The Effect And Mechanism Of Rotating Magnetic Field Ameliorates Experimental Autoimmune Encephalomyelitis In Mice

Purpose: Multiple sclerosis(MS)is an autoimmune disease characterized by T cell infiltration and demyelination of the central nervous system(CNS).Experimental autoimmune encephalomyelitis(EAE)is the classic preclinical animal model of MS and has similar immune and pathological characteristics as MS.Magnetic therapy provides a non-invasive,safe and easy way to directly harm,pain and inflammation and other types of illness.Many years of research by our research group have proved that rotating magnetic field(RMF)can significantly relieve pain and inhibit the development of inflammation.This project aims to investigate whether rotating magnetic field can delay the onset of EAE,and further elaborate the mechanism of RMF to alleviate EAE.Experimental method: Mice were immunized with the MOG35-55 peptide to establish an EAE model.Treatment was started with RMF(0.2 T,4 Hz)on day 1 after immunization.Mice were weighed daily and scored for disease to assess disease progression.On the 10 th day,spleen cells and lymph node cells were taken for MOG-specific lymphocyte proliferation experiments to evaluate the effect of RMF on lymphocyte-specific proliferation.Mice were sacrificed at the peak of onset,and the spinal cord,spleen,and lymph nodes were removed for subsequent experiments.Spinal cord tissues were subjected to full transcriptomics sequencing;spinal cord sections were subjected to HE staining to evaluate inflammatory cell infiltration and LFB staining to observe demyelination;splenocytes and lymph node cells were subjected to flow cytometry to examine the proportion of CD4 + cells and CD8 + cells And CD4 + cell subtypes;RT-PCR and immunofluorescence carry out the expression of related chemokines and cytokines.Results: 1.The MOE-induced EAE mouse model was successfully constructed.After continuous treatment with RMF for 20 days,the weight loss of EAE mice,disease score and disease incidence were significantly reduced.2.Locomotor behavioral experiments showed that the movement distance and average speed of EAE mice within 30 minutes were significantly reduced compared with the Sham group(P <0.01).Continuous RMF treatment could significantly improve the exercise ability of EAE mice(with EAE Compared with the group,P <0.05),but has not yet returned to normal levels(compared with the Sham group,P <0.001).3.The whole transcriptome sequencing results of spinal cord tissue showed that RMF mainly acts on the immune system of EAE mice,and the differential genes mainly focus on the following pathways: cytokine-cytokine receptor interaction,Th17 cell differentiation,antigen processing and presentation,Th1 and Th2 cell differentiation and other signaling pathway systems.However,RMF exposure had little effect on MOG-stimulated lymphocyte-specific proliferation.However,RMF exposure had little effect on MOG-stimulated lymphocyte-specific proliferation.4.Effects of RMF on peripheral lymphocyte migration in EAE mice: After RMF treatment,the peripheral lymphoid tissues of EAE mice were enlarged,such as the spleen(compared with EAE group,P <0.001).The cell count showed that the absolute number of total lymphocytes and CD4 T cells in the spleen and lymph nodes of EAE mice increased significantly(compared to the EAE group,P <0.05),and the proportion of CD4+ T cells decreased by flow cytometry(compared to the EAE group,P <0.01).Spinal cord sections of EAE mice also showed that CD4 + cells were significantly Decreased(compared with EAE group,P <0.001).5 The results of sequencing and RT-PCR showed that RMF treatment down-regulated the expression of CCL-2,CCL-3 and CCL-5 in the spinal cord of EAE mice(compared with EAE group,P <0.05).At the same time,RMF treatment also down-regulated the expression of CXCL-1,CXCL-2,CXCL-9 and CXCL-10 in EAE mice(compared with EAE group,P <0.05).6.Effect of magnetic field on immunosuppressive function in EAE mice: RMF treatment increased the proportion of regulatory T(Treg)cells in spleen and lymph nodes of EAE mice(compared with EAE group,P <0.01).The results of sequencing and RT-PCR showed that RMF treatment up-regulated Foxp3 and TGF-β expression(compared with EAE group,P <0.05).RMF treatment inhibited the proportion of T helper 1(Th1)(compared with EAE group,P <0.01)and T helper 17(Th17)(compared with EAE group,P <0.05)cell in EAE mice.And sequencing and RT-PCR results showed that the expressions of Rorc(compared with EAE group,P <0.01)and T-bet(compared with EAE group,P <0.05)were down-regulated.7.Effect of RMF on immune factor production in EAE mice: Peripheral blood cytokine detection experiments showed that RMF treatment significantly reduced the levels of IFN-γ and IL-17 A in peripheral blood of EAE mice(compared with EAE group,P <0.05),but had no effect on several other cytokines.Spinal cord section results showed that RMF treatment caused IFN-γ + cells(compared with EAE group,P <0.05)and IL-17 + cells(compared with EAE group,P <0.001)in EAE mice decreased significantly.Sequencing and RT-PCR results showed that RMF treatment significantly inhibited the expression of CD4,IFN-γ,and IL-17(compared to the EAE group,P <0.05).Conclusion: The experimental results show that RMF(0.2 T,4 Hz)treatment can prevent EAE disease progression,including reducing the severity of the disease and delaying the onset of the disease.RMF exposure had no effect on MOG-stimulated lymphocyte-specific proliferation.A possible mechanism is that RMF improves EAE disease by inhibiting the migration of CD4 + T cells to the spinal cord and improving the imbalance between treg cells and Th1 / Th17 cells.As a mild physical therapy,RMF may become an effective intervention for clinical treatment of MS,which significantly improves the prognosis and quality of life.