Binding of opsonized immune complexes to Erythrocyte crlicd35 inhibits TNF-a production by restricting immune complex uptake by macro phages

Abstract

Previous studies have shown that children suffering from severe malaria have elevated concentrations of TNF-a, increased levels of circulating immune complexes (ICs) and decreased levels of complement regulatory proteins (mainly CRl /CD35 and DAF/ CD55) on their erythrocytes. The cross-linking of FcR on the macrophages has been shown to cause activation and subsequently induce release of pro-inflammatory cytokines which could lead to malarial anemia, a major complication of P. falciparum and an important cause of child mortality and morbidity. In this study, it was postulated that the erythrocytes of individuals suffering from or at the risk of severe malarial anemia have reduced levels of complement regulatory proteins and this compromises their ability to mop out circulating ICs. As a result, a lot of ICs remain in circulation, engage the macrophages and induce the secretion of TNF-a which is associated with malarial anemia. Using anti-CRI monoclonal antibody, erythrocyte CRI copy numbers were determined by flow cytometry and cryopreserved erythrocytes from a cross sectional study in Kombewa were categorized as low, medium 'and high expressers. 15 individuals from each cohort were selected and IC binding capacity determined by flow cytometry. Using an in vitro model system, macrophages were stimulated with a cocktail of erythrocytes and pre-opsonized BSA-anti-BSA ICs, loaded erythrocytes, supernatants and relevant controls. At the end of 8-hour incubation period, the supernatants were harvested and ELISA done to determine the levels of TNF-a present. The data generated in this study indicated that the IC binding capacity was influenced by the CRI copy number and it was complement dependent. The data did show that the erythrocytes inhibit IC-induced TNF-a production by macrophages and that the buffering capacity was in a manner proportional to the level of CRI. Also, the erythrocytes soaked with ICs stimulated macrophages more than plain erythrocytes though the stimulation was not in a manner proportional to CRI. Based on the findings it was concluded that erythrocyte CRI may act as a dynamic buffering system which prevents ICs from stimulating macrophages to release TNF-a which is implicated in the pathogenesis of severe malaria. Also, the CRI enables the erythrocytes to soak in ICs and in the process makes the erythrocytes to become stimulatory leading to secretion of TNF-a by the macrophages