The Role And Mechanism Of Agmatine In Experimental Autoimmune Encephalomyelitis

BackgroundMultiple sclerosis(MS) is a chronic autoimmune inflammatory demyelinating disease of the central nervous system characterized by inflammatory cell infiltration,demyelination, axonal damage and neurologic disability and subsequent neurodegeneration. MS is still an incurable disease, currently available therapies are partially effective at reducing the number of relapses and slowing the progression of disability. Myelin Oligodendrocyte Glycoprotein(MOG)-induced experimental autoimmune encephalomyelitis(EAE) is the most commonly used mouse model for MS. Agmatine(Agm) is an endogenous neuromodulator. Bone morphogenetic protein(BMP) are elevated in EAE. Agm treatment improved neurological through modulating the BMP2、4、7 expressions following spinal cord injury. Agm would be a potential tool for MS/EAE. Currently, there is little research about it.ObjectiveTo demonstrate effect of Agm on neuron injury induced with glutamate, to investigate the therapeutic role of Agm in EAE and clarify the mechanism behind it.Methods(1)Establish glutamate-induced neurol model and neuron-astrocyte co-culture system.(2)Neuron–astrocyte co-cultures were stimulated by Agm and glutamate.(3)Agm stimulates astrocytes and neuron. ELISA to detect the expression of BMP.( 4) C57BL/6 mice were immunized with 200 ug MOG35-55 emulsified with Complete Freund′s Adjuvant(CFA). Pertussistoxin(PT) was intraperitoneally injected at 300 ng/per mouse at day 0 and 2 post immunization(p.i.).(5)Different doses of Agm was intraperitoneally injected to EAE in different time, control group by PBS.(6)MOG-EAE and control mice were sacrificed at peak mean clinical sevrity(20 days postimmuniza tion). The expression of BMP2/4/7, p Smad1/5/8, GFAP, Olig2,MAP2, CD11b/c, NF-κBp65, p-IκB-α, IκB-α were tested by werstern blotting, q PCR or Immunofluorescence.All experiments were repeated at least three times. Data were expressed as mean±standard deviation(SD), analysis between two groupswere performed by Student’s t-test. One-way analysis of variance(ANOVA) with Dunnett’s multiple comparisons test was applied to compare the mean values between 3 groups. For clinical score comparisons between different EAE groups, continuous analysis of variance were conducted. When P value was less than 0.05 the difference was considered as stastistic significantly.Result(1) The protective of Agm on neurons are dose dependent.(2) The expression of BMP7 in astrocytes and neurons are elevated afterstimulated by Agm.(3) We successful induced the EAE mice model using MOG35-55 emulsified withCFA followed by PTX intraperitoneally injection.(4) Compared with the PBS group, treatment with Agm have lower percentage ofBMP2/4/7, NF-κBp65, p-IκB-α in the spinal cord(P 0.01).(5) Compared with the PBS group, the expression of IκB-αwas increased aftertreatment with Agm(P 0.05).(6) Compared with the PBS group, Agm can significantly increase the expressionof neurons and oligodendrocytes(P 0.01) and reduced the expression ofmicroglia in the spinal cord(P 0.01).Conclusion(1) Agm by regulating the expression of BMP7 can protect neurons damagecaused by glutamate.(2) MOG35-55 emulsified with CFA followed by PTX intraperitoneally injectioncould induce EAE mice mode in C57BL/6 mice.(3) Agm can effectively relieve the symptoms of EAE by reducing theexpression of BMP2、7.(4) Agm can effectively relieve symptoms of EAE by inhibiting the activation ofmicroglia and increasing the expression of oligodendrocytes and neurons.