Department of Oral Biology, Yonsei University College of Dentistry, Seoul 03722, Korea.
Department of Molecular Bioscience, School of Bioscience and Biotechnology, Kangwon National University, Chuncheon 24341, Korea.
Int J Mol Sci. 2020 Oct 16;21(20):7679. doi: 10.3390/ijms21207679.
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.
Y-27632 是一种选择性 Rho 相关卷曲螺旋形成激酶（ROCK）抑制剂。研究表明，Y-27632 可诱导多种神经元细胞的轴突生长。然而，将轴突生长与 Y-27632 联系起来的确切分子机制尚不完全清楚。在这项研究中，我们研究了 Y-27632 在 PC12 细胞中诱导轴突生长的能力，并评估了信号级联。ROS 清除剂如 N-乙酰半胱氨酸（NAC）和 Trolox 可抑制 Y-27632 诱导的轴突生长。此外，Y-27632 诱导的轴突生长不受 NADPH 氧化酶 1（NOX1）敲低或 NOX 抑制剂二苯基碘（DPI）的触发。表达显性负 Rac1 突变体（Rac1N17）抑制 Rho 家族 GTPase Rac1，该酶受 ROCK 的负调控，可阻止 Y-27632 诱导的轴突生长。此外，Rac1 抑制剂 NSC23766 可阻止 Y-27632 诱导的 AKT 和 p21 激活激酶 1（PAK1）激活。用 MK2206 抑制 AKT 可抑制 Y-27632 诱导的 PAK1 磷酸化和轴突生长。总之，我们的结果表明，Rac1/NOX1 依赖性 ROS 生成和随后激活 AKT/PAK1 级联反应有助于 Y-27632 诱导的 PC12 细胞轴突生长。
