The Establishment Of Experimental Autoimmune Encephalomyelitis Model In Mice And Exploration Of Curcumin Therapy

Objective1.Establish and evaluate the EAE model;2.To investigate the intervention effect and preliminary mechanism of curcumin on the mice of experimental autoimmune encephalomyelitis.MethodsSPF female C57BL/6 mice were randomly divided into 5 groups: normal control group,EAE model group,EAE-DMSO group,EAE-rapamycin group,EAE-curcumin group.In addition to the normal control group(Control),the other groups induced EAE model in C57BL/6 mice by oligodendrocyte glycoprotein 35-55(MOG35-55),through mouse body weight change,neurological function score and pathological HE staining.And the results of the Weil staining of the myelin sheath,the EAE model was initially screened.The EAE mice were further treated with the drugs rapamycin and curcumin.The pathological HE staining and Weil myelin staining of the spinal cord tissue of each experimental group were observed,and the expression of related proteins and serum inflammatory factors were detected and quantified.Results1.Weight change and neurological function score of mice in each experimental group: Weight change evaluation found that the weight of control group and EAE group increased 7 days after immunization,and the trend was not different;14 days after immunization,the weight of EAE group increased,but the weight of EAE group increased slowly;then,the weight of EAE group decreased after 21 days of immunization,and the weight of control group continued to rise until 30 days after immunization,respectively.The weight of mice in EAE group tended to be stable,but the average weight of mice in EAE group was significantly lower than that in Control group(P < 0.05).Neurological score showed that the control group had normal activity,good mental status and no special clinical symptoms;while the EAE modelgroup had an average onset time of(8.83 + 0.41)days,with the first clinical symptoms of gray fur,tail inability to move freely,slow or slight gait abnormality;and the average time to the peak of onset was(20.5 + 0.84)days,with an average of(20.5 + 0.84)days.The clinical score was(2.34 + 0.28)and the highest was(3.42 +0.38).Compared with EAE model group,mice in curcumin group and rapamycin group could delay the onset of the disease.The average symptom score and the highest symptom score were significantly different(p < 0.05),but there was no significant difference between the two drug treatment groups(p > 0.05).2.Pathological observation of mice in each experimental group: Compared with the normal control group,EAE group and DMSO group had obvious inflammatory infiltration.The curcumin group and rapamycin group had significantly reduced inflammatory infiltration compared with EAE group,but turmeric There was no significant difference in inflammatory cell infiltration between the two groups compared with the rapamycin group.Weil myelin staining results: in the control group,the myelin sheath was well preserved and neatly arranged;however,the blue-black myelin staining in the EAE group was significantly reduced compared with the control group and the myelin sheath with different degrees of punctate white vacuoles appeared.In the missing area,the extent and extent of myelin loss in the curcumin group and the rapamycin group were significantly reduced compared with EAE mice,and the degree of myelin loss was less in the rapamycin group than in the rapamycin group.3.The expression of AKT,mToR and LC3-II in the spinal cord tissues before and after treatment with curcumin showed that the expression of Akt and m-TOR protein in the EAE group and the DMSO group was significantly higher than that in the normal group,while the curcumin group and the thunder were compared with the control group.The increase in the papamycin group was not significant.The expression of LC3-II was positive in the spinal cord tissue of the normal control group,but decreased in the EAE group and the DMSO group,but not in the curcumin group and the rapamycin group.4.The changes of AKT/mTOR autophagy pathway in EAE mice before and after treatment with curcumin showed that p-AKT,p-mTOR and P62 were highly expressed in the spinal cord of EAE group compared with the control group,while LC3-II and Beclin-1 was low expression.In contrast,compared with the EAE group,p-AKT,p-mTOR,and P62 were down-regulated in the curcumin group and rapamycin group,while LC3-II and Beclin-1 expression were significantly increased(except p-AKT,P <0.05),and there was no statistical difference between the two drug treatment groups(P>0.05).In addition,there was no statistical difference in non-phosphorylated Akt and TOR proteins between the EAE group and the curcumin group.5.The effect of curcumin treatment on plasma cytokine levels in EAE mice showed that the expression levels of IFN-?,IL-17 and TNF-? in EAE group were significantly higher than those in normal group,and the difference was statistically significant(P<0.05),while the concentrations of IFN-? and IL-17 in the curcumin-treated group and the rapamycin group were significantly lower than those in the EAE model group(P<0.05);compared with the rapamycin group,curcumin treatment The expression levels of IL-17 and IFN-? decreased slightly,while TNF-? increased,but there was no significant difference(P>0.05).Conclusion1.The EAE mouse model is successfully established in 6-8 weeks old female SPF C57BL/6mice induced by myelin oligodendrocyte glycoprotein MOG35-55;2.Curcumin treatment significantly relieve the clinical symptoms and central nervous system inflammation in EAE mice;3.Curcumin may protect the EAE by affecting the expression of autophagy in EAE mice by affecting the AKT-mTOR autophagy signaling pathway,and further balancing the changes of central nervous system and peripheral autophagy and inflammation.