Ischemic preconditioning via epsilon protein kinase C activation requires cyclooxygenase-2 activation in vitro.
The signaling pathway of cyclooxygenase-2 (COX-2) induction following ischemic preconditioning (IPC) in brain remains undefined. To determine role of COX-2 in ischemic preconditioning, we used two in vitro models: mixed cortical neuron/astrocyte cell cultures and organotypic hippocampal slice cultures. We simulated IPC by exposing cell or slice cultures to 1 h or 15 min of oxygen/glucose deprivation (OGD), respectively, 48 h prior to ischemia. To mimic ischemia in vitro, we exposed cell or slice cultures to OGD of 4 h or 40 min, respectively. In cell cultures, these experiments revealed that COX-2 induction peaked at 24 h following IPC in cell culture. Inhibition of COX-2 activation with 50 microM NS-398 (a COX-2 selective inhibitor) abolished IPC-mediated neuroprotection in both in vitro models. Next, we tested whether epsilon protein kinase C (epsilonPKC) and extracellular signal regulated kinase 1/2 (ERK1/2) activation was involved in IPC-mediated neuroprotection and COX-2 expression in cell culture. Cell cultures were treated with an epsilonPKC-specific activating peptide (psiepsilonRACK, 100 nM) for 1 h, and 48 h later were exposed to OGD. epsilonPKC activation increased ERK1/2 phosphorylation and COX-2 induction and conferred neuroprotection similar to IPC. Additionally, inhibition of either epsilonPKC or ERK1/2 activation abolished COX-2 expression and neuroprotection due to ischemic preconditioning. These results demonstrate a crucial role for the epsilonPKC-->ERK1/2-->COX-2 pathway in the induction of neuroprotection via ischemic preconditioning.
Kim
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Cerebral Vascular Disease Research Center, Department of Neurology and Neuroscience Program, University of Miami Miller School of Medicine, Miami, FL 33101, USA.
Raval
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AP
Defazio
R A
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Perez-Pinzon
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MA
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000502:physiology*; D018929:Cell Culture Techniques; D016923:Cell Death; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation; D006624:Hippocampus / Q000098:blood supply / Q000502:physiology*; D019194:Ischemic Preconditioning; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D008297:Male; D016202:N-Methylaspartate / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D009924:Organ Culture Techniques; D051744:Protein Kinase C-epsilon / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015424:Reperfusion
D052246:Cyclooxygenase 2 Inhibitors; D016202:N-Methylaspartate; D007770:L-Lactate Dehydrogenase; D051546:Cyclooxygenase 2; D051744:Protein Kinase C-epsilon