Unresponsiveness of MS-Teff to Treg control is mediated by IL-6.

<p>(A) We defined Teff as Treg-depleted PBMC stimulated with anti-CD3 mAb in presence or absence of Treg and determined IL-6 production in supernatants on day three. Boxes represent pooled results of IL-6 secretion of 14 donors from either MS patients (black) or HC (white). Median and interquartile ranges are depicted, statistical significance was determined by Mann-Whitney-test, P-values to Teff ***p<0.001. (B) IL-6R expression within PBMC from HC (grey) or MS (black) was determined by flow cytometry. Negative control as dashed lines, average percentages ± SEM of n=4 different donors are shown. (C) MS-Teff (black) or HC-Teff (white) with Treg were stimulated with anti-CD3 mAb in presence (squares) or absence (circles) of anti-IL6R mAb. Curves show percentage of proliferation in presence of Treg normalized to Teff (n=4), P-values to MS-Teff * p<0.05 ** p<0.01, *** p<0.001. (D) Left: PKB/c-Akt phosphorylation was determined by flow cytometry within CD3⁺ Teff from MS (solid) or HC (dashed). Grey histogram depicts isotypic control of MS. One experiment of n=4 is shown; MS-MFI 366-629; HC-MFI 210-386. Right: MS-Teff (black) with Treg were stimulated with anti-CD3 mAb in presence (diamond) or absence (circle) of Akt-VIII inhibitor. Proliferation of HC (white) served as control. Curves show percentage of proliferation with Treg normalized to Teff (n=4), P-values to MS-Teff * p<0.05, *** p<0.001.