title: Involvement of the mitochondrial death pathway in chemopreventive benzyl isothiocyanate-induced apoptosis.
abstract: In the present study, we studied the molecular mechanism underlying cell death induced by a cancer chemoprotective compound benzyl isothiocyanate (BITC). The cytotoxic effect of BITC was examined in rat liver epithelial RL34 cells. Apoptosis was induced when the cells were treated with 20 mum BITC, characterized by the appearance of phosphatidylserine on the outer surface of the plasma membrane and caspase-3 activation, whereas no caspase activation and propidium iodide incorporation into cell were detected with 50 mum BITC that induced necrosis. The mitochondrial death pathway was suggested to be involved in BITC-induced apoptosis because the treatment of cells with BITC-induced caspase-9-dependent apoptosis and mitochondrial transmembrane potential (Delta Psi m) alteration. We demonstrated here for the first time that BITC directly modifies mitochondrial functions, including inhibition of respiration, mitochondrial swelling, and release of cytochrome c. Moreover, glutathione depletion by diethyl maleate significantly accelerated BITC-triggered apoptosis, suggesting the involvement of a redox-dependent mechanism. This was also implicated by the observations that intracellular accumulation of reactive oxygen species, including superoxide (O(2)) and hydroperoxides (HPOs), was indeed detected in the cells treated with BITC and that the intracellular HPO level was significantly attenuated by pretreatment with N-acetylcysteine. The treatment with a pharmacological scavenger of O(2), Tiron, also diminished the HPO formation by approximately 80%, suggesting that most of the HPOs were H(2)O(2) derived from the dismutation of O(2). These results suggest that BITC induces apoptosis through a mitochondrial redox-sensitive mechanism.
authors.affiliation: Laboratory of Food and Biodynamics, Nagoya University Graduate School of Bioagricultural Sciences, Nagoya 464-8601, Japan. yossan@agr.nagoya-u.ac.jp
authors.forename: Yoshimasa
authors.initials: Y
authors.lastname: Nakamura
authors.forename: Makiko
authors.initials: M
authors.lastname: Kawakami
authors.forename: Akira
authors.initials: A
authors.lastname: Yoshihiro
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Miyoshi
authors.forename: Hajime
authors.initials: H
authors.lastname: Ohigashi
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Kawai
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Osawa
authors.forename: Koji
authors.initials: K
authors.lastname: Uchida
mesh_terms: D000818:Animals; D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D017209:Apoptosis; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003574:Cytochrome c Group / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000187:drug effects; D005434:Flow Cytometry; D017879:Isothiocyanates / Q000494:pharmacology*; D008099:Liver / Q000166:cytology / Q000187:drug effects; D008564:Membrane Potentials; D008928:Mitochondria / Q000378:metabolism / Q000473:pathology*; D009336:Necrosis; D010084:Oxidation-Reduction; D010718:Phosphatidylserines / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D013997:Time Factors
chemical_list: D016588:Anticarcinogenic Agents; D003574:Cytochrome c Group; D017879:Isothiocyanates; D010718:Phosphatidylserines; D017382:Reactive Oxygen Species; C031403:benzyl isothiocyanate; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases