TLR4⁺ immune cells induce epithelial NF-κB signaling in the cecal mucosa upon LPS exposure. Mice were i.v. injected with LPS. Cecal explants were imaged at 1 h.p.inj. by two-photon microscopy. (A–C) Representative images and quantification of epithelial NF-κB activation in the indicated mice (A, n = 4–7) or BMCs (B, n = 5; C, n = 5). Each circle represents one mouse. Black line: median. **, P ≤ 0.01 by Mann–Whitney U test. (D) Small intestinal organoids were treated with 5, 50, or 500 ng/ml LPS and imaged for ∼1 h. Representative images of one organoid over time (top) and quantification of NF-κB activation (bottom; relative change). Each circle represents one organoid at the given time (minutes after start of the treatment, n = 7). Lines connect data points from the same organoid. Red dashed line: 50% activation threshold. Black dotted line: no change. Scale bars: 50 µm. Combined data of two (C and D), three (A), or four (B) independent experiments.