Monocytes recruited into the alveolar air space of mice show a monocytic phenotype but upregulate CD14

The evaluation of monocytes recruited into the alveolar space under both ph ysiological and inflammatory conditions is hampered by difficulties in disc riminating these cells from resident alveolar macrophages (rAMs). Using the intravenous injected fluorescent dye PKH26, which accumulated in rAMs with out labeling blood leukocytes, we developed a technique that permits the id entification, isolation, and functional analysis of monocytes recruited int o lung alveoli of mice. Alveolar deposition of murine JE, the homologue of human monocyte chemoattractant protein (MCP)-1 (JE/MCP-1), in mice provoked an alveolar influx of monocytes that were recovered by bronchoalveolar lav age and separated from PKH26-stained rAMs by flow cytometry. Alveolar recru ited monocytes showed a blood monocytic phenotype as assessed by cell surfa ce expression of F4/80, CD11a, CD11b, CD18, CD49d, and CD62L. In contrast, CD14 was markedly upregulated on alveolar recruited monocytes together with increased tumor necrosis factor-alpha message, discriminating this monocyt e population from peripheral blood monocytes and rAMs. Thus monocytes recru ited into the alveolar air space of mice in response to JE/MCP-1 keep pheno typic features of blood monocytes but upregulate CD14 and are "primed" for enhanced responsiveness to endotoxin with increased cytokine expression.