Figure 3.

LPS-stimulated monocytes release ATP that can affect T cells. A, LPS-stimulated monocytes release excessive ATP. PBMCs were stained with anti-CD4-allophycocyanin antibodies. Cells were allowed to attach to fibronectin-coated glass-bottom coverslip dishes. Then they were labeled with the fluorescent membrane–anchoring ATP probe 2-2Zn. Immune synapse formation between monocytes and CD4 T cells was initiated by stimulation with anti-CD3 antibodies, and ATP release in response to LPS (10 ng/ml) was observed with live-cell fluorescence microscopy. Representative images of n = 7–10 T cell/monocyte conjugates derived from three different experiments are shown; ×100 objective (NA 1.4). B, PBMCs release more ATP than monocytes in response to LPS. Purified human monocytes or PBMC cultures were stimulated with LPS (10 ng/ml), and ATP concentrations in the cell supernatant were measured at the indicated times. Data are means ± S.D. (error bars) of n = 4 (monocytes) or 6 (PBMCs) experiments. * and #, p < 0.05 versus no LPS controls, one-way ANOVA.