Nature of the cardiomyocyte injury induced by lipid hydroperoxides.
OBJECTIVE
As a result of oxidative stress to membrane lipid matrix, the peroxidation of polyunsaturated fatty acids induced the transient formation of lipid hydroperoxides (ROOH). The aim of this study was to evaluate the damaging effects of ROOH on the cardiac cell and the link between the alterations observed and intracellular calcium overload.


METHODS
Necrosis of cultured rat cardiac cells was determined by measuring the release of lactate dehydrogenase (LDH). In guinea-pig papillary muscles, action potential (AP) and isometric tension were recorded with standard microelectrodes and a transducer, respectively. The reactive oxygen species (ROS) scavenging properties of tested compounds were determined using a cell-free model of lipid photoperoxidation.


RESULTS
15(S)-HpETE (15(S)-hydroperoxyeicosatetraenoic acid), an arachidonic acid hydroperoxide, induced a concentration-dependent loss of cardiomyocytes membrane integrity. The release of LDH induced by 15(s)-HpETE (30 microM) was prevented by a ROS scavenger, BW755C (10 microM), but not by a sarcolemmal calcium channel blocker, Amlodipine (10 microM), or a calcium overload protective agent, R56865 (10 microM). Cardiomyocytes necrosis induced by calcium paradox was prevented by Amlodipine (10 microM) and R56865 (10 microM), but not by BW755C (10 microM). Superfusion of papillary muscles with 15(S)-HpETE (20 microM) induced a membrane depolarization and a marked reduction in the AP amplitude and duration. Concomitantly, a transient positive inotropic effect and a progressive rise in diastolic tension were observed. These alterations were maximal after 15 min and associated with delayed after-depolarizations (DADs) and after-contractions. Every alteration was inhibited by BW755C (10 microM) and R56865 (30 microM), but not by Amlodipine (1 microM). Ryanodine (1 microM), a blocker of sarcoplasmic reticulum calcium channel, only prevented the appearance of DADs and after-contractions. Only BW755C exhibited ROS scavenging properties.


CONCLUSIONS
ROOH induced enzyme leakage and electromechanical alterations in cardiac cells. These effects of ROOH implicated oxidative mechanisms and resulted in an intracellular calcium overload.
Thollon
C
C
Division Pathologies Cardiaques et Vasculaires, Institut de Recherches Servier, Suresnes, France.
Iliou
J P
JP
Cambarrat
C
C
Robin
F
F
Vilaine
J P
JP
D015772:4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine / Q000494:pharmacology; D000200:Action Potentials / Q000187:drug effects; D017311:Amlodipine / Q000494:pharmacology; D000818:Animals; D052160:Benzothiazoles; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D016166:Free Radical Scavengers / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism*; D006168:Guinea Pigs; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D015289:Leukotrienes / Q000494:pharmacology*; D008054:Lipid Peroxides / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D009336:Necrosis; D010880:Piperidines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D016897:Respiratory Burst; D012433:Ryanodine / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D014662:Vasoconstrictor Agents / Q000494:pharmacology*
D052160:Benzothiazoles; D002121:Calcium Channel Blockers; D016166:Free Radical Scavengers; D005609:Free Radicals; D015289:Leukotrienes; D008054:Lipid Peroxides; D010880:Piperidines; D013844:Thiazoles; D014662:Vasoconstrictor Agents; D012433:Ryanodine; D017311:Amlodipine; D015772:4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; C025086:15-hydroperoxy-5,8,11,13-eicosatetraenoic acid; C054551:R 56865; D007770:L-Lactate Dehydrogenase; D002118:Calcium