1. Notch levels in niche cells
Notch levels in niche cells Left: Cross‐sectional view of murine intestinal crypt bottoms with co‐immunofluorescence (co‐IF) showing intermingled LGR5‐EGFP+ (green) CBCs and lysozyme+ (LYZ, red) Paneth cells. Scale bar: 50 μm. Right: Schematic illustration of a niche pattern in both longitudinal and cross‐sectional views of a crypt.RT–qPCR quantification of Notch signaling components in CBC and Paneth cell populations. The experiment was performed in triplicate and presented mean ± SEM (***P ≤ 0.001, **P ≤ 0.01, *P ≤ 0.05; Student's t‐test).Representative FACS plots of organoids treated with DMSO, JAG1 (embedded in Matrigel), EDTA or DAPT for 48 h, including gated analysis to isolate CD24high/SSChigh Paneth cells and LGR5‐EGFP+ CBCs according to an established protocol (Sato et al, 2011c).RT–qPCR quantification of Notch signaling components in CBCs and Paneth cell populations after organoids were treated with Matrigel‐embedded JAG1 (top), EDTA (middle), or DAPT (bottom). The experiments were performed in triplicate and presented mean ± SEM (**P ≤ 0.01, *P ≤ 0.05; Student's t‐test).Western blot analysis of Notch signaling components from conditions described in (D). Actin was used as a loading control.Schematic illustration of lateral inhibition and positive feedback between neighboring cells. Transparent colors and dotted lines represent low expression/activity levels. Source data are available online for this figure.
Kai‐Yuan Chen et al. Mol Syst Biol 2017;13:927
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