Establishment and Evaluation of Experimental Autoimmune Encephalomyelitis (EAE) Model

I. Background and Significance of EAE Research

Experimental autoimmune encephalomyelitis (EAE) is a model for autoimmune diseases mediated by CD4+ T cells. This model simulates the process of autoimmune damage in the human central nervous system, providing an important tool for studying the pathological mechanisms of demyelinating diseases such as multiple sclerosis (MS).

From a pathological perspective, the EAE model closely resembles human MS. Both exhibit progressive destruction of myelin within the central nervous system, accompanied by characteristic changes such as inflammatory cell infiltration and glial cell activation. This similarity makes the EAE model an indispensable experimental platform in neuroimmunology research, widely used in exploring pathogenesis, drug screening, and evaluating treatment strategies.

II. Selection of Experimental Animals and Strain Characteristics Analysis

The selection of experimental animals is crucial during the establishment of the EAE model. Various animal species can be used for EAE modeling, including rodents (mice, rats), lagomorphs (rabbits), primates (monkeys), among others. There are significant differences in sensitivity to EAE across different species due to genetic backgrounds and immune response characteristics.

Lewis and DA strains of rats show high susceptibility to EAE; their disease progression closely resembles that seen during acute phases in human MS. Among mouse strains, PL/J and SJL/J mice are commonly used models due to their Th1-type immune response advantage. Notably, although C57BL/6 mice have relatively low sensitivity to EAE, they have become a preferred strain for inducing chronic EAE models with MOG35-55 peptide because they possess clear genetic backgrounds and stable breeding performance.

III. Methods for Establishing the EAE Model & Technical Details

(A) Active Immunization Induction Method Active immunization induction is one of the most common methods for establishing an EAE model. This method involves mixing myelin antigens with adjuvants before injecting them into animals to induce an autoimmune response. Commonly used myelin antigens include myelin basic protein (MBP), proteolipid protein (PLP), and myelin oligodendrocyte glycoprotein (MOG). The choice of these antigens needs optimization based on research objectives and animal strains. The emulsification process requires special attention to technical details; after mixing antigen with complete Freund's adjuvant (CFA) at appropriate ratios through repeated aspiration forms a stable oil-in-water emulsion whose quality directly affects immunogenicity—an ideal emulsion should appear uniformly milky white without spreading when dropped into water. Injection typically employs subcutaneous multi-point injections at lymphoid-rich areas like back regions or groins. (B) Passive Transfer Experiment Technique Passive transfer experiments involve transferring lymphocytes from sensitized donor animals into healthy recipients from similar strains to achieve disease transfer directly demonstrating specific immune cells' roles in developing EAE—a powerful tool for investigating disease mechanisms requiring strict control over donor selection, cells purification techniques, and transferred cell quantities ensuring reproducibility throughout experiments.

IV.Evaluation System For The Established EAEModel n (A) Clinical Neurological Function Scoring Criteria Kono scoring system serves as standard clinical criteria assessing severity levels classified into five grades: Grade 1 indicates reduced tail tension; Grade 2 shows hind limb weakness; Grade 3 reveals obvious hind limb paralysis; Grade 4 progresses towards forelimb paralysis alongside hind limbs while grade five represents near-death status observations must occur daily at fixed times performed trained personnel minimizing subjective errors . n (B) Pathological Histological Detection HE staining allows observation inflammation infiltrations within CNS typical pathological alterations associated with eae includes perivascular inflammatory cuffs ,myelin loss axonal injury luxol fast blue stains specifically display integrity whereas bielschowsky silver stain aids assessment degree axonal damages combining pathology indicators analyzing clinical symptoms comprehensively . n (C) Immunological Detection Indicators Flow cytometry analyzes variations amongst subsets present eae typically observe increased th17 proportions decreased treg counts furthermore cytokine detection elisa quantifies expression levels key inflammatory factors like il-17 ifng offering objective basis evaluating activity degrees diseases n ### V.C57BL/6 Mouse MOG35-55 Induced EAEModel Construction Plan n (A) Pre-experimental Preparations Select six-to-eight-week-old female c57bl/6 mice weighing between eighteen twenty-five grams housing under spf conditions ensuring absence specific pathogen infections prepare complete freund’s adjuvant beforehand incorporating heat-killed Mycobacterium tuberculosis H37Ra strain final concentration controlled around mg/ml store pertussis toxin prepared PBS suitable concentrations reserved use . n (B) Implementation Of Immune Protocol After anesthetizing mice inject m og35 -55/cfa emulsion subcutaneously both sides back fifty microliters each point four injection sites perform intraperitoneal administration pt x two hundred ng/mouse day immunity forty-eight hours later boost immunity seventh day identical injection methods maintain sterile operations avoiding non-specific infections impacting results . n **(C) Observation Data Collection Record weight fluctuations clinical signs post-immunization typical eae course consists latent phase days onset period days recovery stage beyond twenty days endpoint usually set thirty days following immunizations tissue collection subsequent testing all procedures comply ethical standards necessary humane endpoints implemented \ ### VI.Application Prospects Research Value Of The Established EAEModel As classic tools researching multiple sclerosis,e ae holds significance both fundamental translational medicine fields optimizing methodologies evaluation systems researchers simulate accurately processes underlying human ailments future advancements gene editing technologies development chimeric mouse models will enhance role further understanding ms pathogenic mechanisms developing novel therapeutic strategies treatments utilizing eaemodel provide reliable platforms assess efficacy immunomodulatory drugs stem-cell therapies biological agents additionally investigate interactions environmental factors genetic background contributing autoimmunity providing theoretical foundations prevention.