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(Hypertension. 1997;29:668-672.)
© 1997 American Heart Association, Inc.

Articles

N-Acetylcysteine Does Not Influence the Activity of Endothelium-Derived Relaxing Factor In Vivo

Mark A. Creager; Mary-Anne Roddy; Kimberly Boles; Jonathan S. Stamler

the Vascular Medicine and Atherosclerosis Unit of the Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, Mass.

	Abstract

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Nitric oxide forms complexes with an array of biomolecular carriers that retain biological activity. This reactivity of nitric oxide in physiological systems has led to some dispute as to whether endothelium-derived relaxing factor is nitric oxide or a closely related adduct thereof, such as a nitrosothiol. In vitro bioassays used to address this question are limited by the exclusion of biological thiols that are requisite for nitrosothiol formation. Thus, the purpose of this study was to obtain insight into the identity of endothelium-derived relaxing factor in vivo. We reasoned that if endothelium-derived relaxing factor is nitric oxide, infusion of physiological concentrations of thiol would potentiate its bioactivity by analogy with effects seen in vitro, whereas nitrosothiol would be resistant to such modulation. We used venous-occlusion plethysmography to study forearm blood flow in normal subjects. Methacholine (0.3 to 10 µg/min) and nitroglycerin (1 to 30 µg/min) were infused via the brachial artery to elicit endothelium-dependent and endothelium-independent vasodilation, respectively. Dose-response determinations were made for each drug before and after an intra-arterial infusion of the reduced thiol, N-acetylcysteine, at rates estimated to achieve a physiological concentration of 1 mmol/L. Methacholine increased forearm blood flow in a dose-dependent manner. Infusion of N-acetylcysteine did not change the sensitivity (ED₅₀, 1.7 versus 1.7 µg/min, P=NS) or maximal response to methacholine. In contrast, thiol increased the sensitivity to nitroglycerin (ED₅₀, 4.7 versus 2.8 µg/min, P<.01). Thus, conflicting with reports in vitro, thiol does not modulate endothelium-derived relaxing factor responses in vivo. These data indicate that sulfhydryl groups are not a limiting factor for endothelium-derived relaxing factor responses in forearm resistance vessels in normal humans and are in keeping with reports that nitrosothiol contributes to endothelium-derived relaxing factor bioactivity in plasma and vascular smooth muscle. Potentiation of the effects of nitroglycerin by N-acetylcysteine can be attributed to its enhanced biotransformation to an endothelium-derived relaxing factor equivalent, such as nitrosothiol. These observations support the notion of an equilibrium between nitric oxide and nitrosothiol in biological systems that may be influenced by redox state.

Key Words: endothelium-derived relaxing factor • nitric oxide • N-acetylcysteine • N^G-monomethyl-L-arginine • forearm blood flow

	Introduction

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The term EDRF was coined to account for the observation that the endothelium liberates a relaxing substance on stimulation with acetylcholine.¹ Later studies in bioassay systems in vitro have shown that EDRF possesses physicochemical and biological properties that closely resemble NO·.^{2 3} Perhaps the most distinctive feature of this molecule is that it exerts its biological activity by virtue of its chemical reactivity, as opposed to the more traditional "noncovalent" interactions of biological mediators with their receptors.⁴ It is the intrinsic reactivity of NO·, resulting in the formation of surrogates possessing similar bioactivity, that has led to the dispute over whether EDRF is actually NO itself or a closely related adduct thereof such as an RS-NO.^{5 6 7} Indeed, both Furchgott et al⁸ and Wei and Kontos⁹ have concluded that EDRF behaves more like RS-NO than NO· on interaction with vascular smooth muscle, eg, in a system replete with physiological thiol. In support of these claims, in vitro studies demonstrate that reconstitution of systems with physiological concentrations of thiol potentiates the activity of EDRF through formation of RS-NO.^{10 11 12 13} Moreover, S-nitrosothiols have recently been identified in vivo^{14 15 16 17} and appear to be more potent smooth muscle relaxants than NO itself.^{12 18 19 20 21}

The purpose of this study was to obtain insight into the identity of EDRF in vivo. We reasoned that if EDRF activity derives solely from NO·, then thiol supplementation with N-acetylcysteine would enhance its activity, ie, through RS-NO formation, by analogy with effects seen in vitro. In contrast, if EDRF (bioactivity) exists largely in adduct form in vivo, then thiol infusion should have little enhancing effect. In best attempts to mimic the original experiments that Furchgott et al performed in vitro,¹ endothelium-dependent relaxation was induced with intra-arterial infusions of methacholine. We also administered nitroglycerin to address related mechanistic issues.

