Cell Growth and Antimicrobial Activity of Mouse Peritoneal Macrophages in Response to Glucocorticoids, Choleragen and Lipopolysaccharide

ABSTRACT Normal, thioglycolate (TG)-stimulated and BCG-activated mouse peritoneal macrophages were cultivated in vitro with the conditioned medium of mouse L-929 cells (L-CM). The TG- and BCG-macrophages rapidly proliferated, whereas normal macrophages grew much more slowly. Certain immunomodulators, glucocorticoids at physiological concentrations, choleragen at 10 pg/ml, and Escherichia coli lipopolysaccharide (LPS) at 10 pg/ml inhibited the growth of the TG-macrophages. We devised a simple, quantitative assay method for candidacidal activity of macrophages. The normal mouse macrophage monolayers formed in 96-multiwell tissue culture plates were infected with serially diluted Candida parapsilosis , and an end-point of dilution, whereby candida cells added were found to be completely destroyed, was determined after a 48 hr incubation period. An addition of the L-CM to 20% of the culture medium, stimulated the killing activity more than 128-fold, compared with no addition of L-CM. In the medium containing the L-CM, macrophages spread very well on the plastic with several dendrites, whereas the cells spread poorly and gradually cytolysed in the medium lacking L-CM. It was found that E. coli LPS at 1-10 μg/ml and muramyl dipeptide at 100 μg/ml stimulated the activity 4 to 16 times. Macrophages treated with 1 μM dexamethasone and 10 ng/ml choleragen had a reduced activity (50% of the control). An application of this method to destroying other kinds of microbes, measuring the activity of other phagocytes, and screening of immunomodulators was discussed.