Single-cell RNA sequencing of ILCs and T cells reveals enrichment of NK-like T cells in pediatric and adult inflammatory bowel disease

Background and aims:  Innate lymphoid cells (ILCs) are the innate counterparts of adaptive T cells, which together execute specialized effector programs in the intestinal mucosa. Understanding how unique and common dysregulation of the innate and adaptive lymphocytes contribute to inflammatory bowel disease (IBD) pathogenesis is crucial for development of novel and efficient therapies. Methods: We used flow cytometry to extensively characterize ILCs in pediatric (p)IBD patients (n=12) and controls (n=9), as well as single-cell RNA sequencing to determine the heterogeneity and inflammation-specific imprinting of ILCs, NK cells and T cells (n=6). Key findings were extended to adult IBD colonic mucosa (n=8) using flow cytometry. Results: We report a dysregulated mucosal ILC composition in pIBD that correlates to inflammatory activity. ILCs show pronounced and unique transcriptional changes in inflammation, particularly those related to antigen-presentation, while several interferon-associated transcripts are upregulated across lymphocyte lineages in inflammation. Furthermore, we identify two transcriptionally distinct T cell subsets, which are dysregulated in inflammation. This includes a subset transcribing KLRF1 and other NK-associated transcripts, that were enriched in inflamed pIBD mucosa. A corresponding NK-like NKp80+ T cell population, expressing high levels of NK cell receptors and cytotoxic molecules, are also accumulated in adult IBD. Conclusions: Using a combination of high-dimensional single-cell analysis on the transcriptional and protein level, we provide a comprehensive characterization of ILCs and T cells in pediatric and adult IBD. Our findings contribute to increased mechanistic understanding of intestinal inflammation and identify promising targets for IBD therapy, including the NK cell receptor NKp80. Single-cell RNA sequencing of ILCs and T-cells in matched noninflamed and inflamed colonic tissues of 6 pediatric IBD patients **The submitter declares that the raw data cannot be deposited due to patient privacy concerns.**