| Popis: |
Around one quarter of the world’s population is infected with helminth parasites, which are masters at modulating their host immune system. Expansion of the T regulatory cell (Treg) compartment is seen in many helminth infections and may prevent T helper 2 (Th2) cells from mediating worm clearance. As the Treg and Teffector cell balance may determine resistance versus susceptibility, this thesis focuses on these populations during infection. The model helminth Heligmosomoides polygyrus was studied in two mouse strains with different susceptibilities to infection, Th2-biased BALB/c mice which expel infection, and Th1-skewed C57BL/6 with a more regulatory response that fails to clear. Using single cell RNA sequencing and flow cytometric analysis, the CD4+ T cell populations within the two strains were compared. At the RNA level, Th2 cells fell into 5 subsets, some showing signs of anergy, and others with stronger TCR, co-stimulation and cytokine signalling; overall BALB/c T cells were more highly activated. However, both strains showed high levels of regulatory IL-10 signalling when infected. Treg populations were also characterised, defining 5 subsets. Tregs from infected BALB/c were more transcriptionally active than their naïve counterparts, although C57BL/6 mice showed greater proportions of Maf+ and Ly6e+ subsets that might explain their susceptibility. Treg responses were further characterised at the protein level using flow cytometry, including some markers suggested by subset delineation in scRNAseq analysis. There was a marked increase in co-expression of GATA3 with Foxp3, consistent with the Th2 environment. However, in contrast to published work, RORγt+ Tregs were not more frequent and were not affected by blockade of IL-6 in vivo. The expression of co-inhibitory receptors CTLA-4, PD-1 and TIGIT were also assessed on Tregs and Teffector cells, with CTLA-4 upregulated on both Tregs and Teffector cells post infection whereas PD-1 and TIGIT were only upregulated on effectors. Finally, a novel approach was taken to identify the epitopes recognised by CD4+ T cells during H. polygyrus infection, using the novel mouse line Nr4a3-Tocky which allows temporal and sensitive tracking of TCR engagement and thus antigen recognition. However, only a small proportion of CD4+ T cells recognised excretory-secretory (HES) antigens, even when isolated from H. polygyrus experienced mice, and there were components in HES which activated naive CD4+ T cells. More studies are required to further understand the antigen specific response to H. polygyrus. Overall, the thesis demonstrates that the adaptive response to H. polygyrus differs in resistant versus susceptible individuals at the transcriptomic, signalling and cell population levels; further functional investigation into the dynamics and role of each subset is required in future research. |
