S-Nitrosothiols signal hypoxia-mimetic vascular pathology
Lisa A. Palmer, … , Timothy Macdonald, Benjamin Gaston
Lisa A. Palmer, … , Timothy Macdonald, Benjamin Gaston
Published September 4, 2007
Citation Information: J Clin Invest. 2007;117(9):2592-2601. https://doi.org/10.1172/JCI29444.
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S-Nitrosothiols signal hypoxia-mimetic vascular pathology

Abstract

NO transfer reactions between protein and peptide cysteines have been proposed to represent regulated signaling processes. We used the pharmaceutical antioxidant N-acetylcysteine (NAC) as a bait reactant to measure NO transfer reactions in blood and to study the vascular effects of these reactions in vivo. NAC was converted to S-nitroso-N-acetylcysteine (SNOAC), decreasing erythrocytic S-nitrosothiol content, both during whole-blood deoxygenation ex vivo and during a 3-week protocol in which mice received high-dose NAC in vivo. Strikingly, the NAC-treated mice developed pulmonary arterial hypertension (PAH) that mimicked the effects of chronic hypoxia. Moreover, systemic SNOAC administration recapitulated effects of both NAC and hypoxia. eNOS-deficient mice were protected from the effects of NAC but not SNOAC, suggesting that conversion of NAC to SNOAC was necessary for the development of PAH. These data reveal an unanticipated adverse effect of chronic NAC administration and introduce a new animal model of PAH. Moreover, evidence that conversion of NAC to SNOAC during blood deoxygenation is necessary for the development of PAH in this model challenges conventional views of oxygen sensing and of NO signaling.

Authors

Lisa A. Palmer, Allan Doctor, Preeti Chhabra, Mary Lynn Sheram, Victor E. Laubach, Molly Z. Karlinsey, Michael S. Forbes, Timothy Macdonald, Benjamin Gaston

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Figure 3

SNOAC is formed from NAC in blood ex vivo and in vivo.

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SNOAC is formed from NAC in blood ex vivo and in vivo. (A) The SNOrbc in...
(A) The SNOrbc in heparinized LV blood (black bars), measured by reductive chemiluminescence (11), was lower than normal following 3 weeks of treatment with 10 mg/ml NAC (n = 3–4 each). In the same mice, plasma SNOAC levels (gray bars; measured by MS) increased from undetectable to approximately 2 μM over the same time (*P < 0.05). (B) Serum SNOAC, measured by MS, formed in NAC-treated mice (3 weeks). Left: liquid chromatogram; right: MS spectrum. NAC-treated mice had a SNOAC peak (m/z 193; red) coeluting with 15N-labeled SNOAC standard (m/z 194; black) that was absent in untreated animals (green) and was not detected in NAC-treated mice after serum pretreatment with HgCl2 to displace NO+ from the thiolate (blue). (C) Oxygenated erythrocytes were deoxygenated ex vivo (argon; ref. 11) in the presence of 100 μM NAC; supernatant SNOAC was measured by MS (above). SNOAC concentration increased with oxyhemoglobin (Oxy Hb) desaturation (co-oximetry: inset), being maximal at 59.3% saturation (blue), less at 77.2% saturation (green), and undetectable at 98% saturation. (D) SNOAC (filled circles) accumulated as the concentration of S-nitrosothiol–modified Hb (SNOHb; open circles) and oxyhemoglobin saturation (Hb SO2; blue line) both decreased in heparinized whole blood using argon with 5% CO2 (pH 7.3) in a tonometer. Both the increase in SNOAC and the loss of SNOrbc between 0 and 20 minutes were significant (P < 0.01 by ANOVA followed by pairwise comparison to the maximum value; n = 3). #SNOAC levels were below the limit of detection when the oxyhemoglobin saturation was greater than 80%.