Endothelial Lipoxygenase Metabolites and Vascular Tone

内皮脂氧合酶代谢物和血管张力

• 批准号: 8470696
• 负责人: WILLIAM BRYSON CAMPBELL
• 金额: $ 36.41万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8470696
• 关键词: Acetylcholine; Acids; Address; Agonist; Angiotensin II; Angiotensins; Apamin; Arachidonate 15-Lipoxygenase; Arachidonic Acids; Arteries; Binding; Binding Sites; Biochemical; Biological Assay; Blood Pressure; Blood Vessels; Calcium; Cardiovascular Diseases; Cell membrane; Cells; Cholesterol; Contractile Proteins; Coupled; Diabetes Mellitus; Eicosatrienoic Acid; Endothelial Cells; Endothelium; Endothelium-Dependent Relaxing Factors; Epoprostenol; Functional disorder; G-substrate; GTP Binding; GTP-Binding Proteins; Goals; Guanosine Triphosphate; Health; Heart failure; Heterotrimeric GTP-Binding Proteins; Hypertension; Hypoxia; Innovative Therapy; Interleukins; Knowledge; Link; Lipoxygenase; Mediating; Membrane; Methods; Molecular; Molecular Mechanisms of Action; Monomeric GTP-Binding Proteins; Myocardial Ischemia; Myosin Light Chains; Nitric Oxide; Nitric Oxide Synthase; Norepinephrine; Oryctolagus cuniculus; Pathway interactions; Potassium; Potassium Channel; Prostaglandin-Endoperoxide Synthase; Proteins; Regulation; Relaxation; Rho-associated kinase; Role; Signal Pathway; Smooth Muscle Myocytes; Testing; Therapeutic; Vascular Diseases; Vasoconstrictor Agents; Vasodilator Agents; angiotensin hypertension; blood pressure regulation; constriction; cyclooxygenase 2; guanine nucleotide binding protein; improved; insight; myosin phosphatase; novel; novel strategies; novel therapeutic intervention; patch clamp; phospholipase inhibitor; receptor; relaxing factor; vasoconstriction

DESCRIPTION (provided by applicant): Endothelial cells release nitric oxide, prostaglandin I2 and endothelium-derived hyperpolarizing factor (EDHF) that reduce vascular tone and counteract vasoconstriction. Ischemic heart disease, heart failure and hyperten- sion are associated with vasoconstriction that is attributed, in part, to reductions in endothelium-dependent di- lation. Therapeutic approaches restoring endothelial function represent promising new treatments for cardio- vascular diseases with a major impact on health. We have identified a 15-lipoxygenase (LO) metabolite of ara- chidonic acid, 11(R),12(S),15(S)-trihydroxy-eicosatrienoic acid (11,12,15-THETA), that acts as an EDHF. It relax arteries, in part, by activating apamin-sensitive, small conductance, calcium-activated potassium (SKCa) channels and hyperpolarizing smooth muscle cells (SMCs). It also acts by inhibition of RhoA activation. 11,12,15-THETA mediates a portion of the endothelium-dependent relaxations to acetylcholine (ACh), in- creases in flow and AA and opposes the action of vasoconstrictors. We will test the hypothesis that 11,12,15- THETA mediates relaxations to ACh and AA by a unique combination of mechanisms: (1) activating a hetero- trimeric guanine nucleotide binding protein (G protein) that opens SKCa channels resulting in hyperpolarization of SMCs and (2) inhibiting the activation of the small GTPase RhoA reducing the calcium sensitivity of contrac- tile proteins. These studies will indicate that 11,12,15-THETA is an EDHF and an endothelium-derived calcium (Ca) desensitizer. 11(R),12(S),15(S)-THETA increases GTP335S binding to rabbit arterial membranes implicat- ing a G protein-coupled binding site/receptor. Using biochemical and molecular methods, we will identify the G protein(s) that is coupled to 11,12,15-THETA action. Using patch clamp and bioassay methods, we will char- acterize the role of the G protein in the regulation of SKCa channel activity, hyperpolarization and dilation by 11,12,15-THETA. These studies will provide new insights into the molecular mechanism of action of 11,12,15- THETA and the regulation of vascular SKCa channel activity. Agonists such as norepinephrine and angiotensin promote constriction by activating the RhoA-Rho kinase cascade that sensitizes contractile proteins to Ca. 11,12,15-THETA decreases the formation of the active form of RhoA, RhoA-GTP. These studies will test the hypothesis that 11,12,15-THETA causes relaxation by inhibiting RhoA activation resulting in decreased Rho kinase activity increasing myosin light chain phosphatase activity and decreasing phosphorylated myosin light chain. These studies will access the effect of 11,12,15-THETA on RhoA activity in SMCs and arteries and de- termine the contribution of RhoA inhibition to the relaxations to 11,12,15-THETA. The role of a G protein in RhoA inhibition by 11,12,15-THETA will be determined. These studies will identify two new mechanisms of action for the endothelium-derived relaxing factor 11,12,15-THETA and provide a new therapeutic approach to enhancing endothelial function.

