Investigating lymphocyte development in the foetal thymus and modulation by the melanocortin 5 receptor

Lymphocyte development and activation are critical for the generation of effective protective immunity. Innate lymphoid cells and T cells play central roles in orchestrating bespoke immune reactions by producing specialised repertoires of immuno-regulatory cytokines. ILC2 and natural killer (NK) cells are generated in the bone marrow and thymus, whilst T cell development is restricted to the thymus. Once in the tissues, these cells respond to microenvironmental cues that refine their responses, for example, via receptor-mediated cell-cell contacts, soluble cytokines and neuropeptides. The co-development of ILC2, NK cells and T cells within a 5-day window in the foetal thymus provides a tractable system for studying their genesis and differentiation. Using combinatorial multicolour transcription factor reporter mouse lines, flow cytometry and confocal imaging I investigated the temporal stages of thymic innate lymphoid cell development. I found that NK cell progenitors (NKp) were first visible at embryonic (E) day E13.5, followed by the appearance of ILC2 progenitors (ILC2p) and developing T cells. Notably, the NKp and ILC2p formed clusters in the vicinity of medullary thymic epithelial cells (mTEC). To provide greater molecular insight into this cellular niche, single-molecule RNA fluorescence in situ hybridisation (smFISH) was performed on E17.5 foetal thymus to visually identify gene expression of 100 genes with single-cell resolution. This highlighted potential foetal thymus niche factors that may support innate lymphoid cell development. In the periphery ILC2, NK cells and T cells can respond to niche-derived factors to help initiate or potentiate immunity. Recent reports have highlighted the importance of tissue-resident stromal cells and neurons in this process, especially for ILC2. By analysing bulk RNAseq gene expression data, I identified Mc5r, encoding the fifth melanocortin receptor (Mc5r), as expressed more highly by ILC2 than ILC3. Treatment of mice with the neuropeptide alpha-melanocortin-stimulating hormone (αMSH), an agonist for melanocortin receptors, induced increases in the proportions of lung NK cells and T cells. The generation of a mouse line in which Mc5r was deleted in all lymphocytes (Il7r^CreMc5r^fl/fl mice) confirmed that Mc5r is required for normal NK cell and T cell representation in the lung at basal state, after IL-25 or IL-33 alarmin challenge and after viral respiratory infection of mice by the Pneumonia Virus of Mice.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2022

Author dc:creator

• Heycock, Morgan

Advisor dc:contributor.advisor

• McKenzie, Andrew

• immunology
• lung
• mouse
• thymus

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/8d660b56-15f6-4eea-90e3-a7a4b87c8f41/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.96659

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/349905