain reaction; pregnancy; protein protein interaction; tissue /cell culture; voltage gated channel; western blottings

DESCRIPTION (provided by applicant): Understanding the mechanisms regulating myometrial intracellular free Ca (Cai) dynamics is critical to understanding the control of myometrial activity. Signal-regulated calcium entry (also called capacitative entry) is broadly defined here to include both receptor- and store-operated entry. This pathway has been implicated in regulation of membrane potential and tone in smooth muscle and may play a role in the regulation of myometrial contractile activity by hormones, growth factors and cytokines. Hence regulated calcium entry may participate in control of uterine contractile activity during pregnancy and parturition. We found that rat and human myometrial cells respond to more than one signal-regulated calcium stimulus and express a number of TrpC mRNAs and proteins implicated in this type of calcium entry. The overall goal of this study is to understand the role of TrpC proteins in the regulation of myometrial Cai dynamics and control of contractile activity. Aim 1 will determine the relative importance of signal-regulated Ca entry in the control of myometrial Cai dynamics, measured by fluorescence imaging, and to contractile activity in uterine strips. Aim 2 will delineate the contribution of specific TrpC proteins to signal-regulated calcium entry and Cai dynamics in human myometrial cells. The effects of both overexpression and suppression of TrpC expression on myometrial cell Cai dynamics will be determined and linked to specific responses. Aim 3 will examine the regulation of TrpC expression and of signal-regulated Ca entry by other signaling pathways in myometrium. These studies will utilize Q-RT-PCR, immunoblot and Ca imaging approaches in human myometrial cells and the pregnant rat model. Aim 4 will examine the influence of TrpC subtype expression on TrpC association with myometrial plasma membrane and possible effects of signal-regulated Ca entry on activity of other membrane proteins. These studies will use scanning confocal microscopy and subcellular fractionation approaches. These studies promise new insights into the role of this tightly regulated calcium entry mechanism in the control of myometrial Cai dynamics and myometrial contraction and may lead to design of new approaches for controlling myometrial activity during pregnancy and labor.

描述（由申请人提供）：了解调节子宫肌层细胞内游离 Ca (Cai) 动态的机制对于了解子宫肌层活性的控制至关重要。信号调节的钙进入（也称为电容性进入）在这里被广泛定义为包括受体操纵的进入和钙池操纵的进入。该途径与平滑肌膜电位和张力的调节有关，并且可能在激素、生长因子和细胞因子对子宫肌层收缩活性的调节中发挥作用。因此，调节钙的进入可能参与妊娠和分娩期间子宫收缩活动的控制。我们发现大鼠和人类子宫肌层细胞对不止一种信号调节的钙刺激作出反应，并表达许多与这种类型的钙进入有关的 TrpC mRNA 和蛋白质。本研究的总体目标是了解 TrpC 蛋白在调节子宫肌层动态和控制收缩活动中的作用。 目标 1 将确定信号调节的 Ca 进入在控制子宫肌层 Cai 动态（通过荧光成像测量）以及子宫条收缩活动中的相对重要性。 目标 2 将描述特定 TrpC 蛋白对信号调节的钙进入和人类子宫肌细胞中的 Cai 动力学的贡献。 TrpC 表达的过度表达和抑制对子宫肌细胞 Cai 动力学的影响将被确定并与特定反应联系起来。 目标 3 将检查子宫肌层中其他信号通路对 TrpC 表达和信号调节 Ca 进入的调节。这些研究将在人类子宫细胞和怀孕大鼠模型中利用 Q-RT-PCR、免疫印迹和 Ca 成像方法。 目标 4 将检查 TrpC 亚型表达对 TrpC 与子宫肌膜质膜关联的影响，以及信号调节 Ca 内流对其他膜蛋白活性的可能影响。这些研究将使用扫描共焦显微镜和亚细胞分级方法。这些研究为了解这种严格调节的钙进入机制在控制子宫肌层动态和子宫肌层收缩中的作用提供了新的见解，并可能导致设计出在怀孕和分娩期间控制子宫肌层活动的新方法。