The Potential Of Fasudil In The Treatment Of Eae And Anti-inflammatory Effects

Objective In the present study, we try to observe the therapeutic potential of Rho kinaseinhibitor Fasudil in C57BL/6 mice with the experimental autoimmune encephalomyelitis (EAE )models induced by myelin oligodendrocyte glycoprotein peptides 35-55(MOG35-55),and explorethe possible mechanisms in the treatment of EAE.Methods Chronic EAE was induced by MOG35-55peptide in female C57BL/6 mice,which were randomly divided into EAE group, Fasudil early-treated group and Fasudillate-treated group. Fasudil was injected intraperitoneally on day 3 post-immunization (Fasudilearly-treated group) or at onset of symptoms (Fasudil late-treated group) respectively. Animalswere weighed and evaluated for clinical score every other day in a blinded fashion by at leasttwo investigators. On day 28 after immunization, mice were sacrificed, brains and spinal cordswere obtained for HE staining and CD4+T cell staining. Protein extracts from brains and spinalcords were collected for the measurement of p-MYPT1 and NF-kB by Western blot. The datawere analysed by GraphPad Prism 4 software.Results 1. Fasudil delayed onset of EAE and improved clinical symptoms. The incidenceof EAE (28.6%) in Fasudil early-treated mice was decreased as compared with EAE control(100%) and Fasudil late-treated mice (78.6%). In EAE group, mean onset date was 12.57±1.55,mean maximum score is 2.50±1.13. Fasudil late-treatment slightly delayed onset (13.45±2.07),and declined maximum clinical score (1.36±1.06, p<0.05). Obviously, Fasudil early-treatmentis able to delay its onset (16.50±4.20, p<0.05), and obviously declined maximum clinical score(0.32±0.70, p<0.01). We also observed that the treatment of Fasudil at induction or onset phasesof EAE can reduce body weight loss as compared with that of EAE mice.2. Fasudil declined the infiltration of inflammatory cells in brains and spinal cords. HEstaining showed marked inflammatory cell infiltration in spinal cords of EAE mice. The numberof infiltrating cells in spinal cords demonstrated a marked decrease in Fasudil late-treated mice,especially in Fasudil early-treated mice. Immunohistochemistry showed marked inflammatorycell infiltration in spinal cords of EAE mice. The number of infiltrating cells in spinal cordsdemonstrated a marked decrease in Fasudil late-treated mice, especially in Fasudil early-treatedmice.3. Fasudil inhibited the expression of p-MYPT1 in spinal cords and brains. The expression ofp-MYPT1 in spinal cord from both Fasudil late- and early-treated mice was significantlysuppressed, as compared with EAE control mice. The expression of p-MYPT1 in brain isrelatively low. Thus, the expression of p-MYPT1 was significantly reduced only in brain of micewith Fasudil-early-treated mice. 4．Fasudil decreased the level of NF-κB in spinal cords and brains. The expression ofNF-kB in spinal cords exhibited a marked decrease in both Fasudil late-treated mice andearly-treated mice. In EAE control group, the expression of NF-κB in spinal cords wassignificantly higher than that in brains. Fasudil late- and early-treatment slightly decreased theexpression of NF-κB in brains, but there was no statistical significanc.Conclusion In EAE, Fasudil treatment, both late and early stages of EAE, exhibitstherapeutic potential, but Fasudil early treatment is more effective. The therapeutic effect ofFasudil may be associated with the inhibition of inflammatory cell infiltration, p-MYPT1 andNF-κB in the CNS, thereby reducing the inflammatory microenvironment caused by theinflammatory cytokines, and achieving the treatment and prevention in the occurrence anddevelopment of EAE.