190 Mucosal gene expression reflects tissue specificity, inflammatory status and disease subtypes Principal component analysis (PCA) showed that gene transcriptional patterns could be stratified by biopsy location (ileum vs. colon), inflammatory status (non-inflamed vs. inflamed) and IBD subtype (CD vs. UC) in the first two components (Figure 2A), consistent with previous observations.13 Tissue location and inflammatory status were significantly associated with the first two PCs (biopsy location, ileum vs. colon: PWilcoxon=2.87x10-12; biopsy inflammatory status, P=7.15x10-27), whereas disease/control status (CD vs. UC vs. controls) was associated with the second PC (P=2.14x10-16). Inflammation-associated gene expression showed overlap between inflamed biopsies from ileal CD, colonic CD and UC (Figure 2B). Differential expression analyses between non-IBD controls, non-inflamed and inflamed biopsies in all these three groups revealed 3,157, 3,486, and 6,710 differentially expressed genes (DEGs), respectively (FDR<0.05) (Supplementary Table S1). These DEGs fall mainly within interleukin signaling, neutrophil degranulation and extracellular matrix (ECM) organization pathways (FDRFisher<0.05, Extended Data Figure S1). Overlapping results from all three differential expression analyses identified 1437 shared DEGs, including DUOX2, MUC1, JAK2, OSM and IL17A (Figure 2C). We also observed an enrichment of these DEGs in IBD- associated genomic loci (PFisher=9.6x10-9).2 Figure 2 | Mucosal host gene expression patterns in intestinal tissue from patients with IBD and controls. a, Principal component analysis, labeled by tissue location (ileum/colon), inflammatory status (noninflamed/inflamed) and disease diagnosis (control/CD/UC), shows that variation in host gene expression can be significantly explained by tissue location and inflammatory status. b, Venn diagram of inflammationassociated genes from three comparisons: 1) ileal tissue from controls vs. non-inflamed tissue from patients with CD vs. inflamed tissue from patients with CD, 2) colonic tissue from controls vs. non-inflamed tissue from patients with CD vs. inflamed tissue from patients with CD and 3) colonic tissue from controls vs. noninflamed tissue from patients with UC vs. inflamed tissue from patients with UC (all FDR <0.05). c, Relevant examples of four inflammation-associated genes, DUOX2, JAK2, MUC1 and IL17A, illustrating the presence of tissue inflammation (FDR <0.05). d, Relevant examples of inflammation-associated genes differentially expressed between patients with CD and UC (keeping tissue location and inflammatory status constant) showing higher expression of HDAC9 (histone deacetylase 9) and HEY1 (hairy/enhancer-of-split related with YRPW motif protein 1) in patients with CD and higher expression of VNN1 (pantetheinase) and APOB (apolipoprotein B) in patients with UC. e, Analysis of pathways associated with either the presence of CD (orange) or UC (purple) demonstrates that genes upregulated in CD are mainly associated with Notch-1 signaling, whereas pathways upregulated in UC are mainly related to vitamin and cofactor metabolism, SLC-mediated transmembrane transport and intracellular protein modification. Pathways were annotated using the Reactome pathway database. CDi, inflamed tissue from patients with Crohn’s disease. CD-non, non-inflamed tissue from patients with Crohn’s disease. FDR, false discovery rate. PC, principal component. UCi, inflamed tissue from patients with ulcerative colitis. UC-non, non-inflamed tissue from patients with ulcerative colitis. Chapter 6
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