ANG II OPPOSES INS MEDIATED VASORELAXATION & GLU UTILZA

ANG II 反对 INS 介导的血管舒张

• 批准号: 7185818
• 负责人: James Russell Sowers
• 金额: $ 27.26万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-10 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7185818
• 关键词: 1-Phosphatidylinositol 3-Kinase; ANG gene; Address; Adipocytes; Adipose tissue; Angiotensin II; Angiotensins; Binding; Binding Proteins; Blood Vessels; Cardiovascular Diseases; Characteristics; Chemosensitization; Complement; Condition; Data; Diabetes Mellitus; Dominant-Negative Mutation; Endothelial Cells; Enzymes; Fatty acid glycerol esters; Generations; Glucose; Hypertension; IGF-1 Signaling Pathway; Impairment; In Vitro; Insulin; Insulin Resistance; Insulin-Like Growth Factor I; Investigation; Knowledge; Laboratories; Liquid Chromatography; Mediating; Metabolism; Modeling; Myosin ATPase; Myosin Light Chains; NAD(P)H oxidase; Na(+)-K(+)-Exchanging ATPase; Nitric Oxide; Nitric Oxide Synthase; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Oxidative Stress; Pathogenesis; Pathology; Pathway interactions; Patients; Peptides; Phosphoric Monoester Hydrolases; Phosphorylation; Play; Post-Translational Protein Processing; Production; Protein Overexpression; Protein phosphatase; Proto-Oncogene Proteins c-akt; Rattus; Reactive Oxygen Species; Relaxation; Resistance; Rhea; Rho-associated kinase; Rodent Model; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Site; Skeletal Muscle; Smooth Muscle Myocytes; Somatomedins; Testing; Tissue Model; Tissues; Transgenic Organisms; Vascular Endothelial Cell; Vasodilation; angiogenin; autocrine; base; glucose disposal; glucose transport; human CYBA protein; in vivo; insulin sensitivity; normotensive; receptor; response; tandem mass spectrometry

Insulin (INS) resistance and hypertension often coexist, and frequently progress to diabetes and cardiovascular disease. INS and its structurally and functionally similar peptide, insulin-like growth factor (IGF-1), both have been shown to promote vasorelaxation, as well as glucose (GLU) disposal. Recent studies in our laboratory suggest that these actions of INS/ IGF-1 are mediated through the activation of phosphatidylinositol- 3-kinase (PI3-K) and the downstream, protein kinase B (Akt) signaling cascade. This signaling pathway increases: nitric oxide (NO) production by endothelial cells (EC), vascular smooth muscle cell (VSMC)- sodium pump (Na +, K+-ATPase) and myosin bound phosphatase (MBP) activity and GLU transport in skeletal muscle and adipocytes. Thus, functional alterations in these INS/ IGF-1 signaling pathways are likely to play an important role in these pathologies. In this regard, there is emerging evidence that angiotensin II (Ang II) interferes with this signaling, resulting in a state of resistance to INS/ IGF-1 mediated vasorelaxation and GLU transport; however, the role of intermediary signaling molecules are unclear. We hypothesize that Ang II exerts antagonistic effects on INS / IGF-l-stimulated PI3-K/Akt signaling through the generation of reactive oxygen species (ROS) and activation of RhoA. Further, it is proposed that in states of lNS/ IGF-1 resistance and hypertension there is exaggerated tissue Ang II mediated generation of ROS, and activation of RhoA. To test the main hypothesis and its corollary, we will address the following 2 specific aims: 1. To determine the role of RhoA and ROS in mediating the inhibitory effects of Ang II on the Akt-mediated actions of INS and IGF-1 involved in NO metabolism in EC; VSMC Na+,K+-ATPase; MBP activation; and also in the potentiation of GLU transport in skeletal muscle and adipose tissue. 2. To ascertain the role or Ang H-mediated RhoA/ROS stimulation in the impairment of GLU disposal and vasorelaxation in INS-resistant and Ang II overexpressing rodent models of hypertension. Using primary cultures of EC and VSMC, we will examine the role of Ang II-stimulated RhoA and ROS on NO metabolism in EC and VSMC Na +, K+-ATPase and MBP activity. To delineate the role of Akt, RhoA, and ROS signaling in mediating the counterregulatory actions of Ang II and INS/ IGF-1, we will interrupt each of these signaling pathways by transfecting VSMC with dominant negative constructs of Akt and RhoA, and antisense expression of p22phox, one of the critical components of NAD(P)H oxidase. We will profile and quantify site-specific phosphorylation and ROS-induced posttranslational modifications of the signaling molecules (i.e., Akt, RhoA, phoxes) and enzymes (i.e., eNOS, sodium pump, MBP) using liquid chromatography-tandem mass spectrometry. These studies will be complemented with in vivo and ex vivo studies of vasculature and skeletal muscle/adipocytes in rodent models of hypertension and INS resistance as well as normotensive controls. This proposed investigation should significantly enhance our knowledge regarding the role of Ang II in the pathogenesis of INS resistance and hypertension

