Integrative molecular profiling of autoreactive CD4 T cells in autoimmune hepatitis.
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.
['Renand|Amédée|A|', 'Cervera-Marzal|Iñaki|I|', 'Gil|Laurine|L|', 'Dong|Chuang|C|', 'Garcia|Alexandra|A|', 'Kervagoret|Erwan|E|', 'Aublé|Hélène|H|', 'Habes|Sarah|S|', 'Chevalier|Caroline|C|', 'Vavasseur|Fabienne|F|', 'Clémenceau|Béatrice|B|', 'Cardon|Anaïs|A|', 'Judor|Jean-Paul|JP|', 'Mosnier|Jean-François|JF|', 'Tanné|Florence|F|', 'Laplaud|David-Axel|DA|', 'Brouard|Sophie|S|', 'Gournay|Jérôme|J|', 'Milpied|Pierre|P|', 'Conchon|Sophie|S|']
D000328:Adult D001323:Autoantibodies D001324:Autoantigens D001402:B-Lymphocytes D015496:CD4-Positive T-Lymphocytes D018984:Epitopes, T-Lymphocyte D005260:Female D019693:Hepatitis, Autoimmune D006801:Humans D007156:Immunologic Memory D008297:Male D008875:Middle Aged D061026:Programmed Cell Death 1 Receptor D011948:Receptors, Antigen, T-Cell D054380:Receptors, CXCR5 D017423:Sequence Analysis, RNA D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7
Autoreactive CD4 T cells Autoimmune hepatitis Single-cell RNA-seq Autoreactive TCR clonotypes Soluble liver antigen