L-arginine metabolism in diabetes-induced coronary dysfunction

描述（由申请人提供）：我们的总体目标是确定β-淀粉酶活性在糖尿病诱导的心血管疾病中的特定作用。这些研究将为开发新的策略以预防导致糖尿病心脏病的血管损伤奠定基础。内皮细胞功能障碍是糖尿病血管并发症的显著特征。我们的初步数据显示，糖尿病大鼠冠状动脉血管舒张功能受损和血管周围纤维化/肥大，这与活性氧（ROS）的形成和增加的精氨酸酶活性有关，这将L-arg从eNOS转移到鸟氨酸途径。我们的数据还表明，高葡萄糖和ROS会增加血管紧张素转换酶I的表达，他汀类HMG CoA还原酶抑制剂和补充性L-瓜氨酸（L-cit）可阻断糖尿病增加ROS、诱导血管紧张素转换酶表达和活性以及导致冠状动脉纤维化和血管舒张受损的作用。这些效应可以通过研究来解释，研究表明糖尿病和高葡萄糖诱导的ROS激活小G蛋白RhoA，从而激活Rho激酶（ROCK）。ROCK反过来激活eNOS酶并抑制eNOS，而NO的产生抑制eNOS酶/鸟氨酸途径，他汀类药物增强eNOS活性、NO形成并抑制RhoA/ROCK的激活。此外，我们的研究表明，L-cit，增加L-arg水平和抑制NOS活性，保持L-arg可用性比L-arg本身更好。基于这些数据，我们假设糖尿病和高血糖通过RhoA/ROCK介导的辅酶A酶/鸟氨酸通路的激活引起血管功能障碍。降低的L-精氨酸对eNOS的可用性使酶解偶联以形成超氧化物而不是NO，这减少了血管舒张。酶/鸟氨酸途径的加速活性诱导血管周围纤维化、肥大，导致血管硬化。将使用分子、药理学和生物化学方法的组合来检查该模型，以检验以下用于糖尿病动物、糖尿病患者的血管和血液以及培养细胞的实验的假设。1)定义糖尿病激活胰蛋白酶的分子机制。2)检测糖尿病是否通过激活血管紧张素转换酶导致EC依赖性血管舒张功能受损。3)检测糖尿病是否通过激活血管酶导致血管纤维化、肥大和硬化。4)测试糖尿病是否通过激活RhoA/ROCK来激活谷胱甘肽酶/鸟氨酸途径。公共卫生相关性：冠状动脉血管性心脏病可导致心脏病发作，是一个主要的健康问题，这是更糟糕的人谁有糖尿病。了解糖尿病中减少心脏血流量的因素和事件顺序对于预防和治疗这种疾病至关重要。限制酶的作用，可以防止心脏获得足够的血液，并导致心脏血管增厚和僵硬，为减少这种疾病的发生提供了很大的希望。

DESCRIPTION (provided by applicant): Our overall aim is to define the specific role of arginase activity in diabetes-induced cardiovascular disease. These studies will set the stage for developing new strategies to prevent the vascular injury that leads to diabetic heart disease. Endothelial cell (EC) dysfunction is a prominent feature in the vascular complications of diabetes. Our preliminary data show impaired coronary vasorelaxation and perivascular fibrosis/hypertrophy in diabetic rats, which are associated with reactive oxygen species (ROS) formation and increased arginase activity, which diverts L-arg from eNOS to the ornithine pathway. Our data also show that high glucose and ROS increase arginase I expression and that statin HMG CoA reductase inhibitors and supplemental L-citrulline (L-cit) block the actions of diabetes in increasing ROS, inducing arginase expression and activity and causing coronary fibrosis and impaired vasorelaxation. These effects are explained by studies showing that diabetes and high glucose-induced ROS activate the small G protein RhoA, which activates Rho kinase (ROCK). ROCK in turn activates arginase and inhibits eNOS, whereas NO production inhibits the arginase/ornithine pathway and statins enhance eNOS activity, NO formation and inhibit activation of RhoA/ROCK. Additionally, our studies indicate that L-cit, which increases L-arg levels and inhibits arginase activity, maintains L-arg availability for NOS better than L-arg itself. Based on these data, we hypothesize that diabetes and high glucose cause vascular dysfunction by RhoA/ROCK-mediated activation of the arginase/ornithine pathway. Decreased L-arginine availability to eNOS uncouples the enzyme to form superoxide rather than NO which reduces vasodilation. Accelerated activity of the arginase/ornithine pathway induces perivascular fibrosis, hypertrophy, leading to vascular stiffening. This model will be examined using a combination of molecular, pharmacological and biochemical approaches to test the following hypotheses for experiments in diabetic animals, vessels and blood from diabetic patients and cultured cells. 1) Define the molecular mechanisms by which diabetes activates arginase. 2) Test whether diabetes causes impaired EC-dependent vasorelaxation by activating arginase. 3) Test whether diabetes causes vascular fibrosis, hypertrophy and stiffening by activating arginase. 4) Test whether diabetes activates the arginase/ornithine pathway by activation of RhoA/ROCK. PUBLIC HEALTH RELEVANCE: Coronary vascular heart disease which can cause heart attacks is a major health problem, which is much worse in people who have diabetes. Understanding the factors and sequence of events in diabetes which reduce blood flow to heart is critical to preventing and treating this disease. Limiting actions of an enzyme which can prevent the heart from getting enough blood and cause thickening and stiffness of heart blood vessels offers much promise for reducing the incidence of this disease.