	Methods

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Subjects
We studied 29 normal subjects (16 men and 13 women). Their ages ranged from 22 to 60 years and averaged 42±2 years. Each subject provided a complete history and underwent a physical examination and laboratory analysis to exclude individuals with hypertension, hypercholesterolemia, diabetes mellitus, overt evidence of atherosclerosis, and hematologic renal or hepatic dysfunction. This study was approved by the Human Research Committee at Brigham and Women's Hospital, and each subject gave written informed consent.

Experimental Protocol
All studies were conducted in a temperature-controlled (23°C) room. Alcohol, caffeine, and cigarettes were prohibited within 12 hours of the study. With local anesthesia and sterile conditions, a polyethylene catheter was inserted into a brachial artery of each subject for blood pressure determination and drug infusion. Subjects rested at least 30 minutes after catheter placement so a stable baseline could be established before data collection.

We studied the effect of N-acetylcysteine on endothelium-dependent vasodilation to methacholine chloride and on endothelium-independent vasodilation to nitroglycerin in 20 subjects. During the control period, measurements of FBF and blood pressure were repeated every 10 minutes until stable. Dextrose (5%) was infused intra-arterially at a rate of 0.4 mL/min during the control period. The protocol we used to examine forearm vascular reactivity has been reported previously.²² FBF was measured during intra-arterial infusion of the endothelium-dependent vasodilator methacholine chloride at doses of 0.3, 1, 3, and 10 µg/min, each for 3 minutes, delivered at a rate of 0.4 mL/min. The endothelium-independent vasodilator nitroglycerin was administered in doses of 1, 3, 10, and 30 µg/min, each for 3 minutes, at a rate of 0.4 mL/min. The order of methacholine chloride and nitroglycerin administration was randomized for each subject. Basal conditions were reestablished before each intervention. Drug doses were chosen to achieve decreases in forearm vascular resistance without causing systemic effects. Dose-response curves were generated for each drug infusion.

After baseline conditions were reestablished, subjects were given N-acetylcysteine (Parvolex, Glaxo) intra-arterially at a dose of 11.6 mg/min. The final estimated concentration of 1 mmol/L approximated that shown previously in in vitro studies to potentiate nitroglycerin-induced vasodilation.^{13 23} Hemodynamic measurements were repeated after N-acetylcysteine was infused for 20 minutes. Thereafter, during concurrent administration of N-acetylcysteine, subjects again received intra-arterial infusions of methacholine chloride and nitroglycerin.

To confirm that methacholine chloride induced endothelium-dependent vasodilation via NO release, we assessed the effect of the NO synthase antagonist L-NMMA in nine subjects. In these individuals, FBF was measured during intra-arterial infusion of methacholine chloride (0.3 to 10 µg/min), before and during intra-arterial infusion of L-NMMA, at a dose of 2 mg/min.

Hemodynamic Measurements
Bilateral FBF was determined by venous-occlusion strain-gauge plethysmography using calibrated mercury-in-Silastic strain gauges (DE Hokanson, Inc) and expressed as milliliters per 100 mL tissue per minute.²⁴ Each arm was supported above heart level. Venous-occlusion pressure averaged 34±1 mm Hg. Circulation to the hand was prevented by inflation of a wrist cuff to suprasystolic pressure before each FBF determination. Determination of FBF comprised at least five separate measurements performed at 10- to 15-second intervals. By measuring blood flow in the infused arm, one can determine the direct effect of the vasoactive drug. By measuring blood flow in the noninfused arm, one can be assured that systemic effects have not occurred if no change in blood flow develops during the drug infusion. Blood pressure was measured via an intra-arterial cannula attached to a blood pressure transducer aligned to an amplifier on a physiological recorder (Gould, Inc). Heart rate was determined from the simultaneously obtained electrocardiographic signal and calculated from the R-R interval.

Statistical Analysis
Results are presented as mean±SE. Single-factor repeated measures ANOVA followed by a Newman-Keuls post hoc test was used for comparison of the effect of each vasoactive drug before and after administration of N-acetylcysteine or L-NMMA. Statistical significance was accepted at the 95% confidence interval (P<.05).

