title: Caspase-8 activation precedes alterations of mitochondrial membrane potential during monocyte apoptosis induced by phagocytosis and killing of Staphylococcus aureus.
abstract: Human peripheral blood monocytes become apoptotic following phagocytosis and killing of Staphylococcus aureus. Although this type of monocyte apoptosis is known to be initiated by Fas-Fas ligand (FasL) interactions, the downstream signaling pathway has not been determined. In this work the involvement of mitochondria and the kinetics of caspase-8 and caspase-3 activation after phagocytosis of S. aureus were studied. Caspase-8 activity was measured in cell lysates by using the fluorogenic substrate Ac-IETD-AFC. Active caspase-3 levels and mitochondrial membrane potential (Deltapsi(m)) were measured in whole cells by flow cytometry using monoclonal antibodies reacting with activated caspase-3 and chloromethyl-X-rosamine, respectively. The results show that caspase-8 was activated shortly after phagocytosis of bacteria. Caspase-8 activation was followed by progressive disruption of Deltapsi(m), which is associated with the production of reactive oxygen intermediates. The irreversible caspase-8 inhibitor zIETD-FMK prevented the disruption of Deltapsi(m) and the release of cytochrome c from S. aureus-exposed monocytes. Caspase-3 activation occurred following disruption of Deltapsi(m). These results strongly suggest that apoptosis of monocytes that have phagocytosed and killed S. aureus is driven by the Fas-FasL-initiated pathway, which is typical for type II cells.
authors.affiliation: Department of Immunology, Faculty of Biotechnology, Jagiellonian University, Cracow, Poland.
authors.forename: Kazimierz
authors.initials: K
authors.lastname: Weglarczyk
authors.forename: Jarosław
authors.initials: J
authors.lastname: Baran
authors.forename: Marek
authors.initials: M
authors.lastname: Zembala
authors.forename: Juliusz
authors.initials: J
authors.lastname: Pryjma
mesh_terms: D017209:Apoptosis; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D007113:Immunity, Innate; D066298:In Vitro Techniques; D008564:Membrane Potentials; D008928:Mitochondria / Q000378:metabolism; D009000:Monocytes / Q000276:immunology / Q000382:microbiology / Q000473:pathology / Q000502:physiology*; D010587:Phagocytosis; D017382:Reactive Oxygen Species / Q000378:metabolism; D013211:Staphylococcus aureus / Q000276:immunology / Q000472:pathogenicity*
chemical_list: D017382:Reactive Oxygen Species; C505407:CASP3 protein, human; C505530:CASP8 protein, human; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases