Abstract
ObjectiveMitochondrial dysfunction underlies the development of various cardiovascular diseases. Previously, we reported the presence of mitochondrial nitric oxide synthase (mtNOS) in cardiomyocytes and for the first time observed that NOS inhibitors decreased the state IIIu respiration in isolated cardiac mitochondria but had no significant effect on basal/state II or ATP production. These effects of NOS inhibitors on mitochondrial respiration in isolated mitochondria were significantly different than the effects on mitochondrial respiration in intact cells suggesting novel functional role of mtNOS. Arginase regulates the levels of arginine available for the local nitric oxide synthases, thereby regulating the NO production. Cardiomyocytes specifically express arginase II localized in mitochondria and has been shown to regulate cardiomyocyte contractility. We hypothesized that inhibition of arginase II increases respiration in isolated murine cardiac mitochondria in vitro.MethodsCardiac mitochondria were isolated from three‐month‐old C57bl6 (n= 4–5) mice and treated with specific arginase inhibitor, S‐(2‐boronoethyl)‐L‐cysteine hydrochloride, S‐(2‐boronoethyl)‐L‐cysteine, BEC (100μM) for 30 minutes in assay buffer containing 10 mM pyruvate and 2 mM malate. Mitochondria (5 μg/well) were plated into cell plates and oxygen consumption rate (OCR) was measured using Agilent Seahorse XFe24 analyzer after serial injections of ADP (5 mM), oligomycin (5 μM), FCCP (5 μM) and antimycin and rotenone (10 μM and 2 μM). Respiratory parameters were reported as OCR (pico moles of oxygen/minute).ResultsArginase inhibition did not significantly alter the basal/state II respiration (106.2 ± 6.2 vs 98.8 ± 1.6) or state IVo (87.1 ± 8.0 vs 80.9 ± 7.7) or proton leak (53.2 ± 11.6 vs 41.7 ± 5.7). Interestingly, arginase inhibition increased the ADP‐induced or state III respiration by 25% (329.9 ± 18.7 vs 263.5 ± 20.2, p=0.05) and ATP production by 30% (237.5 ± 13.5 vs 182.7 ± 15.5, p=0.03) compared to untreated mitochondria. Uncoupler‐induced or state IIIu respiration (state IIIu) showed strong trend towards increase by 33% (237.5 ± 13.6 vs 182.7 ± 15.5, p=0.059) in isolated mitochondria treated with arginase inhibitor.ConclusionsArginase II negatively regulates the mitochondrial respiration in the heart and thus along with mtNOS it may play critical role in cardiovascular physiology and pathophysiology.Support or Funding InformationSupport: National Institute of Health: National Institute of General Medical Sciences and National Institute of Neurological Disorders and Stroke (Katakam: R01NS094834).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
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