	Results

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N-Acetylcysteine did not affect basal hemodynamic measurements. No significant changes in FBF (2.8±0.3 versus 2.9±0.3 mL/100 mL tissue per minute), mean blood pressure (79±1 versus 80±1 mm Hg), or heart rate (58±2 versus 58±2 beats per minute) were observed after administration of this drug.

Methacholine chloride increased FBF in these normal subjects (n=20) before and during coadministration of N-acetylcysteine (Fig 1). The FBF dose-response relationship did not differ significantly between the two treatment periods. Before N-acetylcysteine, the ED₅₀ of methacholine was 1.7±0.4 µg/min. This did not change after N-acetylcysteine was added (1.7±0.4 µg/min). Furthermore, the FBF response to the highest dose of methacholine chloride did not differ before and after coadministration of N-acetylcysteine (13.2±1.0 versus 14.1±1.4 mL/100 mL tissue per minute, respectively; P=NS). No changes in FBF occurred in the noninfused arm during either treatment period. Furthermore, intra-arterial infusion of methacholine chloride caused no change in blood pressure or heart rate before or during coadministration of N-acetylcysteine.

View larger version (18K): [in this window] [in a new window]   Figure 1. FBF response to methacholine chloride before and during coadministration of N-acetylcysteine. The dose-response relationship did not differ significantly between treatment periods. Values are mean±SE.

Nitroglycerin also increased FBF in a dose-dependent manner before and during coadministration of the reduced thiol N-acetylcysteine (Fig 2). In contrast to methacholine chloride, however, the forearm vasodilator response was shifted to the left during coadministration of N-acetylcysteine. The ED₅₀ of nitroglycerin was higher before administration of N-acetylcysteine than during its coadministration (4.7±0.5 versus 2.8±0.3 µg/min, P<.01). The change in FBF was significantly greater at nitroglycerin doses of 1 and 3 µg/min. The maximal dose of nitroglycerin caused comparable increases in FBF before and during coadministration of N-acetylcysteine.

View larger version (19K): [in this window] [in a new window]   Figure 2. FBF response to nitroglycerin before and during coadministration of N-acetylcysteine. N-Acetylcysteine significantly increased the sensitivity of nitroglycerin; *P<.01. Values are mean±SE.

To confirm that the vasodilator effects of methacholine in vivo in humans are mediated by endothelium-derived NO, we administered L-NMMA to nine subjects. L-NMMA significantly attenuated the FBF response to methacholine (P=.01). The ED₅₀ of methacholine was less before infusion of L-NMMA than during its coadministration (1.8±0.2 versus 2.8±0.3 µg/min, P<.01).

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
The new information derived from these experiments is that thiol supplementation with N-acetylcysteine enhances the FBF response to the exogenous NO donor nitroglycerin but not to methacholine chloride, an endothelium-dependent vasodilator that stimulates EDRF release. These findings suggest that EDRF (bioactivity) exists at least in part in adduct form, primarily as a nitrosothiol. The following discussion puts our findings in perspective with prior studies that have attempted to define EDRF.

There has been a fair amount of controversy over the identity of EDRF. Much of this dispute has centered on the question of whether EDRF is NO· itself or a closely related adduct thereof, such as RS-NO.^{2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20} Reflection on these studies suggests that the apparent controversy may be largely explained by differences in methodology, as the identity of EDRF appears to depend critically on assay conditions: Where thiol (ie, the substrate for adduct formation) is excluded from (bio)assays, the (bio)activity of the system exists mainly in the form of free NO·.^{8 12 25 26} However, when thiol is present at physiological concentrations (0.5 to 10 mmol/L), the activity of EDRF is increased significantly through formation of RS-NO.^{10 11 12 13} The critical importance of thiol concentration in the regulation of EDRF responses is underscored by the potential for lower concentrations of thiol (<100 µmol/L) to inhibit NO·.²¹ In this context, the finding that thiol has little, if any, effect on EDRF responses in vivo strongly argues that NO bioactivity exists predominantly in adduct form under normal physiological conditions. These observations are in keeping with the reports that RS-NO predominates over other bioactive NO· equivalents in plasma^{14 15 16} and best account for EDRF bioactivity on its interaction with vascular smooth muscle.^{8 9 10} Indeed, in microvessels in which EDRF exerts physiological control, the activity of RS-NO is superior to that of NO·.¹⁹ Thus, the findings in our study are essentially identical to those performed in vitro under the same redox conditions—specifically, in systems replete with thiol.

