Fig. 4

BrQ alleviates EAE symptoms in a CD8⁺ T cell-dependent manner. A Diagram illustrating the administration scheme of anti-CD8 or an isotype control in EAE mice. The EAE mice were treated daily by gavage with either 200 µL of vehicle (0.5% CMC) or 30 mg/kg of BrQ, starting from day 3 post-immunization. B The EAE clinical scores were assessed in mice treated with vehicle or BrQ (30 mg/kg), with or without CD8 depletion. ^####p < 0.0001 by two-way ANOVA. C Statistical analysis was performed on the EAE onset day, peak EAE score, and cumulative clinical scores (n = 6–8 per group). Data are mean ± SEM. *p < 0.05, **p < 0.01 (one-way ANOVA test). D-E The number of Th1 and Th17 cells infiltrating the CNS were quantified in mice treated with vehicle or BrQ (30 mg/kg), with or without CD8 depletion, n = 6 per group. Data are mean ± SEM. **p < 0.01, ***p < 0.001, (one-way ANOVA test). F-G The splenocytes and leukocytes harvested from EAE mice on day 10 postimmunization were restimulated with MOG_35–55 (20 µg/ml) for 48 h and 72 h in vitro, respectively. Evaluation of recall responses of antigen-specific CD4⁺ T cell was performed using FACS analysis (n = 6). Data are mean ± SEM. **p < 0.01 (Student’s t test). H Isolated CD8⁺ T cells from the draining lymph nodes of C57BL/6 mice were labeled with CFSE. The proliferation of CD8⁺ T cells was assessed via flow cytometry after treatment with 20 ng/ml IL-2, 5 µg/ml CD3/CD28, and 0.25 µM BrQ for 3 days. I The total leukocytes from C57BL/6 mice were labeled with CFSE. The proliferation of CD8⁺ T cells was evaluated by flow cytometry after treatment with 20 ng/ml IL-2, 5 µg/ml CD3/CD28 and varying concentrations of BrQ for 3 days. Data are mean ± SEM. **p < 0.01, ****p < 0.0001 versus vehicle group by one-way ANOVA test. The experiments were repeated twice with consistent results