胰岛素抵抗(Ins)与高血压常常并存，并经常进展为糖尿病和心血管疾病。胰岛素及其结构和功能相似的多肽，胰岛素样生长因子(IGF-1)，都被证明可以促进血管松弛和葡萄糖(GLU)的释放。我们实验室最近的研究表明，INS/IGF-1的这些作用是通过激活磷脂酰肌醇-3-激酶(PI3-K)和下游的蛋白激酶B(Akt)信号级联来介导的。这一信号通路增加了内皮细胞(EC)产生一氧化氮(NO)、血管平滑肌细胞(VSMC)-钠泵(Na+，K+-ATPase)和肌球蛋白结合磷酸酶(MBP)活性以及骨骼肌和脂肪细胞的GLU转运。因此，这些INS/IGF-1信号通路的功能改变可能在这些病理过程中发挥重要作用。在这方面，越来越多的证据表明，血管紧张素II(Ang II)干扰了这一信号转导，导致对INS/IGF-1介导的血管松弛和GLU运输产生抵抗；然而，中介信号分子的作用尚不清楚。我们推测，Ang II通过产生ROS和激活RhoA来拮抗INS/IGF3-L刺激的PI3-K/Akt信号转导通路。进一步提出，在LNS/IGF-1抵抗和高血压状态下，存在组织血管紧张素II介导的ROS的产生和RhoA的激活。为了验证主要假说及其推论，我们将探讨以下两个具体目标：1.确定RhoA和ROS在Ang II介导的INS和IGF-1抑制血管内皮细胞NO代谢的INS和IGF-1、VSMC Na+，K+-ATPase、MBP激活以及促进骨骼肌和脂肪组织GLU转运中的作用。2.探讨Ang H介导的RhoA/ROS刺激在Ins抵抗和Ang II高表达的高血压大鼠GLU处置和血管松弛障碍中的作用。利用原代培养的EC和VSMC，我们将检测Ang II刺激的RhoA和ROS对EC和VSMC的NO代谢、Na+，K+-ATPase和MBP活性的影响。为了阐明Akt、RhoA和ROS信号在介导Ang II和Ins/IGF-1的反调节作用中的作用，我们将通过将Akt和RhoA的显性负结构和NAD(P)H氧化酶的关键成分之一p22Phox的反义表达导入VSMC来阻断这些信号通路。我们将利用液相色谱-串联质谱仪对信号分子(如Akt、RhoA、Phoxes)和酶(如eNOS、钠泵、MBP)的位点特异性磷酸化和ROS诱导的翻译后修饰进行描述和定量。这些研究将与高血压和胰岛素抵抗啮齿动物模型以及正常血压对照组的血管系统和骨骼肌/脂肪细胞的体内和体外研究相补充。这项拟议的研究将显著提高我们对血管紧张素转换酶II在胰岛素抵抗和高血压发病机制中的作用的认识。