Biological thiols can influence the activity of nitrogen oxides in several ways. The prevailing biochemical mechanism will depend on the identity and concentration of thiol, the identity of the NO donor compound, and the assay in question. Notwithstanding this complexity, the effects of thiol can be broadly divided into those that influence common aspects of nitrogen oxide metabolism and those specific to each NO donor. This is well illustrated in the case of nitroglycerin. Thiol potentiation of its vasodilator activity may derive from the enhanced conversion to RS-NO,^{18 23 27} by the scavenging of oxyradicals that inactivate NO·,^{9 16} and/or through facilitating the activation of guanylate cyclase, the enzyme responsible for vasorelaxation.^{28 29} The physiological contributions of these different mechanisms have not previously been elucidated. From the present study, we conclude that the potentiation of the actions of nitroglycerin in vivo is mediated by the enhanced biotransformation to RS-NO and not by the alternative proposed mechanisms, which would have otherwise potentiated the effects of EDRF as well. In this regard, reports of thiol potentiation of RS-NO activity¹² are probably an artifact of in vitro systems in which contaminant metals promote the breakdown of RS-NO, yielding NO·: By complexing these metals, thiols increase the lifetime of RS-NO, giving the appearance of enhancing their potency.³⁰ It is noteworthy that S-nitrosocysteine is exceptionally susceptible to this bioassay artifact.³⁰

The studies on nitroglycerin bring to light one insurmountable limitation of our work, namely, the inability to test the effects of N-acetylcysteine on NO itself. Specifically, our contentions would be strengthened by the direct demonstration that N-acetylcysteine does not influence the bioactivity of exogenous NO. However, we know of no way to administer NO directly to humans or to avoid its propensity to form adducts once administered.

Our results are further complemented by the recent finding that thiol does not modify the hemodynamic effects of nitroprusside [Fe(CN₅)NO],³¹ which, much like RS-NO, contains the NO group in adduct form (in this case, directly coordinated to a metal center).⁴ Taken together, these studies highlight the common functions of thiols and transition metals in stabilizing NO bioactivity.⁴ S-Nitrosothiols and iron-nitrosyl derivatives are also envisioned to play roles in transporting and targeting NO· to specific effector sites. Supporting this notion, low molecular weight thiols facilitate the release of the NO group from more stable protein RS-NO in plasma^{14 32} and from iron-nitrosyl reservoirs in the endothelial cell cytosol.^{33 34} Thiols may thereby deliver the NO moiety to target sites on the cell surface,^{35 36 37} where NO· release is known to be catalyzed by reactions that are both enzymatic^{38 39} and chemically controlled,^{39 40 41} and to extracellular sites,³³ presumably for localized action. The high concentration of thiol in biological systems and the covalent nature of the S-NO bond rationalize the apparent predominance of RS-NO over the other nitrosylated forms.⁴ Indeed, early studies have shown thiol-dependent conversion of nitroprusside to RS-NO.¹⁸

The biochemical mechanisms of RS-NO formation in vivo are not clear. NO does not react well with thiols directly but requires oxidative activation to a species with NO⁺ character.^{4 27} This formal oxidation of NO· can occur by way of its reaction with oxygen, superoxide, or transition metals.^{4 30} NO/O₂ reactions are probably facilitated in membranes. Whether such pathways of RS-NO formation are enzymatically regulated in vivo remains to be established. Recent work by Malinski and Taha³⁷ shows that NO· "chemi-absorbs" (forms chemical adducts) to the plasma membranes of endothelial cells upon cell activation. Systems rich in thiol would be predicted to support transfer of the NO moiety from the plasma membrane, thereby providing an additional mechanism of RS-NO synthesis in vivo. Conversely, in the absence of thiol, NO· itself will be liberated from the cell surface, perhaps explaining the apparent prevalence of NO· in standard in vitro bioassays.^{8 12} Both mechanisms of NO release (ie, heterolytic transfer of NO⁺ and homolytic fission of NO·) are likely to occur in vivo with the balance weighted by redox state. Notwithstanding this caveat, RS-NO may dictate biological responses even when present at lower concentrations than NO because nitrosothiols are often more potent.^{12 18 19 20 30}

