Pulmonary macrophages of mice in the steady state were isolated by lavage with PBS containing EDTA and subsequent enzymatic digestion of tissue with pronase and DNA-ase. By this method, the total pulmonary macrophage population was obtained in two cell suspensions, one with a pure population of pulmonary alveolar macrophages (PAM) and the other with a mixed population of pulmonary alveolar and pulmonary tissue macrophages (PTM). The morphological, cytochemical, and functional characteristics of both PAM and PTM were like those of mature tissue macrophages except for the presence of C3 receptors. These receptors were almost absent on PAM and present on a larger number of cells in the mixed population of PAM and PTM. The total pulmonary macrophage population of mice in the steady state is approximately equal to 2 x 10(6), of which about 93% are PAM and about 7% are PTM. In labeling experiments with 3H-thymidine, the low in vitro labeling indices (less than 3%) for both PAM and the mixture of PAM and PTM, showed that both are essentially nondividing cells. In vivo labeling studies showed an increase in the number of labeled macrophages that can only be attributed to labeled monocytes migrating into the lungs. Additional evidence was provided by a decrease in the labeling indices of pulmonary macrophages when mice were treated with hydrocortisone acetate, which causes a severe monocytopenia, thus preventing monocyte influx into the lungs. Confirmation of the bone marrow origin was obtained in mice labeled after x-irradiation with partial bone marrow shielding: labeled pulmonary macrophages were found in the exposed lungs. In all experiments, the labeling indices were identical in the two macrophage populations isolated. These results show that the influx of monocytes is the source of cell renewal for the pulmonary macrophages. No indications for an interstitial division or maturation compartment in the lung were found. Quantitation of the efflux of labeled monocytes from the blood, and the number of labeled pulmonary macrophages, showed that in the steady state about 15% of the monocytes leaving the circulation become pulmonary macrophages and that the turnover time of pulmonary macrophages is approximately equal to 27 d.

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