Figure 1.
Adoptively transferred encephalitogenic T cells infiltrate the LM and brain, but not the dura, during the establishment of EAE. (A) Clinical scores were tracked in SJL/J mice in which EAE was induced by adoptive transfer of 10 million CFSE-labeled Th17-skewed encephalitogenic T cells. EAE mice developed clinical disability, including paralysis (reflected by increasing clinical score), and experienced weight loss. (B–D) At various time points throughout the disease, mice were euthanized, and the dura, LM, brain, blood, spleen, and BM from the skull and femur/tibia were collected and analyzed by flow cytometry. Results are expressed as the absolute number of cells in the whole tissue or 100 μl of blood (B and C) or as a percentage of CD3+CFSE+ cells (D). The percentage of CFSE+ T cells that expressed the phenotype CD44hiCD62L− was quantified in the blood, LM, and brain at days 7 and 11 of EAE (D). Results in B are data from one experiment wherein we did not collect or analyze brain tissue or BM, and results in C and D are data from two independent experiments wherein we analyzed brain tissue and BM in addition to the tissues analyzed in Fig. 1 B. Clinical scores in A are representative of all three independent experiments in B–D. A total of 3–12 mice were used per time point in A–D, as indicated in the graph legends. Error bars indicate mean ± SEM. For B (blood and spleen) and C (spleen), a Kruskal–Wallis test followed by Dunn’s multiple comparison test was conducted to assess statistical significance from naïve. For B and C (CNS) and D, a mixed-effects analysis followed by Dunnett’s multiple comparison test was conducted to assess statistical significance from the dura (B and C) or blood (D) within time points. For C (BM), a two-way ANOVA followed by Sidak’s multiple comparison test was used to assess statistical significance between the leg and skull BM within time points. In all cases, statistical significance was denoted as * = P < 0.05, ** = P < 0.005, *** = P < 0.0005, **** = P < 0.0005, and no label = not significant. The two variables compared in the mixed-effects analysis/two-way ANOVA were days postadoptive transfer (time) and tissue type (B: dura versus LM; C: femur/tibia BM versus skull BM or dura versus LM and brain; D: blood versus LM and brain).

Adoptively transferred encephalitogenic T cells infiltrate the LM and brain, but not the dura, during the establishment of EAE. (A) Clinical scores were tracked in SJL/J mice in which EAE was induced by adoptive transfer of 10 million CFSE-labeled Th17-skewed encephalitogenic T cells. EAE mice developed clinical disability, including paralysis (reflected by increasing clinical score), and experienced weight loss. (B–D) At various time points throughout the disease, mice were euthanized, and the dura, LM, brain, blood, spleen, and BM from the skull and femur/tibia were collected and analyzed by flow cytometry. Results are expressed as the absolute number of cells in the whole tissue or 100 μl of blood (B and C) or as a percentage of CD3+CFSE+ cells (D). The percentage of CFSE+ T cells that expressed the phenotype CD44hiCD62L− was quantified in the blood, LM, and brain at days 7 and 11 of EAE (D). Results in B are data from one experiment wherein we did not collect or analyze brain tissue or BM, and results in C and D are data from two independent experiments wherein we analyzed brain tissue and BM in addition to the tissues analyzed in Fig. 1 B. Clinical scores in A are representative of all three independent experiments in B–D. A total of 3–12 mice were used per time point in A–D, as indicated in the graph legends. Error bars indicate mean ± SEM. For B (blood and spleen) and C (spleen), a Kruskal–Wallis test followed by Dunn’s multiple comparison test was conducted to assess statistical significance from naïve. For B and C (CNS) and D, a mixed-effects analysis followed by Dunnett’s multiple comparison test was conducted to assess statistical significance from the dura (B and C) or blood (D) within time points. For C (BM), a two-way ANOVA followed by Sidak’s multiple comparison test was used to assess statistical significance between the leg and skull BM within time points. In all cases, statistical significance was denoted as * = P < 0.05, ** = P < 0.005, *** = P < 0.0005, **** = P < 0.0005, and no label = not significant. The two variables compared in the mixed-effects analysis/two-way ANOVA were days postadoptive transfer (time) and tissue type (B: dura versus LM; C: femur/tibia BM versus skull BM or dura versus LM and brain; D: blood versus LM and brain).

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