Amédée Renand1, Iñaki Cervera-Marzal2, Laurine Gil2, Chuang Dong2, Alexandra Garcia1, Erwan Kervagoret1, Hélène Aublé3, Sarah Habes4, Caroline Chevalier3, Fabienne Vavasseur3, Béatrice Clémenceau5, Anaïs Cardon1, Jean-Paul Judor1, Jean-François Mosnier6, Florence Tanné7, David-Axel Laplaud8, Sophie Brouard1, Jérôme Gournay9, Pierre Milpied2, Sophie Conchon10. 1. Université de Nantes, CHU Nantes, Inserm, Centre de Recherche en Transplantation et Immunologie, UMR 1064, ITUN, Nantes, France. 2. Aix Marseille Université, CNRS, Inserm, Centre d'Immunologie de Marseille-Luminy, CIML, Marseille, France. 3. Centre d'Investigation Clinique gastro-nutrition, CHU Nantes, Nantes, France. 4. Service Hepato-gastro-entérologie et Assistance Nutritionnelle, CHU Nantes, Nantes, France. 5. Université de Nantes, CHU Nantes, Inserm, CRCINA, Nantes, France. 6. Université de Nantes, CHU Nantes, Inserm, Centre de Recherche en Transplantation et Immunologie, UMR 1064, ITUN, Nantes, France; Service Anatomie et Cytologie Pathologiques, CHU Nantes, Nantes, France. 7. Service d'hépato gastroentérologie, CHU Cavale Blanche, Brest, France. 8. Université de Nantes, CHU Nantes, Inserm, Centre de Recherche en Transplantation et Immunologie, UMR 1064, ITUN, Nantes, France; Service de Neurologie et CIC 1413, CHU Nantes, Nantes, France. 9. Service Hepato-gastro-entérologie et Assistance Nutritionnelle, CHU Nantes, Nantes, France; Institut des Maladies de l'Appareil Digestif, IMAD, CHU Nantes, Nantes, France. 10. Université de Nantes, CHU Nantes, Inserm, Centre de Recherche en Transplantation et Immunologie, UMR 1064, ITUN, Nantes, France. Electronic address: sophie.conchon@univ-nantes.fr.
Abstract
BACKGROUND & AIMS: In most autoimmune disorders, crosstalk of B cells and CD4 T cells results in the accumulation of autoantibodies. In autoimmune hepatitis (AIH), the presence of anti-soluble liver antigen (SLA) autoantibodies is associated with reduced overall survival, but the associated autoreactive CD4 T cells have not yet been characterised. Herein, we isolated and deeply characterised SLA-specific CD4 T cells in patients with AIH. METHODS: We used brief ex vivo restimulation with overlapping SLA peptides to isolate and phenotype circulating SLA-specific CD4 T cells, and integrative single-cell RNA-seq (scRNA-seq) to characterise their transcriptome and T-cell receptor (TCR) repertoire. Autoreactive TCRs were cloned and used to identify dominant SLA-derived epitopes. SLA-specific CD4 T cells were tracked in peripheral blood through TCR sequencing to identify their phenotypic niche. We further characterised disease-associated peripheral blood T cells by high-content flow cytometry in 42 patients with AIH and 17 controls with non-alcoholic steatohepatitis. RESULTS: Autoreactive SLA-specific CD4 T cells were only detected in patients with anti-SLA autoantibodies and had a memory PD-1+CXCR5-CCR6-CD27+ phenotype. ScRNA-seq revealed their pro-inflammatory/B-helper profile. SLA81-100 and SLA177-204 contain dominant T-cell epitopes. Autoreactive TCR clonotypes were predominantly found in the memory PD-1+CXCR5-CD4 T cells, which were significantly increased in the blood of patients with AIH and supported B-cell differentiation through IL-21. Finally, we identified specific T-cell phenotypes linked to disease activity and IgG level during AIH. CONCLUSIONS: We provide a deep characterisation of rare circulating autoreactive CD4 T cells and identify their peripheral reservoir in AIH. We also propose a specific phenotype of autoreactive T cells related to AIH disease activity, which will be essential to track, delineate, and potentially target these pathogenic cells. LAY SUMMARY: One principal characteristic of autoimmune hepatitis (AIH), like for many other autoimmune diseases, is the accumulation of autoantibodies produced by B lymphocytes following their interaction with autoreactive CD4 T lymphocytes. In this study, we identified and characterised with high resolution these CD4 T cells. This will be essential to track, delineate, and potentially target them during AIH.
BACKGROUND & AIMS: In most autoimmune disorders, crosstalk of B cells and CD4 T cells results in the accumulation of autoantibodies. In autoimmune hepatitis (AIH), the presence of anti-soluble liver antigen (SLA) autoantibodies is associated with reduced overall survival, but the associated autoreactive CD4 T cells have not yet been characterised. Herein, we isolated and deeply characterised SLA-specific CD4 T cells in patients with AIH. METHODS: We used brief ex vivo restimulation with overlapping SLA peptides to isolate and phenotype circulating SLA-specific CD4 T cells, and integrative single-cell RNA-seq (scRNA-seq) to characterise their transcriptome and T-cell receptor (TCR) repertoire. Autoreactive TCRs were cloned and used to identify dominant SLA-derived epitopes. SLA-specific CD4 T cells were tracked in peripheral blood through TCR sequencing to identify their phenotypic niche. We further characterised disease-associated peripheral blood T cells by high-content flow cytometry in 42 patients with AIH and 17 controls with non-alcoholic steatohepatitis. RESULTS: Autoreactive SLA-specific CD4 T cells were only detected in patients with anti-SLA autoantibodies and had a memory PD-1+CXCR5-CCR6-CD27+ phenotype. ScRNA-seq revealed their pro-inflammatory/B-helper profile. SLA81-100 and SLA177-204 contain dominant T-cell epitopes. Autoreactive TCR clonotypes were predominantly found in the memory PD-1+CXCR5-CD4 T cells, which were significantly increased in the blood of patients with AIH and supported B-cell differentiation through IL-21. Finally, we identified specific T-cell phenotypes linked to disease activity and IgG level during AIH. CONCLUSIONS: We provide a deep characterisation of rare circulating autoreactive CD4 T cells and identify their peripheral reservoir in AIH. We also propose a specific phenotype of autoreactive T cells related to AIH disease activity, which will be essential to track, delineate, and potentially target these pathogenic cells. LAY SUMMARY: One principal characteristic of autoimmune hepatitis (AIH), like for many other autoimmune diseases, is the accumulation of autoantibodies produced by B lymphocytes following their interaction with autoreactive CD4 T lymphocytes. In this study, we identified and characterised with high resolution these CD4 T cells. This will be essential to track, delineate, and potentially target them during AIH.