Fig 4.

TGEV infection triggers inflammatory responses via the NF-κB pathway in apical-out intestinal organoids. (A) Apical-out organoids were infected with TGEV for 24 and 48 h, then P-JUN, P-P65, total P65, cytoplasmic P65, nuclear P65, IκBα, and TGEV N were detected by Western blotting. (B) The density ratios of P-JUN, P-P65, total P65, cytoplasmic P65, nuclear P65, and IκBα were calculated with ImageJ. (C) TGEV-infected or mock-infected apical-out organoids at 48 h were stained with TGEV N and P65 and analyzed by confocal microscopy; scale bar: 5 µm. (D and E) Apical-out organoids were infected with TGEV followed by BAY11 (2 µM) treatment for 48 h. Subsequently, TGEV viral load (D) and mRNA levels of TNF-α, IL-8, IL-6, and IL-1β (E) were detected by RT-qPCR. Results are presented as mean ± SD of data from three independent experiments *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001, determined by two-tailed Student’s t test.