Protective role of nitric oxide synthase against ischemia-reperfusion injury in guinea pig myocardial mitochondria
| Autor: | Hidetsugu Otsuka-Murakami, Wei Liu, Michiko Fujita, Yoshihiro Hotta, Toshiki Masumizu, Michio Yajima, Norio Kawai, Naohisa Ishikawa, Junichi Nakagawa, Masahiro Kohno |
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| Rok vydání: | 1999 |
| Předmět: |
Male
Free Radicals Nitric Oxide Synthase Type III Guinea Pigs Immunoblotting Myocardial Reperfusion Injury Arginine Nitric Oxide Mitochondria Heart Nitric oxide Superoxide dismutase chemistry.chemical_compound medicine Animals Humans Nitric Oxide Donors Enzyme Inhibitors Pharmacology chemistry.chemical_classification Reactive oxygen species Nitrates ATP synthase biology Superoxide Dismutase Myocardium Penicillamine Electron Spin Resonance Spectroscopy Heart Free Radical Scavengers Hydrogen Peroxide Hydrogen-Ion Concentration medicine.disease Immunohistochemistry Molecular biology Nitric oxide synthase NG-Nitroarginine Methyl Ester chemistry Biochemistry biology.protein Calcium Female Nitric Oxide Synthase Reactive Oxygen Species Spin Trapping Reperfusion injury Peroxynitrite |
| Zdroj: | European Journal of Pharmacology. 380:37-48 |
| ISSN: | 0014-2999 |
| DOI: | 10.1016/s0014-2999(99)00531-2 |
| Popis: | In guinea-pig myocardial mitochondria preparation, lowering the Ca2+ concentration or pH level in the perfusate rapidly elevated the fura-2 Ca2+ signal ([Ca2+]m). Pretreatment with 10(-4) M L-Arg inhibited the rapid [Ca2+]m influx, whereas administration of 10(-4) M L-NAME did not, suggesting some association between nitric oxide (NO*) synthase (NOS) activation and Ca2+ kinetics in mitochondria. Immunoblotting analysis showed that endothelial (e)-NOS was present in mitochondria, but not inducible (i)-NOS or brain (b)-NOS. Electron microscopy observations revealed that the e-NOS antibody-reactive site in the mitochondria was the inner cristae. The production of reactive oxygen species and NO* in isolated mitochondria was detected by the spin trapping technique with electron paramagnetic resonance (EPR) spectrometry. Pretreatment with 10(-5) M S-nitroso-N-acetyl-DL-penicillamine (SNAP) and 10(-5) M 3-[2-Hydroxy-1-(1-methylethyl)-2-nitrosohydrazino]-1-propananin e (NOC 5), which spontaneously generate NO*, completely inhibited the [Ca2+]m uptake. In addition, N-morpholino sydnonimine hydrochloride (SIN-1) (10(-5) M), which simultaneously generates NO* as well as *O2- and peroxynitrite anion (ONOO-), inhibited the increase in [Ca2+]m. ONOO- (3 x 10(-4) M) itself also inhibited this increase. Pretreatment with the *O2(-)-scavenger manganese superoxide dismutase or catalase (200 units/ml) completely inhibited the increase in [Ca2+]m caused by lowering of either the Ca2+ concentration or the pH in the perfusate. These results suggested that the formation of reactive oxygen species promoted the [Ca2+]m influx. The agents that inhibited the [Ca2+]m influx improved contractility even in Langendorff preparations after ischemia. Based on these findings, we concluded that e-NOS exists in mitochondria and that NO* may play an important protective role in reperfusion cardiac injury after ischemia, by inhibiting the Ca2+ influx into mitochondria which are otherwise damaged by *O2-. |
| Databáze: | OpenAIRE |
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