描述（由申请人提供）：内皮细胞释放一氧化氮、前列腺素 I2 和内皮源性超极化因子 (EDHF)，从而降低血管张力并抵消血管收缩。缺血性心脏病、心力衰竭和高血压与血管收缩有关，部分原因是内皮依赖性扩张的减少。恢复内皮功能的治疗方法代表了对健康产生重大影响的心血管疾病的有前途的新疗法。我们已经鉴定出花生四烯酸的 15-脂氧合酶 (LO) 代谢物 11(R),12(S),15(S)-三羟基二十碳三烯酸 (11,12,15-THETA)，它充当EDHF。它在一定程度上通过激活 apamin 敏感的小电导钙激活钾 (SKCa) 通道和超极化平滑肌细胞 (SMC) 来放松动脉。它还通过抑制 RhoA 激活发挥作用。 11,12,15-THETA 介导部分内皮依赖性乙酰胆碱 (ACh) 松弛，增加血流和 AA，并对抗血管收缩剂的作用。我们将测试以下假设：11,12,15-THETA 通过独特的机制组合介导 ACh 和 AA 的松弛：(1) 激活异源三聚鸟嘌呤核苷酸结合蛋白（G 蛋白），打开 SKCa 通道，导致 SKCa 通道超极化。 SMC 和 (2) 抑制小 GTP 酶 RhoA 的激活，降低收缩蛋白的钙敏感性。这些研究表明 11,12,15-THETA 是一种 EDHF 和内皮源性钙 (Ca) 脱敏剂。 11(R),12(S),15(S)-THETA 增加 GTP335S 与兔动脉膜的结合，暗示 G 蛋白偶联结合位点/受体。使用生化和分子方法，我们将鉴定与 11,12,15-THETA 作用偶联的 G 蛋白。使用膜片钳和生物测定方法，我们将表征 G 蛋白在 11,12,15-THETA 调节 SKCa 通道活性、超极化和扩张中的作用。这些研究将为11,12,15-THETA的分子作用机制和血管SKCa通道活性的调节提供新的见解。去甲肾上腺素和血管紧张素等激动剂通过激活 RhoA-Rho 激酶级联来促进收缩，该级联使收缩蛋白对 Ca 敏感。 11,12,15-THETA 减少 RhoA 活性形式 RhoA-GTP 的形成。这些研究将检验以下假设：11,12,15-THETA 通过抑制 RhoA 激活导致 Rho 激酶活性降低，增加肌球蛋白轻链磷酸酶活性并减少磷酸化肌球蛋白轻链，从而引起放松。这些研究将了解 11,12,15-THETA 对 SMC 和动脉中 RhoA 活性的影响，并确定 RhoA 抑制对 11,12,15-THETA 松弛的贡献。将确定 G 蛋白在 11,12,15-THETA 抑制 RhoA 中的作用。这些研究将确定内皮源性舒张因子11,12,15-THETA的两种新作用机制，并提供增强内皮功能的新治疗方法。