Exploring the interface between trained immunity and infection-driven granulopoiesis

Abstract

Trained immunity describes the innate immune system’s ability to ‘remember’ previous inflammatory challenges. Training involves adaptations in innate cells and their progenitors wherein, upon secondary exposure to inflammatory stimuli, trained cells are poised to respond more strongly/efficiently. Stimulated haematopoietic stem and progenitor cells (HSPCs) impart a trained phenotype to newly generated myeloid cells through supporting demand-adapted myelopoiesis. At the time of starting this thesis, no published literature had described the contribution of neutrophils to trained immunity. Previous work in the Hall lab has shown that neutrophils generated by one mode of demand-adapted myelopoiesis, emergency granulopoiesis (EG), display enhanced bactericidal activity compared to neutrophils generated at homeostasis. In light of recent advancements in the field of trained immunity around the involvement of HSPCs, it was reasoned that infectionexperienced HSPCs generate neutrophils with a trained phenotype. To test this hypothesis, the conserved biology and high-resolution in vivo imaging potential of larval zebrafish was exploited. The data presented in this thesis further demonstrates that EG-generated neutrophils display enhanced functionality. Results show that EG-generated neutrophil enhanced bactericidal activity is nonspecific, and that elevated reactive nitrogen species production may contribute to this phenotype. RNA-sequencing analysis suggests that the induction of mitochondrial reactive oxygen species and the exploitation of fatty acid catabolism are likely involved in driving the enhanced bactericidal phenotype exhibited by EG-generated neutrophils. Furthermore, through developing a unique allotransplant protocol, infection-experienced HSPCs were shown to generate neutrophils with enhanced bacterial killing capacity compared to infection-naïve HSPCs. This provides the first in vivo evidence that directly links infection-driven granulopoiesis with the generation of neutrophils with an enhanced antibacterial phenotype. The trained immune phenotype was further explored in larval zebrafish by utilising β-glucan, a potent inducer of trained immunity, and bacterial infection to stimulate training. Results show that training confers non-specific, long-lasting protective benefits to larvae evident of a trained phenotype and their enhanced expression of the neutrophil-attracting chemokine cxcl8a likely plays a protective role. Taken together, the results presented in this thesis demonstrate that infection-driven granulopoiesis and trained immunity operate in larval zebrafish to protect against subsequent infectious challenge providing exciting avenues for further exploration.