Fig. 3

LLOME and particle-induced lysosomal rupture causes the release of active CTSS from macrophages. a CTSS activity in lysates measured as increased RFU, produced from the proteolytic cleavage of the reporter substrate, Z-VVR-AMC (50 μM). Lysates were generated from BMDMs which were left untreated or treated with LLOME (0.5 mM) for the indicated times. Ctss ^−/− cells were used as a control to determine background levels of substrate turnover not due to CTSS, n = 3 +/− standard error. b Extracellular turnover of Z-VVR-AMC (50 μM) by CTSS in supernatants harvested from BMDMs treated with LLOME (0.5 mM) for 0.5-2 h, n = 4 +/− standard error. c Extracellular turnover of Z-VVR-AMC (50 μM) by CTSS in supernatants harvested from BMDMs treated with LLOME (0. 5 mM) for 16 h, n = 3 +/− standard error. d Cell supernatants were incubated with biotin-PEG-LVG-DMK (10 uM) to label secreted mature CTSS from BMDMs treated with LLOME (0.5 mM) for 6 and 16 h. Supernatants from ctss ^−/− cells which had also been treated with LLOME (0.5 mM) for 16 h were used as a control to confirm the identity of labeled CTSS. Blot is representative of 3 independent experiments. e Extracellular turnover of Z-VVR-AMC (50 μM) by CTSS in supernatants harvested from BMDMs treated for 4 h with alum (500 μg/mL), silica (250 μg/mL) or PSNPs (500 μg/mL), n = 3 +/− standard error