From Monocyte to Macrophage;: In Vitro Differentiation of Human Monocytes to Macrophages and Characterization of Macrophage Properties
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- Institutt for biologi 
Monocytes and macrophages are critical effectors and regulators of human host defence and inflammation. The enlightenment provided by good model systems for studying cellular mechanisms related to monocytes and macrophages is crucial. Human peripheral blood monocytes may be differentiated to viable macrophages in vitro, in the presence of M-CSF. In this study, four different methods for isolating monocytes from whole blood were compared on the basis of several decisive factors, such as yield, purity, morphology, viability and practicality. In addition, subpopulations of monocytes and monocyte-derived macrophages were flowcytometrically classified on the basis of the surface markers CD14 and CD16. Using microscopy, flow cytometry and RT-qPCR, the monocyte-derived macrophages were subsequently characterized at different time points based on morphology and the known macrophage-specific markers CD71 and CD206. Monocytes and macrophages are heterogeneous in terms of characteristics and functional capacities due to various stimuli. In this study, functional properties of cryopreserved, elutriation-purified monocytes and monocyte-derived macrophages exposed to wild type and mutant (LPS-deficient) Neisseria meningitidis (Nm) as well as purified LPS from Nm were comparatively investigated in vitro using flow cytometry, RT-qPCR and protein activity measurements. After 10 days of cultivation with M-CSF, Nm-exposed monocyte-derived macrophages possessed the ability of phagocytosing opsonized bacteria, displayed procoagulant activity, and they expressed TNF-α and IL-1β mRNAs as well as TNF-α protein, but not IL-1β protein. This implies that we have established an in vitro model system for differentiation of monocytes to macrophages, which may be used for studying cellular mechanisms related to macrophages.