Mechanism Of Treg/Th17Cell For Demyelinated Optic Neuritis

Objective: This proposal tried to establish experimental autoimmune encephalomyelitis (EAE) and explore the immunomodulatory effects of T helper (Treg), Th17cell in the model of optic neuritis. Make the immune mechanism of demyelinating optic neuritis clear.Methods:1. EAE model was established to observe clinical symptoms and visual electrophysiological changes of optic neuritis in mice.2. Hematoxylin-eosin staining was used to observe histological changes in the mouse models of EAE. Immunohistochemical staining was used to detect optic nerve axon damage markers β-amyloid peptide pression (β-APP) protein expression quantity changes. Flow cytometric assay was used to detecte the rate of Treg cells in mouse splenocyte. Reverse transcription PCR (RT-PCR) was used to detect the dynamic expression of forkhead/winged helix transcription factor p3(Foxp3), transforming growth factor-β (TGF-β), interleukin-1β (IL-1β), IL-17, IL-10in optic nerve tissue.Results:1. After immunization11days, neurological symptoms of EAE mice started to appear. Hematoxylin-eosin staining showed optic nerve tissue infiltration of inflammatory cytokines. Immunohistochemical staining showed enhancement of axonal injury markers β-APP positive staining.2. Flash visual evoked potential (F-VEP) test showed7,11,14,19,23and28days, compared with normal mice, P1prepatents of model group were prolonged after modeling, the differences were statistically significant (t=4.487,15.203,16.364,11.540,11.959,16.163, all at P＜0.05). when7days after modeling, the difference of N1-P1amplitudes between normal group and model group was no statistical significance (t=-0.992, P=0.378), while11,14,19,23and28days after modeling, compared with normal mice, N1-P1amplitudes of model mice were significantly lower, the differences were statistically significant (all at P＜0.05).3. Flow cytometric assay showed that the count of CD4+CD25+T cells and Foxp3+CD4+CD25+T cells in mouse splenocyte from model groups had reduced slightly compared with normal group since early until the peak of the morbidity. Compared with the control group, when11,14,19,23and28days after modeling, the count of CD4+CD25+T cells and the number of Treg cells were decreased, the differences were statistically significant (F=74.968,51.680P＜0.05).4, The expressions of TGF-β, IL-1β, IL-17, Foxp3, IL-10mRNA among different time pionts had stastistic significant difference (F=12.721,15.015,14.343,69.374,68.290, all at P=0.000). Compared with the normal group, when11days,14days after modeling, TGF-β mRNA levels were significantly increased, and when19days,23days after modeling, IL-1β mRNA levels were significantly increased, and when7days,11days after modeling, IL-17mRNA levels were significantly increased, and when7,11,14,19,23,28days, Foxp3mRNA levels were significantly lower, and when19,23,28days, IL-10mRNA levels were significantly lower, the differences were statistically significant (all at P＜0.05).Conclusions:1. Demyelinating optic neuritis was established successfully by EAE model.2. Th17cells launch demyelinating optic neuritis immune injury, and Thl subgroup performs the function of maintaining inflammatory lesions, Treg cell subgroup is abnormal much earlier than Th2subgroup. The imbalance of Th17/Treg cell may be involved in the pathogenesis of demyelinating optic neuritis.