Fig. 8
OPCs express myelin peptide on MHC class I and are cytotoxic targets in vivo. C57BL/6 PDGFRα-CreER × Rosa26-YFP were kept on a CPZ diet for 4 weeks. After 3 weeks, 4-HT was injected to induce Cre recombination in PDGFRα expressing cells and MOG35-55 reactive T cells were AT. Cells were isolated ex vivo and analyzed by flow cytometry. a The OPC population was distinguished based on YFP− and YFP+ expression and analyzed for caspase 3/7 activity, quantification below; CPZ (gray) and CPZ+MOG35-55 AT (blue). b–d Alternatively, C3HeB/FeJ donor mice were immunized with whole recombinant rat MOG1-125 to induce EAE, and myelin-specific CD4 T cells were isolated ex vivo and reactivated with peptide. Syngeneic recipient mice were fed a CPZ diet for 6 weeks before AT. b H2Kk (left) [No CPZ vs. AT mean diff = −15.2, 95% CI = −24.7–(−)5.6, CPZ vs. CPZ+AT mean diff = −19.3, 95% CI = −29.4–(−)9.3], and Fas (right) (No CPZ vs. AT mean diff = −38.1, 95% CI = −49.6–(−)26.7, CPZ vs. CPZ+AT mean diff = −37.4, 95% CI = −49.5–(−)25.3) expression from the OPC population was determined by flow cytometry analysis with quantification below each flow histogram; no AT (gray; n = 8,8,4), MOG97-114 AT (blue; n = 9,10,5), CPZ+no AT (gray; n= 8,12,4), and CPZ + MOG97-114 AT (blue; n = 8,17,4). Statistical significance was determined by one-way ANOVA analysis followed by Tukey’s multiple comparisons. Error bars represent the standard deviation. c Caspase 3/7 activity was determined by flow cytometry [No CPZ vs. AT mean diff = −36.4, 95% CI = −55.4–(−)17.5, CPZ vs. CPZ+AT mean diff = −39.9, 95% CI = −59.9–(−)20.0]. Further population analysis was done from the caspase 3/7 active population by Live/Dead staining (top), H2Kk expression (middle) (Diff between means = 13.7 ± 3.3, 95% CI = 6.7–20.7) and FAS expression (bottom) (Diff between means = 14.1 ± 3.9, 95% CI = 4.8–23.4). d H2Kk-MBP79-87 staining from OPCs with FMO staining control and quantification below. At peak disease flow cytometry was completed on whole brain tissue to assess intracellular granzyme b staining in target cells (No CPZ vs. AT mean diff = −7.9, 95% CI = −11.3 to −4.5, CPZ vs. CPZ + AT mean diff = −6.9, 95% CI = −9.5 to −4.3)
