Myeloid cell differentiation in pigs
Creators: name: Waddell, Lindsey A.
Publisher: name: The University of Edinburgh
Year: 2024
Subjects: pig immune system development
phagocytosis
bone marrow-derived macrophages
ADGRE1
macrophage markers
BMDMs
classical dendritic cells
cDC cell-specific molecule expression

description: Pigs are an important species in which to carry out research, not only in terms of their worldwide importance as an agricultural species, but also for economical and food supply reasons. Through many previous studies they have also been proven a valuable model of human health and the response to pathogens, and a number of studies have focussed upon the immune response. The key cell types studied within this thesis are bone marrow–derived macrophages (BMDMs) and classical dendritic cells (cDCs). There has been great debate within the scientific literature surrounding the similarities and differences in the function of these two cell types within the immune response. They are believed to arise from a common myeloid progenitor and are driven down separate paths of differentiation by the presence or absence of the growth factors CSF1 or FLT3L respectively. BMDMs are essential in the destruction of pathogens through phagocytosis, and cDCs play a vital role in initiating the immune response through communication with other cells involved within the immune response. This thesis focuses upon the determination of BMDM and cDC cell-specific molecule expression, and the investigation of the potential functions of these two cell types, as driven by separate growth conditions. This is assisted by the production and characterisation of novel mouse-anti porcine monoclonal antibodies (mAbs) which recognise ADGRE1 and CSF1 cell surface proteins. These have been identified as key molecules during the differentiation of myeloid cells in both rodents and humans, but due to the lack of porcine-specific reagents their role in this species remained largely undetermined. Cell-type specific expression of a number of cell surface molecules was determined on BMDMs and cDCs by flow cytometry. The molecules CD14, CD16, CD172a, ADGRE1 and CD203a were found to be expressed at differing levels by both cell types, but CSF1 and MERTK were predominately expressed only by BMDMs. Thus, selective expression of these could be used to determine differential roles in an immune response of BMDMs in comparison to cDCs. Through the creation and use of ADGRE1 and CSF1 mAbs in this project these molecules were both identified as molecules which were associated with the differentiation of bone marrow progenitor cells to BMDMs, with cell surface expression levels shown to increase over the differentiation process. ADGRE1 expression was also observed on mature tissue macrophage populations of the liver and lung by immunohistochemistry as well as FLT3L-derived cDCs. CSF1 was shown to be highly expressed on BMDMs and only at low levels on cDCs, suggesting this mAb’s potential as a distinguishing marker between these cell subsets. The results produced during this thesis provide insight into the differing functions of BMDMs and cDCs, providing potential differentiation markers of each cell type and suggesting future targets for vaccine or species-specific novel reagent development. All of these findings provide vital information for the progression of understanding of the porcine immune response.