Caspase-8 activation precedes alterations of mitochondrial membrane potential during monocyte apoptosis induced by phagocytosis and killing of Staphylococcus aureus.
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.
Weglarczyk
Kazimierz
K
Department of Immunology, Faculty of Biotechnology, Jagiellonian University, Cracow, Poland.
Baran
Jarosław
J
Zembala
Marek
M
Pryjma
Juliusz
J
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*
D017382:Reactive Oxygen Species; C505407:CASP3 protein, human; C505530:CASP8 protein, human; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases