GSK-3β activation mediates Nogo-66-induced inhibition of neurite outgrowth in N2a cells

The axons of the adult mammalian brain and spinal cord fail to regenerate after injury, and it has been suggested that Nogo-66 could prevent CNS axon repair. However, the mechanism of Nogo-66 inhibiting neurite outgrowth remains unknown. Our previous results indicated that protein kinase B (PKB) is involved in the inhibition of the neurite outgrowth by Nogo-66. Glycogen synthase kinase-3 beta (GSK-3 beta) is implicated in many processes in the nervous system, including differentiation, specification, polarity, plasticity and axon growth. In addition, GSK-3 beta is one of the most important molecules downstream of PKB. In the present study, we report on the role of GSK-3 beta signaling on Nogo-66-treated mouse neuroblastoma N2a cells. Nogo-66 reduced the phosphorylation of GSK-3 beta at Ser9 in N2a cells. In contrast, pretreatment with SB216763, a specific inhibitor of GSK-3 beta, resulted in an amelioration of neurite outgrowth by Nogo-66, compared with the Nogo-66 alone group (P < 0.05). Moreover, we performed RNA interference experiments to knock down GSK-3 beta expression levels in N2a cells via transient transfection of shRNA plasmids. The inhibition of neurite outgrowth by Nogo-66 was subdued in shRNA cells, compared to the non-RNAi cells (P < 0.05). Taken together, these data suggest that GSK-3 beta is involved in the inhibition by Nogo-66 of neurite outgrowth in N2a cells. (C) 2011 Elsevier Ireland Ltd. All rights reserved.