Abstract
Bacillus Calmette-Guerin (BCG) is the only effective tuberculosis (TB) vaccine and has been studied and used for over a century, yet TB remains the deadliest bacterial infection, killing and infecting millions of people globally each year. Though the mechanisms of BCG-induced protection are still not entirely understood, recent studies have indicated that training of innate immune cells is more important than previously thought, contradicting previous notions of protection derived primarily from memory CD4+ T cells. BCG-trained innate immune cells exhibit a variety of alterations in cell signalling, metabolism, transcription, and epigenetics establishing a ‘trained immunity’ phenotype that has improved immunological performance and can last for months. Thus training critically important innate immune cells involved in fighting a Mycobacterium tuberculosis (Mtb) infection such as alveolar macrophages (AM) could improve the effectiveness of the BCG vaccine. To further understand metabolic reprogramming of BCG-trained AMs, glycolytic activity was compared between lung cells from BCG vaccinated and unvaccinated mice with the intention of using sorted AMs, and AM subpopulations in the future. The work presented in this thesis further develops a flow-cytometry panel designed to identify murine AM subpopulations for future fluorescence-activated cell sorting (FACS) and describes the optimisation of the XF Glycolysis Stress Test for murine lung cells. Lung cells from both BCG-vaccinated and unvaccinated mice were shown to have similar basal metabolic activity at 4 weeks post vaccination, however, lung cells from vaccinated mice demonstrated an enhanced glycolytic response to glucose. Interestingly, the XF Glycolysis Stress Test suggests lung cells from BCG-vaccinated and unvaccinated mice have no glycolytic reserve. This thesis lays the groundwork for studying the metabolic activity of BCG-trained murine AM subpopulations which will inform future studies to understand the mechanisms behind BCG training and ultimately BCG-induced protection against TB.