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| Status |
Public on May 29, 2020 |
| Title |
Integrative molecular profiling of autoreactive CD4 T cells in autoimmune hepatitis |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
Background & Aims: In most autoimmune disorders, crosstalk of B cells and CD4 T cells results in the accumulation of autoantibodies targeting specific self-antigen like the Soluble Liver Antigen (SLA or SepSecs) in autoimmune hepatitis (AIH). But the associated autoreactive CD4 T cells have not been characterized yet. Here we isolated and deeply characterized SLA-specific CD4 T cells in AIH patients. Methods: We used brief ex vivo re-stimulation with overlapping SLA-derived peptides to isolate and phenotype circulating SLA-specific CD4 T cells, and integrative single-cell RNA-seq (scRNA-seq) to characterize their transcriptome and T cell receptor (TCR) repertoire. Identified auto-reactive TCRs were reconstituted cloned and expressed in T cell hybridoma to identify dominant SLA-derived CD4 T cell epitopes. SLA-specific CD4 T cells were tracked in peripheral blood through TCR sequencing, to identify their phenotypic niche. We further characterized disease-associated peripheral blood T cells by high content flow cytometry in an additional cohort of n=42 AIH patients and n=17 non-alcoholic steatohepatitis (NASH) controls. Results: Frequency of autoreactive SLA-specific CD4 T cells was associated with anti-SLA autoantibodies and had a memory CD45RA-CD27+PD-1+CXCR5-CCR6- phenotype. ScRNA-seq revealed their pro-inflammatory/B-Helper profile (IL21, IFNG, TIGIT, CTLA4, NR3C1, CD109, KLRB1 and CLEC2D). SLA81-100 and SLA177-204 contain dominant T cell epitopes. Autoreactive TCR clonotypes were restricted to the memory PD-1+CXCR5- CD4 T cells which was significantly increased in the blood of AIH patients and supported B cell differentiation through IL-21. Finally, we identified a specific phenotype (CD45RA-PD-1+CD38+CXCR5-CD127-CD27+) of T cells linked to disease activity and IgG response during AIH. Conclusions: This work provides for the first time a deep characterization of rare circulating autoreactive CD4 T cells and the identification of their peripheral reservoir in AIH. We also propose a generic phenotype of pathogenic T cells related to AIH disease activity which will be essential to track, delineate and potentially target those pathogenic cells.
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| Overall design |
This dataset contains data from SLA-specific and C.alb-specific CD4 T cells sorted from human peripheral blood of SLA-positive AIH patients after in vitro peptide restimulation. Single-cells were FACS-sorted into several 96-well plates and analyzed by 5'-end single-cell RNAseq following the FB5P-seq protocol. The samples were processed and sequenced in three separate experiments (experiment 1: samples 028-03, 128-01, 018-10; experiment 2: samples 018-13, 004-05, 018-04; experiment 3: sample 051-04).
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| Contributor(s) |
Milpied P, Gil L, Dong C, Renand A |
| Citation(s) |
32649971 |
| Submission date |
Apr 24, 2020 |
| Last update date |
Aug 28, 2020 |
| Contact name |
Pierre Milpied |
| E-mail(s) |
milpied@ciml.univ-mrs.fr
|
| Organization name |
Centre d'Immunologie de Marseille-Luminy (CIML)
|
| Lab |
Integrative B cell immunology
|
| Street address |
Parc Scientifique de Luminy, case 906
|
| City |
Marseille |
| ZIP/Postal code |
13288 |
| Country |
France |
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| Platforms (1) |
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| Samples (14)
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| Relations |
| BioProject |
PRJNA627921 |
| SRA |
SRP258323 |
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