| Summary: | 碩士 === 國立陽明大學 === 腦科學研究所 === 100 === Ischemic stroke is one of the leading causes of mortality and disability worldwide. Sonic hedgehog (SHH) is a morphogen involved in embryonic neural development. Our previous studies demonstrated that exogenous SHH carries beneficial effects in middle cerebral artery occlusion (MCAO) animal model of ischemic stroke. However, it is still unknown whether SHH can directly protect primary culture of cortical neurons against oxygen-glucose deprivation (OGD). Granulocyte macrophage colony-stimulating factor (GM-CSF), a hematopoietic cytokine, may protect brain against ischemia. Our previous data indicated that GM-CSF was altered in SHH-treated rat brains subjected to MCAO/reperfusion by using cytokine antibody array screening. In the present study, we test whether GM-CSF mediates SHH-dependent neuroprotection against OGD in rat cortical culture. We found that rat cortical neurons pretreated with SHH for 8 h were resistant to subsequent exposure to OGD based on LDH assay, Hoechst staining and immunocytochemistry. SHH also enhanced astrocytes expression of glial fibrillary acidic protein (GFAP) and carried protective effects against OGD-induced cell injury in a 72-h cotreatment paradigm. Furthermore, SHH significantly induced the levels of GM-CSF mRNA and protein at early phase of 2-4 h. GM-CSF protein decreased following OGD exposure. More importantly, SHH restored GM-CSF expression down-regulated by OGD, and the atrophied dendrites of neurons upon OGD exposure were recovered by SHH in both preconditioning and cotreatment paradigms. Moreover, SHH-induced GM-CSF was expressed on both neurons and astrocytes in a cotreatment paradigm. In addition, GM-CSF neutralizing antibody partially abolished SHH induced-protection in cortical cells exposed to OGD for 72 h. However, exogenous GM-CSF failed to prevent cortical cultures from OGD-induced injury in our experimental paradigm. Taken together, our results indicate that SHH carried direct beneficial effects on OGD-exposed cortical neurons and restored the expression levels of GM-CSF that was down-regulated by OGD exposure in both preconditioning and cotreatment paradigms; further, GM-CSF partially mediated SHH-induced protection against OGD injury in a 72-h cotreatment paradigm.
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