Effect of CV159-Ca(2+)/calmodulin blockade on redox status hepatic ischemia-reperfusion injury in mice evaluated by a newly developed in vivo EPR imaging technique.
1,4-Dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinedicarboxylic acid methyl 6-(5-phenyl-3-pyrazolyloxy)hexyl ester (CV159) exhibits selective blocking of Ca(2+)/calmodulin and inhibits Ca(2+) overloading in living organisms. The effects of this antagonist in mice with hepatic ischemia-reperfusion injury were investigated using electron paramagnetic resonance imaging (EPRI) and ex vivo EPR (x-band EPR) techniques. The EPRI determined that the 3-carbamoyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl half-life in CV159-treated mice was significantly shorter than that in untreated mice and was almost equal to that in the sham group. Both the cytosolic and the mitochondrial superoxide scavenging activities in CV-treated mice were significantly higher than that in untreated mice. Faint staining of the anti-superoxide dismutase antibody and strong staining of anti-inducible nitric oxide synthase antibody were observed in the liver of control group. In contrast to these findings, immunostaining of these antibodies in the liver of CV159-treated mice were reversed compared to control group. Western blotting showed that CV159 contributed to the high superoxide dismutase expression and low expression of inducible nitric oxide synthase. The alanine aminotransferase level in CV159-treated mice significantly decreased in comparison to that observed in the untreated mice. We conclude that CV159 retains its organ-reducing activity against radicals in hepatic reperfusion injury, which is mediated by the inhibition of Ca(2+) overloading.
Kobayashi
Hiromichi P
HP
Department of Gastroenterological and General Surgery, St. Marianna School of Medicine, Kawasaki, Kanagawa, Japan.
Watanabe
Taiji
T
Oowada
Shigeru
S
Hirayama
Aki
A
Nagase
Sohji
S
Kamibayashi
Masato
M
Otsubo
Takehito
T
D000410:Alanine Transaminase / Q000097:blood; D000818:Animals; D002118:Calcium / Q000037:antagonists & inhibitors*; D002147:Calmodulin / Q000037:antagonists & inhibitors*; D004095:Dihydropyridines / Q000494:pharmacology*; D004578:Electron Spin Resonance Spectroscopy; D007150:Immunohistochemistry; D008099:Liver / Q000098:blood supply / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D052247:Nitric Oxide Synthase Type II / Q000032:analysis; D010084:Oxidation-Reduction; D015427:Reperfusion Injury / Q000378:metabolism* / Q000473:pathology; D013482:Superoxide Dismutase / Q000032:analysis; D000072105:Superoxide Dismutase-1
D002147:Calmodulin; D004095:Dihydropyridines; C055634:CV 159; D052247:Nitric Oxide Synthase Type II; C000606291:Sod1 protein, mouse; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D000410:Alanine Transaminase; D002118:Calcium