title: Role of calcium-induced mitochondrial hydroperoxide in induction of apoptosis of RBL2H3 cells with eicosapentaenoic acid treatment.
abstract: Eicosapentaenoic acid (EPA) was previously shown to induce caspase-independent apoptosis in rat basophilic leukemia cells (RBL2H3 cells) by translocation of apoptosis-inducing factor (AIF) [Free Radic Res (2005) 39, 225-235]. Here, we attempted to investigate the mechanism of EPA-induced apoptosis. A rapid and sustained increase in calcium was observed in mitochondria at 2 h after the addition of EPA prior to apoptosis. Coincidently, hydroperoxide was generated in the mitochondria after exposure to EPA. Production of mitochondrial hydroperoxide was significantly reduced by ruthenium red, an inhibitor of mitochondrial calcium uniporter, and BAPTA-AM, a cytoplasmic calcium chelator, indicating that generation of hydroperoxide is triggered by an accumulation of calcium in the mitochondria. The production of mitochondrial hydroperoxide was markedly attenuated by overexpression of phospholipid hydroperoxide glutathione peroxidase (PHGPx) in the mitochondria. Apoptosis was therefore, significantly prevented through inhibition of mitochondrial hydroperoxide generation with mitochondrial PHGPx, ruthenium red or BAPTA-AM. However, accumulation of calcium in the mitochondria was not prevented by mitochondrial PHGPx although apoptosis was blocked, indicating that elevated calcium does not directly induce apoptosis. Taken together, our results show that calcium-dependent hydroperoxide accumulation in the mitochondria is critical in EPA-induced apoptosis.
authors.affiliation: Kitasato University, School of Pharmaceutical Sciences, 5-9-1 Shirokane, Minato-ku, Tokyo, 108-8641, Japan.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Koumura
authors.forename: Chika
authors.initials: C
authors.lastname: Nakamura
authors.forename: Yasuhito
authors.initials: Y
authors.lastname: Nakagawa
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D045744:Cell Line, Tumor; D045304:Cytochromes c / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D015118:Eicosapentaenoic Acid / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic; D005979:Glutathione Peroxidase / Q000235:genetics / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D051381:Rats; D012430:Ruthenium Red / Q000494:pharmacology
chemical_list: D012430:Ruthenium Red; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D045304:Cytochromes c; D015118:Eicosapentaenoic Acid; D006861:Hydrogen Peroxide; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D005979:Glutathione Peroxidase; D002118:Calcium