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Nitrite augments tolerance to ischemia/reperfusion injury via the modulation of mitochondrial electron transfer.

Shiva S, Sack MN, Greer JJ, Duranski M, Ringwood LA, Burwell L, Wang X, MacArthur PH, Shoja A, Raghavachari N, Calvert JW, Brookes PS, Lefer DJ, Gladwin MT - J. Exp. Med. (2007)

Bottom Line: Although the mechanism of nitrite-mediated cytoprotection is unknown, NO is a mediator of the ischemic preconditioning cell-survival program.This cytoprotection is associated with increases in mitochondrial oxidative phosphorylation.These data suggest that nitrite dynamically modulates mitochondrial resilience to reperfusion injury and may represent an effector of the cell-survival program of ischemic preconditioning and the Mediterranean diet.

View Article: PubMed Central - PubMed

Affiliation: Vascular Medicine Branch, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
Nitrite (NO(2)(-)) is an intrinsic signaling molecule that is reduced to NO during ischemia and limits apoptosis and cytotoxicity at reperfusion in the mammalian heart, liver, and brain. Although the mechanism of nitrite-mediated cytoprotection is unknown, NO is a mediator of the ischemic preconditioning cell-survival program. Analogous to the temporally distinct acute and delayed ischemic preconditioning cytoprotective phenotypes, we report that both acute and delayed (24 h before ischemia) exposure to physiological concentrations of nitrite, given both systemically or orally, potently limits cardiac and hepatic reperfusion injury. This cytoprotection is associated with increases in mitochondrial oxidative phosphorylation. Remarkably, isolated mitochondria subjected to 30 min of anoxia followed by reoxygenation were directly protected by nitrite administered both in vitro during anoxia or in vivo 24 h before mitochondrial isolation. Mechanistically, nitrite dose-dependently modifies and inhibits complex I by posttranslational S-nitrosation; this dampens electron transfer and effectively reduces reperfusion reactive oxygen species generation and ameliorates oxidative inactivation of complexes II-IV and aconitase, thus preventing mitochondrial permeability transition pore opening and cytochrome c release. These data suggest that nitrite dynamically modulates mitochondrial resilience to reperfusion injury and may represent an effector of the cell-survival program of ischemic preconditioning and the Mediterranean diet.

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Acute nitrite treatment protects mitochondria against I/R injury. (A) Model of in vitro mitochondrial damage showing preanoxic (green) and postanoxic rates in the absence (blue) and presence (red) of nitrite. Arrow denotes the addition of 10 μM nitrite. Respiration rates (B) and ATP generation rates (C) before anoxia (green) and after anoxia in the presence of saline (blue) and nitrite (red). (D) Quantification of rate of ATP generation before and after anoxia. (E) Recovery of respiration rate with increasing concentrations (0–100 μM) of nitrite added during anoxia. All experiments are means ± SEM of at least n = 3 independent experiments. *, P < 0.01. RLU, relative light units.
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fig4: Acute nitrite treatment protects mitochondria against I/R injury. (A) Model of in vitro mitochondrial damage showing preanoxic (green) and postanoxic rates in the absence (blue) and presence (red) of nitrite. Arrow denotes the addition of 10 μM nitrite. Respiration rates (B) and ATP generation rates (C) before anoxia (green) and after anoxia in the presence of saline (blue) and nitrite (red). (D) Quantification of rate of ATP generation before and after anoxia. (E) Recovery of respiration rate with increasing concentrations (0–100 μM) of nitrite added during anoxia. All experiments are means ± SEM of at least n = 3 independent experiments. *, P < 0.01. RLU, relative light units.

Mentions: The lack of evidence for nitrite-dependent regulation of mitochondrial protein expression suggested that the protective effect of nitrite on mitochondria was most likely caused by the posttranslational modification of existing mitochondrial proteins. We hypothesized that if posttranslational modification was indeed responsible, mitochondrial protection of respiration and ATP synthesis should be observed even with acute nitrite treatment of mitochondria in vitro. To test this, we subjected isolated mitochondria from untreated rats to 30 min of anoxia followed by reoxygenation in the presence and absence of nitrite (Fig. 4 A). As predicted, nitrite-treated (10 μM) mitochondria had both higher complex II–dependent state 3 respiration (78 ± 12 nmol O2/min/mg; Fig. 4, A and B) and ATP production (16 ± 6 relative light units/s; Fig. 4, C and D) rates than control mitochondria (54 ± 4 nmol O2/min/mg and 9.8 relative light units/s, respectively) after anoxia/reoxygenation. Interestingly, nitrite-dependent protection from the anoxia-induced drop in respiration rate showed a biphasic dose–response curve (0–100 μM), with peak protection occurring at a concentration of 12.5 μM (Fig. 4 E). This biphasic response is identical in shape and concentration to previously observed dose–response curves for nitrite-mediated cytoprotection in vivo, and the concentration of 12.5 μM nitrite conferring peak protection to the mitochondria parallels the tissue concentration of nitrite shown to orchestrate peak protection in vivo (8).


