Role of endogenous hydrogen peroxide in the development of nitrate tolerance

doi:10.1016/j.vph.2006.10.007

Vascular Pharmacology

Volume 46, Issue 4, April 2007, Pages 247-252

https://doi.org/10.1016/j.vph.2006.10.007 Get rights and content

Abstract

The present study was designed to test the hypothesis that hydrogen peroxide plays a role in the development of nitrate tolerance. Isolated rat aortic rings were suspended in organ chambers for isometric tension recording. The rings were incubated with (tolerant) and without (control) nitroglycerin (10^− 4 M) for 90 min, followed by repeated rinsing for 1 h. Hydrogen peroxide release in control and tolerant tissues was measured fluorimetrically using amplex red. Nitroglycerin (10^− 9–10^− 4 M) caused concentration-dependent relaxations in control (− logEC₅₀ = 7.15 ± 0.1) and tolerant rings (− logEC₅₀ = 5.83 ± 0.1) contracted with norepinephrine. Nitrate tolerance was evident by a > 20-fold rightward shift in the nitroglycerin concentration–response curve in tissues exposed previously to nitroglycerin for 90 min. Incubation of the rings with the superoxide dismutase (SOD)-mimetic, tempol (10^− 4 M), during the 90-min exposure period to nitroglycerin caused a leftward shift in the nitroglycerin concentration–response curve in tolerant rings (− logEC₅₀ = 6.84 ± 0.2), but had no effect on the response to nitroglycerin in control rings. Treatment of the rings with catalase (1200 U/ml) or ebselen (1.5 × 10^− 5 M), a glutathione peroxidase-mimetic, during the 90-min exposure period to nitroglycerin resulted in a further rightward shift in the nitroglycerin concentration–response curve in tolerant rings (− logEC₅₀ = 5.41 ± 0.1 and 4.98 ± 0.1; catalase and ebselen respectively), without altering the response to nitroglycerin in control rings. In the presence of catalase, the effect of tempol on nitrate tolerance was abolished (− logEC₅₀ = 5.46 ± 0.1). Hydrogen peroxide release was reduced by ∼ 64% in nitrate tolerant tissues when compared to control. The decrease in hydrogen peroxide release was completely reversed by treatment with tempol, whereas treatment with ebselen caused a further decrease in hydrogen peroxide release in nitrate tolerant tissues. Addition of hydrogen peroxide (3 × 10^− 5 M) to nitrate tolerant rings caused a leftward shift in the nitroglycerin concentration–response curve in tolerant rings (− logEC₅₀ = 7.18 ± 0.3), but had no effect on the response to nitroglycerin in control rings. These results suggest that nitrate tolerance is associated with decreased endogenous formation of hydrogen peroxide, which attenuates nitrate tolerance development. SOD-mimetics may reduce nitrate tolerance, in part, by increasing the formation of hydrogen peroxide.

Introduction

Nitroglycerin is widely used in the treatment of patients with ischemic heart disease and other cardiovascular disorders. The therapeutic benefits of nitroglycerin are thought to be due to vasodilation, which results from the formation of nitric oxide via enzymatic metabolism of nitroglycerin (Chen et al., 2005, McDonald and Bennett, 1990). A major limitation to the use of nitroglycerin in long term therapy is that continuous exposure to nitroglycerin rapidly leads to a decline in efficacy, i.e. nitrate tolerance (Fung, 2004, Munzel et al., 2005, Parker and Parker, 1998).

Although the mechanisms underlying nitrate tolerance are incompletely understood, it is now recognized that nitroglycerin induces oxidative stress, which, in turn, contributes to nitrate tolerance (Munzel et al., 1995). Indeed, nitroglycerin increases the formation of superoxide anions, which can directly inactivate nitric oxide and nitroglycerin-metabolizing enzymes (Munzel et al., 1995). Moreover, free radical scavengers, such as ascorbic acid, reduce oxidative stress and attenuate nitrate tolerance (Bassenge et al., 1998, Daniel and Nawarskas, 2000). Superoxide anions spontaneously gain an electron to form hydrogen peroxide, a reaction that is accelerated by intracellular and extracellular forms of superoxide dismutase in the blood vessel wall (Faraci and Didion, 2004). The possible modulating influence of endogenously produced hydrogen peroxide on nitrate tolerance has not been investigated; thus, the present study was designed to examine the role of hydrogen peroxide in the development of nitrate tolerance.

Section snippets

Tissue preparation

Male Sprague–Dawley rats (Zivic, Pittsburgh, PA), 4–5 weeks old, were exsanguinated under halothane anesthesia, in accordance with a protocol approved by the Institutional Animal Care and Use Committee of North Dakota State University. The thoracic aorta was isolated, carefully trimmed of adventitial fat and connective tissue, and cut into rings 3–4 mm in length. Each ring was suspended in water-jacketed organ chambers filled with 25 ml of physiological salt solution (PSS). The solution was

Organ chamber studies

Nitroglycerin caused concentration-dependent relaxations in endothelium-intact aortic rings contracted with norepinephrine (Fig. 1, Fig. 2, Fig. 3, Fig. 4). In untreated control tissues, the − log (M) EC₅₀ for nitroglycerin was 7.15 ± 0.1. The concentration–response curve to nitroglycerin was shifted to the right in rings exposed previously to nitroglycerin (10^− 4 M) for 90 min. Under these conditions, the − log (M) EC₅₀ for nitroglycerin was 5.83 ± 0.1 (P < 0.05 vs. control), resulting in a > 20-fold

