Serum amyloid P component binds to late apoptotic cells and mediates their uptake by monocyte‐derived macrophages

Abstract

Objective Some pentraxins, such as C-reactive protein, bind to apoptotic cells and are involved in the clearance of these cells. We undertook this study to determine whether serum amyloid P component (SAP; a pentraxin that, when deficient in mice, results in lupus-like disease) binds to apoptotic cells and to assess the functional consequences of SAP binding for their phagocytosis by macrophages. Methods Human peripheral blood monocytes were isolated and cultured for 7 days to obtain monocyte-derived macrophages. Jurkat cells were irradiated with ultraviolet B to induce apoptosis. After 4 hours, a mean ± SEM of 54.0 ± 5.1% of these cells stained with annexin V and were propidium iodide negative (early apoptotic [EA] cells). After 24 hours, 77.3 ± 2.7% of cells stained positive with both annexin V and propidium iodide (late apoptotic [LA] cells or secondary necrotic cells). EA and LA cells were incubated with fluorescein isothiocyanate–labeled SAP in the presence or absence of Ca²⁺, and binding was measured by flow cytometry. Phagocytosis was tested by incubation of macrophages with EA or LA cells in the presence of normal human serum (NHS) and quantified as a phagocytosis index (PI; number of Jurkat cells internalized by 100 macrophages). Experiments were repeated with SAP-depleted serum and after reconstitution with increasing concentrations of SAP. Results The majority of LA cells did bind SAP in the presence of Ca²⁺, whereas EA cells did not. SAP depletion of NHS resulted in a 50% decrease in the PI for LA cells, and complete restoration of the PI could be demonstrated with SAP reconstitution up to 100 μg/ml. SAP depletion had no effect on phagocytosis of EA cells. Conclusion SAP binds to LA cells and is involved in the phagocytosis of these cells by human monocyte–derived macrophages. This may have consequences for diseases such as systemic lupus erythematosus, in which phagocytosis of apoptotic cells is decreased.