FIG 2.

F. tularensis infection of macrophages increases PGE₂ synthesis, COX-2 protein levels, and cPLA₂ phosphorylation. (A and B) RAW264.7 macrophages (A) or BMDMs (B) were inoculated with LVS at an MOI of 200:1. At 0, 1, 4, 6, 12, 18, and 24 h postinoculation, supernatants were collected, and PGE₂ concentrations were determined. The data are the means and SEM from three independent experiments. (C) RAW264.7 macrophages were inoculated with LVS at an MOI of 200:1. At 0, 1, 2, 4, 6, 12, 18, and 24 h postinoculation, whole-cell lysates were collected and analyzed for COX-2 protein levels by Western blotting (left). The band intensities (right) were measured in each experiment using LiCor Odyssey infrared imaging system software and were normalized to the loading control (tubulin). The graph shows the band intensities from three independent experiments. (D) BMDMs were inoculated with LVS at an MOI of 200:1. At 0, 15, 30, 60, and 120 min postinoculation whole-cell lysates were collected and analyzed for cPLA₂ phosphorylated on Ser⁵⁰⁵ by Western blotting (left). The band intensities (right) were measured in each experiment using LiCor Odyssey infrared imaging system software and were normalized to the loading control (tubulin).The graph shows the band intensities from three independent experiments. Statistical differences (*, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001) compared to the 0-h sample were determined by Dunnett's multiple comparison test.