Background FcgammaIIa (CD32) has been identified as the functional high affinity CRP receptor on leukocytes (monocytes and neutrophils). Studies on the effects of CRP on neutrophils show that receptor activation signals to downstream functional changes in phagocytosis, cytokine release and generation of reactive oxygen species. The effects of CRP on monocytes which are key players in rheumatoid chronic inflammation, have not been studied.
Objectives To investigate the phenotypic and functional consequences of CRP on peripheral blood monocytes in vitro.
Methods Peripheral whole bloods (PWB) from consenting individuals, with or without RA (ARA criteria), were collected into citrate and kept on ice. Mononuclear cells (MNC) and monocytes (MON) were isolated by differential centrifugation using lymphoprep and Dynal negative isolation kit respectively, using sigmacoted tubes to minimise artefactual activation. Cells were exposed to CRP from 0–200 microg for up to 30 min at 37°C and analysed for (a) CD11b and CD32 expression by flow cytometry (PWB and MNC); (b) adhesion to LPS-activated human umbilical vein endothelial cells (MON) and (c) release of reactive oxygen species (MON).
Results CD14+ve rheumatoid monocytes expressed more than double the level of CD11b (8.644 + 0.57), compared with those from healthy controls (4.17 + 0.4), p = 0.0043. There was little difference in CD32 expression. Exposure of PWB and MNC’s from normal healthy subjects to increasing concentrations of CRP led to a dose dependent increase in CD11b expression from 5.7 + 0.1 to 12 + 0.6 at 200 microg CRP (p < 0.01); this was not enhanced by LPS (500 ng/ml). However, not all normals were responders to CRP: 46% of subjects (n = 13) showed no change in CD11b levels. Monocyte adhesion to HUVECs as determined using BCECF loaded cells relative to U937 monocytes, significantly altered following treatment with CRP for 30 mins at 37°C. Release of superoxide anion (MON) was elicited by 200 but not 20 microg CRP, as determined by cytochrome C reduction.
Conclusion CRP can effect monocyte activation ex vivo, and induce phenotypic changes similar to those observed in rheumatoid patients. The variation in response from normal subjects may reflect different CD32 genotypes. A single nucleotide polymorphism in FcgammaRIIa which encodes histidine or arginine at position 131, strongly influences both IgG2a and CRP binding1 and may identify individuals with functionally discrete responses to CRP. Allele specific response to CRP may be important in monocyte activation and extravasation during rheumatoid arthritis and complicating vascular disease.
Stein MP. C-reactive protein binding to Fc gamma RIIa on human monocytes and neutrophils is allele specific. J Clin Invest. 2000;105:369–76
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