Fasudil Attenuated 6-OHDA Cytotoxicity in PC12 Cells through Inhibition of JAK/STAT and Apoptosis Pathways

Background: 6-Hydroxydopamine (6-OHDA) is widely used to induce neurotoxicity and investigate the mechanisms of Parkinson disease. 6-OHDA causes cell injury through various mechanisms including oxidative stress, inflammation and apoptosis. The selective Rho-kinase inhibitor, fasudil displays neuroprotective effects in several neurodegenerative disorders. The aim of this study was to assess the protective effect of fasudil in PC12 cytotoxicity induced by 6-OHDA. Methods: PC12 cells were exposed to 5, 10, 25, and 50 mu M of fasudil concentrations. After 24 h, the IC50 value of 6-OHDA (150 mu M) was added. Twenty-four hours later, the viability of cells was evaluated via MTT assay and the formation of reactive oxygen species (ROS) was measured by the fluorimetric method. At the 50 mu M concentration of fasudil, with or without 6-OHDA, the changes of protein levels including STAT3, P-STAT3, JAK2, P-JAK2, and caspase-3 were determined via western blotting. Results: Our results showed that 6-OHDA increased the intracellular level of ROS, reduced cell viability, upregulated p-STAT3/STAT3 and p-JAK2/JAK2 ratios and significantly raised cleaved caspase-3 in comparison to control group. Furthermore, pretreatment of cells with fasudil (50 mu M) for 24 h could reverse all changes induced by 6-OHDA. Conclusion: 6-OHDA caused cytotoxicity in PC12 cells through inducing oxidative stress and activating JAK/STAT and apoptosis pathways, while pretreatment with fasudil exhibited protec-tive effect on 6-OHDA-induced neurotoxicity via the inhibition of oxidative stress and preven-tion of these pathways.