Research Article

Clearance of Apoptotic Cells by Macrophages Induces Regulatory Phenotype and Involves Stimulation of CD36 and Platelet-Activating Factor Receptor

Figure 3

Efferocytosis requires lipid raft integrity. BMDM were treated with βCD (1 mM) or αCD (1 mM) for 10 min before addition of apoptotic thymocytes (10 per macrophages) for 90 min for phagocytosis. After washing for removal of noningested targets, coverslips were stained with haematoxylin/eosin and the phagocytic indexes (derived by multiplying the percentage of macrophages containing at least one ingested target by the number of targets ingested by the macrophages) were assessed by cell counting under an optical microscope. Values are mean ± SEM of at least three independent experiments ( versus control) (a). In parallel, BMDM were incubated with apoptotic thymocytes (10 per macrophage) for 20 min before addition of lysis buffer. Cells lysates were subjected to immunoprecipitation/immunoblotting assays as described in Section 2 using antibodies to PAFR or CD36 and flotillin-1. Protein expression was quantified by the AlphaEaseFC software V3.2 beta. The autoradiographs show one representative experiment, and graph data are presented as mean ± SEM of three experiments ( ) (b).