FIGURE 6.

Extracellular Ca²⁺ is required for Tat_HXB2-induced IL-10 production from monocytes. Monocytes were pretreated with 10 nm FS2, 1 μm nimodipine (both Ca_L inhibitors), 1 μm xestospongin C, 1 μm TMB-8 (inhibitors of intracellular calcium stores stimulated by inositol 1,4,5-triphosphate and ryanodine, respectively), 2 mm ethylenediaminetetraacetic acid (EDTA) or vehicle control or placed in Ca²⁺-free media for 1 h prior to incubation with 10 nm Tat_HXB2, 2 μm Tat_1–45, or 10 nm Tat_93In for 4 h, after which cells were washed and incubated in fresh complete media for a further 20 h, and supernatants were collected and analyzed for IL-10. The Ca_L inhibitors, the Ca²⁺ chelator EDTA, and incubation in Ca²⁺-free media all abrogated the IL-10 response elicited by Tat_HXB2, Tat_93In, and Tat_1–45, whereas the inhibitors of Ca²⁺ from inositol 1,4,5-triphosphate-regulated stores or caffeine-sensitive ryanodine receptor-regulated intracellular stores had no effect. The bars represent means ± S.E. from three independent experiments carried out in triplicate.