Single-Cell Analyses of Colon and Blood Reveal Distinct Immune Cell Signatures of Ulcerative Colitis and Crohn's Disease.

BACKGROUND & AIMS Studies are needed to determine the mechanisms of mucosal dysregulation in patients with inflammatory bowel diseases (IBD) and differences in inflammatory responses of patients with ulcerative colitis (UC) vs Crohn's disease (CD). We used mass cytometry (CyTOF) to characterize and compare immune cell populations in the mucosa and blood from patients with IBD and without IBD (controls) at single-cell resolution. METHODS We performed CyTOF analysis of colonic mucosa samples (n=87) and peripheral blood mononuclear cells (PBMCs, n=85) from patients with active or inactive UC or CD and controls. We also performed single-cell RNA-sequencing, flow cytometry, and RNA in situ hybridization analyses to validate key findings. We used random forest modeling to identify differences in signatures across subject groups. RESULTS Compared with controls, colonic mucosa samples from patients with IBD had increased abundances of HLA-DR+CD38+ T cells, including T-regulatory cells that produce inflammatory cytokines; CXCR3+ plasmablasts; and IL1B+ macrophages and monocytes. Colonic mucosa samples from patients with UC were characterized by expansion of IL17A+ CD161+ effector memory T cells and IL17A+ T-regulatory cells; expansion of HLA-DR+CD56+ granulocytes; and reductions in type 3 innate lymphoid cells. Mucosal samples from patients with active CD were characterized by IL1B+HLA-DR+CD38+ T cells, IL1B+TNF+IFNG+ naïve B cells, IL1B+ dendritic cells (DCs), and IL1B+ plasmacytoid DCs. PBMCs from patients with active CD differed from those of active UC in that the PBMCs from patients with CD had increased IL1B+ T-regulatory cells, IL1B+ DCs and IL1B+ plasmacytoid DCs, IL1B+ monocytes, and fewer group 1 innate lymphoid cells. Random forest modeling differentiated active UC from active CD in colonic mucosa and blood samples; top discriminating features included many of the cellular populations identified above. CONCLUSIONS We used single-cell technologies to identify immune cell populations specific to mucosa and blood samples from patients with active or inactive CD and UC and controls. This information might be used to develop therapies that target specific cell populations in patients with different types of IBD.

[1]  Matthew E. Ritchie,et al.  A divergent transcriptional landscape underpins the development and functional branching of MAIT cells , 2019, Science Immunology.

[2]  M. Clatworthy,et al.  Phenotypically distinct neutrophils patrol uninfected human and mouse lymph nodes , 2019, Proceedings of the National Academy of Sciences.

[3]  J. C. Love,et al.  Highly Efficient, Massively-Parallel Single-Cell RNA-Seq Reveals Cellular States and Molecular Features of Human Skin Pathology , 2019, bioRxiv.

[4]  Aviv Regev,et al.  Intra- and Inter-cellular Rewiring of the Human Colon during Ulcerative Colitis , 2019, Cell.

[5]  Samuel J. S. Rubin,et al.  Mass cytometry reveals systemic and local immune signatures that distinguish inflammatory bowel diseases , 2019, Nature Communications.

[6]  Shiliang Ma,et al.  B Cell Dysfunction Associated With Aging and Autoimmune Diseases , 2019, Front. Immunol..

[7]  Takehiro Mitsuishi.  Correlation between histological findings and endoscopic findings in patients with ulcerative colitis: Basal plasmacytosis is an important finding suggesting active inflammation , 2018, JGH open : an open access journal of gastroenterology and hepatology.

[8]  R. Hendriks,et al.  Frequencies of circulating regulatory TIGIT+CD38+ effector T cells correlate with the course of inflammatory bowel disease , 2018, Mucosal Immunology.

[9]  M. Merad,et al.  High-dimensional immune phenotyping and transcriptional analyses reveal robust recovery of viable human immune and epithelial cells from frozen gastrointestinal tissue , 2018, Mucosal Immunology.

[10]  Alex K. Shalek,et al.  Allergic inflammatory memory in human respiratory epithelial progenitor cells , 2018, Nature.

[11]  Ang Lin,et al.  Granulocytes: New Members of the Antigen-Presenting Cell Family , 2017, Front. Immunol..

