Fig. 5

TUDCA inhibits ER stress induced by B. pseudomallei. a Effect of TUDCA on the cell viability. RAW264.7 cells were treated with TUDCA (0, 20, 100, 200, and 500 μM) for 24 h. b The effect of TUDCA on B. pseudomallei culture growth. B. pseudomallei were treated with 100 μM TUDCA for different time points. At respective time points optical density (OD) was measured at 600 nm. c Representative TEM images of ER in RAW264.7 cells. Cells were treated with TUDCA (100 μM), 4-PBA (250 μM) or PBS for 24 h, and infected with B. pseudomallei (MOI = 10) for 8 h. White arrowheads indicate the B. pseudomallei, and black arrowheads indicate the ER. d Immunofluorescence assay of calreticulin in RAW264.7 cells. Cells were treated as described above and stained with anti-calreticulin antibody (red), anti-B. pseudomallei antibody (green) or DAPI (blue). Scale bar is 10 μm. e Percentage of cells with fragmented calreticulin signal area, and the area of calreticulin as a function of the total cell area, were calculated in TUDCA, 4-PBA and PBS-treated cells (n > 50 cells per experiment). f The average of intracellular B. pseudomallei in each cell were counted. g The mRNA expression of Bip, CHOP in RAW264.7 cells infected with B. pseudomallei (MOI = 10) for 8 h. h Immunoblot analysis of Bip, CHOP, elF2α and p-elF2α levels after B. pseudomallei infection (MOI = 10) for 8 h. i Cells were infected with 10 MOI for 8 h, 2.5% agarose gel image shows unspliced and spliced (XBP1(u) and XBP1(s)) mRNA species. The full-length blots or gels are presented in Supplementary Fig. S4 and S5. Results are representative of 3 independent experiments. *P < 0.05, **P < 0.01