NLRP3 inflammasome activation induced by the pknF Mtb mutant is dependent on chloride efflux but independent of calcium influx.

BMDMs were either left untreated or treated with chloride channel blocker, DIDS (100μM) or with BAPTA-AM (30μM), Ca²⁺ chelator and then infected with CDC1551 Mtb wild-type and ΔpknF mutant. Culture supernatants were harvested at 20 hpi and analyzed for (A, B) IL-1β secretion by ELISA. BMDMs were either left uninfected (UI) or infected with different CDC1551 Mtb strains (Mtb, ΔpknF mutant and complement ΔpknF::pknF) at an MOI of 10 for 4h. BMDMs were primed with LPS (1μg/ml) for 4 h and stimulated with ATP (5mM, 30 min) and used as positive control for inducing NLRP3 inflammasome activation. Intracellular calcium mobilization was analyzed at (C) 4 hpi and (D) 6 hpi by Fluo-Forte Calcium Assay kit. Data are representative of three independent experiments. Error bars represent mean ± SEM; **, p<0.01, ****, p<0.0001, ns (non-significant).

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