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Cytochrome P450–generated reactive metabolites cause mitochondrial permeability transition, caspase activation, and apoptosis in rat hepatocytes
Author(s) -
Haouzi Delphine,
Lekéhal Mounia,
Moreau Alain,
Moulis Claude,
Feldmann Gérard,
Robin MarieAnne,
Lettéron Philippe,
Fau Daniel,
Pessayre Dominique
Publication year - 2000
Publication title -
hepatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.488
H-Index - 361
eISSN - 1527-3350
pISSN - 0270-9139
DOI - 10.1053/jhep.2000.9034
Subject(s) - mitochondrial permeability transition pore , cytochrome c , programmed cell death , apoptosis , glutathione , apoptosome , mitochondrion , caspase , biochemistry , microbiology and biotechnology , caspase 3 , biology , chemistry , cytosol , enzyme
Although cytochrome P‐450 (CYP)‐generated reactive metabolites can cause hepatocyte apoptosis, the mechanism of this effect is incompletely understood. In the present study, we assessed the hepatotoxicity of skullcap, a diterpenoid‐containing herbal remedy. Male rat hepatocytes were incubated for 2 hours with skullcap diterpenoids (100 μg/mL). This treatment decreased cell glutathione and protein thiols and increased cell [Ca 2+ ]. This activated Ca 2+ ‐dependent tissue transglutaminase, forming a cross‐linked protein scaffold, and also opened the mitochondrial permeability transition pore, causing outer mitochondrial membrane rupture, increased cytosolic cytochrome c, activation of procaspase 3, internucleosomal DNA fragmentation, and ultrastructural features of apoptosis. Cell death was increased by a CYP3A inducer (dexamethasone) or a sulfur amino acid‐deficient diet increasing glutathione depletion. In contrast, cell death was prevented by decreasing CYP3A activity (with troleandomycin), preventing glutathione depletion (with cysteine or cystine), blocking Ca 2+ ‐modulated events (with calmidazolium), preventing mitochondrial permeability transition (with cyclosporin A), or inhibiting caspase 3 (with acetyl‐Asp‐G u‐Va‐Asp‐a dehyde). Both calmidazolium and cyclosporin A also prevented the increase in cytosolic cytochrome c and procaspase 3 activation. In conclusion, CYP3A activates skullcap diterpenoids into reactive metabolites that deplete cellular thiols and increase cell [Ca 2+ ]. This activates Ca 2+ ‐dependent transglutaminase and also opens the mitochondrial permeability transition pore, causing outer mitochondrial membrane rupture, cytochrome c release, and caspase activation. Preventing mitochondrial permeability transition pore opening and/or caspase activity blocks apoptosis, showing the fundamental role of these final events in metabolite‐mediated hepatotoxicity.