Discussion

Recent studies indicate that hydrogen peroxide, a reactive oxygen species, may play a role in vascular biology. Depending on the experimental conditions, hydrogen peroxide has been shown to cause both vasoconstriction and vasodilation (Gil-Longo and Gonzalez-Vazquez, 2005), and it has been suggested as a putative endothelium-derived hyperpolarizing factor (EDHF) (Matoba et al., 2002, Yada et al., 2003). The present study demonstrates a novel role for endogenously formed hydrogen peroxide in the

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Effects of tempol and redox-cycling nitroxides in models of oxidative stress
2010, Pharmacology and Therapeutics
Tempol is a redox-cycling nitroxide that promotes the metabolism of many reactive oxygen species (ROS) and improves nitric oxide bioavailability. It has been studied extensively in animal models of oxidative stress. Tempol has been shown to preserve mitochondria against oxidative damage and improve tissue oxygenation. Tempol improved insulin responsiveness in models of diabetes mellitus and improved the dyslipidemia, reduced the weight gain and prevented diastolic dysfunction and heart failure in fat-fed models of the metabolic syndrome. Tempol protected many organs, including the heart and brain, from ischemia/reperfusion damage. Tempol prevented podocyte damage, glomerulosclerosis, proteinuria and progressive loss of renal function in models of salt and mineralocorticosteroid excess. It reduced brain or spinal cord damage after ischemia or trauma and exerted a spinal analgesic action. Tempol improved survival in several models of shock. It protected normal cells from radiation while maintaining radiation sensitivity of tumor cells. Its paradoxical pro-oxidant action in tumor cells accounted for a reduction in spontaneous tumor formation. Tempol was effective in some models of neurodegeneration. Thus, tempol has been effective in preventing several of the adverse consequences of oxidative stress and inflammation that underlie radiation damage and many of the diseases associated with aging. Indeed, tempol given from birth prolonged the life span of normal mice. However, presently tempol has been used only in human subjects as a topical agent to prevent radiation-induced alopecia.
Prevention of nitroglycerin tolerance in vitro by T0156, a selective phosphodiesterase type 5 inhibitor
2008, European Journal of Pharmacology
The efficacy of nitroglycerin as a vasodilator is limited by tolerance, which develops shortly after treatment begins. The present study aims to examine whether T0156, a newly developed potent and selective inhibitor of phosphodiesterase type 5 (PDE5), could attenuate the tolerance to nitroglycerin on rat aortas. Rat aortic rings were suspended in organ bath for the measurement of changes in isometric tension and nitrate tolerance was acutely induced by preceding exposure for 90 min to 30 µM nitroglycerin. Concentration–response curves to nitroglycerin were obtained on aortic rings pre-contracted with phenylephrine. Pre-exposure of rings with or without endothelium to nitroglycerin reduced the relaxations to nitroglycerin. The tissue levels of cyclic GMP were measured by enzyme immunoassay kit. Treatment with T0156 inhibited and prevented the reduced relaxation and cyclic GMP levels in response to nitroglycerin in tolerant rings. In contrast, nitroglycerin-induced tolerance was unaffected by cilostazol (PDE3 inhibitor) and rolipram (PDE4 inhibitor). Finally, incubation of aortic rings with thromboxane prostanoid receptor antagonist, cyclooxygenase inhibitor, or endothelin ET_A receptor antagonist did not inhibit the development of tolerance. The present results suggest that nitroglycerin tolerance may involve an increased activity of PDE5 but not PDE3 or PDE4 isoforms in vascular smooth muscle cells since T0156 prevents the development of tolerance. Thromboxane A₂, cyclooxygenase (COX)-dependent prostaglandins and endothelin 1 play little role in the acute induction of nitroglycerin tolerance.
LA-419, a nitric-oxide donor for the treatment of cardiovascular disorders
2009, Current Opinion in Investigational Drugs
Chemistry and antihypertensive effects of tempol and other nitroxides
2008, Pharmacological Reviews

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Role of endogenous hydrogen peroxide in the development of nitrate tolerance

Abstract

Introduction

Section snippets

Tissue preparation

Organ chamber studies

Discussion

Eur. J. Pharmacol.

Vascul. Pharmacol.

J. Am. Coll. Cardiol.

Biochem. Biophys. Res. Commun.

J. Am. Coll. Cardiol.

Mech. Ageing Dev.

Roles of superoxide, peroxynitrite, and protein kinase C in the development of tolerance to nitroglycerin

J. Pharmacol. Exp. Ther.

Dietary supplement with vitamin C prevents nitrate tolerance

J. Clin. Invest.

An essential role for mitochondrial aldehyde dehydrogenase in nitroglycerin bioactivation

Proc. Natl. Acad. Sci. U. S. A.

Identification of the subcellular site for nitroglycerin metabolism to nitric oxide in bovine coronary smooth muscle cells

J. Pharmacol. Exp. Ther.

Heterozygous deficiency of manganese superoxide dismutase in mice (Mn−SOD+/−): a novel approach to assess the role of oxidative stress for the development of nitrate tolerance

Mol. Pharmacol.