Fig. 5: Induction of high IL-1β by M. tuberculosis 4I2 results from enhanced inflammasome activation.

a Gene expression heatmap representing cytokine expression for non-infected BMDMs (NI, yellow) and BMDMs infected with M. tuberculosis isolate 4I2 (dark red) or M. tuberculosis isolate 6C4 (blue). z-scores are color coded as shown. b C57BL/6 WT BMDMs were generated and infected with M. tuberculosis isolates 4I2 (solid circles) or 6C4 (open squares), and at the indicated time points, the cell cultures were lysed, RNA extracted, converted to cDNA and subjected to real-time PCR. c C57BL/6 WT or TLR2 deficient BMDMs were generated and infected with M. tuberculosis isolates 4I2 (solid circles) or 6C4 (open squares). Twenty-four hours post-infection the culture supernatants were harvested and the amount of IL-1β quantified by ELISA. d THP-1-ASC-GFP monocytes were PMA-differentiated and left uninfected (NI) or infected with M. tuberculosis isolates 4I2 (solid bars) or 6C4 (open bars) at different MOI, as indicated. Six hours post-infection, cells were fixed and stained with DAPI prior to image acquisition. The percentage of Speck-positive cells was determined for the different conditions. C57BL/6 WT BMDMs were generated and infected with M. tuberculosis isolates 4I2 (e, f) or 6C4 (g, h) in the absence (0) or presence of increasing doses of NLRP3 (MCC950; e, g) or Caspase-1 (VX-765; f, h) inhibitors. BMDMs were infected with increasing MOI of M. tuberculosis isolate 6C4 (I), with a combination of both isolates at different doses (j) or with M. tuberculosis isolate 6C4 in the absence (6C4) or presence (6C4 + ATP) of 5 mM of ATP (k). e–k Twenty-four hours post-infection the culture supernatants were harvested and the amount of IL-1β quantified by ELISA. Unless indicated, an MOI of 2 was used for infections. Represented is the mean ± SEM of triplicate wells for one experiment representative of two (b–j); or for one experiment (k). Statistical analysis was performed using two-tailed unpaired Student’s t-test (c, k) or One-way ANOVA (d–i) (* p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001).