Buskens, C. J. | bioRxiv Science

bioRxiv2026

UNRAVELLING A HIDDEN SUBTYPE: MULTIOMICS REVEAL PSORIASIS-LIKE SIGNATURE WITH SURGICAL RELEVANCE IN A SUBSET OF PERIANAL FISTULIZING CROHN'S DISEASE

Background and aimsPerianal fistulizing Crohns disease (pCD) affects 20% of patients with Crohns disease (CD) and severely impacts quality of life. Current therapies fail to provide sustained relief, and the molecular underpinnings of pCD remain poorly understood. This study aimed to elucidate the molecular landscape of perianal fistulas through multiomic profiling. MethodsPaired fistula-tract and adjacent rectal mucosal biopsies were collected from 63 patients (48 with pCD, 15 with cryptoglandular fistula [CPTGL]). Longitudinal sampling generated 101 unique molecular profiles, comprising of both RNA sequencing (RNA-seq) and 16S rRNA sequencing from both tissue sites, followed by integrative multiomics and gene network analyses. ResultsUnsupervised clustering revealed three patient clusters primarily defined by host gene expression, with minimal contribution from microbial profiles. Fistulae in clusters 1 and 2 showed strong immune activation and epithelial-mesenchymal transition (EMT). In contrast, cluster 3 fistulae displayed keratinization and metabolic reprogramming resembling psoriatic skin, together with reduced JAK-STAT signalling. Cluster 3 was enriched for patients classified as TOpClass:2a, who are more suitable for surgical repair (p = 0.02). Conversely, cluster 2, characterized by rectal keratinization and EMT in both fistula and rectum, showed the highest MRI inflammatory-mass score (p = 0.02) and a greater risk of subsequent ileostomy (Kaplan-Meier; p = 0.008). An independent RNAseq dataset validated the keratinization signature in a subset of pCD fistulae. ConclusionIntegrated multiomic analysis identified distinct molecular subtypes of pCD with surgical and therapeutic relevance. These findings refine the molecular understanding of pCD and support a precision medicine approach. What You Need to Know?O_ST_ABSBACKGROUND AND CONTEXTC_ST_ABSO_LIPerianal fistulas affect 1 in 5 Crohns disease patients and have a severe negative impact on the quality of life of the patients. C_LIO_LIMost advanced IBD therapies are not efficacious for perianal Crohns disease. C_LIO_LISurgical repair is not suitable for all patients. C_LI NEW FINDINGSO_LIThis study is the first to delineate molecularly defined patient subtypes in perianal fistulizing Crohns disease. It further characterizes a psoriasis-like keratinization program in a subset of fistulas. Finally, it identifies molecular features and histological indicators that may explain-and potentially help predict-favorable surgical outcomes in selected patients C_LI LIMITATIONSO_LIFunctional and mechanistic studies will be required to further dissect the drivers and dynamics of the epithelial remodeling identified. C_LI CLINICAL RESEARCH RELEVANCEO_LIRaises the possibility of refining of existing clinical stratification using molecular markers for improved therapeutic management and outcome. C_LI BASIC RESEARCH RELEVANCEO_LIProposes keratinization as an opposing molecular process to EMT within the perianal fistula tract. C_LIO_LIIdentifies robust gene signatures associated with EMT and keratinization in the fistula for further experimental and clinical studies. C_LI

immunology↗

bioRxiv2025

Nuclear Calprotectin mediates Epithelial Wound Healing Defects in Crohns Disease related Fistula

Wound healing is critical to homeostasis in particular in tissues exposed to environmental insults such as the skin and the intestine. At the intersection of these tissues a particular form of wounding occurs in the form of perianal fistula, i.e. abnormal tracts connecting the intestine with the skin. Importantly, a natural disparity is found in healing efficiency in perianal fistula depending on the underlying medical condition, which is currently not well understood. Using this disparity, we modelled appropriate and defective wound healing by comparing Crohns disease related fistula (poor healing) and non-IBD/idiopathic fistula (well healing). We show that during wound healing, cytokines TNF and IL6 in conjunction with TGF-{beta} induce differentiation of columnar intestinal mucosa into squamous epithelium with high expression of keratins 5 and 13 and WFDC2. Specifically in poorly healing wounds, cytokines including IL17, IL22 and IFN{gamma} induced expression of S100A8/9 in this intestinal derived tissue. Surprisingly, S100A8/9 was not released but rather functioned as a transcriptional (co)regulator, affecting downstream inflammatory mediators such as C3 and CXCL17. These data identify separate immune pathways contributing to healing in general and deficient healing, allowing for more targeted intervention approaches.

cell biology↗

bioRxiv2024

Tuft cells act as regenerative stem cells in the human intestine

In mice, intestinal tuft cells have been described as a long-lived, post-mitotic cell type of which two distinct subsets have been identified, named tuft-1 and tuft-21. By combining analysis of primary human intestinal resection material and intestinal organoids, we identify four distinct human tuft cell states, two of which overlap with their murine counterparts. We show that tuft cell development depends on the presence of Wnt ligands, and that tuft cell numbers rapidly increase upon interleukin (IL)-4 and IL-13 exposure, as reported previously in mouse2-4. This occurs through proliferation of pre-existing tuft cells, rather than through increased de novo generation from stem cells. Indeed, proliferative tuft cells occur in vivo both in fetal and in adult human intestine. Single mature proliferating tuft cells can form organoids that contain all intestinal epithelial cell types. Unlike stem- and progenitor cells, human tuft cells survive irradiation damage and retain the ability to generate all other epithelial cell types. Accordingly, organoids engineered to lack tuft cells fail to recover from radiation-induced damage. Thus, tuft cells represent a damage-induced reserve intestinal stem cell pool in humans.

cell biology↗