Effects of ?1?acid glycoprotein onpolymorphonuclear leukocytes ?involvement of cell surface receptors

Abstract: Alpha1?acid glycoprotein (AGP) is a highly glycosylated lipid?binding acute?phaseprotein. Although the exact mechanisms are unknown, several studies havesuggested that AGP may regulate the function of neutrophils and hence modulateinflammatory responses. The general aim of this thesis was to investigate if AGP isable to mediate intracellular signalling in neutrophils through binding to specificreceptors.Measurements of intracellular calcium concentration showed that AGP elicited asmall rise in [Ca2+]i in neutrophils that was markedly enhanced by pre?treatmentwith anti?L?selectin antibodies. In contrast, desialylation of AGP reduced the Ca2+mobilizing capacity significantly. The AGP?induced Ca2+ signal was mediatedthrough Src tyrosine kinases, PLC and PI3K which suggests involvement of cellsurface receptors. Indeed, AGP was shown to bind to, and mediate Ca2+ signallingthrough, sialic acid binding immunoglobulin?like lectin (Siglec)?5 and/or ?14.Increased fucosylation of AGP is common during acute?phase reactions. We showthat hyperfucosylated AGP has a diminished Ca2+ signalling capacity compared tonormally fucosylated AGP. This could be due to a reduced capacity of AGP tointeract with Siglec?5/?14 since it is known that the presence of fucose residues onsialylated glycans has a negative effect on Siglec?5/?14 affinity. AGP was alsodemonstrated to bind to the neutrophil proteins S100A8 and S100A9. In additionwe show that AGP?bound hydroxyeicasotetraenoic acids (HETEs) induce increasesin [Ca2+]i in neutrophils through binding to the leukotriene B4 receptor BLT2. Wepropose a two?step binding model where AGP binds to Siglec?5/?14 on L?selectinactivated neutrophils. This may orient AGP in a way that assists an interactionbetween AGP and the neutrophil membrane which favours transfer of AGP?boundHETEs to the BLT2 receptor.In conclusion, these data gives new insights regarding how AGP interacts with andmediates signalling in human neutrophils and supports the view of AGP as beingan acute phase reactant with immunomodulatory properties.