Division of Clinical Neuroscience, Chiba University Center for Forensic Mental Health, Inohana, Chiba, Japan.
Eur J Pharmacol. 2010 Dec 1;648(1-3):67-73. doi: 10.1016/j.ejphar.2010.09.007. Epub 2010 Sep 18.
ROCK, a serine/threonine protein kinase that has been identified as a Rho GTP-binding protein, is a promising target for neuropsychiatric disorders. The selective ROCK inhibitor Y-27632 has been shown to induce neurite outgrowth in PC12 cells. However, the precise cellular and molecular mechanisms underlying ROCK inhibition-induced neurite outgrowth are not fully understood. In this study, we examined the roles of cellular signaling pathways in the potentiation of nerve growth factor (NGF)-induced neurite outgrowth by Y-27632. Y-27632 significantly potentiated NGF (2.5 ng/ml)-induced neurite outgrowth in PC12 cells, in a concentration-dependent manner. Furthermore, another ROCK inhibitor, H-1152, and the Rho inhibitor botulinum exoenzyme C3 also potentiated NGF (2.5 ng/ml)-induced neurite outgrowth. The effects by Y-27632 were antagonized by co-administration of inositol 1,4,5-trisphosphate (IP(3)) receptor antagonists (xestospongin C or 2-aminoethoxydiphenylborate (2-APB)). Moreover, the potentiation by Y-27632 was blocked by co-administration of the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 or an Akt inhibitor. In contrast, the specific inhibitors of phospholipase C (PLC-γ), p38MAPK, c-Jun N-terminal kinase (JNK), and the Ras/Raf/mitogen-activated protein kinase (MAPK) signaling pathways did not affect the potentiation of NGF-induced neurite outgrowth by Y-27632. The results of double-staining immunocytochemistry suggested that both ROCK1 and type-1 IP₃ receptors may be co-localized in the cell body of PC12 cells. In conclusion, these findings suggest that IP₃ receptors and PI3K-Akt signaling pathways might be involved in the mechanisms of potentiation of NGF-induced neurite outgrowth by ROCK inhibitors.
ROCK 是一种丝氨酸/苏氨酸蛋白激酶，已被鉴定为 Rho GTP 结合蛋白，是神经精神疾病的一个有前途的靶点。选择性 ROCK 抑制剂 Y-27632 已被证明可诱导 PC12 细胞的轴突生长。然而，ROCK 抑制诱导的轴突生长的确切细胞和分子机制尚不完全清楚。在这项研究中，我们研究了细胞信号通路在 Y-27632 增强神经生长因子（NGF）诱导的轴突生长中的作用。Y-27632 以浓度依赖的方式显著增强了 PC12 细胞中 NGF（2.5ng/ml）诱导的轴突生长。此外，另一种 ROCK 抑制剂 H-1152 和 Rho 抑制剂肉毒梭菌外毒素 C3 也增强了 NGF（2.5ng/ml）诱导的轴突生长。Y-27632 的作用被肌醇 1,4,5-三磷酸（IP3）受体拮抗剂（xestospongin C 或 2-氨基乙氧基二苯硼酸盐（2-APB））共同给药所拮抗。此外，Y-27632 的增强作用被磷脂酰肌醇 3-激酶（PI3K）抑制剂 LY294002 或 Akt 抑制剂共同给药所阻断。相比之下，PLC-γ、p38MAPK、c-Jun N-末端激酶（JNK）和 Ras/Raf/丝裂原活化蛋白激酶（MAPK）信号通路的特异性抑制剂并不影响 Y-27632 对 NGF 诱导的轴突生长的增强作用。双重染色免疫细胞化学的结果表明，ROCK1 和 1 型 IP₃ 受体可能在 PC12 细胞的胞体中共定位。总之，这些发现表明，IP₃ 受体和 PI3K-Akt 信号通路可能参与了 ROCK 抑制剂增强 NGF 诱导的轴突生长的机制。
