Binding and phagocytosis of opsonized SRBC by primary rat hepatocyte cultures at different time points after Ad-mediated FcγRIIA cDNA gene transfer. Primary hepatocytes were infected with AdNull and AdFcγRIIA at 10 moi. After 48 hours, cells were incubated with IgG-coated 51Cr-labeled SRBC for 10, 30, and 60 minutes. As a control, the cells were incubated with nonopsonized SRBC for 60 minutes. To evaluate binding of IgG-coated SRBC to primary hepatocytes, cells were washed with PBS and then lysed by incubation with 0.5% SDS for 10 minutes. To evaluate SRBC phagocytosis by the primary hepatocytes, cells were washed with PBS and the SRBC bound to the cell surface were lysed by incubation with hypotonic lysis buffer for 1 minute. The cells were lysed by incubation with 0.5% SDS for 10 minutes and the radioactivity of lysate was quantified. (A) Binding of opsonized SRBC at 10, 30, and 60 minutes to hepatocytes that were not infected (control; •), infected with AdNull (▪), or infected with AdFcγRIIA (▴). Also indicated as controls are parallel cultures of cells incubated with nonopsonized SRBC for 60 minutes with uninfected hepatocytes (○), AdNull-infected hepatocytes (□), and AdFcγRIIA-infected hepatocytes (▵). (B) Phagocytosis of opsonized RBC at 10, 30, and 60 minutes. The symbols are identical to that in (A). For all data, shown are the means of activity (dpm)/well from 3 measurements ± standard error.
