Characterising the Function of CD300e Immunoregulatory Molecule in CD300e-/- Mice

Abstract

The CD300 family of immunoglobulin-like molecules have immune regulatory functions. Including both inhibitory and triggering receptors, which are highly conserved within the family as well as across species. Establishing their individual function has been slowed by the lack of well characterised and specific reagents, particularly monoclonal antibodies. Most members of the CD300 family are expressed on myeloid cells including dendritic cells (DCs), monocytes and macrophages. The most restricted member of the family is the CD300e molecule. It is only known to be expressed on human monocytes and on a subpopulation of DCs. As there are no antibodies available to mouse CD300e, I generated a novel CD300e-deficient (CD300e-/-) mouse strain as an alternative approach to study CD300e biology. In addition, I produced a mouse anti-mouse CD300e monoclonal antibody (mCD300e mAb) to facilitate study of CD300e in mice.

This thesis describes the development of mCD300e mAb using a hybridoma production technique and provides a detailed characterisation of the changes of immune cell populations and their functions resulting from CD300e deletion. Our data show that CD300e-/- mice have leukopenia, reduced cellularity of immune organs, and disruption of BM niches accompanied with reduced CD169+ER-HR3+ macrophages. Using BM chimera approaches, the results suggest the involvement of CD300e in Ly6C- monocyte differentiation, erythropoiesis, and indirectly B cell lymphopoiesis. Due to its selective expression in myeloid cells, I investigated the functions of CD300e using BM-derived macrophages (BMDM) by further testing a variety of monocyte and macrophage functions. I found in the absence of the CD300e molecule, macrophage functions are altered including phagocytosis of apoptotic cells and production of pro-inflammatory cytokine (TNF-⍺, IL-6, IL-10) and chemokines (CXCL1, CCL17, CCL22) in the M1 macrophages. Finally, a mouse model of thioglycolate-induced peritonitis was used to study the effect of CD300e deficiency on migration ability of myeloid cells during inflammation. Reduced infiltration of neutrophils and monocytes was observed in CD300e-/- mice one day post injection. Collectively, this thesis demonstrates that the CD300e molecule contributes to functional BM microenvironment and immune function of macrophages.