Phagocytosis of unopsonized zymosan particles by trypsin-sensitive and β-glucan-inhibitable receptors on bone marrow-derived murine macrophages

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Abstract
Murine bone marrow cells, plated at 4 X 10(4) cells/well and cultured in 50% fibroblast CM, yielded pure populations of large, individual, adherent cells that were phagocytic and morphologically indistinguishable from macrophages. Adherent macrophages appeared in small numbers with 24 h of culture, increased to maximal cell numbers within 10 days of culture, and remained at these cell densities for at least 11 weeks in culture. The capacities of adherent macrophages to ingest unopsonized zymosan particles and EsIgG, at inputs of 1.25 X 10(7) targets, were expressed by 7 and 40% of the cells derived from 24-hour cultures, respectively, were increased at nearly identical rates to comparable maximal levels within 10-14 days of culture and were exhibited by essentially all adherent cells derived from 2-11-week cultures. The percentage of adherent macrophages from twelve 3-6-week cultures ingesting greater than or equal to 1, greater than or equal to 6 and greater than or equal to 10 zymosan particles was 89 +/- 5, 47 +/- 11 and 14 +/- 9% (mean +/- SD, n = 12), respectively, and the percentage ingesting greater than or equal to 1, greater than or equal 6 and greater than or equal to 10 EsIgG was 86 +/- 5, 49 +/- 10 and 14 +/- 8%, respectively. Incubation of adherent macrophages with mannan-free ss-glucan particles at inputs of 5 X 10(5)-5 X 10(7)/ml initiated a phagocytic response comparable to that obtained with the same doses of zymosan particles which contained mannan and beta-glucan. Preincubation of adherent macrophages with 100 micrograms/ml of a fully soluble beta-glucan, laminarin, and solubilized barley beta-glucan reduced subsequent macrophage phagocytosis of greater than or equal to 6 zymosan particles by 53 and 40%, respectively. In contrast, yeast alpha-mannan was less than 1% as active, and 10 mg/ml reduced the number of adherent macrophages ingesting greater than or equal to 1 zymosan particles by 64%. At concentrations as high as 2 mg/ml, laminarin and barley beta-glucan had no effect on Fc receptor-mediated ingestion of EsIgG, and mannan at 20 mg/ml also failed to inhibit EsIgG ingestion. Pretreatment of adherent macrophages with 20 micrograms/ml of trypsin reduced the number of cells ingesting greater than or equal to 1 zymosan particles from 89 to 10% and those ingesting greater than or equal to 6 zymosan from 43 to 0%, whereas pretreatment with as much as 100 micrograms/ml of trypsin failed to decrease macrophage ingestion of EsIgG.(ABSTRACT TRUNCATED AT 400 WORDS)

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