Regulation of vascular maturation/regression in diabetes

描述(由申请人提供)：糖尿病心肌梗死患者的高死亡率可能与心肌血管生成障碍和冠脉侧支生长障碍有关。我们的长期目标是明确糖尿病心脏血管异常成熟和血管生成障碍的分子机制(S)。这项修订后的提案将调查血管生成素(Ang)/Tie-2和apelin途径在糖尿病相关血管成熟异常和毛细血管退化中的可能中断。我们的实验室发现糖尿病小鼠血管生成素-2(Ang-2)和脯氨酸羟基酶-2(PHD2)的表达持续增加，Ang-1/Tie-2和HIF-11/apelin的表达下降。我们先前证明糖尿病小鼠心肌血管成熟受损；暗示Angiopoietins/Tie-2系统的破坏有利于Ang-2，从而导致未成熟血管形成和毛细血管倒退，这可能是糖尿病心脏血管生成受损的新机制。我们的总体假设是，糖尿病通过涉及Ang-2和PHD2激活的机制扰乱Ang-1/Tie-2和Apelin通路；这些异常导致糖尿病心脏血管异常成熟和毛细血管退缩。具体目标1将确定高血糖干扰血管成熟和毛细血管退缩的机制(S)，重点是Ang-2在Ang-1/Tie-2和apelin途径中断中的作用。利用心脏微血管内皮细胞(EC)、共培养的EC-SMC球体和分离自野生型(WT)或糖尿病db/db小鼠的小鼠主动脉外植体，我们将确定：(I)高糖诱导的Ang-2过量是否扰乱了Ang-1/Tie-2信号转导并抑制了Ang-1诱导的apelin表达；以及(Ii)Ang-2和apelin之间的相互作用对于高糖条件下血管生成和血管退行性变的调控至关重要。在特定的目标2中，我们将在体内心肌缺血模型中确定Ang-2和PHD2的激活在糖尿病相关的血管成熟和血管生成中断以及促进血管退化中的作用。利用Ang-2缺陷和PHD2条件性基因敲除的糖尿病小鼠模型，我们将确定Ang-2缺陷或PHD2内皮细胞缺失是否拯救受损的apelin表达，使未成熟新生血管正常化，并改善心肌血管生成。在具体目标3中，我们将进一步确定全身应用apelin是否能挽救受损的血管生成信号，使未成熟的新生血管正常化，并增加糖尿病心脏的心肌血管生成。我们的研究将为开发有针对性的减少Ang-2和PHD2激活的治疗提供一个框架，以改善或逆转糖尿病血管成熟和血管生成的异常，这些异常是糖尿病状态的特征。

DESCRIPTION (provided by applicant): Impairment of myocardial angiogenesis and coronary collateral growth may contribute to high mortality in diabetic myocardial infarction. Our long-term goal is to define the molecular mechanism(s) responsible for abnormal vascular maturation and impairment of angiogenesis in the diabetic hearts. This revised proposal will investigate a possible disruption in the angiopoietins (Ang)/Tie-2 and apelin pathway in abnormal diabetes- associated vascular maturation and capillary regression. Our laboratory has shown a sustained increase in angiopoietin-2 (Ang-2) and prolyl hydroxylase-2 (PHD2) expression, and reduced Ang-1/Tie-2 and HIF- 11/apelin expression in diabetic mice. Our previous demonstration of impaired myocardial vessel maturation in diabetic mice; implicate that disruption of angiopoietins/Tie-2 system in favor of Ang-2, which leading to immature vessel formation and capillary regression, might be a novel mechanism responsible for impaired angiogenesis in diabetic hearts. Our overall hypothesis is that diabetes disrupts Ang-1/Tie-2 and apelin pathway by a mechanism involving Ang-2 and PHD2 activation; and these abnormalities lead to abnormal vascular maturation and capillary regression in diabetic hearts. Specific Aim 1 will define the mechanism(s) by which hyperglycemia interferes with vascular maturation and capillary regression with a focus on the role of Ang-2 in the disruption of Ang-1/Tie-2 and apelin pathway. Using heart microvascular endothelial cells (EC), co-cultured EC-SMC spheroids and mouse aortic explants isolated from wild type (WT) or diabetic db/db mice, we will determine whether: (i) high glucose-induced excess of Ang-2 disrupts Ang-1/Tie-2 signaling and attenuates Ang-1-induced apelin expression; and (ii) interactions between Ang-2 and apelin are critical for the regulation of angiogenesis and vascular regression under high glucose conditions. In specific aim 2, we will determine the role of Ang-2 and PHD2 activation in diabetes-associated disruption of vascular maturation and angiogenesis and promotion of vessel regression in an in vivo model of myocardial ischemia. Using Ang-2 deficient and PHD2 conditional knockout diabetic mice models, we will determine whether deficiency of Ang-2 or endothelial cell deletion of PHD2 rescues impaired apelin expression, normalizes immature neovessels, and improves myocardial angiogenesis. In specific aim 3, we will further determine whether systemic administration of apelin rescues impaired angiogenic signaling, normalizes immature neovessels, and increases myocardial angiogenesis in diabetic hearts. Our studies will provide a framework for the development of a targeted therapeutic reduction in Ang-2 and PHD2 activation to ameliorate or reverse the abnormalities in diabetic vessel maturation and angiogenesis that characterizes the diabetic state.