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.
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