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. Author manuscript; available in PMC: 2015 May 1.
Published in final edited form as: Int J Med Microbiol. 2014 Feb 6;304(0):384–392. doi: 10.1016/j.ijmm.2014.01.002

Figure 4. IL-1β production by macrophages infected with isolated E. coli depends on the NLRP3 inflammasome.

Figure 4

(A) Bone marrow-derived macrophages from Nlrp3−/−,Asc−/− or WT mice cells were primed with LPS for 4 h. After that, cells were infected with E. coli isolated from CD (CD1-a, CD2-a, CD6-b, CD6-r, CD8-a) patients, control (C7-a) or HS strain (MOI=10) for 1 h and then treated with gentamicin. Twenty-four h.p.i, supernatants were analysed for IL-1β TNF-α and (C) IL-6 secretion by ELISA. (D) As an indicator of caspase-1 activation, content of caspase-1 p20 subunit was determined in WT or Nlrp3−/− macrophages primed with LPS for 4 h, infected with CD2-a or NRG857c strains (MOI=10) for two h and then treated with gentamicin overnight. Extracts were prepared from cell and supernatant, and immunobloted using a caspase-1 antibody. Arrows denote procaspase-1 and its processed p20 subunit. Graph bar shows content of caspase-1 p20 subunit related to procaspase-1 in each condition expressed as the fold induction in infected to non-infected cells. Graph represents mean and standard errors of three independent experiments (E) Bacterial clearance ability of Nlrp3−/− or WT macrophages infected with CD2-a or HS strains was evaluated at 3, 24 and 48 h.p.i. Graph shows the ratio of gentamicin-resistant bacteria to initial uptake (3 h.p.i). Graph represents mean and standard errors of three independent experiments, performed in duplicate. No significant differences were found in bacterial clearance between Nlrp3−/− and WT macrophages, infected with the same strain, using Fisher’s exact test with a two-sided p-value.