FIG 5.

Sialostatin L2 does not inhibit caspase-1-mediated cytokine secretion during uric acid stimulation of macrophages. (A to C) BMDMs (1 × 10⁶) from WT and nox2^−/− mice were primed with LPS (100 ng/ml) overnight and stimulated with uric acid (50 μg/ml) for 4 h in the presence or absence of SL2 (5 μM). SL2 was added to the cell culture 30 min before uric acid stimulation. IL-1β (A) and IL-6 (C) were measured by ELISA. (B) IL-18 (p18) and mature caspase-1 (p20) in the supernatants of LPS-primed cells were measured by Western blotting (IB). β-Actin, pro-IL-1β, and caspase-1 were detected in the cell lysate. (D) BMDMs (1 × 10⁶) from wild-type (WT) and nox2^−/− mice were stimulated with A. phagocytophilum (MOI = 5) for 18 h in the presence or absence of sialostatin L2 (SL2; 5 μM). IL-1β secretion was measured by ELISA. Cytokine measurements were taken in triplicate and presented as means ± SEM. *, P < 0.05 by Student's t test comparing nontreated and SL2-treated cells. (−), nonstimulated cells; NS, not significant. Experiments were repeated twice. Numbers below the Western blot panels show quantitation by densitometry and indicate fold differences compared to WT, non-SL2-treated samples.