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Department of Medicine, Emergency Resuscitation Center, University of Chicago, Chicago, Illinois 60637
Reactive oxygen species (ROS) and nitric
oxide (NO) are implicated in induction of ischemic
preconditioning. However, the relationship between these
oxidant signals and opening of the mitochondrial ATP-dependent
potassium (KATP) channel during early preconditioning is
not fully understood. We observed preconditioning protection by
hypoxia, exogenous H2O2, or PKC activator PMA
in cardiomyocytes subjected to 1-h ischemia and 3-h
reperfusion. Protection was abolished by KATP channel
blocker 5-hydroxydecanoate (5-HD) in each case, indicating that these
triggers must act upstream from the KATP channel.
Inhibitors of NO synthase abolished protection in preconditioned cells,
suggesting that NO is also required for protection. DAF-2 fluorescence
(NO sensitive) increased during hypoxic triggering. This was amplified
by pinacidil and inhibited by 5-HD, indicating that NO is generated
subsequent to KATP channel activation. Exogenous NO during
the triggering phase conferred protection blocked by 5-HD. Exogenous NO
also restored protection abolished by 5-HD or
N
-nitro-L-arginine methyl ester
in preconditioned cells. Antioxidants given during pinacidil or NO
triggering abolished protection, confirming that ROS are generated by
KATP channel activation. Coadministration of
H2O2 and NO restored PMA-induced protection in
5-HD-treated cells, indicating that ROS and NO are required downstream
from the KATP channel. We conclude that ROS can trigger preconditioning by causing activation of the KATP channel,
which then induces generation of ROS and NO that are both required for preconditioning protection.
hydrogen peroxide; nitric oxide; ischemia; cardiomyocytes
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