B-Arrestins and GPCR Kinases in Vascular Function/Growth

描述（由申请人提供）：G蛋白偶联受体（gpcr），包括儿茶酚胺和血管紧张素II的受体，调节血管反应性，包括血管收缩和血管舒张，以及血管平滑肌（VSM）细胞的有丝分裂和迁移。高血压患者的血管反应性可能受到干扰，而手术旁路或动脉血管成形术后再狭窄后，VSM有丝分裂发生和迁移的改变是病理性内膜增生的特征。GPCR激活后，GPCR被7种GPCR激酶（grk）中的一种磷酸化，然后b-抑制素的两种同型中的一种被募集到受体上。b-阻滞蛋白结合立体阻断进一步向G蛋白传递信号，导致受体脱敏和信号衰减。b-arrestins也在信号传导中发挥积极作用，作为适配器和支架组织GPCR介导的MAP激酶级联反应的激活，如细胞外信号调节激酶（erk1 /2）。这些MAP激酶调节VSM细胞和其他细胞类型的有丝分裂和迁移。我们的研究小组已经培育出了grk和b-阻滞分别被敲除的小鼠。我们将利用这些动物及其VSM细胞来验证通过b-arrestins和GRKs调控GPCR信号对正常血管稳态至关重要的中心假设。我们的具体目标是：1)利用分离的主动脉环分析清醒和麻醉血压反应和血管反应性，阐明b-抑制蛋白和GRK基因敲除小鼠的血管表型；2)通过测定A) B -arrestins和GRKs通过内源性GPCR脱敏第二信使信号的特异性，以及B) B -arrestins和GRKs在GPCR刺激VSM细胞ERK激活、增殖和迁移中的作用，阐明B -arrestins和GRKs在野生型和敲除小鼠离体动脉和静脉VSM细胞中通过内源性GPCR信号传导的作用；3)确定特异性b-停搏蛋白或GRKs的缺失是否会改变小鼠静脉移植手术或动脉损伤后体内增生性内膜增生。这些实验有可能导致限制静脉移植失败和再狭窄的新策略的发展。

DESCRIPTION (provided by the applicant): G protein-coupled receptors (GPCRs) including those for catecholamines and angiotensin II regulate vascular reactivity, including vasoconstriction and vasodilation, as well as vascular smooth muscle (VSM) cell mitogenesis and migration. Vascular reactivity may be perturbed in hypertension, whereas altered VSM mitogenesis and migration characterize pathological intimal hyperplasia following surgical bypass or restenosis after arterial angioplasty. Following GPCR activation, GPCRs are phosphorylated by one of seven GPCR kinases (GRKs) and then one of two isoforms of b-arrestin is recruited to the receptor. b-arrestin binding sterically interdicts further signaling to G proteins, leading to receptor desensitization and attenuation of signaling. b-arrestins also play positive roles in signaling, serving as adapters and scaffolds to organize GPCR- mediated activation of MAP kinase cascades, such as the extracellular signal regulated kinases (ERK 1/2). These MAP kinases regulate mitogenesis and migration of VSM cells and other cell types. Our group has developed mice in which the GRKs and b-arrestins have been individually knocked out. We will utilize these animals, and VSM cells from them, to test the central hypothesis that regulation of GPCR signaling by b-arrestins and GRKs is critical for normal vascular homeostasis. Our specific aims are 1) To elucidate the vascular phenotype of b-arrestin and GRK knockout mice by analyzing conscious and anesthetized blood pressure responses and vascular reactivity using isolated aortic rings; 2) To elucidate the roles of b-arrestins and GRKs in signaling via endogenous GPCRs in isolated arterial and venous VSM cells from wild type and knockout mice by determining both A) the specificity of b-arrestins and GRKs in desensitizing second messenger signaling via endogenous GPCRs and B) the roles of b-arrestins and GRKs in GPCR stimulated ERK activation, proliferation, and migration of VSM cells; and 3) To determine if the loss of specific b-arrestins or GRKs alters in vivo proliferative intimal hyperplasia following mouse vein-graft surgery or arterial injury. These experiments have the potential to lead to the development of new strategies for limiting vein graft failure and restenosis.