Shaping of the regulatory T cell repertoire and function in the intestine

Kopplin, Lydia; Spehr, Marc (Thesis advisor); Pabst, Oliver (Thesis advisor)

Aachen : RWTH Aachen University (2022)
Dissertation / PhD Thesis

Dissertation, RWTH Aachen University, 2022

Abstract

The complex interplay of tolerogenic and proinflammatory T cells at mucosal sites plays a crucial role in health and disease. Regulatory T cells (Tregs) are at the centre of this complex system and the focus of extensive research. If the balance between Tregs and proinflammatory T cells is disrupted, chronic disorders like inflammatory bowel disease (IBD) are often the consequence. Multiple immune pathways have been implicated in IBD, in particular dysregulation of colonic Tregs. Although Treg biology has been a burning area of interest for many decades, it is currently not understood whether the suppression of inflammation by colonic Tregs is dependent on cognate antigen recognition or acts through T cell receptor (TCR)-independent mechanisms. Furthermore, the relationship of Tregs and effector T cell populations (Teff) to each other and the microbiota is still poorly understood. To address these questions, we modified the classical adoptive T cell transfer colitis model in mice by transferring harmonized collections of T cells to multiple Rag2-deficient individuals. The in vitro expansion of Tregs or naïve T cells allowed us to track clonally identical (harmonized) T cell populations in the large intestine lamina propria (LI LP) of individual recipient mice and to analyse the selection- and expansion process of Tregs and Teff cells. Our data demonstrate that expansion of Tregs in the colon is highly dependent on TCR-specificity, similarly to effector T cell subsets. A small fraction of clones dominated the Treg repertoire between different individuals, suggesting that expansion is driven by cognate TCR engagement.Moreover, we were able to investigate the relationship of Teff cells to peripherally induced Tregs (pTregs). Although both populations arise from the same naïve T cell progenitors, pTreg- and Teff repertoires are largely distinct indicating that TCR specificity has an effect on the T cell phenotype. This finding was further underlined through the adoptive transfer of in vitro induced Tregs (iTregs) into Rag2-deficient mice: the TCR repertoires of stable iTregs and unstable ex-iTregs were distinct. Dominant clones were not shared between those two subpopulations highlighting a role of the TCR in Treg lineage commitment. Similarly, single cell sequencing paired with single cell TCR repertoire analysis allowed us to combine phenotype information with TCR clonality. Cells of a clone clustered together in unifold manifold approximation and projection (UMAP) and showed distinct gene expression profiles compared to other T cell clones. This suggests a role of the TCR in initiating a specific T cell fate. Interestingly, in accordance with several recent studies, the transcriptional profiles of the T cells did not reflect the established T helper cell/Treg signatures. Furthermore, alterations of the microbiota-composition of recipients by antibiotic-treatment, or the additional colonization with a single dominant bacterial species, were directly reflected in changes in Teff- and pTreg repertoires. This suggests that microbial antigen is an important source for selection and expansion of both Teff and Tregs in the intestine. In perspective, the novel model of harmonized T cell repertoires will allow us to further narrow down the source of antigen by reducing the complexity of the antigenic environment. Targeted colonization or exclusive diets can be used to address the impact of microbial- or food antigen on selection and expansion of colonic Tregs and Teff. Furthermore, the model can be used to investigate the role of the TCR in lineage commitment. These insights may lead to the development of new therapeutic approaches directly addressing patients` Treg differentiation, stability and function.