Abstract Background Perianal fistulizing Crohn’s disease (PFCD) involves breakdown of mucosal surfaces and tunneling with transitions of epithelium from the internal opening through stroma and to the perianal skin. EMT is widely-accepted to be involved in the process of fistulization and has most commonly been studied in rectal epithelium. However, due to challenges in sampling these often millimeter-wide tracts, the identity of perianal fistula tract epithelium in patients with Crohn’s disease remains incompletely characterized. Methods We screened 130 proctectomies performed at our institution for patients with PFCD from 2009 through 2021. Of these, 81 proctectomies had formalin-fixed paraffin-embedded tissue available stored by our pathology department. Sections were scored for presence of fistula, quality of fistula tract sections (i.e. with the least fragmentation), and presence of internal and/or external openings. We performed spatial transcriptomics (Visium 10X) on formalin-fixed paraffin-embedded sections from three representative patients with PFCD. Space-ranger output files, including the H5 file and relevant sequencing outputs, were used to compile objects in R software according to the Seurat spatial workflow. Data was normalized using SC-Transform before UMAP clustering and PCA analysis. Anatomy was carefully reviewed and regions containing epithelia were selected for using the 10X Genomics Loupe browser v.4.0. The barcodes of the aggregate areas and non-aggregate areas were extracted as csv files and used for comparison by the SetIdent function in Seurat. Results We annotated fistula tract lining, anal epithelium, rectal epithelium, and perianal skin in proctectomy specimens of patients with inflamed and non-inflamed perianal fistula tracts (Figure 1B and 1D). Fistula tract epithelium clusters with anal squamous epithelium (Figure 1A). Rectal epithelium, on the other hand, was transcriptionally distant (Figure 1A). Anal transitional zone keratin KRT13 is upregulated in perianal fistula tract lining, whereas rectal keratin KRT8 was not present (Figure 1C). This was true for both inflamed and non-inflamed tracts. Conclusion Our data show that the epithelium of perianal fistula tracts are more closely related anal squamous epithelium than rectal columnar cells. These findings suggest that, in addition to rectal transition, anal squamous cell transition may play a role in perianal fistula pathogenesis. References: 1. Bataille F, Klebl F, Rümmele P, Schroeder J, Farkas S, Wild PJ, Fürst A, Hofstädter F, Schölmerich J, Herfarth H, Rogler G. Morphological characterisation of Crohn’s disease fistulae. Gut. 2004 Sep;53(9):1314-21. doi: 10.1136/gut.2003.038208. Erratum in: Gut. 2004 Nov;53(11):1722. PMID: 15306592. 2. Bataille F, Rohrmeier C, Bates R, Weber A, Rieder F, Brenmoehl J, Strauch U, Farkas S, Fürst A, Hofstädter F, Schölmerich J, Herfarth H, Rogler G. Evidence for a role of epithelial mesenchymal transition during pathogenesis of fistulae in Crohn’s disease. Inflamm Bowel Dis. 2008 Nov;14(11):1514-27. doi: 10.1002/ibd.20590. PMID: 18626977. 3. Rizzo G, Rubbino F, Elangovan S, Sammarco G, Lovisa S, Restelli S, Pineda Chavez SE, Massimino L, Lamparelli L, Paulis M, Maroli A, Roda G, Shalaby M, Carvello M, Foppa C, Drummond SP, Spaggiari P, Ungaro F, Spinelli A, Malesci A, Repici A, Day AJ, Armuzzi A, Danese S, Vetrano S. Dysfunctional Extracellular Matrix Remodeling Supports Perianal Fistulizing Crohn’s Disease by a Mechanoregulated Activation of the Epithelial-to-Mesenchymal Transition. Cell Mol Gastroenterol Hepatol. 2023;15(3):741-764. doi: 10.1016/j.jcmgh.2022.12.006. PMID: 36521659. Conflict of interest: Wong, Serre-Yu: Other: Research contract and support from Takeda/Trinetx and Eli Lilly and advisory board for Bristol Meyers Squibb Lin, Meng-Ju: No conflict of interest Wu, Cheng-Lin: No conflict of interest Cao, Wenqing: No conflict of interest Isda, Vladislav: No conflict of interest Israel, Yonatan: No conflict of interest Cho, WonKyung: No conflict of interest Shenoy, Shabari: No conflict of interest Colombel, Jean-Frédéric: Grant: AbbVie, Janssen Pharmaceuticals, Takeda, Prometheus and Bristol Myers Squibb Lectures from: AbbVie, Roche and Takeda Other: AbbVie, Amgen, AnaptysBio, Allergan, Apini, Arena Pharmaceuticals, Astellas, Boehringer Ingelheim, Bristol Myers Squibb, candidrx Celgene, Celltrion, Clearview Curogen, Eli Lilly, Envision Pharma Ferring Pharmaceuticals, Galmed Research, Glaxo Smith Kline, Roche, Janssen Pharmaceuticals, Kaleido Biosciences, Immunic, Iterative Scopes, Landos, Microba Life Science, Merck, Mirador, Novartis, Otsuka Pharmaceutical, Owkin, Pfizer, Protagonist Therapeutics, Sanofi, Sun Pharma, Takeda, Teva, TiGenix, and is holding stock options in Intestinal Biotech Development Plietz, Michael: No conflict of interest Hahn, Sue: No conflict of interest Khaitov, Sergey: none Polydorides, Alexandros: No conflict of interest Lu, Catherine P.: No conflict of interest
Wong et al. (Thu,) studied this question.