Figure 1: Squamous epithelium contributes to wound healing in colitis.

A-B) Surface representations after 3D imaging of chemically cleared, flattened colonic tissue after DSS injury reveal a distinct epithelium replacing colonic crypts, proximal to the dentate line (orange) but distal to the margin (green) of colonic crypts. C-D) H&E images show that this long-lasting “neo-epithelium” (C) has a broadly squamous structure with rete pegs (*) but lacks a fully differentiated surface (#). The neo-epithelium does not exist in the absence of injury (D). E) Squamous neo-epithelium of colon (SNEC) persists as a KRT14+ tissue structure 15 months after injury (n=3 mice), as shown in whole-mount images. F) Plot of SNEC size (length of incursion from the dentate line) associated with multiple cycles of DSS (n=5–7 mice/condition). G) 3D reconstruction of SNEC surface and abluminal views shows distinct morphology compared to colonic crypts. H) Immunostaining reveals high expression of P63 and SOX2 in squamous cells at the dentate line and in rete pegs, but exclusion from colonic epithelium (n=4 mice). Orange arrows: dentate line. I-K) Apoptotic targeting in Sox2-DTA mice with 4-hydroxytamoxifen (4-OHT) enemas retards body weight recovery (I), leaving ulcers (J,K) in the colon after DSS colitis (n=5 mice/condition, t-test). L) Retrospective analysis of squamous epithelium in colons of Il10−/− mice. H&E photomicrographs show that young (8 wks-old) Il10−/− mice lacking inflammation were devoid of SNEC. Orange arrow marks the dentate line. M) However, older mice (>36 wks-old) with severe distal colitis and buried columnar dysplasia exhibit SNEC. N) Colitis (as defined by histological injury score >0) was a prerequisite for finding SNEC. Each dot represents a mouse. O) Linear regression of animal age and histological colitis score modeled the size of the SNEC tissue. Error bars: SE. See also Figures S1–S3.