Caspase-3 inhibition partially protects oxidant production in apoptotic human neutrophils.
Apoptotic PMN lose functional activity, which emphasizes the tissue injury limiting potential of PMN apoptosis. Caspase-3 activation is the first step in the execution phase of apoptosis. We hypothesized that PMN functional activity, as evidenced by oxidant production, can be restored in apoptotic PMN by inhibition of caspase-3.


METHODS
To accelerate PMN apoptosis, PMN were UV-irradiated for 15 min as previously described. PMN were pretreated with the caspase-3 inhibitor DEVD-fmk (100 microM) for 30 min prior to UV. PMN apoptosis was quantitated by flow cytometry with CD16 staining. Oxidant production in response to 10 microM PMA was quantitated fluorometrically using the method of Hyslop and Sklar. Caspase-3 activity was quantitated fluorometrically using a commercially available assay.


RESULTS
UV-treated PMN demonstrated a 3-fold increase in caspase-3 activity. This was associated with a significant increase in apoptotic PMN and a 10-fold decrease in oxidant production compared to control PMN. DEVD-fmk blocked increases in caspase-3 activity and significantly reduced PMN apoptosis. Oxidant production was increased 5-fold compared to UV-treated PMN but was still significantly less than control PMN.


CONCLUSIONS
In UV-accelerated PMN apoptosis, inhibition of caspase-3 activity partially protects oxidant production in apoptotic PMN. This suggests that signaling events in the initiation phase of PMN apoptosis, which are proximal to caspase-3 activation, may in part be responsible for loss of oxidant production in apoptotic PMN independent of caspase-3 activity.
Sweeney
J F
JF
Department of Surgery, University of Michigan, and Surgery Service, Ann Arbor VA Medical Center, Ann Arbor, Michigan 48109-0331, USA. josw@umich.edu
Nguyen
P K
PK
Hinshaw
D B
DB
D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism / Q000502:physiology* / Q000528:radiation effects; D009842:Oligopeptides / Q000494:pharmacology*; D016877:Oxidants / Q000096:biosynthesis*; D014466:Ultraviolet Rays
D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D009842:Oligopeptides; D016877:Oxidants; C110772:benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases