Integrative analysis of coronary adaptations to ischemia

这项建议的目标是描绘与冠状动脉侧枝生长或非萌芽血管生成有关的信号。我们的总体观点是，侧枝生长严重依赖于特定生长因子及其受体的随时间变化的表达。在缺血开始时：受炎症、缺氧和/或细胞氧化还原状态调节的血管生成因子(胰岛素样生长因子-1)的表达增加。B.这些血管生成因子的受体的表达增加(IGF-I和-II受体，Flk-1和Flt受体的血管内皮生长因子，转化生长因子受体的血管内皮生长因子，转化生长因子受体的类型I和II)。2.在足以改善闭塞区缺血的侧支过程中：A.切应力调节的血管生成因子(血小板衍生生长因子-B及其受体(PDGF-α和-β受体)和内皮型一氧化氮合酶[eNOS])的表达增加。B.与血管生成和/或细胞增殖相关的生长因子和受体的表达恢复到基线水平。在糖尿病动物模型中：4.冠状动脉侧支循环生长，对短暂、重复的缺血发作的反应是受损的。5.缺血开始时，受缺氧、炎症和氧化还原状态调节的血管生成因子表达减少或缺失，而这些因子的受体表达。慢性冠状动脉闭塞2分钟可诱导冠脉新生血管形成。血管生成将在几个不同的时间进行评估：1.早期反应；2.快速生长阶段；3.维持期；4.最后生长阶段。除了接受重复缺血刺激的动物外，还将对假手术组和赋形剂对照组进行研究。侧支传导将通过测量闭塞区域的心肌功能、侧支循环(放射性微球)和闭塞后反应性充血反应的减少来评估。与血管生成相关的有丝分裂原将从正常和缺血血管区域的心肌间质透析液中检测(细胞增殖、管状形成、Western分析)。Northern分析和逆转录聚合酶链式反应将被用来评估特定转录本在心肌和血管系统中的表达。这种方法结合了从分子到生理水平的一系列技术，将有助于全面了解与冠状动脉血管生成相关的信号，以及糖尿病破坏冠状动脉侧支循环生长的机制。

The goal of this proposal is to delineate the signals involved in coronary collateral growth or non-budding angiogenesis. Our general thesis is that collateral growth is critically dependent on the time-dependent expression of specific growth factors and their receptors. At the onset of ischemia: a expression of angiogenic factors (insulin-like growth factor-1 that are regulated by inflammation, hypoxia, and/or the cellular redox state is increased. b. expression of receptors for these angiogenic factors is increased (IGF-I and -II receptors, flk-1 and flt receptors for VEGF, TGF receptors for VEGF, TGF receptor Types I and II). 2. During collateralization sufficient to ameliorate ischemia in the occluded territory: a. expression of angiogenic factors regulated by shear stress (platelet derived growth factor-B [PDGF-B] and its receptors (PDGF- alpha and -beta receptor), and endothelial nitric oxide synthase [eNOS]) is increased. b. expression of growth factors and the receptors associated with angiogenesis and/or cellular proliferation returns to baseline. In an animal model of diabetes: 4. Coronary collateral growth, in response to brief, repetitive episodes of ischemia is impaired. 5. At the onset of ischemia, expression of the angiogenic factors regulated by hypoxia, inflammation, and the redox state is reduced or absent; whereas, receptors for these factors are expressed. Coronary angiogenesis will be induced in chronically-instrumented dogs by 2 minute repetitive coronary artery occlusions. Angiogenesis will be evaluated at several different times: 1. Early responses; 2. Rapid growth phase; and 3. Maintenance phase; and 4. Final growth phase. Sham and vehicle control groups will be studied in addition to the animals receiving the repetitive ischemic stimuli. Collateral conductance will be evaluated by measurements of myocardial function in the occluded territory, collateral flow (radioactive microspheres), and diminution of reactive hyperemic responses following the occlusions. Mitogens associated with angiogenesis will be assayed from myocardial interstitial dialysate of the normal and ischemic vascular regions (cell proliferation, tube formation, Western analysis). Northern analysis, and reverse transcriptase polymerase chain reaction will be used to evaluate expression of the specific transcripts in the myocardium and vasculature. This approach integrates a range of techniques from this molecular to physiological levels will facilitate a complete understanding the signals associated with coronary angiogenesis and the mechanisms by which diabetes abrogates the growth of the coronary collateral circulation.