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 inhabitants96. The current study examined the affects of growth conditions of Streptomyces anulatus, isolated from indoor air of a moldy building, on the inflammatory potential of spores of this 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 macrophages, reaching a level from 4.2 microM to 39.2 microM depending on the composition of the growth medium of the microbe. Expression of inducible NO synthase (iNOS) was detected in macrophages after exposure to spores collected from all growth media. Production of reactive oxygen species (ROS) was significantly increased only by the highest dose of S. anulatus spores grown on glycerol-arginine agar. Furthermore production of cytokines was affected by growth 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 (TNFalpha) 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. anulatus has a fundamental role in the ability of the spores to induce inflammatory responses and cytotoxicity in mammalian cells.
Hirvonen
M R
MR
National Public Health Institute, Division of Environmental Health, Laboratory of Toxicology, PO Box 95, FIN-70701 Kuopio, Finland. Maija-Riitta.Hirvonen@ktl.fi
Suutari
M
M
Ruotsalainen
M
M
Lignell
U
U
Nevalainen
A
A
D000391:Air Microbiology; D016902:Air Pollution, Indoor; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000502:physiology; D003470:Culture Media / Q000378:metabolism*; D005562:Formazans / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism* / Q000382:microbiology; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D017382:Reactive Oxygen Species / Q000378:metabolism; D013170:Spores / Q000502:physiology; D013302:Streptomyces / Q000254:growth & development* / Q000302:isolation & purification; D013778:Tetrazolium Salts / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D003470:Culture Media; D005562:Formazans; D015850:Interleukin-6; D017382:Reactive Oxygen Species; D013778:Tetrazolium Salts; D014409:Tumor Necrosis Factor-alpha; C079782:MTT formazan; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse