Critical role for caspases 3 and 8 in neutrophil but not eosinophil apoptosis.
BACKGROUND
Apoptosis is a necessary process to control cell numbers in multicellular organisms. In many chronic inflammatory diseases, reduced cell death of different types of granulocytes is one important mechanism for cell accumulation. Here, we studied the role of caspases in neutrophil and eosinophil apoptosis in the presence or absence of granulocyte-macrophage stimulating factor and anti-CD95 monoclonal antibodies, respectively.


METHODS
Granulocytes were isolated from human blood using standard protocols. Immunoblot and functional studies with cell-permeable specific peptide inhibitors were performed to analyze caspase involvement. Fas receptor and Fas ligand expression was analyzed by RT-PCR, flow cytometry, and immunoblotting. Cell death was analyzed by ethidium bromide exclusion test.


RESULTS
Caspases 3 and 8 are critically involved in the regulation of neutrophil apoptosis in vitro. In contrast, these two caspases did not appear to play a major role in the regulation of eosinophil apoptosis. However, the broad-range caspase inhibitor VAD prevented eosinophil death, indicating that caspases are also involved within the apoptotic machinery of eosinophils. Functional inhibitor studies suggested that caspase 9 is crucial for both caspase 3 and 8 activation, at least in neutrophils. In contrast, spontaneous apoptosis of neutrophils or eosinophils is unlikely to be the consequence of Fas ligand/Fas receptor molecular interactions.


CONCLUSIONS
The data of this study indicate differences in the usage of caspases between neutrophils and eosinophils.
Daigle
I
I
Swiss Institute of Allergy and Asthma Research, Davos, Switzerland.
Simon
H U
HU
D000911:Antibodies, Monoclonal / Q000276:immunology; D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002478:Cells, Cultured; D004789:Enzyme Activation; D004804:Eosinophils / Q000201:enzymology / Q000502:physiology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D018375:Neutrophil Activation; D009504:Neutrophils / Q000201:enzymology / Q000502:physiology*; D019014:fas Receptor / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D061945:Caspase Inhibitors; D019014:fas Receptor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases