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 ac tivity. Caspase-1 and caspase-3 have both been connected to apoptosis in ot her 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 oligonucleos omal DNA fragmentation,,vas 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 o f cleavage of procaspase-3 to active forms by immunoblots and by cleavage o f the fluorogenic substrate DEVD-AFC, In contrast, a caspase-1 family inhib itor, YVAD-CMK, did not protect monocytes from apoptosis, and the fluorogen ic substrate YVAD-AFC failed to show an increase in activity in apoptotic m onocytes, When cultured with LPS (1 mu g/ml), monocyte apoptosis was preven ted, as was the activation of caspase-3, Unexpectedly, LPS did not change b aseline caspase-1 activity. These findings link spontaneous monocyte apopto sis to the proteolytic activation of caspase-3.