Nitrite augments tolerance to ischemia/reperfusion injury via the modulation of mitochondrial electron transfer.

Shiva S, Sack MN, Greer JJ, Duranski M, Ringwood LA, Burwell L, Wang X, MacArthur PH, Shoja A, Raghavachari N, Calvert JW, Brookes PS, Lefer DJ, Gladwin MT - J. Exp. Med. (2007)

Acute nitrite treatment protects mitochondria against I/R injury. (A) Model of in vitro mitochondrial damage showing preanoxic (green) and postanoxic rates in the absence (blue) and presence (red) of nitrite. Arrow denotes the addition of 10 μM nitrite. Respiration rates (B) and ATP generation rates (C) before anoxia (green) and after anoxia in the presence of saline (blue) and nitrite (red). (D) Quantification of rate of ATP generation before and after anoxia. (E) Recovery of respiration rate with increasing concentrations (0–100 μM) of nitrite added during anoxia. All experiments are means ± SEM of at least n = 3 independent experiments. *, P < 0.01. RLU, relative light units.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2118713&req=5

fig4: Acute nitrite treatment protects mitochondria against I/R injury. (A) Model of in vitro mitochondrial damage showing preanoxic (green) and postanoxic rates in the absence (blue) and presence (red) of nitrite. Arrow denotes the addition of 10 μM nitrite. Respiration rates (B) and ATP generation rates (C) before anoxia (green) and after anoxia in the presence of saline (blue) and nitrite (red). (D) Quantification of rate of ATP generation before and after anoxia. (E) Recovery of respiration rate with increasing concentrations (0–100 μM) of nitrite added during anoxia. All experiments are means ± SEM of at least n = 3 independent experiments. *, P < 0.01. RLU, relative light units.
Mentions: The lack of evidence for nitrite-dependent regulation of mitochondrial protein expression suggested that the protective effect of nitrite on mitochondria was most likely caused by the posttranslational modification of existing mitochondrial proteins. We hypothesized that if posttranslational modification was indeed responsible, mitochondrial protection of respiration and ATP synthesis should be observed even with acute nitrite treatment of mitochondria in vitro. To test this, we subjected isolated mitochondria from untreated rats to 30 min of anoxia followed by reoxygenation in the presence and absence of nitrite (Fig. 4 A). As predicted, nitrite-treated (10 μM) mitochondria had both higher complex II–dependent state 3 respiration (78 ± 12 nmol O2/min/mg; Fig. 4, A and B) and ATP production (16 ± 6 relative light units/s; Fig. 4, C and D) rates than control mitochondria (54 ± 4 nmol O2/min/mg and 9.8 relative light units/s, respectively) after anoxia/reoxygenation. Interestingly, nitrite-dependent protection from the anoxia-induced drop in respiration rate showed a biphasic dose–response curve (0–100 μM), with peak protection occurring at a concentration of 12.5 μM (Fig. 4 E). This biphasic response is identical in shape and concentration to previously observed dose–response curves for nitrite-mediated cytoprotection in vivo, and the concentration of 12.5 μM nitrite conferring peak protection to the mitochondria parallels the tissue concentration of nitrite shown to orchestrate peak protection in vivo (8).

Bottom Line: Although the mechanism of nitrite-mediated cytoprotection is unknown, NO is a mediator of the ischemic preconditioning cell-survival program.This cytoprotection is associated with increases in mitochondrial oxidative phosphorylation.These data suggest that nitrite dynamically modulates mitochondrial resilience to reperfusion injury and may represent an effector of the cell-survival program of ischemic preconditioning and the Mediterranean diet.

View Article: PubMed Central - PubMed

Affiliation: Vascular Medicine Branch, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
Nitrite (NO(2)(-)) is an intrinsic signaling molecule that is reduced to NO during ischemia and limits apoptosis and cytotoxicity at reperfusion in the mammalian heart, liver, and brain. Although the mechanism of nitrite-mediated cytoprotection is unknown, NO is a mediator of the ischemic preconditioning cell-survival program. Analogous to the temporally distinct acute and delayed ischemic preconditioning cytoprotective phenotypes, we report that both acute and delayed (24 h before ischemia) exposure to physiological concentrations of nitrite, given both systemically or orally, potently limits cardiac and hepatic reperfusion injury. This cytoprotection is associated with increases in mitochondrial oxidative phosphorylation. Remarkably, isolated mitochondria subjected to 30 min of anoxia followed by reoxygenation were directly protected by nitrite administered both in vitro during anoxia or in vivo 24 h before mitochondrial isolation. Mechanistically, nitrite dose-dependently modifies and inhibits complex I by posttranslational S-nitrosation; this dampens electron transfer and effectively reduces reperfusion reactive oxygen species generation and ameliorates oxidative inactivation of complexes II-IV and aconitase, thus preventing mitochondrial permeability transition pore opening and cytochrome c release. These data suggest that nitrite dynamically modulates mitochondrial resilience to reperfusion injury and may represent an effector of the cell-survival program of ischemic preconditioning and the Mediterranean diet.

Show MeSH
Related in: MedlinePlus