| Objective: Multiple sclerosis (MS) is an inflammatory demyelination disease of the central nervous system (CNS) that causes relapsing and progressive neurological impairment. There are many factors which are associated with the pathogenesis of MS. At present the pathogenesis of MS is still unclear, and there has no specific remedy for it. So it is our main aim that we investigate the pathogenesis of MS and seek effective therapy for MS by setting up an animal model. Recently more and more studies indicate that oxidative stress plays an important role in demyelination and axonal injury. Many studies demonstrate that there are the concentration of ferri ion and oxidative stress in MS patients and EAE models. Edaravone is a free radical scavenger which potently reduces hydroxyl radicals (OH-). It has been used as an anti-oxidative agent in acute ischemic brain disorders. Recently, there are some observations suggest that the active involvement of microglia may have a pathological role in both demyelinating and neurodegenerative disorders. Thus, agents that suppress the production of NO and ROS by activated microglia may be useful in the treatment of such pathological conditions. Edaravone has anti-oxidant and neuroprotective effects on neuronal cell death induced by SIN-1 and activated microglia. It may function as a neuroprotective agent counteracting oxidative neurotoxicity arising from activated microglia,as occurs in either inflammatory or neurodegenerative disorders of the central nervous system.In our study, we administrate edaravone to the rats of experimental autoimmune encephalomyelitis (EAE)-an animal model of MS, in order to investigate the therapeutic potential in treatment of neuroinflammatory diseases like MS. Meanwhile, we checked out the expression of Nrf2 and HO-1 in spinal cord, which could help us to investigate the mechanism of oxidative stress in MS/EAE.Methods: Ninety healthy female Wistar rats weighing 180-200g were divided randomly into five groups: normal control group(18 rats), EAE group(18 rats), dexamethasone(DXM) group(18 rats), low dose of edaravone group(18 rats) and high dose of edaravone group(18 rats). All the groups were divided into three groups: 7 day group, 16 day group, 23 day group, separately. All the experimental rats were immunized subcutaneously in the four footpads and back with emulsion 0.5ml, which including fresh guinea pig spinal cord homogenate (GPSCH) as antigen, emulsified with an equal volume of complete Freund's adjuvant (CFA) containing Mycobacterium tuberculosis 6mg/ml. Clinical signs of EAE were assessed a minimum of twice daily by two investigators. Scores were assigned on the basis of the following symptoms: 1, tail weakness; 2, tail weakness plus limb asthenia; 3, mild limb paralysis; 4, severe limb paralysis; 5, moribund/dead. EAE group, DXM group, low dose of edaravone group and high dose of edaravone group were injected intraperitoneal respectively NS 0.5ml/d, DXM 5mg·kg-1·d-1, edaravone 3mg·kg-1·d-1, edaravone 10mg·kg-1·d-1. These treatments were started on the first day of immunization and continued daily for the duration of the experiment, and EAE group left untreated. During the experiment, the mean maximal score of animals at different time point and the incidence of disease were observed as results.Rats were sacrificed after anesthesia with intraperitoneal injection. Tissues of the brain and spinal cord were fixed with 4% formalin less than 7 days, then the tissues were embedded in paraffin and sectioned at 5μm thickness. The pathological changes of tissue sections were observed under light microscopy after HE staining and trichrome staining. Immunohistochemistry were used to analyse the expression of Nrf2 and HO-1.Results:1 The morbidity of disease in different groupMorbidity of high dose of edaravone group and DXM group were significantly lower than those of EAE group, and the differences were statistically significant(P <0.05).2 Clinical profile of EAE in different groupNeurological deficits scores of high dose of edaravone group and DXM group were significantly lower than those of EAE group, and the differences were statistically significant(P <0.05).3 Neuropathological findingsThe results demonstrated that some monocytes infiltration was observed in the tissues of brains and spinal cords before the clinical signs emerging. The degree of infiltration was associated with severity of EAE. The extent of inflammation of high dose of edaravone group and DXM group were significantly lower than those of EAE group, and the differences were statistically significant(P <0.05).4 The immunoposition cells of Nrf2 and HO-1 in spinal cord : The immunoposition cells of Nrf2 and HO-1 in other groups were all higher than control groups, and the high dose of edaravone group' is highest, and the differences were statistically significant(P <0.05); The differences between EAE group and low dose of edaravone group were not statistically significant(P>0.05).Conclusions:1 The expression of Nrf2 and HO-1 in EAE group was up-regularted , but that was more obvious in high dose of edaravone groups.2 Edaravone reduces the morbidity, and protects the rats from the severity of the disease.3 Edaravone can reduce the lymphocyte infiltration and inflammation in the CNS of EAE rats.4 Edaravone played an important role on oxidative stress by up-regularting the expression of Nrf2 and HO-1.5 Edaravone has good safty and tolerance compared with DXM. These results suggest that edaravone may be used in the demyelinating diseases like MS in the future.
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