Protective Effect of Mitogen- and Stress-Activated Protein Kinase on the Rats with Focal Ischemia-Reperfusion Injury.

Abstract

Mitogen- and stress-activated protein kinase (MSK) is a recently identified nuclear cAMP-regulated enhancer B (CREB) and histone H3 kinase that responds to both mitogen- and stress-activated protein kinases. This study was designed to investigate the protective effect of MSK on the rats with focal ischemia-reperfusion injury. The rat model was established by inserting thread into the middle cerebral artery. The protein expression was measured by immunoblotting. The localization of MSK was measured by immunofluorescence assay. Highly-differentiated pheochromocytoma 12 (PC12) is used as a sympathetic neuron-like cell line and treated with glutamate to induce neurotoxicity. MSK was knocked down and overexpressed by siRNA and MSK over-expressing vector, respectively. The cell viability was measured by cell counting kit (CCK-8) assay. The coronal sections were isolated and stained with 2, 3, 5-triphenyltetrazolium chloride (TTC) to determine infarct volume. Finally, astrocytes were separated from cerebral cortexes of normal rats to analyze the effects of MSK on inflammatory response. In the rats with focal ischemia-reperfusion injury, the expression of MSK was reduced, reaching the lowest level at 3d after ischemia-reperfusion, and then recovered gradually. MSK was found mainly localized in neurons and astrocytes. The expression levels of caspase-3, caspase-8, caspase-9, and INOS showed the opposite trend with respect to MSK. Further analysis showed that overexpression of MSK exerted a protective effect on glutamate-induced neurotoxicity through inhibiting apoptosis of PC12 cells, as well as decreased the infarct size in rat with focal ischemia-reperfusion injury. On the contrary, knockdown of MSK showed opposite results. Finally, MSK suppressed LPS-induced inflammatory response by decreasing the expression of inducible nitric oxide synthase (INOS) and increasing the expression of interleukin-10 (IL-10) in astrocytes from cerebral cortexes of normal rats. In conclusion, MSK exerted a protective effect on rat with focal ischemia-reperfusion injury through its anti-apoptotic effect on neurons and anti-inflammatory effect on astrocytes.