[12]  ZhangPengfei,et al.  CD19loCD27hi Plasmablasts Suppress Harmful Th17 Inflammation Through Interleukin 10 Pathway in Colorectal Cancer , 2017 .

[13]  D. Sachar,et al.  Update on anti-tumor necrosis factor agents and other new drugs for inflammatory bowel disease , 2017, British Medical Journal.

[14]  J. McCluskey,et al.  MAIT cells launch a rapid, robust and distinct hyperinflammatory response to bacterial superantigens and quickly acquire an anergic phenotype that impedes their cognate antimicrobial function: Defining a novel mechanism of superantigen-induced immunopathology and immunosuppression , 2017, PLoS biology.

[15]  A. Griffiths,et al.  Mucosal Expression of Type 2 and Type 17 Immune Response Genes Distinguishes Ulcerative Colitis From Colon-Only Crohn's Disease in Treatment-Naive Pediatric Patients. , 2017, Gastroenterology.

[16]  M. Neurath.  Current and emerging therapeutic targets for IBD , 2017, Nature Reviews Gastroenterology &Hepatology.

[17]  Kenji Suzuki,et al.  Possible involvement of mucosal-associated invariant T cells in the progression of inflammatory bowel diseases. , 2017, Biomedical research.

[18]  Zhiyong Wu,et al.  CD19loCD27hi Plasmablasts Suppress Harmful Th17 Inflammation Through Interleukin 10 Pathway in Colorectal Cancer. , 2017, DNA and cell biology.

[19]  Z. Vadasz,et al.  FoxP3 Expression in Macrophages, Cancer, and B Cells—Is It Real? , 2017, Clinical Reviews in Allergy & Immunology.

[20]  James M. Dahlhamer,et al.  Prevalence of Inflammatory Bowel Disease Among Adults Aged ≥18 Years - United States, 2015. , 2016, MMWR. Morbidity and mortality weekly report.

[21]  Hao Chen,et al.  Cytofkit: A Bioconductor Package for an Integrated Mass Cytometry Data Analysis Pipeline , 2016, PLoS Comput. Biol..

[22]  J. Salleron,et al.  A practical guide to assess the Nancy histological index for UC , 2016, Gut.

[23]  S. Ziegler,et al.  CD4+ Group 1 Innate Lymphoid Cells (ILC) Form a Functionally Distinct ILC Subset That Is Increased in Systemic Sclerosis , 2016, The Journal of Immunology.

[24]  C. Fiocchi,et al.  Immunopathogenesis of IBD: current state of the art , 2016, Nature Reviews Gastroenterology &Hepatology.

[25]  C. Buskens,et al.  Interleukin-12 and -23 Control Plasticity of CD127(+) Group 1 and Group 3 Innate Lymphoid Cells in the Intestinal Lamina Propria. , 2015, Immunity.

[26]  Piet Demeester,et al.  FlowSOM: Using self‐organizing maps for visualization and interpretation of cytometry data , 2015, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[27]  H. Mittrücker,et al.  CD38 Is Expressed on Inflammatory Cells of the Intestine and Promotes Intestinal Inflammation , 2015, PloS one.

[28]  A. Regev,et al.  Spatial reconstruction of single-cell gene expression , 2015, Nature Biotechnology.

[29]  T. Lumley,et al.  gplots: Various R Programming Tools for Plotting Data , 2015 .

[30]  Mark M. Davis,et al.  CD161 Defines a Transcriptional and Functional Phenotype across Distinct Human T Cell Lineages , 2014, Cell reports.

[31]  J. Salleron,et al.  Comparing histological activity indexes in UC , 2014, Gut.

[32]  M. Lederman,et al.  Circulating CD4+ and CD8+ T cells are activated in inflammatory bowel disease and are associated with plasma markers of inflammation , 2013, Immunology.

[33]  A. Ganser,et al.  IFN-γ Production by Allogeneic Foxp3+ Regulatory T Cells Is Essential for Preventing Experimental Graft-versus-Host Disease , 2012, The Journal of Immunology.

[34]  Marc Vandemeulebroecke,et al.  Secukinumab, a human anti-IL-17A monoclonal antibody, for moderate to severe Crohn's disease: unexpected results of a randomised, double-blind placebo-controlled trial , 2012, Gut.

