Fig. 5.

IL-1β processing in macrophages is partially dependent on CTSS. ELISA analysis of IL-1β release in supernatants from: a Wildtype and ctss ^−/− LPS-primed peritoneal macrophages and (b) Wildtype and ctss ^−/− LPS-primed BMDMs, which were pre-treated with Z-VAD-FMK (5uM) for 1 h prior to stimulation with LLOME (0.5 mM) for 16 h, n = 3 +/− standard error. c Caspase 1 activity measured in lysates generated from wildtype and ctss ^−/− BMDMs, untreated or treated with LLOME (0.5 mM) for 2 h. Activity was measured by RFU generated from caspase 1 mediated hydrolysis of Z-YVAD-AMC (50 μM). Data presented as increased RFU in lysates from treated cells relative to untreated control, n = 5 +/− error. d Confocal microscopy of active caspase in wildtype (WT) and ctss ^−/− peritoneal macrophages, treated for 2 h with LLOME (0.5 mM). Cells were incubated for 1 h with FAM-YVAD-FMK prior to washing, fixing and nuclear staining. Top panel: FLICA probe (green) + dapi (blue). Bottom panel: FLICA probe only (green). Images representative of 3 independent experiments, scale bar = 40 μM. e Western blot of the pro-form and active form of IL-1β in lysates generated from wildtype and ctss ^−/− LPS-primed BMDMs treated with LLOME (0.5 mM) for 5 h, before being harvested, lysed and blotted for IL-1β