The Preventive Treatment Of EAE In Wistar Rats By Using Matrine And The Mechasnism Of Action

Objective:In order to investigate the preventive treatment and potential mechanism of matrine (MAT) in experiment autoimmune encephalomyelitis (EAE), we observed the effects of matrine on the rat model of EAE, which include clinical symptoms of EAE, the degree of histopathologic changes in spinal cord, the expression of nitric oxide(NO)/inducible nitric oxide synthase (iNOS) system in rats blood and spinal cord, the influence of interleukin-17(IL-17)/interleukin-23(IL-23) axis.Methods:1 Induction of animal Model:The EAE models were induced in female Wistar rats by being immuned with guinea pig spinal cord homogenate (GPSCH) extracts together with complete Freund adjuvant (CFA). The clinical symptoms of EAE rats,body weights and the actions were observed every day after immunization.2 Animal groups and administration:Forty,Clean,female,Wistar rats were randomly divided into four equal group:EAE model group (M), low dose MAT treatment group (MAT-L), high dose MAT treatment group (MAT-H) and normal control group (N).M group and N group were intraperitoneally injected with 6.7 mL·kg-1 normal saline injection for 16 uninterrupted days after immunization, at the same time MAT-L group were treated MAT at 150 mg·kg-1,and MAT-H group received MAT at 200 mg·kg-1 by intraperitoneally injected for 16 uninterrupted days after immunization. 3 Sample collections:the rats were killed at the 17th day postimmunization.The blood was collected via posteriororbital venous plexus approach and then the serum was separated. The spinal cord were removed after heartperfusion and stored in 4% paraformaldehyde solution.4 Detection of indexes:The degree of histopathologic changes in spinal cord was assessed directly by hematoxylin-eosin (HE) and chromotrope-2R-brilliant green (C-2R-BG). Nitrate reductase method was used for analyzed for NO in serum level.iNOS and iNOSmRNA in spinal cord were determined by immunohisto-chemical techniques(IHC) and Reverse transcription-polymerase chain reaction (RT-PCR)respectively.Enzyme linked immunosorbent assay(ELISA)method was used for determination of IL-17 and IL-23 in serum level.Results:1 Incidence rate and the changes of clinical score, body weight:There was no ill rat in N group;the incidence rate of MAT-H and MAT-L groups were lower than M group (P>0.05); comparing to MAT-H group, the incidence rate of MAT-L group is higher (P>0.05). The body weight of N group was increased constantly in the whole experimental period, the mean weight change of M,MAT-H and MAT-L group were declined;the mean weight change of MAT-H and MAT-L groups were lower than M group (P<0.05); the mean weight change of MAT-H group and MAT-L group have no significantly differences(P>0.05).Compared to the MAT-H and MAT-L group, the clinical scores of M group was significantly increased(P<0.05); the clinical scores of MAT-H group and MAT-L group have no significantly differences(P>0.05).2 The observation of histopathologic changes:There were no inflammatory infiltration and demyelination in N group rats,but the M,MAT-H and MAT-L group rats all have inflammatory infiltration and demyelination; compared to M group, the inflammatory infiltration and demyelination in MAT-H and MAT-L group were reduced(P<0.01); the inflammatory infiltration and demyelination in MAT-H group was reduced than MAT-L group(P>0.05).3 The changes of NO in serum:Compared to N group,the expression of NO in the M,MAT-H and MAT-L group are increased(P<0.05); compared to M group, the expression of NO in MAT-H and MAT-L group is declined(P<0.01); however, the expression of NO in MAT-H group and MAT-L group have no significantly differences(P>0.05).4 The expression of iNOS and iNOSmRNA in spinal cord:Compared to N group,the expression of iNOS and iNOSmRNA in the M,MAT-H and MAT-L group are increased(P<0.05); compared to the M group, the expression of iNOS in MAT-H group is declined(P<0.01),the expression of iNOSmRNA in MAT-H and MAT-L group is declined(P<0.05); however, the expression of iNOSmRNA in MAT-H group and MAT-L group have no significantly differences(P>0.05).5 The expression of IL-17,IL-23 in serum:Compared to N group,the expression of IL-17,IL-23 in the M,MAT-H and MAT-L group are increased(P<0.01); compared to M group, the expression of IL-17,IL-23 in MAT-H and MAT-L group is declined(P<0.01,P<0.05); however, the expression of IL-17,IL-23 in MAT-H group and MAT-L group have no significantly differences(P>0.05).Conclusion:MAT possesses obvious preventive treatment to EAE model in Wistar rats, which were induced by being immuned with spinal cord extracts to together with complete Freund adjuvant.the possible mechanism of action is related to inhibit the expression of NO/iNOS system in rats blood and spinal cord,suppress IL-17/IL-23 axis through the down-regulating of IL-17,IL-23 expression,thus improve the clinical symptom of EAE rats.