The Therapeutic Comparision Of WAR-5, Fasudil And The Exploration Of Their Mechanisms

Objective:To compare the therapeutic effect of WAR-5 and Fasudil in experimental autoimmune encephalomyelitis(EAE), and explore their possible mechanisms, providing experimental basis for further clinical treatment.Methods:Female C57BL/6 mice were immunized with myelin oligodendrocyte glycoprotein peptides 35-55(MOG35-55) to establish chronic EAE model. Mice were randomly divided into three groups: WAR-5 treatment, Fasudil treatment and EAE control.WAR-5 or Fasudil was intraperitoneally injected from day 3 post-immunization(p.i) to day 27 p.i.Animals clinical score was evaluated. On day 28 p.i. animals were sacrificed and spinal cords were obtained for HE staining, myelin staining, CD4, CD68, NGF, BDNF and synaptophysin immunohistochemistry staining. Protein extracts from brains and spleens were collected for the measurement of ROCK II, i NOS and Arg-1 by Western blot.Results:1.Both WAR-5 and Fasudil delayed onset of EAE and improved clinical symptoms,with similar therapeutic effect.2. WAR-5 more significantly inhibited the expression of ROCK II than Fasudil(P<0.05).3. The effect of WAR-5 is similar with Fasudil, WAR-5 improved demyelination and enhanced synaptophysin protein expression in spinal cord. Mice with EAE showed extensive myelin loss(demyelination) in spinal cords. When compared with EAE control,WAR-5 or Fasudil-treated mice had a significant improvement in the extent of demyelination(P<0.001, P<0.01). Immunohistochemistry showed the decrease of synaptophysin protein expression in EAE control mice.4. The effect of WAR-5 is similar with Fasudil, WAR-5 inhibited inflammation in spinal cord. HE staining showed that WAR-5 and Fasudil declined the infiltration of inflammatory cells in spinal cords( P<0.01). Immunohistochemistry showed that EAE control mice contained high numbers of CD4+ T cells and CD68+ macrophages, and the numbers of these cells were significantly reduced in mice treated with WAR-5 or Fasudil(P<0.001).5. Both WAR-5 and Fasudil inhibited the expression of i ONS(P<0.05), increased the expression of Arg-1(P<0.01, P<0.05), shifts M1 to M2 phenotype.6. Both WAR-5 and Fasudil significantly enhanced expression of NGF(P<0.05) and BDNF(P<0.05, P<0.01) in spinal cord compared with control EAE mice.7. Compared with Fasudil, WAR-5 had a relatively weaker vasodilator effect and cell toxicity than Fasudil.Conclusion:Both WAR-5 and Fasudil delayed onset of EAE and improved clinical symptoms,with similar therapeutic effect. WAR-5 and Fasudil can significantly suppress the inflammatory reaction in EAE and their possible mechanisms may be related to downregulating the expression of ROCK II, declining the infiltration of inflammatory cells in spinal cords and shifting M1 to M2 phenotype macrophages. Meanwhile, WAR-5and Fasudil enhanced the expression of NGF and BDNF to promote axonal regeneration.In addition, WAR-5 has a relatively weaker vasodilator effect and cell toxicity than Fasudil.