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 Ca(2+), 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 Ca(2+), 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 microg/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.