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Stricturing Crohn’s Disease Single-Cell RNA Sequencing Reveals Fibroblast Heterogeneity and Intercellular Interactions

2023; Elsevier BV; Volume: 165; Issue: 5 Linguagem: Inglês

10.1053/j.gastro.2023.07.014

1528-0012

Pranab K. Mukherjee, Quang Tam Nguyen, Jiannan Li, Shuai Zhao, Stephen M. Christensen, Gail West, Jyotsna Chandra, Ilyssa O. Gordon, Sinan Lin, Jie Wang, Ren Mao, D. Czarnecki, Carla Rayan, Idan Goren, Suhanti Banerjee, Prerna Kotak, Thomas Plesec, Samir Lal, Thomas Fabre, Shoh Asano, Kathryn Bound, Kevin M. Hart, Chanyoung Park, Robert Martinez, Ken Dower, Thomas A. Wynn, Shaomin Hu, Nayden G. Naydenov, Martin Decaris, Scott Turner, Stefan D. Holubar, Scott R. Steele, Claudio Fiocchi, Andrei I. Ivanov, Kellie M. Kravarik, Florian Rieder,

Single-cell and spatial transcriptomics

Background & Aims Fibroblasts play a key role in stricture formation in Crohn's disease (CD) but understanding its pathogenesis requires a systems-level investigation to uncover new treatment targets. We studied full-thickness CD tissues to characterize fibroblast heterogeneity and function by generating the first single-cell RNA sequencing (scRNAseq) atlas of strictured bowel and providing proof of principle for therapeutic target validation. Methods We performed scRNAseq of 13 fresh full-thickness CD resections containing noninvolved, inflamed nonstrictured, and strictured segments as well as 7 normal non-CD bowel segments. Each segment was separated into mucosa/submucosa or muscularis propria and analyzed separately for a total of 99 tissue samples and 409,001 cells. We validated cadherin-11 (CDH11) as a potential therapeutic target by using whole tissues, isolated intestinal cells, NanoString nCounter, next-generation sequencing, proteomics, and animal models. Results Our integrated dataset revealed fibroblast heterogeneity in strictured CD with the majority of stricture-selective changes detected in the mucosa/submucosa, but not the muscle layer. Cell-cell interaction modeling revealed CXCL14+ as well as MMP/WNT5A+ fibroblasts displaying a central signaling role in CD strictures. CDH11, a fibroblast cell-cell adhesion molecule, was broadly expressed and up-regulated, and its profibrotic function was validated using NanoString nCounter, RNA sequencing, tissue target expression, in vitro gain- and loss-of-function experiments, proteomics, and knock-out and antibody-mediated CDH11 blockade in experimental colitis. Conclusions A full-thickness bowel scRNAseq atlas revealed previously unrecognized fibroblast heterogeneity and interactions in CD strictures and CDH11 was validated as a potential therapeutic target. These results provide a new resource for a better understanding of CD stricture formation and open potential therapeutic developments. This work has been posted as a preprint on Biorxiv under doi: 10.1101/2023.04.03.534781.