DOP38 A rare KRT17+ population emerges from the epithelium in the inflamed intestinal mucosa of patients with Ulcerative Colitis

Abstract

Abstract Background KRT17 is an epithelial "alarmin" that contributes to the early host innate immune response to insults. We previously found that KRT17 transcription, not detectable in the healthy colon, is up-regulated in the intestinal mucosa of patients with active ulcerative colitis (UC). Here, we aimed to characterize the epithelial cells expressing KRT17 in UC mucosa and explore the contribution of the intestinal environment in promoting the emergence of this cell population. Methods We conducted single-cell RNAseq (scRNA-seq) on the epithelial compartment of intestinal biopsies from a cohort of non-inflammatory bowel disease (IBD) controls and patients with active UC. Analysis was performed using our own-curated pipeline in Seurat1. Spatial protein localization was analysed by immunofluorescence on fixed intestinal samples. To measure the modulation of KRT17 and associated markers, ex vivo air-liquid interface (ALI) epithelial cultures derived from human non-IBD organoids were used. The signatures of UC-derived organoids2 were mapped onto the scRNA-seq dataset. Results ScRNA-seq analysis identified a rare KRT17+ epithelial cell population emerging in active UC characterized by the expression of KRT17 (Fig. 1A) and other alarmins (KRT6A, KRT16, S100A8), metalloproteinases (MMP1, MMP10), and inflammatory markers (SAA1, DUOX2…). KRT17 protein, absent in non-IBD colon, was expressed in the epithelium of the inflamed UC mucosa exhibiting a patchy pattern, and marked epithelial clusters with both columnar and squamous-like morphology (Fig. 1B). The ALI culture closely recapitulated the differentiated intestinal epithelium in homeostasis by showing a columnar aspect, high expression of differentiation markers, and low or undetectable expression of stem/proliferation or KRT17+-cell markers. Exposure of the ALI culture to hypoxic stress (by re-submersion in expansion medium) induced the expression of KRT17+ cell-specific and inflammation markers (Fig. 1C), and the acquisition of a squamous morphology (Fig. 1D). IBD-related IFN-ɣ and IL-22 cytokines could not fully replicate this phenotype (Fig. 1E). Lastly, we observed that genes up-regulated in UC, compared to non-IBD organoids, were predominantly expressed within the KRT17+ population (Fig. 1F). Conclusion Our results suggest that a novel KRT17+ epithelial population may arise from the intestinal cells of inflamed UC mucosa under the influence of specific environmental factors. This population could contribute to the persistence of the disease over time.