In vitro effects of nitrogen dioxide on the release of nitric oxide by bovine alveolar macrophages

Bovine alveolar macrophages (BAM) were assessed for nitric oxide-production after exposure to synthetic air or NO2 [0.2 ppm] for 2 h and subsequent treatment with lipopolysaccharide (LPS) [1 mu g/ml]. Nitric oxide (NO) release, measured as nitrite in the supernatant, was quantified at different time points. LPS-stimulated BAM showed elevated nitrite levels after 24 h. Pre-treatment with NO2 resulted in higher nitrite levels in comparison to air exposed BAM. Some of the BAM preparations resulted in pre-activated macrophages that produce high amounts of NO without LPS treatment. Within these cells LPS treatment reduced the nitrite levels in comparison to untreated control incubations from the same preparation. These pre-activated BAM exhibited no uniform effect after NO2-exposure. In some experiments nitrite levels of pre-activated control BAM were decreased, while in others no influence could be observed. NO-production by pre-activated BAM or after I,PS-treatment could be inhibited by N-G-monomethyl-L-arginine (L-NMMA) [1 mM]. In order to investigate, whether NO2-exposure alters inducible NO-synthase mRNA expression, we now perform reverse transcription polymerase chain reaction (RT-PCR) of iNOS mRNA. This method will be a good tool to elucidate whether NO2-exposure can modulate iNOS mRNA expression. (C) 1998 Elsevier Science Ireland Ltd. All rights reserved.