Caveolin-1, Caveolae and Placental Angiogenesis

描述(申请人提供)：总体假设是小凹-1(Cav-1)与血管内皮生长因子(VEGFR)受体(VEGFR)直接和/或间接相互作用，将VEGFR启动的信号事件，即RAS-Raf-MEK1-ERK2/1和PI3K/Akt信号模块集中在内皮细胞(EC)的小窝中，从而调控VEGF对EC增殖、迁移和分化的调控。此外，Cav-1/Caveolae可能通过改变一氧化氮(NO)在EC中的生物利用度来调节胎盘血管生成。为了解决这一假设，将使用绵羊胎儿胎盘动脉EC模型和人胎盘毛细血管内皮细胞系。具体目的1：进一步阐明VEGFRs(即KDR、Flt-1和NP-1)在体外和体内是否与Cav-1相关，从而定位于小窝，以及外源性Cav-1的过度表达或内源性Cav-1的靶向下调是否调节配体依赖的VEGFR激活。具体目的2：确定靶向下调内源性Cav-1或过度表达外源性Cav-1是否会改变血管内皮生长因子对细胞周期进入、增殖、迁移和分化的刺激。具体目的3：确定血管内皮生长因子是否激活小窝中的RAS/Raf/ERK2/1和PI3K/Akt信号模块。特异性目的4：探讨下调Cav-1或Cav-1过表达是否调节血管内皮细胞生长因子对MAPK和PI3K/Akt信号通路的刺激，以及这些信号通路在血管内皮生长因子诱导的细胞周期进入、细胞增殖、迁移和分化中的作用。具体目的5：确定靶向下调内源性Cav-1或过度表达外源性Cav-1是否改变eNOS活性，从而改变NO的生物利用度，进而调节胎盘血管生成。这些研究将对我们理解小窝、血管内皮生长因子/血管内皮生长因子受体、内皮细胞和血管生成生物学产生重大影响，这些都是我们首次在临床上与胎盘血管生成和血管扩张相关的生物学重要领域。当人们认为子宫胎盘内皮细胞对妊娠的适应性，特别是胎儿胎盘和子宫胎盘血流的增加与胎儿生长和存活率直接相关时，这项研究的最终临床意义是显而易见的，并且这些机制在先兆子痫和IUGR等病理性妊娠中是功能失调的。

DESCRIPTION (provided by applicant): The overall hypothesis is that caveolin-1 (cav-1) interacts with vascular endothelial growth factor (VEGF) receptors (VEGFR) directly and/or indirectly to concentrate VEGFR initiated signaling events, i.e., Ras-Raf-MEK1-ERK2/1 and PI3K/Akt signaling modules, in the caveolae of endothelial cells (EC), thereby modulating VEGF regulation of EC proliferation, migration, and differentiation. Moreover, cav-1/caveolae may regulate placental angiogenesis by modifying the bioavailability of nitric oxide (NO) in EC. To address this hypothesis, an ovine fetoplacental artery EC model and a human placental capillary endothelial cell line will be used. Specific Aim 1: To further clarify if VEGFRs (i.e., KDR, Flt-1, and NP-1) are physically associated with cav-1 in vitro and in vivo, thus localized in the caveolae, and if overexpression of exogenous cav-1 or targeted down-regulation of endogenous cav-1 regulates ligand-dependent VEGFR activation. Specific Aim 2: To determine if targeted down-regulation of endogenous cav-1 or overexpression of exogenous cav-1 alters VEGF stimulation of cell cycle entry, proliferation, migration and differentiation. Specific Aim 3: To determine if VEGF activates the Ras/Raf/ERK2/1 and PI3K/Akt signaling modules in the caveolae. Specific Aim 4: To delineate if down-regulation of cav-1 or cav-1 overexpression modulates VEGF stimulation of MAPK and PI3K/Akt signaling pathways and the role of these pathways in VEGF-induced cell cycle entry, cell proliferation, migration, and differentiation. Specific Aim 5: To determine if targeted down-regulation of endogenous cav-1 or overexpresison of exogenous cav-1 alters eNOS activity thereby altering the bioavaibility Iof NO, which in turn regulates placental angiogenesis. These studies will have a great impact on our understanding of caveoli, VEGF/VEGFR, endothelial, and angiogenesis biology, all are biologically important fields we have integrated in the proposal clinically relevant to placental angiogenesis and vasodilatation for the first time. The ultimate clinical importance of this research is evident when one considers that uteroplacental endothelial adaptations to pregnancy, especially the rises in fetoplacental and uteroplacental perfusion, are linked directly to fetal growth and survivability and that these mechanisms are dysfunctional in pathologic pregnancies such as preeclampsia and IUGR.