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: 7467919
• 负责人: BARBARA M SANBORN
• 金额: $ 17.62万
• 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 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7467919
• 关键词: 1,2-diacylglycerol; ATP phosphohydrolase; Abbreviations; Address; Affect; Animal Model; Birth; Calcium; Calcium ion; Cell Fractionation; Cell membrane; Cells; Cyclic AMP-Dependent Protein Kinases; Diglycerides; EGF gene; Endoplasmic Reticulum; Epidermal Growth Factor; Exhibits; Fluorescence; Gene Silencing; Glycerol; Goals; Growth Factor; Hormones; Human; Image; Immunoblotting; Individual; Inositol; Intracellular Membranes; Laser Scanning Confocal Microscopy; Lead; Link; Measures; Mediating; Membrane Potentials; Membrane Proteins; Modeling; Myometrial; Numbers; Pathway interactions; Patient currently pregnant; Phospholipase C; Play; Polymerase Chain Reaction; Pregnancy; Property; Protein C; Protein Kinase C; Protein Overexpression; Proteins; Rattus; Regulation; Relative (related person); Research Personnel; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Pathway; Signal Transduction; Small Interfering RNA; Smooth Muscle; Stimulus; Testing; Tissues; Uterine Contraction; cytokine; design; fluorescence imaging; inositol-1,4,5-triphosphate receptor; insight; myometrium; novel strategies; programs; receptor; response; small hairpin RNA; voltage

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）：了解调节子宫内细胞内游离CA（CAI）动力学的机制对于理解控制肌层活性的控制至关重要。信号调节的钙输入（也称为电容输入）在此广泛定义，包括受体和商店操作的入口。该途径与平滑肌的膜电位和音调有关，并可能在调节激素，生长因子和细胞因子的肌层收缩活性中起作用。因此，受调节的钙进入可能会参与怀孕和分娩期间子宫收缩活动的控制。我们发现大鼠和人肌层细胞对多种信号调节的钙刺激反应，并表达与这种类型的钙进入有关的多种TRPC mRNA和蛋白质。这项研究的总体目标是了解TRPC蛋白在调节肌层CAI动力学和收缩活性控制中的作用。 AIM 1将确定信号调节的CA进入在控制子宫肌体CAI动力学，通过荧光成像和子宫条中收缩活性测量的相对重要性。 AIM 2将描述特定TRPC蛋白对人肌层细胞中信号调节的钙进入和CAI动力学的贡献。过表达和TRPC表达抑制对肌层细胞CAI动力学的影响将被确定并与特定响应有关。 AIM 3将检查TRPC表达和信号调节的CA进入的调控，该CA通过子宫肌层中的其他信号通路进行调节。这些研究将利用人肌层细胞和怀孕大鼠模型中的Q-RT-PCR，免疫印迹和CA成像方法。 AIM 4将检查TRPC亚型表达对TRPC与肌层质膜的关联的影响，以及信号调节的Ca进入对其他膜蛋白活性的可能影响。这些研究将采用扫描共聚焦显微镜和亚细胞分级方法。这些研究有助于对这种严格调节的钙进入机制在控制肌层CAI动力学和肌层收缩中的作用的新见解，并可能导致设计新方法来控制怀孕和劳动期间的肌层活动。