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. 2009 Jun 8;206(6):1303–1316. doi: 10.1084/jem.20090299

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© 2009 Pöllinger et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jem.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

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Figure 3. — Inflammatory cell infiltrates in RR-EAE lesions. (A–C) Cellular infiltrate into the CNS of sick TCR¹⁶⁴⁰ mice (score 3) is composed of macrophages, T cells, and B cells. CNS mononuclear cells were isolated from a sick TCR¹⁶⁴⁰ mouse and stained against CD11b and CD45.1 (A), together with CD8 and CD4 (B) or together with CD19 and B220 (C). Cells in B and C were analyzed among gated CD45.1⁺CD11b⁻ cells (red region as indicated). (D–F) Activation and Th1/Th17 cytokine expression of infiltrating CD4⁺ T cells. (D) CNS infiltrate cells express CD25, but not CD62L, and partially down-modulate their TCR (Vα8.3 and Vβ4). Activation status and TCR expression was compared between splenocytes (left) and CNS-isolated cells (right). (E) CD4⁺CD3⁺ T cells infiltrating the CNS of sick TCR¹⁶⁴⁰ mice predominantly expressed the pathogenic TCR composed of Vα8.3 and Vβ4 chains in a low expression level (TCR^low) compared with the spleen. Numbers indicate the percentage of stained cells in the respective quadrant or region. (F) Intracellular cytokine staining after stimulation with PMA/ionomycin in brefeldin A. Th1 (IFN-γ⁺/IL-17⁻) and Th17 (IFN-γ⁻/IL-17⁺) cells are enriched in CNS infiltrates of sick TCR¹⁶⁴⁰ (score 3.5) mice compared with splenocytes. D–F were analyzed among gated CD45.1⁺CD4⁺ cells. Flow cytometry data are representative of three to five sick TCR¹⁶⁴⁰ mice analyzed in three to five independent experiments.