Fig. 4: MM01 inhibits ASC-dependent inflammasome activity in vitro.

A IL-1β secretion was evaluated by ELISA following activation of the NLRP3 inflammasome with LPS (100 ng/ml) and nigericin (Nig; 10 µM). Cells were treated with MM01 at 10 µM. B Measurement of LDH release into the extracellular medium under the above-described conditions. C THP-1 cells were stimulated as described above, and supernatants (SN) and pellets were analyzed by immunoblotting for IL-1β and cleaved caspase-1. A representative blot is shown. IL-1β measured by ELISA (D) and LDH release (G) were evaluated upon activation of the NLRP1 inflammasome with LPS (100 ng/ml) and MDP (50 µg/ml) in THP-1 cells. Cells were treated with MM01 at 10 µM. IL-1β secretion (E) and LDH release (H) were analyzed upon stimulation of the AIM2 inflammasome with poly (dA:dT) (200 μg/ml) in PMA-differentiated THP-1 cells treated or not with MM01 at 10 µM. IL-1β (F) and LDH secretion (I) were evaluated upon NLRC4 activation mediated by Salmonella typhimurium in THP-1 cells. Cells were treated with MM01 at 10 µM. MM01 inhibits LPS/Nig stimulation of isolated human PBMCs. IL-1β (J) and LDH (K) release were evaluated upon activation of the NLRP3 inflammasome with LPS (100 ng/ml) and nigericin (10 µM) and +/−MM01 at 10 µM in human PBMCs. L Western blot of crosslinked cytosolic pellets from PBMCs stimulated as described above. In all cases, asterisks represent significant differences to the stimulated control as determined by a one-way ANOVA with Tukey’s multiple comparisons test **p < 0.05; ***p < 0.001. All data expressed as mean ± SD of three independent experiments.