Figure 2

Induction of macrophage IL-10 by cytokine-stimulated T cells (Tck) is PI3K- and p70S6K-dependent. Tck were co-cultured with M-CSF-primed macrophages at a T:macrophage ratio of 5:1. Before co-culture, macrophages were treated for 1 hour with PI3K inhibitor LY294002 (a) or wortmannin (b) or with the p70S6K inhibitor rapamycin (d). Bars show mean IL-10 concentrations in triplicate culture supernatants ± SD, for a representative of N = 5 replicate experiments. Western blot analysis of activated phospho-PKB (c) shows that the downstream effector PKB was activated by Tck and that the effect was abrogated by wortmannin (500 nM) and LY294002 (5 μM). Lane 1, macrophage control; 2, Tck control; 3, macrophage+Tck; 4, macrophage+Tck+wortmannin (500 nM); 5, macrophage+Tck+LY294002 (5 μM). In addition, Tck activate macrophage p70S6K, which is PI3K-independent (e). Lane 1, macrophage control; 2, macrophage+Tck; 3, macrophage+Tck+wortmannin (500 nM); 4, macrophage+Tck+LY294002 (5 μM). Data are representative of N = 3 replicate experiments. M-CSF = macrophage-colony-stimulating factor; p70S6K = p70 S6-kinase; p85S6K = p85 S6-kinase (nuclear isoform of p70 S6-kinase); phospho-p70S6K = phosphorylated p70 S6-kinase; (phospho-)PKB = (phosphorylated) protein kinase B; PI3K = phosphatidylinositol 3-kinase. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001.