[35]  P. Klenerman,et al.  Human MAIT and CD8αα cells develop from a pool of type-17 precommitted CD8+ T cells. , 2012, Blood.

[36]  P. Moayyedi,et al.  Efficacy of Biological Therapies in Inflammatory Bowel Disease: Systematic Review and Meta-Analysis , 2011, The American Journal of Gastroenterology.

[37]  D. Littman,et al.  Characterization of interleukin-17-producing regulatory T cells in inflamed intestinal mucosa from patients with inflammatory bowel diseases. , 2011, Gastroenterology.

[38]  K. Tominaga,et al.  Increased numbers of immature plasma cells in peripheral blood specifically overexpress chemokine receptor CXCR3 and CXCR4 in patients with ulcerative colitis , 2011, Clinical and experimental immunology.

[39]  G. Martino,et al.  IL-17– and IFN-γ–Secreting Foxp3+ T Cells Infiltrate the Target Tissue in Experimental Autoimmunity , 2010, The Journal of Immunology.

[40]  Pedro M. Valero-Mora,et al.  ggplot2: Elegant Graphics for Data Analysis , 2010 .

[41]  Nikesh Kotecha,et al.  Web‐Based Analysis and Publication of Flow Cytometry Experiments , 2010, Current protocols in cytometry.

[42]  Hadley Wickham,et al.  ggplot2 - Elegant Graphics for Data Analysis (2nd Edition) , 2017 .

[43]  S. Brand.  Crohn’s disease: Th1, Th17 or both? The change of a paradigm: new immunological and genetic insights implicate Th17 cells in the pathogenesis of Crohn’s disease , 2009, Gut.

[44]  T. Nomura,et al.  Functional delineation and differentiation dynamics of human CD4+ T cells expressing the FoxP3 transcription factor. , 2009, Immunity.

[45]  B. Müller-Myhsok,et al.  Role of the novel Th17 cytokine IL‐17F in inflammatory bowel disease (IBD): Upregulated colonic IL‐17F expression in active Crohn's disease and analysis of the IL17F p.His161Arg polymorphism in IBD , 2008, Inflammatory bowel diseases.

[46]  Geoffrey E. Hinton,et al.  Visualizing Data using t-SNE , 2008 .

[47]  Kenneth G. C. Smith,et al.  Competence and competition: the challenge of becoming a long-lived plasma cell , 2006, Nature Reviews Immunology.

[48]  L. Öhman,et al.  Functional CD4+CD25high regulatory T cells are enriched in the colonic mucosa of patients with active ulcerative colitis and increase with disease activity , 2006, Inflammatory bowel diseases.

[49]  Clare Baecher-Allan,et al.  MHC Class II Expression Identifies Functionally Distinct Human Regulatory T Cells1 , 2006, The Journal of Immunology.

[50]  R. Lechler,et al.  Acquisition of HLA‐DR and Costimulatory Molecules by T Cells from Allogeneic Antigen Presenting Cells , 2005, American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons.

[51]  T. Giese,et al.  Peripheral and intestinal regulatory CD4+ CD25(high) T cells in inflammatory bowel disease. , 2005, Gastroenterology.

[52]  Robert P. Sheridan,et al.  Random Forest: A Classification and Regression Tool for Compound Classification and QSAR Modeling , 2003, J. Chem. Inf. Comput. Sci..

[53]  G. Mateu-Figueras,et al.  Isometric Logratio Transformations for Compositional Data Analysis , 2003 .

[54]  H. Ziegler-Heitbrock,et al.  CD14 is expressed and functional in human B cells , 1994, European journal of immunology.

[55]  R. Macdermott,et al.  Enhand secretion of tumour necrosis factor‐alpha, IL‐6, and IL‐1β by isolated lamina ropria monouclear cells from patients with ulcretive cilitis and Crohn's disease , 1993 .

[56]  S. Ahlstedt,et al.  Neutrophil mucosal involvement is accompanied by enhanced local production of interleukin-8 in ulcerative colitis. , 1993, Gut.

[57]  G. Vanham,et al.  Expression of activation antigens, HLA‐DR and CD38, on CD8 lymphocytes during HIV‐1 infection , 1992, AIDS.

[58]  Y. Mahida,et al.  Enhanced production of interleukin 1-beta by mononuclear cells isolated from mucosa with active ulcerative colitis of Crohn's disease. , 1989, Gut.