Expression of Fc Gamma Receptors on Peripheral Blood Monocytes and Evaluation of their Role in Phagocytosis Using a Novel Method.

By Camilla Holt

Fc gamma receptors (FcR) play an important role in immunity by binding to IgG opsonised antigens and activating the appropriate immune responses. In humans there are three isotypes of FcR; FcRI FcRII and FcRIII. FcRs are also involved in autoimmune disorders and its importance in these diseases has been well established by animal models and patient group studies. As our knowledge of the FcR isotypes, their mechanisms and functions, has increased it has become apparent that a greater understanding of the specific interactions with the different IgG subclasses is needed. In particular how the different FcR:IgG complexes are translated into the different functional responses. In this research group it was previously shown that all FcRs were up-regulated in the synovial tissue from rheumatoid arthritis (RA) patients when compared to healthy controls. To complete this picture and gain a better understanding of the role FcR play in RA we wanted to look at the expression of FcRI, FcRIIa, FcRIIb and FcRIII on peripheral blood monocytes from healthy individuals.

The aim of this study was to investigate the variation in the FcRs in healthy donors and investigate whether they influenced the cells ability to bind and phagocytose IgG subclass specific immune complexes. To do this we used a standard rosetting assay and a newly described phagocytosis assay. FcR expression was analysed by FcR specific antibodies and flowcytometry. We also wanted to compare different methods ofperipheral blood monocyte (PBM) purification and used three methods of PBM purification, plastic adherence, positive MACS and negative MACS. Additionally, two human monocyte cells lines, MonoMac-6 and U-937 were investigated.

Our results show that within an individual the amount of PBM expressing a particular FcR could be predicted only for FcRIIa which was constitutively expressed. The other receptors had varying population sizes of receptor positive PBM. The receptors did have predictable patterns of cell surface expression which are consistent throughout the population. FcRI and FcRIIb are expressed at low and consistent concentrations within an individual, FcRIIa is expressed on the cell surface at high and consistent concentrations, while FcRIII was expressed at variable concentrations. We also showed that the amount of FcRIIa positive cells was the major contributor to phagocytosis of both IgG1 and IgG3 IC while FcRI and FcRIII cell surface concentration was the major contributor to IC binding. These results indicate that specific FcR isotypes effect the cells functional abilities in IgG subclass specific interactions.