Figure EV3. VbP triggers CARD8‐dependent pyroptosis in human myeloid cells.

• A
  Immunoblotting of clonal THP‐1 knockout cell lines of the indicated genotype. * indicates a band at the size of the cleaved CARD8 c‐terminal fragment that was still detectable in the CARD8 knockout cell lines, either indicating unspecific banding or the existence of an isoform that was not affected by the gene targeting approach. One representative blot of three is shown.
• B, C
  THP‐1 cells of the indicated genotype treated with either Pam3CSK4 alone (Pam3, 24 h), Pam3CSK4 (1 h priming) and Val‐boroPro (VbP 23 h), or Pam3CSK4 (4 h priming) and Nigericin (Nig., 4 h). LDH‐activity (B) and IL‐1β concentration (C) in the supernatant were determined by LDH cytotoxicity assay and ELISA, respectively. Individual data points ± SEM from five independent experiments are shown. Statistics indicate significance by two‐way ANOVA: ***P ≤ 0.001; *P ≤ 0.05; ns, not significant. P‐values were corrected for multiple comparisons (Dunnett).