Fig. 3.

PrP fibrils activate caspase-1. Fluorescent inhibitor of caspase-1 was used to detect caspase-1 activation by PrP fibrils by confocal microscopy (a) or by flow cytometry (b). a LPS-primed immortalized macrophages were treated with fibrils (10 and 25 μM). Scale bar 25 μm. b Histogram of fibril-treated (10 μM) (full line), primed-only (dash/dotted line) and untreated macrophages (dotted line). The histogram of unlabeled cells (without fluorescent inhibitor of caspase-1) is shown in gray. c Cleaved caspase-1 (p10) was detected in cell culture media of macrophages stimulated by PrP fibrils and ATP (5 mM). No p10 is detected in untreated or primed only samples. d IL-1β processing induced by PrP fibrils (20, 10, 7, 5, and 3 μM of monomeric PrP in the fibrillar form) in immortalized microglia, deficient in caspase-1 and IL-1R and corresponding wild-type microglia was followed by IL-1β ELISA. Secreted IL-1β concentration was corrected for background (priming only). e IL-1β maturation by PrP fibrils (8.5 μM) in macrophages is decreased upon addition of caspase-1 inhibitor Z-YVAD-FMK. Representative examples of 3 (a), 2 (b), 3 (c, d, e) independent experiments are shown. Error bars SD of duplicate (d) and triplicate (e) wells