Non-invasive measurements of exhaled NO and CO associated with methacholine responses in mice

BACKGROUND: Nitric oxide (NO) and carbon monoxide (CO) in exhaled breath are considered obtainable biomarkers of physiologic mechanisms. Therefore, obtaining their measures simply, non-invasively, and repeatedly, is of interest, and was the purpose of the current study. METHODS: Expired NO (E(NO)) and CO (E(CO)) were measured non-invasively using a gas micro-analyzer on several strains of mice (C57Bl6, IL-10(-/-), A/J, MKK3(-/-), JNK1(-/-), NOS-2(-/- )and NOS-3(-/-)) with and without allergic airway inflammation (AI) induced by ovalbumin systemic sensitization and aerosol challenge, compared using independent-sample t-tests between groups, and repeated measures analysis of variance (ANOVA) within groups over time of inflammation induction. E(NO )and E(CO )were also measured in C57Bl6 and IL-10-/- mice, ages 8–58 weeks old, the relationship of which was determined by regression analysis. S-methionyl-L-thiocitrulline (SMTC), and tin protoporphyrin (SnPP) were used to inhibit neuronal/constitutive NOS-1 and heme-oxygenase, respectively, and alter NO and CO production, respectively, as assessed by paired t-tests. Methacholine-associated airway responses (AR) were measured by the enhanced pause method, with comparisons by repeated measures ANOVA and post-hoc testing. RESULTS: E(NO )was significantly elevated in naïve IL-10(-/- )(9–14 ppb) and NOS-2(-/- )(16 ppb) mice as compared to others (average: 5–8 ppb), whereas E(CO )was significantly higher in naïve A/J, NOS-3(-/- )(3–4 ppm), and MKK3(-/- )(4–5 ppm) mice, as compared to others (average: 2.5 ppm). As compared to C57Bl6 mice, AR of IL-10(-/-), JNK1(-/-), NOS-2(-/-), and NOS-3(-/- )mice were decreased, whereas they were greater for A/J and MKK3(-/- )mice. SMTC significantly decreased E(NO )by ~30%, but did not change AR in NOS-2(-/- )mice. SnPP reduced E(CO )in C57Bl6 and IL-10(-/- )mice, and increased AR in NOS-2(-/- )mice. E(NO )decreased as a function of age in IL-10(-/- )mice, remaining unchanged in C57Bl6 mice. CONCLUSION: These results are consistent with the ideas that: 1) E(NO )is associated with mouse strain and knockout differences in NO production and AR, 2) alterations of E(NO )and E(CO )can be measured non-invasively with induction of allergic AI or inhibition of key gas-producing enzymes, and 3) alterations in AR may be dependent on the relative balance of NO and CO in the airway.