Spontaneous human monocyte apoptosis utilizes a caspase-3-dependent pathway that is blocked by endotoxin and is independent of caspase-1.
Apoptosis is an important mechanism for regulating the numbers of monocytes and macrophages. Caspases (cysteine-aspartate-specific proteases) are key molecules in apoptosis and require proteolytic removal of prodomains for activity. Caspase-1 and caspase-3 have both been connected to apoptosis in other model systems. The present study attempted to delineate what role these caspases play in spontaneous monocyte apoptosis. In serum-free conditions, monocytes showed a commitment to apoptosis as early as 4 h in culture, as evidenced by caspase-3-like activity. Apoptosis, as defined by oligonucleosomal DNA fragmentation, was prevented by a generalized caspase inhibitor, z-VAD-FMK, and the more specific caspase inhibitor, z-DEVD-FMK. The caspase activity was specifically attributable to caspase-3 by the identification of cleavage of procaspase-3 to active forms by immunoblots and by cleavage of the fluorogenic substrate DEVD-AFC. In contrast, a caspase-1 family inhibitor, YVAD-CMK, did not protect monocytes from apoptosis, and the fluorogenic substrate YVAD-AFC failed to show an increase in activity in apoptotic monocytes. When cultured with LPS (1 microgram/ml), monocyte apoptosis was prevented, as was the activation of caspase-3. Unexpectedly, LPS did not change baseline caspase-1 activity. These findings link spontaneous monocyte apoptosis to the proteolytic activation of caspase-3.
Fahy
R J
RJ
Department of Internal Medicine, Division of Pulmonary and Critical Care, The Heart and Lung Institute, Ohio State University, Columbus, OH 43210, USA.
Doseff
A I
AI
Wewers
M D
MD
D017209:Apoptosis / Q000276:immunology*; D020170:Caspase 1 / Q000502:physiology*; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000097:blood / Q000502:physiology*; D002470:Cell Survival / Q000276:immunology; D004789:Enzyme Activation / Q000276:immunology; D006801:Humans; D007375:Interleukin-1 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009000:Monocytes / Q000166:cytology* / Q000201:enzymology* / Q000276:immunology; D011498:Protein Precursors / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology
D061945:Caspase Inhibitors; D007375:Interleukin-1; D008070:Lipopolysaccharides; D011498:Protein Precursors; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1