In summary, we have shown that thiols potentiate the vasorelaxant activity of nitroglycerin but not of endogenous EDRF in vivo. These data indicate that sulfhydryl groups are not a limiting factor for EDRF responses in forearm resistance vessels in normal humans and are in keeping with reports that RS-NO contributes to EDRF bioactivity in plasma and vascular smooth muscle. Potentiation of the effects of nitroglycerin by N-acetylcysteine can be attributed to the enhanced biotransformation of nitroglycerin to an EDRF equivalent, such as RS-NO. Our observations also suggest that an equilibrium exists between NO and RS-NO in biological systems that is influenced by redox state. Various vascular-related disorders, including stroke, peripheral vascular disease, coronary artery disease, hypertension, and shock, are thought to be mediated at least in part through changes in the general redox state and thiol content of vascular tissue. Therefore, imbalance in the equilibrium between NO and its various bioactive adducts in tissues—by disrupting the packaging of NO and its appropriate channeling mechanisms—may have important mechanistic implications for atherothrombotic disease.

	Selected Abbreviations and Acronyms

 

EDRF = endothelium-derived relaxing factor FBF = forearm blood flow L-NMMA = NG-monomethyl-L-arginine NO = nitric oxide NO· = nitric oxide free radical RS-NO = nitrosothiol

	Acknowledgments

 
This work was supported by a Grant-in-Aid from the American Heart Association and Bristol-Myers Squibb. Dr Creager is the recipient of a National Heart, Lung, and Blood Institute Academic Award in Systemic and Pulmonary Vascular Disease (HL-02663). Dr Stamler is a Pew Scholar in the Biomedical Sciences and the recipient of a Clinical Investigator Award from the National Institutes of Health (HL-02582). We gratefully acknowledge Sharon Coleman for technical assistance and Joanne Normandin and Linda Frye for manuscript preparation.

	Footnotes

 
Reprint requests to Mark A. Creager, MD, Cardiovascular Division, Brigham and Women's Hospital, 75 Francis St, Boston, MA 02115.

Received September 15, 1995; first decision October 26, 1995; first decision January 9, 1996;

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Furchgott RF, Zawadzki JV, Cherry PD. Role of the endothelium in the vasodilator response to acetylcholine. In: Vanhoutte PM, Leusen I, eds. Vasodilation. New York, NY: Raven Press Publishers; 1981:49-66.

2. Palmer RMJ, Ferrige AG, Moncada S. Nitric oxide release accounts for the biological activity of endothelium-derived relaxing factor. Nature. 1987;327:524-526.[Medline] [Order article via Infotrieve]

3. Ignarro LJ, Buga GM, Wood KS, Byrns RE, Chaudhuri G. Endothelium-derived relaxing factor produced and released from artery and vein is nitric oxide. Proc Natl Acad Sci U S A. 1987;84:9265-9269.[Abstract/Free Full Text]

4. Stamler JS, Singel DJ, Loscalzo J. Biochemistry of nitric oxide and its redox-activated forms. Science. 1992;258:1898-1902.[Abstract/Free Full Text]

5. Myers PR, Minor RL, Guerra R, Bates JN, Harrison DG. Vasorelaxant properties of the endothelium-derived relaxing factor more closely resemble S-nitrosocysteine than nitric oxide. Nature. 1990;345:161-163.[Medline] [Order article via Infotrieve]

6. Greenberg SG, Wilcox DE, Rubanyi GM. Endothelium-derived relaxing factor released from canine femoral artery by acetylcholine cannot be identified as free nitric oxide by electron paramagnetic resonance spectroscopy. Circ Res. 1990;67:1446-1452.[Abstract/Free Full Text]

7. Angus JA, Cocks TM. Endothelium-derived relaxing factor. Pharmacol Ther. 1989;41:303-351.[Medline] [Order article via Infotrieve]

8. Furchgott RF, Jothianandan D, Khan MT. Comparison of nitric oxide, S-nitrosocysteine and EDRF as relaxants of rabbit aorta. Jpn J Pharmacol. 1992;58:185P-191P.

9. Wei EP, Kontos HA. H₂O₂ and endothelium-dependent cerebral arteriolar dilation. Hypertension. 1990;16:162-169.[Abstract/Free Full Text]

10. Stamler JS, Mendelsohn ME, Amarante P, Smick D, Andon N, Davies PF, Cooke JP, Loscalzo J. N-acetylcysteine potentiates platelet inhibition by endothelium-derived relaxing factor. Circ Res. 1989;65:789-795.[Abstract/Free Full Text]

11. Cooke JP, Stamler JS, Andon N, Davies PF, Loscalzo J. Flow stimulates endothelial cells to release a nitrovasodilator that is potentiated by reduced thiol. Am J Physiol. 1990;28:H804-H812.

12. Feelisch M, tePoel M, Zamora R, Oeussen A, Moncada S. Understanding the controversy over the identity of EDRF. Nature. 1994;368:62-65.[Medline] [Order article via Infotrieve]

13. Stamler JS, Simon DI, Osborne J, Mullins M, Jaraki O, Michel T, Singel D, Loscalzo J. S-nitrosylation of proteins with nitric oxide: synthesis and characterization of biologically active compounds. Proc Natl Acad Sci U S A. 1992;89:444-448.[Abstract/Free Full Text]

14. Stamler JS, Jaraki O, Osborne J, Simon DI, Keaney J, Vita J, Singel D, Valeri CR, Loscalzo J. Nitric oxide circulates in mammalian plasma primarily as an S-nitroso adduct of serum albumin. Proc Natl Acad Sci U S A. 1992;89:7674-7677.[Abstract/Free Full Text]

15. Keaney JF, Simon DI, Stamler JS, Jaraki O, Scharfstein J, Vita JA, Loscalzo J. NO forms an adduct with serum albumin that has endothelium-derived relaxing factor-like properties. J Clin Invest. 1993;91:1582-1589.

16. Gaston B, Reilly J, Drazen JM, Fackler J, Ramdev P, Arnelle D, Mullins ME, Sugarbaker DJ, Chee C, Singel DJ, Loscalzo J, Stamler JS. Endogenous nitrogen oxides and bronchodilator S-nitrosothiols in human airways. Proc Natl Acad Sci U S A. 1993;90:10957-10961.[Abstract/Free Full Text]

17. Stamler JS, Loh E, Roddy MA, Hoffmann KF, Creager MA. Nitric oxide regulates basal systemic and pulmonary vascular resistance in normal humans. Circulation. 1994;89:2035-2040.[Abstract/Free Full Text]

18. Ignarro LJ, Lippton H, Edwards JC, Baricos WH, Hyman AL, Kadowitz PJ, Gruetter CA. Mechanism of vascular smooth muscle relaxation by organic nitrates, nitrites, nitroprusside and nitric oxide: evidence for the involvement of S-nitrosothiols as active intermediates. J Pharmacol Exp Ther. 1981;218:739-749.[Free Full Text]

19. Sellke FW, Myers PR, Bates JN, Harrison DG. Influence of vessel size on the sensitivity of porcine coronary microvessels to nitroglycerin. Am J Physiol. 1990;258:H515-H520.[Abstract/Free Full Text]

20. Gaston B, Drazen J, Jansen A, Sugarbaker DA, Loscalzo J, Richards W, Stamler JS. The relaxation of human bronchial smooth muscle by S-nitrosothiols in vitro. J Pharmacol Exp Ther. 1994;268:978-984.[Abstract/Free Full Text]

21. Liu X, Martin W, Gillespie JS. Role of S-nitrosothiols in non-adrenergic non-cholinergic relaxation of the bovine retractor penis muscle. Endothelium. 1993;1:238. Abstract.

22. Creager MA, Cooke JP, Mendelsohn ME, Gallagher SJ, Coleman SM, Loscalzo J, Dzau VJ. Impaired vasodilation of forearm resistance vessels in hypercholesterolemic humans. J Clin Invest. 1990;86:228-234.

23. Fung H-L, Chong S, Kowaluk E, Hough K, Kakemi M. Mechanisms for the pharmacologic interaction of organic nitrates with thiols: Existence of an extracellular pathway for the reversal of nitrate vascular tolerance by N-acetylcysteine. J Pharmacol Exp Ther. 1988;345:524-531.

