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. Author manuscript; available in PMC: 2016 Feb 15.
Published in final edited form as: J Immunol. 2015 Jan 21;194(4):1763–1775. doi: 10.4049/jimmunol.1401624

Figure 7. Pneumolysin-induced caspase-1 activation and IL-1β secretion by neutrophils requires K+ efflux.

Figure 7

A. Human peripheral blood neutrophils were hkSP primed with hkSP for 3h and stimulated with Ply or PdB for 2h. Total cell contents were extracted using 10% HNO3, and the cell associated K+ concentration was quantified by atomic absorbance spectroscopy. B. FLICA660-YVAD positive murine neutrophils quantified by flow cytometry after treatment with live TIGR4 in presence of increasing concentration of extracellular KCl.; C. hkSP primed neutrophils were stimulated 2h with either Ply (500ng/ml) or WT TIGR4 (50:1) in the presence of 5 mM or 130 mM KCl, and IL-1β secretion was measured. D–F. Neutrophils were primed and stimulated with Ply (500ng/ml), or were stimulated with Nigericin in the presence of the pan-caspase inhibitor ZVAD or the caspase-1 inhibitor YVAD at the indicated concentration (μM), and examined for IL-1β secretion after 2hr (D), intracellular K+ (E) and LDH (F). Histograms are mean ±SD of three samples per treatment condition, and are representative of two similar experiments with neutrophils from different donors. ** p< 0.001, *** p< 0.0001.