Effect of growth medium on potential of Streptomyces anulatus spores to induce inflammatory responses and cytotoxicity in RAW264.7 macrophages

Epidemiological studies have shown an association between microbial growth in buildings and increased risk of respiratory symptoms and disease related to inflammatory reactions in the inhabitants. The current study examined t he affects of growth conditions of Streptomyces anulatus, isolated from ind oor air of a moldy building, on the inflammatory potential of spores of thi s microbe. Spores were harvested from 15 growth media formulations, applied to RAW264.7 macrophages (10(5), 10(6) or 10(7) spores/million cells), and evaluated for the ability to stimulate production of inflammatory mediators and cytotoxicity in these cells 24 h after exposure. Streptomyces anulatus spores induced dose-dependent production of nitric oxide (NO) in macrophag es, reaching a level from 4.2 muM to 39.2 muM depending on the composition of the growth medium of the microbe. Expression of inducible NO synthase (i NOS) was detected in macrophages after exposure to spores collected from al l growth media. Production of reactive oxygen species (ROS) was significant ly increased only by the highest dose of S. anulatus spores grown on glycer ol-arginine agar. Furthermore production of cytokines was affected by growt h medium; the highest dose-dependent levels of interleukin 6 (IL-6) ranged from 900 to 7800 pg/ml, and the levels of tumor necrosis factor alpha (TNF alpha) varied from 490 to 3200 pg/ml. The amount of dead macrophages after the exposure varied from 11% to 96%, depending also on the growth media of the microbe. Altogether, our results suggest that the growth medium of S. a nulatus has a fundamental role in the ability of the spores to induce infla mmatory responses and cytotoxicity in mammalian cells.