Effect of early therapeutic anti-VLA-4 mAb on pathogenic T cell infiltration into the CNS.

<p>(A) Mice were sacrificed one day after last treatment and mononuclear cells isolated from the CNS (brain and spinal cord) of anti-VLA-4 mAb-treated (n = 4) and IgG-treated (n = 4) mice were stained by anti-CD45 and -CD3 mAbs. CNS-infiltrated T cells (CD45⁺CD3⁺) were analyzed by flow cytometry. **; p-value <0.01. (B) Production of pro-inflammatory cytokine by the CNS-infiltrated CD4 T cells in anti-VLA-4 mAb- and IgG-treated mice. CD11b⁺CD45⁺ and CD4⁺ were sequentially gated, and then production of pro-inflammatory cytokines in response to anti-CD3/CD8 mAb in the CD11b⁺CD45⁺CD4⁺ cells was examined. (C) Effect of anti-VLA-4 mAb treatment on CNS-infiltration of GM-CSF⁺Th1 and GM-CSF⁺Th17 cells. CNS-infiltration of GM-CSF⁺TNF-α⁺, GM-CSF⁺IFN-γ⁺, and GM-CSF⁺IL-17⁺ CD4⁺ T cells in anti-VLA-4 mAb (n = 3) and IgG (n = 4)-treated mice were calculated as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099068#s2" target="_blank">Materials and Methods</a>. Data represent mean +/− SEM. Results shown are representative from three experiments.