Maintenance of Peritoneal Macrophages in the Steady State

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Abstract
Resident peritoneal macrophages (Mø) were labeled in situ by intraperitoneal (i.p.) injection of the green fluorescent cell tracking dye PKH-1. After immunofluorescence staining with Mø specific monoclonal antibodies (Mabs) and phycoerythrin (PE) second antibody, the resident Mø were labeled with both the green dye and red Mab label, while recruited Mø were labeled only with the red Mab tag. These populations were distinguished by two-color flow cytometry. PKH-1 labeled resident peritoneal Mø were followed for 1-49 days in mice that received no further treatment (steady state). Dye labeled Mø were still detectable after 49 days in vivo, although their green fluorescence intensity had decreased steadily over time. The decrease in dye intensity was limited to Mø, as the fluorescence intensity of PKH-1 labeled peritoneal lymphocytes did not change. Resident Mø populations were clearly separated from recruited Mø by the intensity of their staining with PKH-1 for up to 28 days. No decrease in the number of resident (dye labeled) peritoneal Mø was observed over 1-28 days. These data indicate that resident peritoneal Mø were not replaced by recruited blood monocytes in the steady state.