Spontaneous in contrast to CD95-induced neutrophil apoptosis is independent of caspase activity

Background: The influence of caspase inhibitors on spontaneous and on CD95- triggered apoptosis was investigated in neutrophils from healthy volunteers and compared with neutrophils from patients with severe sepsis, Methods: To further elucidate the mechanisms of neutrophil apoptosis, isola ted neutrophils from healthy volunteers (n = 9) were either stimulated with the agonistic anti-CD95 antibody (100 ng/mL) or left unstimulated in the p resence or absence of the caspase inhibitors zIETD-fmk (10 mu mol/L), zDEVD -fmk (10 mu mol/L), or zVAD-fmk (20 mu mol/L). Apoptosis was determined by measuring DNA fragmentation and Annexin-V binding in FACS, and caspase-3-li ke activity by DEVD-afc cleavage assay. Results were compared with those fr om patients with severe sepsis (n = 15). Results: Reduced spontaneous neutrophil apoptosis in patients with sepsis ( -48.7%) was completely restored by incubation with agonistic anti-CD95 anti body (p < 0.05), Inhibition of caspases did not influence spontaneous neutr ophil apoptosis in both groups. However, zVAD-fmk reduced anti-CD95 antibod y-induced apoptosis in neutrophils from controls by -22.6% (p < 0.05) and i n patients with sepsis by -43.1% (p < 0.05), Conclusion: These results indicate that spontaneous in contrast to CD95-ind uced neutrophil apoptosis is independent of caspase activity.