Figure 1
T cell induced production of TNF-α by monocytes from healthy donors. (a) Fixed stimulated T cells induce tumour necrosis factor (TNF)-α production by monocytes in a dose-dependent manner. Peripheral-blood T cells (106/ml) were cultured for 48 hours in the presence of immobilized anti-CD3 (3.3 μg/ml) and soluble anti-CD28 (0.8 μg/ml) antibodies. Stimulated T cells were fixed and incubated for 24 hours with freshly isolated monocytes (1.5 × 106/ml) at the indicated ratios. Values are means ± SEM from four different experiments. (b) Cell–cell contact is necessary for T cell-induced production of TNF-α in monocytes. Peripheral-blood T cells (106/ml) were cultured for 48 hours in the presence or absence of immobilized anti-CD3 (3.3 μg/ml) and soluble anti-CD28 (0.8 μg/ml) antibodies. Stimulated (sTc) and resting (rTc) T cells were fixed and incubated for 24 hours with freshly isolated monocytes (1.5 × 106/ml) at a ratio of 7:1 in a transwell system as described in the Materials and methods section. T cells and monocytes were physically separated by the semi-permeable membrane (with insert) or had direct cell–cell contact (without insert). Lipopolysaccharide (LPS; 100 ng/ml) was used as a positive control. Values are means ± SEM for experiments with three different donors. (c) T cell-induced TNF-α production by monocytes is time-dependent. Peripheral-blood T cells (106/ml) were cultured for 48 hours in the presence or absence of immobilized anti-CD3 (3.3 μg/ml) and soluble anti-CD28 (0.8 μg/ml) antibodies. Stimulated (sTc) and resting (rTc) T cells were fixed and incubated for the indicated durations with freshly isolated monocytes (1.5 × 106/ml) at a ratio of 7:1. Values are means ± SEM for experiments with three different donors. Levels of significance: * P < 0.05, *** P < 0.001.