Neovascularization, the natural physiological process of formation of new blood vessels, is extremely important to ameliorate the function of the heart that undergoes ischemic stress. This process is potentially important for the treatment of ischemic heart and limb ischemia which includes formation of capillaries (angiogenesis) and collateral arteries. We demonstrated that ischemic preconditioning (IP) of the heart can trigger angiogenesis. Reactive oxygen species (ROS) derived from gp91phox (Nox2)-containing NAD(P)H oxidase is found to be involved in angiogenic process which involves vascular endothelial growth factor (VEGF) signaling. The role of gp91phox-derived ROS in neovascularization in response to tissue ischemia is unknown. Again to determine this redox signaling we will examine the involvement of principal redox regulated genes, thioredoxin and glutaredoxin in the IP hearts. Discerning the myriad pathways that are possibly associated with this vascular growth process is therefore of utmost importance to develop targeted drugs and bring the therapy from bench to bedside. Previous studies, including ours, also have shown that transient ischemia upregulates VEGF protein in cardiac tissues. Thus, this study will attempt to address an important clinical issue by identifying potential candidates of VEGF signaling in several gene knockout animals such as Flk-1+/-, Flt-1+/-, gp-91 phox-/-, HO-1+/+, Trx-1+/+, Grx1+/+, Grx1-/-. This study will utilize a broad multidisciplinary approach that will combine various techniques, including latest molecular biology techniques, physiology and gene targeting.
Aim 1. Activation and molecular mechanism of the GSK-3beta phosphorylation and beta-catenin translocation followed by pro-angiogenic molecule VEGF expression will be investigated.
Aim 2. Genetically-engineered mice will be used such as MK2-/- to identify down stream target candidates of VEGF signaling through its receptors in IP as well as ischemic reperfused myocardium in vivo Aim 3. ROS mediated stress plays significant role in VEGF induced angiogenesis through Trx and Grx protein.
Aim 4. To gain insights into the mechanisms and to identify candidate genes involved in VEGF- NAD(P)H oxidase signaling in gp91phox-/- homozygous mice we will use Affymetrix Gene chip Analysis. This proposal will provide and will identify important new insights into the cellular and molecular mechanisms of the cardioprotection provided by VEGF signaling through important candidate genes. ? ? ?
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