evidence that the pentose phosphate pathway (PPP)/glucose-6-phosphate dehydrogenase (G6PD) and NADPH redox is involved in modulating contractile function of the coronary (CA) artery. However, the machanism(s) by which G6PD and NADPH modulates contractile function of CA are obscure. Therefore, the primary focus of this proposal will be, to elucidate the signaling pathways involved in mediating the effects of G6PD and NADPH redox on smooth muscle cell L-type Ca2+ currents and CA function. To achieve these goals, we will, in Aim #1 determine if G6PD is active in the sub- cellular fractions of resting and contracting CA, by estimating the rate of glucose oxidation, and the G6PD activity levels by biochemical and radioisotope tracer assays. Furthermore, we will identify mechanism(s) involved in contractile agents-induced-G6PD activation, by investigating the role of PKC and metabolic pathways.
In Aim #2, we will determine whether G6PD mediates L-type Ca2+ channel activity, intracellular Ca2+, and vasomotor tone in resting and contracting CA, by examining L-type Ca2+ function, measure intracellular Ca2+ changes and vasomotor function after inhibiting G6PD with pharmacological agents and siRNA transfection, and in G6PD deficient mouse aorta.
In Aim #3, we will determine whether glucose-6-phosphate dehydrogenase modulates the L-type Ca2+ channel function, intracellular Ca2+, contraction and redox changes, in CA via direct physical interaction with the ion channel proteins (alpha subunit of CaV1.2), by co-immunoprecipitation, co-localization and in-vitro binding assays. Additionally, we will determine whether direct binding of NADP+ or NADPH to the L-type Ca2+ channel protein inactivates the channel and whether changes in the levels of reduced/oxidized glutathione (GSH) or hydrogen peroxide (H2O2), induced by decrease in NADPH levels (due to the inhibition of G6PD activity), modulates L-type Ca2+ channel function, in smooth muscle cells isolated from coronary and aorta of G6PD deficient mouse. The PPP/G6PD and NADPH redox is up-regulated in diabetes, pulmonary hypertension and heart failure, thereby suggesting a potential role for G6PD and NADPH redox in profoundly impairing the contractile function of blood vessels in these diseases. This study, on completion as anticipated, will prove to be useful in developing novel therapies for the treatment of vascular dysfunction in pulmonary hypertension, diabetes, and heart failure. Project Description Page 6
|Rawat, Dhawjbahadur K; Alzoubi, Abdallah; Gupte, Rakhee et al. (2014) Increased reactive oxygen species, metabolic maladaptation, and autophagy contribute to pulmonary arterial hypertension-induced ventricular hypertrophy and diastolic heart failure. Hypertension 64:1266-74|
|Ochi, Rikuo; Chettimada, Sukrutha; Gupte, Sachin A (2014) Poly(ethylene glycol)-cholesterol inhibits L-type Ca2+ channel currents and augments voltage-dependent inactivation in A7r5 cells. PLoS One 9:e107049|
|Chettimada, Sukrutha; Ata, Hirotaka; Rawat, Dhwajbahadur K et al. (2014) Contractile protein expression is upregulated by reactive oxygen species in aorta of Goto-Kakizaki rat. Am J Physiol Heart Circ Physiol 306:H214-24|
|Schneider, Andrew M; Rawat, Dhwajbahadur; Weinstein, L Steve et al. (2012) Effects of laparoscopic Roux-en-Y gastric bypass on glucose-6 phosphate dehydrogenase activity in obese type 2 diabetics. Surg Endosc 26:823-30|
|Gupte, Rakhee S; Ata, Hirotaka; Rawat, Dhawjbahadur et al. (2011) Glucose-6-phosphate dehydrogenase is a regulator of vascular smooth muscle contraction. Antioxid Redox Signal 14:543-58|
|Ata, Hirotaka; Rawat, Dhwajbhadur K; Lincoln, Thomas et al. (2011) Mechanism of glucose-6-phosphate dehydrogenase-mediated regulation of coronary artery contractility. Am J Physiol Heart Circ Physiol 300:H2054-63|
|Wolin, Michael S; Gupte, Sachin A; Mingone, Christopher J et al. (2010) Redox regulation of responses to hypoxia and NO-cGMP signaling in pulmonary vascular pathophysiology. Ann N Y Acad Sci 1203:126-32|
|Wolin, Michael S; Gupte, Sachin A; Neo, Boon Hwa et al. (2010) Oxidant-redox regulation of pulmonary vascular responses to hypoxia and nitric oxide-cGMP signaling. Cardiol Rev 18:89-93|
|Gupte, Sachin A (2010) Targeting the Pentose Phosphate Pathway in Syndrome X-related Cardiovascular Complications. Drug Dev Res 71:161-167|
|Gupte, Rakhee S; Rawat, Dhawjbahadur K; Chettimada, Sukrutha et al. (2010) Activation of glucose-6-phosphate dehydrogenase promotes acute hypoxic pulmonary artery contraction. J Biol Chem 285:19561-71|
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