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 (ENO) and CO (ECO) 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. ENO and ECO 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
ENO 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 ECO 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 ENO by ~30%, but did not change AR in NOS-2-/- mice. SnPP reduced ECO in C57Bl6 and IL-10-/- mice, and increased AR in NOS-2-/- mice. ENO decreased as a function of age in IL-10-/- mice, remaining unchanged in C57Bl6 mice.


CONCLUSIONS
These results are consistent with the ideas that: 1) ENO is associated with mouse strain and knockout differences in NO production and AR, 2) alterations of ENO and ECO 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.
Sethi
Jigme M
JM
Division of Allergy, Pulmonary, Immunology, Critical Care and Sleep Division, Department of Internal Medicine, University of Texas Medical Branch, Galveston TX 77755-1083, USA. jmsethi@upmc.edu
Choi
Augustine M K
AM
Calhoun
William J
WJ
Ameredes
Bill T
BT
D000818:Animals; D001944:Breath Tests; D016085:Bronchoconstrictor Agents / Q000009:adverse effects*; D002245:Carbon Dioxide / Q000378:metabolism*; D045853:Exhalation; D006419:Heme Oxygenase (Decyclizing) / Q000037:antagonists & inhibitors / Q000378:metabolism; D016753:Interleukin-10 / Q000235:genetics / Q000378:metabolism; D048371:MAP Kinase Kinase 3 / Q000235:genetics / Q000378:metabolism; D008297:Male; D016210:Methacholine Chloride / Q000009:adverse effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D048055:Mitogen-Activated Protein Kinase 8 / Q000235:genetics / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D011014:Pneumonia / Q000139:chemically induced* / Q000378:metabolism*; D012044:Regression Analysis
D016085:Bronchoconstrictor Agents; D016210:Methacholine Chloride; D016753:Interleukin-10; D002245:Carbon Dioxide; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496317:Nos2 protein, mouse; C496315:Nos3 protein, mouse; D006419:Heme Oxygenase (Decyclizing); D048055:Mitogen-Activated Protein Kinase 8; D048371:MAP Kinase Kinase 3; C482531:Map2k3 protein, mouse