Altering CO2 during reperfusion of ischemic cardiomyocytes modifies mitochondrial oxidant injury.
OBJECTIVE
Acute changes in tissue CO2 and pH during reperfusion of the ischemic heart may affect ischemia/reperfusion injury. We tested whether gradual vs. acute decreases in CO2 after cardiomyocyte ischemia affect reperfusion oxidants and injury.


METHODS
Comparative laboratory investigation.


METHODS
Institutional laboratory.


METHODS
Embryonic chick cardiomyocytes.


METHODS
Microscope fields of approximately 500 chick cardiomyocytes were monitored throughout 1 hr of simulated ischemia (PO2 of 3-5 torr, PCO2 of 144 torr, pH 6.8), followed by 3 hrs of reperfusion (PO2 of 149 torr, PCO2 of 36 torr, pH 7.4), and compared with cells reperfused with relative hypercarbia (PCO2 of 71 torr, pH 6.8) or hypocarbia (PCO2 of 7 torr, pH 7.9).


RESULTS
The measured outcomes included cell viability (via propidium iodide) and oxidant generation (reactive oxygen species via 2',7'-dichlorofluorescin oxidation and nitric oxide [NO] via 4,5-diaminofluorescein diacetate oxidation). Compared with normocarbic reperfusion, hypercarbia significantly reduced cell death from 54.8% +/- 4.0% to 26.3% +/- 2.8% (p < .001), significantly decreased reperfusion reactive oxygen species (p < .05), and increased NO at a later phase of reperfusion (p < .01). The NO synthase inhibitor N-nitro-L-arginine methyl ester (200 microM) reversed this oxidant attenuation (p < .05), NO increase (p < .05), and the cardioprotection conferred by hypercarbic reperfusion (increasing death to 54.3% +/- 6.0% [p < .05]). Conversely, hypocarbic reperfusion increased cell death to 80.4% +/- 4.5% (p < .01). It also increased reactive oxygen species by almost two-fold (p = .052), without affecting the NO level thereafter. Increased reactive oxygen species was attenuated by the mitochondrial complex III inhibitor stigmatellin (20 nM) when given at reperfusion (p < .05). Cell death also decreased from 85.9% +/- 4.5% to 52.2% +/- 6.5% (p < .01). The nicotinamide adenine dinucleotide phosphate oxidase inhibitor apocynin (300 microM) had no effect on reperfusion reactive oxygen species.


CONCLUSIONS
Altering CO2 content during reperfusion can significantly affect myocardial postresuscitation injury, in part by modifying mitochondrial oxidants and NO synthase-induced NO production.
Lavani
Romeen
R
Emergency Resuscitation Center, Sections of Emergency Medicine and Pulmonary/Critical Care, Department of Medicine, University of Chicago, Chicago, IL, USA.
Chang
Wei-Tien
WT
Anderson
Travis
T
Shao
Zuo-Hui
ZH
Wojcik
Kimberly R
KR
Li
Chang-Qing
CQ
Pietrowski
Robert
R
Beiser
David G
DG
Idris
Ahamed H
AH
Hamann
Kimm J
KJ
Becker
Lance B
LB
Vanden Hoek
Terry L
TL
D000818:Animals; D002245:Carbon Dioxide / Q000097:blood*; D002470:Cell Survival; D002478:Cells, Cultured; D002642:Chick Embryo; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D006935:Hypercapnia / Q000378:metabolism / Q000503:physiopathology; D016857:Hypocapnia / Q000378:metabolism / Q000503:physiopathology; D008928:Mitochondria / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D016877:Oxidants / Q000378:metabolism*; D010313:Partial Pressure; D011090:Polyenes / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D015427:Reperfusion Injury / Q000378:metabolism* / Q000473:pathology / Q000517:prevention & control
D004791:Enzyme Inhibitors; D016877:Oxidants; D011090:Polyenes; D017382:Reactive Oxygen Species; D002245:Carbon Dioxide; D009569:Nitric Oxide; C041573:stigmatellin; D019331:NG-Nitroarginine Methyl Ester