Identification, confirmation and functional relevance of neutrophil maturity in homeostasis, inflammation and disease

Abstract

During chronic inflammation, severe infection, and cancer, increased numbers of neutrophils are produced and released into the peripheral circulation, many of which are postulated to be developmentally immature. Although crucial to anti- microbial defence and having roles in tissue maintenance, neutrophils can worsen pathology by direct mediator production and suppression of other leukocytes. Neutrophil maturation is essential to their development of distinct granules that mediate neutrophil function in host defence. However, the identification and functional capacity of immature neutrophils in disease has not been well characterised. The aim of this thesis was to accurately identify immature neutrophils and investigate their role in mouse models of systemic inflammation and cancer. I characterised the efficiency of the neutrophil specific cell surface marker Ly6G in distinguishing immature (Ly6G- intermediate (Ly6GInt) expressing) from mature (Ly6G-high (Ly6GHi) expressing) neutrophils. Ly6G expression had an increased accuracy and consistency compared to other maturity markers in homeostasis, colony simulating factor-3 (CSF-3), and lipopolysaccharide (LPS) mouse models of haematopoietic stress. Ly6GInt neutrophils had a distinct phenotype and functional capacity compared to Ly6GHi neutrophils. I identified reduced expression of cell surface proteins important to the neutrophil response by Ly6GInt, compared to Ly6GHi neutrophils. Ly6GInt and Ly6GHi neutrophils suppressed T cell proliferation, however, CSF-3 enhanced suppression by Ly6GHi neutrophils only. Ly6GInt neutrophils also had increased intracellular ROS production at steady state. Furthermore Ly6GInt neutrophils were identified in mouse models of colorectal cancer, pancreatic adenocarcinoma, and hepatocellular carcinoma. Here, they correlated with metastasis and metastatic potential. I also found differences in neutrophil development between the bone marrow (BM) and spleen highlighting the requirement for further investigation into the affect of extramedullary haematopoiesis in neutrophil heterogeneity and disease pathology. Overall, differences in the functional capacity of Ly6GInt neutrophils, compared to Ly6GHi, indicate their physiological relevance in disease and could be relevant when considering the effects of immunotherapy.