TCR diversity and the development of regulatory T cells in the human thymus
Year: 2019
Type: dissertation
Abstract: A diverse T cell receptor (TCR) repertoire is essential for the adaptive immune system. T cells have to recognise and react to a huge variety of potential pathogens and at the same time maintain tolerance to self-antigens. TCR repertoire is established in the thymus during T cell development. Previous estimates of TCR diversity have studied the mature repertoire in the peripheral blood, identifying 1-3 x 106 unique TCRβ and 0.5 x 106 TCRα sequences, but the human intrathymic diversity has not been previously measured. In this thesis I provide the first estimate of the total TCR diversity in the human thymus. We have used high-throughput TCR sequencing and detected up to 10.3 x 106 unique TCRβ sequences and 3.7 x 106 TCRα sequences, which sets a new lower limit for TCR diversity. Computational estimators preseq, DivE and Chao2 produced 40 to 70 x 106 unique TCRβ sequences and 60 to 100 x 106 TCRα sequences in the human thymus. Somatic recombination produces a diverse thymic TCR repertoire that includes also the thymocytes instructed to develop into natural regulatory T cells. They are characterised by stable expression of the forkhead/winged-helix transcription factor FOXP3 and demethylation of Treg-specific-demethylated region (TSDR). Regulatory T cells develop as a separate cell lineage in the thymus and commitment occurs already at the CD4+CD8+ double positive (DP) stage. TCR-mediated signalling instructs the selection to the regulatory lineage and cytokines promote Treg development. We have used high-throughput TCR sequencing and methylation analysis of TSDR to track the timing of Treg commitment. Comparing the common TCR sequences showed that regulatory DP CD25+ and CD4SP CD25+ populations were clearly closer to each other than any other population, indicating that they are part of the common clonal and developmental pathway. The methylation analysis of TSDR in the DP and CD4+ single positive (SP) regulatory populations showed no difference between them. Thus a stable Treg phenotype can already be detected at the double positive stage. In this thesis I have studied the role of common γ chain cytokines in regulatory T cell development in the human thymus. Since thymic commitment of human Tregs begins at the double positive stage I have studied the effects and mechanisms of interleukin-2, interleukin-7 and interleukin-15 at the DP phase. Previous studies in murine models have established a two-step model. According to these studies, TCR signalling upregulates interleukin-2 receptor (IL-2R) and increases the responsiveness of developing Tregs to common γ chain cytokines that induce the expression of FOXP3. IL-7 has been considered to have a minor role in Treg development since mature peripheral Tregs express low levels of interleukin-7 receptor (IL-7R). We show that during thymic development DP FOXP3+ thymocytes upregulate IL-7R and are responsive to IL-7. At the DP stage TCR-signalling dictates Treg maturation during positive selection and IL-15 enhances Treg survival. After positive selection IL-2 and IL-7 increase the expression of FOXP3 and anti-apoptotic protein BCL-2 (B cell lymphoma 2) and promote Treg phenotype. DP FOXP3+ cells are highly susceptible to apoptosis but all three cytokines enhance their survival. Suppressing apoptosis is a key mechanism in promoting Treg development at the DP and CD4SP stage in the human thymus.
Source: Työväentutkimus Vuosikirja
Author: Vanhanen, Reetta
Language: English
Topic: T-cell and B-cell Immunology
Subfield: Immunology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2947705356