24. Hokanson DE, Sumner DS, Strandness DE Jr. An electrically calibrated plethysmograph for direct measurement of limb blood flow. IEEE Trans Biomed Eng. 1975;22:25-29.[Medline] [Order article via Infotrieve]

25. Furchgott RF, Khan MT, Jothianandan D. Comparison of properties of nitric oxide and endothelium-derived relaxing factor: some cautionary findings. In: Rubanyi GM, Vanhoutte PM, eds. Endothelium-derived Relaxing Factors. Basel, Switzerland: Karger; 1990:8-21.

26. Jia L, Furchgott RF. Inhibition by sulfhydryl compounds of vascular relaxation induced by nitric oxide and endothelium-derived relaxing factor. J Pharmacol Exp Ther. 1993;267:371-378.[Abstract/Free Full Text]

27. Feelisch M, Noack EA. Correlation between nitric oxide formation during degradation of organic nitrates and activation of guanylate cyclase. Eur J Pharmacol. 1987;139:19-30.[Medline] [Order article via Infotrieve]

28. Ignarro LJ. Biological actions and properties of endothelium-derived nitric oxide formed and released from artery and vein. Circ Res. 1989;65:1-21.[Free Full Text]

29. Stamler JS, Loscalzo J. The antithrombotic effects of organic nitrates. Trends Cardiovasc Med. 1991;1:346-353.

30. Stamler JS. S-nitrosothiols and bioregulatory actions of nitrogen oxides through reactions with thiol groups. Curr Top Microbiol Immunol. 1995;196:19-36.[Medline] [Order article via Infotrieve]

31. Daley WL, Loscalzo J, Vita J, Folts J, Stamler JS. The effects of thiol on the hemodynamic profile of nitroprusside: mechanistic implications in vivo. Circulation. 1992;86(suppl I):I-488. Abstract.

32. Scharfstein JS, Keaney JF, Slivka A, Welch GN, Vita JA, Stamler JS, Loscalzo J. In vitro transfer of nitric oxide between a plasma protein-bound reservoir and low molecular weight thiols. J Clin Invest. 1994;94:1432-1439.

33. Mulsch A, Mordvintcev P, Vanin AF, Busse R. The potent vasodilating and guanylyl cyclase activating dinitrosyl-iron(II) complex is stored in a protein-bound form in vascular tissue and is released by thiols. FEBS Lett. 1993;294:252-256.

34. Vanin AF. Endothelium derived relaxing factor is a nitrosyl iron complex with thiol ligands. FEBS Lett. 1991;289:1-3.[Medline] [Order article via Infotrieve]

35. Lipton SA, Choi YB, Sizheg ZL, Loscalzo J, Singel D, Stamler JS. A redox based mechanism for the neuroprotective and neurodestructive effects of nitric oxide and related nitroso-compounds. Nature. 1993;364:626-632.[Medline] [Order article via Infotrieve]

36. Morris SL, Hansen JN. Inhibition of Bacillus cereus spore outgrown by covalent modifications of a sulfhydryl group by nitrosothiol and iodoacetate. J Bacteriol. 1981;148:465-471.[Abstract/Free Full Text]

37. Malinski T, Taha Z. Nitric oxide release from a single cell measured in situ by a porphyrinic-based microsensor. Nature. 1992;358:676-678.[Medline] [Order article via Infotrieve]

38. Kowaluk EA, Fung HL. Spontaneous liberation of nitric oxide cannot account for in vitro vascular relaxation by S-nitrosothiols. J Pharmacol Exp Ther. 1990;255:1256-1264.[Abstract/Free Full Text]

39. Radomski MW, Rees DD, Durta A, Moncada S. S-nitroso-glutathione inhibits platelet activation in vitro and in vivo. Br J Pharmacol. 1993;107:745-749.[Medline] [Order article via Infotrieve]

40. Simon DI, Stamler JS, Jaraki O, Keaney J, Osborne JA, Francis SA, Single DJ, Loscalzo J. Antiplatelet properties of protein S-nitrosothiols derived relaxing factor. Arterioscler Thromb. 1993;13:791-799.[Abstract/Free Full Text]

41. Askew SC, Butler AR, Flitney FW, Kemp GD, Megson IL. Chemical mechanisms underlying the vasodilator and platelet anti-aggregating properties of S-nitroso-N-acetyl-DL-penicillamine and S-nitrosoglutathione. Bioorg Med Chem. 1995;3:1-9.[Medline] [Order article via Infotrieve]

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