Figure 1.
Repairing Intestinal Epithelium Has a Cellular and Molecular Profile Distinct from the Normal Epithelium
(A) Macroscopic images of the distal part of the colon in an untreated mouse (left) and a mouse 2 weeks post-administration of DSS (right). The demarcated area indicates regions undergoing active re-epithelialization. Scale bar, 2 mm.
(B) H&E staining of homeostatic tissue and tissue in the repair phase at 2 weeks following DSS administration. Scale bar, 100 μm.
(C) The mucosal/submucosal thickness at homeostasis and repair phase (2 weeks). Shown are mean distances ± SEM (n = 4 animals; p = 0.007 based on two-sided Student’s t test).
(D) Sca1 (green) expression in colonic epithelium in homeostatic and repair phase. Sections are counterstained with DAPI (blue). The demarcated line indicates the epithelial structure. Scale bar, 100 μm.
(E) Flow cytometric analysis of cells from the colonic epithelium during homeostasis and in repair phase (2 weeks post-administration of DSS). Diagrams show representative plots for Sca1-PECy5 in live CD45−veCD31−veEpcam+ve cell population.
(F) Quantification of the percentages of Sca1high cells in homeostasis and repair phase. Diagram shows average ± SEM (n = 3; p = 4 × 10−5 based on two-sided Student’s t test).
(G) qRT-PCR analyses in Epcam+Sca1high cells sorted via FACS from the repairing epithelium and in homeostatic epithelial cells (Epcam+). Bars represent average levels ± SEM (n = 3; Ly6a, p = 1.3 × 10−4; Reg3b, p = 9.7 × 10−3; Reg3g, p = 8.5 × 10−3 based on two-sided Student’s t test).
(H) Heatmap analysis of differentially expressed probe sets (fold change > 2.0; FDR < 0.05), comparing the expression profile of epithelial cells isolated from homeostatic tissue and Sca1high cells from the repairing epithelium.
(I) Detection of Lrig1 (green) in homeostasis, early and late repair phase counterstained for E-cadherin (red) with DAPI (blue). Scale bar, 50 μm.
(J) GSEA of repairing and homeostatic epithelium using Lgr5 intestinal stem cell gene signature.
(K) Organoid formation for Epcam+ cells isolated from normal homeostasis, as well as Epcam+ Sca1low and Sca1high cells during tissue repair showing representative images of formed organoids. Bars represent average number of organoids formed ± SEM (n = 3 animals).