Y-27632 Induces Neurite Outgrowth by Activating the NOX1-Mediated AKT and PAK1 Phosphorylation Cascades in PC12 Cells

Abstract

Y-27632 is known as a selective Rho-associated coiled coil-forming kinase (ROCK) inhibitor. Y-27632 has been shown to induce neurite outgrowth in several neuronal cells. However, the precise molecular mechanisms linking neurite outgrowth to Y-27632 are not completely understood. In this study, we examined the ability of Y-27632 to induce neurite outgrowth in PC12 cells and evaluated the signaling cascade. The effect of Y-27632 on the neurite outgrowth was inhibited by reactive oxygen species (ROS) scavengers such as N-acetyl cysteine (NAC) and trolox. Furthermore, Y-27632-induced neurite outgrowth was not triggered by NADPH oxidase 1 (NOX1) knockdown or diphenyleneiodonium (DPI), a NOX inhibitor. Suppression of the Rho-family GTPase Rac1, which is under the negative control of ROCK, with expression of the dominant negative Rac1 mutant (Rac1N17) prevented Y-27632-induced neurite outgrowth. Moreover, the Rac1 inhibitor NSC23766 prevented Y-27632-induced AKT and p21-activated kinase 1 (PAK1) activation. AKT inhibition with MK2206 suppressed Y-27632-induced PAK1 phosphorylation and neurite outgrowth. In conclusion, our results suggest that Rac1/NOX1-dependent ROS generation and subsequent activation of the AKT/PAK1 cascade contribute to Y-27632-induced neurite outgrowth in PC12 cells.