title: Cadmium induces caspase-independent apoptosis in liver Hep3B cells: role for calcium in signaling oxidative stress-related impairment of mitochondria and relocation of endonuclease G and apoptosis-inducing factor.
abstract: Cadmium-induced cellular toxicity has been related to necrosis and/or caspase-dependent apoptosis. In the present study, we show that, on cadmium exposure, the human hepatocarcinoma Hep3B cells undergo caspase-independent apoptosis associated with nuclear translocation of endonuclease G and apoptosis-inducing factor, two mitochondrial apoptogenic proteins. Release of these proteins is likely related to calcium-induced alteration of mitochondrial homeostasis. Indeed, it was first preceded by a rapid and sustained increase in cytoplasmic calcium and then by a coincident loss in mitochondrial membrane potential and production of reactive oxygen species. Bapta-AM (acetoxymethyl ester of 5, 5'-dimethyl-bis (o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid), a calcium chelator, blocked all these events and prevented cadmium-induced apoptosis. Production of reactive oxygen species was inhibited by ruthenium red and rotenone, two mitochondrial inhibitors, and by diphenyleneiodonium, a flavoprotein inhibitor, which also prevented both loss in mitochondrial membrane potential and apoptosis. In addition, Bapta-AM and diphenyleneiodonium were found to almost totally block decreased expression of the mitochondrial anti-apoptotic nuclear factor-kappaB-regulated bcl-x(L) protein in cadmium-treated cells. Taken together, our results show that cadmium induces Hep3B cells apoptosis mainly by calcium- and oxidative stress-related impairment of mitochondria, which probably favors release of apoptosis-inducing factor and endonuclease G.
authors.affiliation: INSERM U620, Détoxication et Réparation Tissulaire, Université de Rennes 1, 2 avenue du Pr. Léon Bernard, 35043 Rennes, France.
authors.forename: Anthony
authors.initials: A
authors.lastname: Lemarié
authors.forename: Dominique
authors.initials: D
authors.lastname: Lagadic-Gossmann
authors.forename: Claudie
authors.initials: C
authors.lastname: Morzadec
authors.forename: Nathalie
authors.initials: N
authors.lastname: Allain
authors.forename: Olivier
authors.initials: O
authors.lastname: Fardel
authors.forename: Laurent
authors.initials: L
authors.lastname: Vernhet
mesh_terms: D021581:Active Transport, Cell Nucleus; D017209:Apoptosis; D015152:Blotting, Northern; D015153:Blotting, Western; D002104:Cadmium / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004706:Endodeoxyribonucleases / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000166:cytology*; D008564:Membrane Potentials; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D009336:Necrosis; D009861:Onium Compounds / Q000494:pharmacology; D018384:Oxidative Stress; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012313:RNA / Q000737:chemistry; D017382:Reactive Oxygen Species / Q000378:metabolism; D012402:Rotenone / Q000494:pharmacology; D012430:Ruthenium Red / Q000494:pharmacology; D013997:Time Factors; D014475:Uncoupling Agents / Q000494:pharmacology; D051020:bcl-X Protein
chemical_list: C494811:BCL2L1 protein, human; D002614:Chelating Agents; D016328:NF-kappa B; D009861:Onium Compounds; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D014475:Uncoupling Agents; D051020:bcl-X Protein; D002104:Cadmium; D012402:Rotenone; D012430:Ruthenium Red; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D012313:RNA; C007517:diphenyleneiodonium; D004247:DNA; D004706:Endodeoxyribonucleases; C052391:endonuclease G; D020169:Caspases; D002118:Calcium