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. 2018 Jan 22;8:2000. doi: 10.3389/fimmu.2017.02000

Figure 3.

Figure 3

Volcanic ash-induced IL-1β production is NLRP3 inflammasome dependent. Lipopolysaccharide (LPS)-primed macrophages from wild-type or knock-out mice (NLRP3−/−, ASC−/−, and Casp-1/11−/−) were stimulated with MRA5/6/99 at indicated concentrations. (A) IL-1β from supernatants (SN) was measured with ELISA. (B) Mature IL-1β (p17) inSN and pro-IL-1β (p35) from cell lysates (CL) of cells stimulated with 1 mg/ml MRA5/6/99 was assessed by western blot. β-actin IgG mAb served as loading control. (C,D) Primary human peripheral blood mononuclear cells (PBMC) were either LPS-primed or left untreated and subsequently stimulated with MRA5/6/99. (C) IL-1α and (D) IL-1β levels in SN were determined by ELISA. Insertion (D) shows the western blot of bioactive IL-1β (p17) detected from SN. (E,F) IL-1α and IL-1β in SN of ash-treated human PBMC in the presence or absence of the NLRP3 inhibitor CP-456,773, assessed by ELISA. Positive controls are poly(dAdT), as a NLRP3-independent inducer, and adenosine triphosphate (ATP) or nigericin, as NLRP3-dependent inducers of IL-1β. Representative data from two independent experiments performed as triplicates are shown. (A) *p ≤ 0.05, compared to NLRP3−/−, ASC−/−, Caspase-1−/−; #p ≤ 0.05, compared to NLRP3−/− and ASC−/−; (C,D) *p ≤ 0.05, compared to none (Ø); (E,F) *p ≤ 0.05, compared to inhibitor (CP-456,773).