Mycobacterium abscessus infection leads to enhanced production of type 1 interferon and NLRP3 inflammasome activation in murine macrophages via mitochondrial oxidative stress
Fig 3
MAB-R-induced mtROS increased IFN-β and IL-1β production via activation of IRF-3 and the NLRP3 inflammasome.
(A) BMDMs or J774A.1 cells were infected with strains of MAB-R, MAB-S or M. smegmatis (Msm) at an M.O.I. of 10 for 24 h. The supernatants of the infected cells were collected, and IL-1β levels were measured by ELISA. (B) Supernatants from J774A.1 cells infected with live or heat-killed (HK) strains of MAB-R, MAB-S or M. smegmatis (Msm) at an M.O.I. of 10 for 24 h were collected, and the levels of IL-1β were measured by ELISA. (C) J774A.1 cells were pre-treated with or without mito-TEMPO (100 μM) and infected with strains of MAB-R, MAB-S or M. smegmatis (Msm) at an M.O.I. of 10 for 24 h. IL-1β and IFN-β cytokine levels in the supernatants of infected cells were analysed by ELISA. (D) Representative immunoblot images and quantitative bar graphs of NLRP3 (110 kDa), IL-1β (pro form, 35 kDa; active form, 17 kDa), p-IRF3 (44~55 kDa), IRF3 (44~55 kDa) and caspase-1 (Casp1; pro form, 45 kDa; active form, 10 kDa) induced by the indicated mycobacteria (M.O.I. of 10) infection for 4 or 24 h in BMDMs pre-treated with or without mito-TEMPO. The expression levels of IL-1β, NLRP3 and caspase-1 were normalized to that of GAPDH. Error bars represent the SD. Statistical significance was determined by ANOVA with Tukey's multiple comparison test (A) and two-tailed Student’s t-test (C and D).