Abstract
T cells are a family of specialised adaptive immune cells which are central to the identification and elimination of viral infections. The dynamic activity of T cells is underpinned by highly regulated metabolic processes, and thus there is a growing interest in the field of immunometabolism as a method to identify T cell dysfunction and provide therapeutic targets, which can modulate their adaptive response.SCENITH is a novel technique that exploits the benefits of flow cytometry whilst producing a metabolic profile for immune cell populations. SCENITH can be utilised without the requirement of immune cell isolation, and as such can give insight into the metabolic activities within the heterogeneity of T cell populations.
Cryopreservation is often performed to facilitate delayed, high-throughput analysis of clinical samples. However, as suggested by previous studies utilising other metabolic assays, this process could result in metabolic defects, leading to misunderstandings about patient clinical samples.
Dengue infection is of increasing importance, and the understanding of the immunopathology behind severe disease is important for the development of therapeutics which can reduce its burden. PD-1 is a marker found on the surface of activated and exhausted T cells which has been identified on seemingly impaired T cells during severe dengue infection. PD-1, when studied previously using other metabolic assays, has been shown to elicit a downregulation of glycolysis and upregulation of fatty acid metabolism, when bound to its complimentary ligand
PDL1.
This thesis aimed to optimise and evaluate SCENITH when using cryopreserved samples, identify the activity of PD-1-mediated metabolism in health donor T cells using SCENITH and interpret the role of PD-1 during severe and non-severe dengue using its metabolic output.
Following cryopreservation, T cells were shown to become less metabolically active upon stimulation, and were shown to become less reliant on oxidative phosphorylation and favouring glycolysis in a cryopreservation time-dependent manner. Despite this, the distinction between the metabolic profiles of T cell populations within each cryopreservation timepoint remained visible.
Upon PD-1 ligation, healthy donor T cells were shown to trend towards an increased reliance on fatty acid metabolism, whilst showing trends towards a decreased dependence on glucose metabolism. Potential anti-PD-1 therapies, Metformin and Nivolumab, demonstrated a moderate capacity in reversing some of the trends observed within T cells during PD-1 ligation, decreasing their capacity for fatty acid metabolism.
Within severe patients with dengue, PD-1 expression of T cells correlated with a decreased capacity for glycolysis, particularly within CD8+ T cells, suggesting there may be impaired activation within these CD8+ PD-1+ populations.
Overall, this thesis’ findings have implications and considerations for future utilisation of SCENITH, whilst also providing some biological insights into the activities of PD-1+ T cells in healthy donors and dengue patients
| Date of Award | 23 Jan 2024 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | Laura Rivino (Supervisor) & Christoph Wuelfing (Supervisor) |