sl-327 and Brain-Ischemia

MEK1/2 is a serine/threonine protein kinase that phosphorylates and activates extracellular signal-responsive kinase (ERK)1/2. In the present study we explored the role of MEK1/2 in ischemic brain injury using a selective MEK1/2 inhibitor, SL327, in mice. C57BL/6 mice were subjected to a 30-min occlusion of the middle cerebral artery (MCAO) followed by reperfusion. Western blot analysis demonstrated the immediate activation of MEK/ERK after reperfusion (within the first 10 min) in the ischemic brain; this activation was dose dependently blocked by SL327 (10-100 mg/kg, i.p.). A single dose of SL327 (100 mg/kg) administered 15 min before or 25 min after the onset of ischemia resulted in 63.6% (n = 18, p < 0.001) and 50.7% (n = 18, p < 0.01) reduction in infarct size, respectively, compared with vehicle-treated mice. Similarly, SL327 significantly reduced neurological deficits 1 to 3 days after reperfusion (n = 12, p < 0.01). The salutary effect of SL327-induced neuroprotection was independent of mitochondrial cytochrome c release or caspase-8-mediated apoptosis; however, SL327 markedly suppressed the levels of active caspase-3 and DNA fragmentation (as a measure of apoptosis) after ischemia/reperfusion. Our data suggest that the inhibition of MEK1/2 results in neuroprotection from reperfusion injury and that this protection may be associated with the reduction in apoptosis.

Topics: Aminoacetonitrile; Animals; Apoptosis; Blotting, Western; Brain Infarction; Brain Ischemia; Caspase 3; Caspases; Cytochrome c Group; DNA Fragmentation; Enzyme Inhibitors; Infarction, Middle Cerebral Artery; Male; MAP Kinase Kinase 1; Mice; Mice, Inbred C57BL; Mitochondria; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Neuroprotective Agents; Phosphorylation; Postural Balance; Protease Inhibitors; Protein Serine-Threonine Kinases; Reperfusion Injury

Activation of the extracellular-signal-responsive kinase (ERK 1/2) by MAP kinase/ERK kinase (MEK1/2) following ischemia/reperfusion in the brain has been associated with cell death since inhibition of MEK1/2 provides neuroprotection in cerebral ischemia injury. Since inflammation has been implicated in ischemic brain injury, the present study investigated whether MEK1/2 modifies expression of two key inflammatory cytokines, IL-1beta and TNFalpha, that have been shown to exacerbate ischemic brain injury. A mouse model of transient cerebral ischemia was deployed to test the effect of selective MEK1/2 inhibitor (SL327) on infarct size and cytokine expression. SL327 (100 mg/kg, i.p.) administered 15 min prior to ischemia resulted in 64% reduction in infarct size over controls (n = 8, P < 0.01). Under the same condition, SL327 significantly reduced peak expression of IL-1beta mRNA (59% reduction compared to vehicle, P < 0.01, n = 4) but not TNF-alpha mRNA. A parallel reduction in IL-1beta protein (67%, P < 0.05, n = 6) was also observed using ELISA analysis. These data suggest that the neuroprotective effect of MEK1/2 inhibition may be mediated by suppression of IL-1beta. The study also demonstrates for the first time that these two cytokines are differentially regulated by kinase mediated signaling pathways.

Topics: Aminoacetonitrile; Animals; Brain; Brain Ischemia; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Interleukin-1; Male; MAP Kinase Kinase 1; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mitogen-Activated Protein Kinase Kinases; Protease Inhibitors; Protein Serine-Threonine Kinases; Reverse Transcriptase Polymerase Chain Reaction; RNA; RNA, Messenger; Signal Transduction; Tumor Necrosis Factor-alpha