A transient, sublethal ischemic interval confers resistance to a subsequent, otherwise lethal ischemic insult, in a process termed ischemic preconditioning.
Poly(ADP-ribose) polymerase-1 (PARP-1) normally functions in DNA repair, but extensive PARP-1 activation is a major cause of ischemic cell death. Because PARP-1 can be cleaved and inactivated by
caspases, we investigated the possibility that
caspase cleavage of PARP-1 could contribute to ischemic preconditioning. Murine cortical cultures were treated with
glucose deprivation combined with 0.5 mm
2-deoxyglucose and 5 mm
azide ("chemical
ischemia") to model the reversible energy failure that occurs during transient
ischemia in vivo. Cortical cultures preconditioned with 15 min of chemical
ischemia showed increased resistance to subsequent, longer periods of chemical
ischemia. These cultures were also more resistant to the PARP-1 activating agent,
N-methyl-N'-nitro-N-nitrosoguanidine, suggesting reduced capacity for PARP-1 activation after preconditioning. Immunostaining for the 89 kDa PARP-1 cleavage fragment and for
poly(ADP-ribose) formation confirmed that PARP-1 was cleaved and PARP-1 activity was attenuated in the preconditioned neurons. Preconditioning also produced an increase in activated
caspase-3 peptide and an increase in
caspase-3 activity in the cortical cultures. A cause-effect relationship between
caspase activation, PARP-1 cleavage, and ischemic preconditioning was supported by studies using the
caspase inhibitor Ac-
Asp-Glu-Val-Asp-
aldehyde (
DEVD-CHO). Cultures treated with
DEVD-CHO after preconditioning showed reduced PARP-1 cleavage and reduced resistance to subsequent
ischemia. These findings suggest a novel interaction between the
caspase- and PARP-1-mediated cell death pathways in which sublethal
caspase activation leads to PARP-1 cleavage, thereby increasing resistance to subsequent ischemic stress.