Quantifying changes in the T cell receptor repertoire during thymic development
- CNRS, France
- Weizmann Institute of Science, Israel
- University College London, United Kingdom
Abstract
One of the feats of adaptive immunity is its ability to recognize foreign pathogens while sparing the self. During maturation in the thymus, T cells are selected through the binding properties of their antigen-specific T-cell receptor (TCR), through the elimination of both weakly (positive selection) and strongly (negative selection) self-reactive receptors. However, the impact of thymic selection on the TCR repertoire is poorly understood. Here, we use transgenic Nur77-mice expressing a T-cell activation reporter to study the repertoires of thymic T cells at various stages of their development, including cells that do not pass selection. We combine high-throughput repertoire sequencing with statistical inference techniques to characterize the selection of the TCR in these distinct subsets. We find small but significant differences in the TCR repertoire parameters between the maturation stages, which recapitulate known differentiation pathways leading to the CD4+ and CD8+ subtypes. These differences can be simulated by simple models of selection acting linearly on the sequence features. We find no evidence of specific sequences or sequence motifs or features that are suppressed by negative selection. These results favour a collective or statistical model for T-cell self non-self discrimination, where negative selection biases the repertoire away from self recognition, rather than ensuring lack of self-reactivity at the single-cell level.
Data availability
All code for reproducing the figures of this paper can be found at https://github.com/statbiophys/ thymic_development_2022.git. The data has been deposited on the SRA as BioProject ID PRJNA804508 http://www.ncbi.nlm.nih.gov/bioproject/804508.
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thymic selection repertoires miceNCBI BioProject, PRJNA804508.
Article and author information
Author details
Aleksandra M Walczak
Funding
CNRS-Weizmann (80 prime)
- Francesco Camaglia
European Research Council (COG 724208)
- Aleksandra M Walczak
Agence Nationale de la Recherche (ANR-19-CE45-0018)
- Thierry Mora
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Frederik Graw, Heidelberg University, Germany
Ethics
Animal experimentation: The experiment was carried out using three 6-weeks old male inbred Nur77-GFP/Foxp3-mCherry (C57BL/6 background). The cross was bred and maintained at the Weizmann institute. All animals were handled according to Weizmann Institute's Animal Care guide- lines, in compliance with national and international regulations. This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All of the animals were handled according to approved institutional animal care and use committee (IACUC) protocols ({\#}21661115-2) of the Weizmann Institute of Science. The protocol was approved by the Committee on the Ethics of Animal Experiments of the Weizmann Institute of Science. Every effort was made to minimize suffering.
Version history
- Preprint posted: February 15, 2022 (view preprint)
- Received: July 5, 2022
- Accepted: January 18, 2023
- Accepted Manuscript published: January 20, 2023 (version 1)
- Version of Record published: February 16, 2023 (version 2)
Copyright
© 2023, Camaglia et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Quantifying changes in the T cell receptor repertoire during thymic development
eLife 12:e81622.
https://doi.org/10.7554/eLife.81622
Further reading
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- Cell Biology
- Immunology and Inflammation
Cytokine polyfunctionality is a well-established concept in immune cells, especially T cells, and their ability to concurrently produce multiple cytokines has been associated with better immunological disease control and subsequent effectiveness during infection and disease. To date, only little is known about the secretion dynamics of those cells, masked by the widespread deployment of mainly time-integrated endpoint measurement techniques that do not easily differentiate between concurrent and sequential secretion. Here, we employed a single-cell microfluidic platform capable of resolving the secretion dynamics of individual PBMCs. To study the dynamics of poly-cytokine secretion, as well as the dynamics of concurrent and sequential polyfunctionality, we analyzed the response at different time points after ex vivo activation. First, we observed the simultaneous secretion of cytokines over the measurement time for most stimulants in a subpopulation of cells only. Second, polyfunctionality generally decreased with prolonged stimulation times and revealed no correlation with the concentration of secreted cytokines in response to stimulation. However, we observed a general trend towards higher cytokine secretion in polyfunctional cells, with their secretion dynamics being distinctly different from mono-cytokine-secreting cells. This study provided insights into the distinct secretion behavior of heterogenous cell populations after stimulation with well-described agents and such a system could provide a better understanding of various immune dynamics in therapy and disease.
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- Immunology and Inflammation
- Medicine
Background:
Prinflammatory extracellular chromatin from neutrophil extracellular traps (NETs) and other cellular sources is found in COVID-19 patients and may promote pathology. We determined whether pulmonary administration of the endonuclease dornase alfa reduced systemic inflammation by clearing extracellular chromatin.
Methods:
Eligible patients were randomized (3:1) to the best available care including dexamethasone (R-BAC) or to BAC with twice-daily nebulized dornase alfa (R-BAC + DA) for seven days or until discharge. A 2:1 ratio of matched contemporary controls (CC-BAC) provided additional comparators. The primary endpoint was the improvement in C-reactive protein (CRP) over time, analyzed using a repeated-measures mixed model, adjusted for baseline factors.
Results:
We recruited 39 evaluable participants: 30 randomized to dornase alfa (R-BAC +DA), 9 randomized to BAC (R-BAC), and included 60 CC-BAC participants. Dornase alfa was well tolerated and reduced CRP by 33% compared to the combined BAC groups (T-BAC). Least squares (LS) mean post-dexamethasone CRP fell from 101.9 mg/L to 23.23 mg/L in R-BAC +DA participants versus a 99.5 mg/L to 34.82 mg/L reduction in the T-BAC group at 7 days; p=0.01. The anti-inflammatory effect of dornase alfa was further confirmed with subgroup and sensitivity analyses on randomised participants only, mitigating potential biases associated with the use of CC-BAC participants. Dornase alfa increased live discharge rates by 63% (HR 1.63, 95% CI 1.01–2.61, p=0.03), increased lymphocyte counts (LS mean: 1.08 vs 0.87, p=0.02) and reduced circulating cf-DNA and the coagulopathy marker D-dimer (LS mean: 570.78 vs 1656.96 μg/mL, p=0.004).
Conclusions:
Dornase alfa reduces pathogenic inflammation in COVID-19 pneumonia, demonstrating the benefit of cost-effective therapies that target extracellular chromatin.
Funding:
LifeArc, Breathing Matters, The Francis Crick Institute (CRUK, Medical Research Council, Wellcome Trust).
Clinical trial number:
