Preliminary data using isolated mouse heart EC (MHEC) in vitro, coronary microvessel ex vivo, and LAD ligation surgery in vivo, we demonstrate in the current application that short-term (4-12 weeks) temporal increase in EC-specific ROS induce AMPK-eNOS-mediated endothelium-dependent coronary vasodilatation, and AMPK-mTOR-mediated protective autophagy and PGC1α/HIF1α-induced expression of the mitochondrial antioxidant enzymes in MHEC. However, sustained (>16 weeks) increase in EC-ROS resulted in inhibition of MHEC proliferation, marked reduction in endothelium-dependent coronary vasodilatation and nitro-tyrosine-mediated mitochondrial injury-induced autophagy. This proposal will examine a central hypothesis that whereas short-term temporal (4-12 weeks) increase (1.8-fold) in Nox2-derived ROS induces AMPK-eNOS-, AMPK-mTOR-, and AMPK-PGC1α-mediated improvement in EC function, sustained increase in EC-ROS beyond 16 weeks leads to excessive ONOO, mitochondrial injury and mitophagy resulting in endothelial dysfunction. We will test 3 Aims: Specific Aim 1: To elucidate the molecular mechanisms by which EC-specific short-term (16 weeks) EC-specific increase in ROS results in mitochondrial injury and mitophagy leading to coronary endothelial dysfunction. We will use isolated MHEC and coronary vessels to determine mitochondrial ROS, membrane potential, mitophagy and coronary vasodilatation.Specific Aim 3: To determine whether short-term increase in EC-ROS increases vessel density in ischemic myocardium through activation of AMPK-induced mitochondrial antioxidant defense mechanism, and EC survival. Using LAD ligation model in animals with high ROS exposure for 16 weeks, we will determine vessel density and EC survival/apoptosis in ischemic myocardium (CD31, BrdU and TUNEL), and cardiac function by P-V loop. Outcomes of the proposed studies will help develop therapeutic modalities utilizing regulated modulation of ROS in vascular diseases.
|Effective start/end date||07/01/2014 → 06/30/2017|