Skip to main content
. 2019 Mar;189(3):604–618. doi: 10.1016/j.ajpath.2018.11.015

Figure 1.

Figure 1

NF-κB is activated in the intestine of mice within 12 hours of exposure to a necrotizing enterocolitis (NEC) protocol, and myeloid cells express NF-κB–target genes on NEC induction. A: Less than 12-hour–old heterozygous NF-κB–reporter mice were subjected to NEC protocol and imaged every 12 hours with an IVIS camera. Luciferin (0.1 mg, intraperitoneally) was administered 10 minutes before each image acquisition. Top panel: Representative bioluminescence images from two mice obtained at time 0 (T0) and then every 12 hours during exposure to the NEC protocol are shown. Bottom panel: Abdominal area bioluminescence images were quantified, and results are presented as count-fold increase over baseline. Three to five pups were used per time point. B: Bioluminescence of NEC or dam-fed (DF) control intestines obtained at 24 and 48 hours. DF and NEC pups were euthanized simultaneously, and intestines were immediately dissected by two different investigators and placed on a petri dish to be immediately imaged. White arrows indicate the small intestine and the colon. C: Heterozygous NF-κB–reporter pups were placed on NEC protocol or allowed to be DF for different time periods, and luciferase activity of homogenized small-intestinal tissues was measured by in vitro assay. Luciferase activity normalized to protein concentration [relative light units (RLU)/μg] is presented. D: Percentage of specific leukocyte subsets from intestinal lamina propria of 48-hour DF control (open bars) and NEC pups (closed bars) determined by flow cytometry. Cells were gated on live, single, CD45+ cells. Cd11c+ refers to Cd11c+MHC-II+. Cd11b+ refers to Cd11b+Ly6g−. E: Absolute counts determined by flow cytometry. Gates were made on live, single cells and evaluated for CD45 expression or further gated on CD45+Ly6g−SSClo Cd11b+ cells. F: Inflammatory gene expression of isolated Cd11b+ cells from 12- to 24-hour NEC (closed bars) or control (open bars) pups, as measured by real-time PCR. G: Inflammatory gene expression of isolated Cd11b+ cells from either 48-hour NEC (closed bars) or control (open bars) pups, as measured by real-time PCR. Data are expressed as means ± SEM. n = 3 per group (D); n = 6 per group (F); n = 5 per group (G). ∗P ≤ 0.05, ∗∗P ≤ 0.01, and ∗∗∗P ≤ 0.001. Scale bar = 1 cm (B). D0, day 0.