Oropharyngeal origin of markers in exhaled breath

Abstract: Normal NO formation in the human airways occurs primarily in the nasal airways, where it is catalyzed by inducible NO synthase (iNOS), and in the oropharyngeal tract, via as yet not fully defined pathways. This NO can be detected in exhaled breath and when inflammation is present in the airways, for example in asthma, the concentration of NO is increased. Although most studies on non-invasive measurements of airway inflammation have focused on NO in exhaled breath, there has also been an increasing interest in measurements of compounds in exhaled breath condensate. For example, levels of nitrite and ammonia in condensate are altered in patients with asthma. Finding out where in the respiratory tract exhaled aerosol particles and gaseous products primarily originate is an important challenge, as is elucidation of to what extent the oropharyngeal compartment contributes to markers of physiological status. We wanted to determine the contribution of NO from the nasal, oral and lower airway compartment both during exhalation and inhalation. We also investigated low levels of exhaled NO in smokers to determine whether this effect is due to an effect in the oropharyngeal tract. Furthermore, we wanted to elucidate the chemical and anatomical origin of nitrite and nitrate measured in nasal and exhaled breath condensate and their relation to NO formation. Finally, we wished to study the chemical and anatomical origin of ammonia measured in exhaled breath condensate and its possible influence on pH. We show a substantial contribution of both nasal and oral NO during both exhalation and inhalation, and that nasal NO output is higher during inhalation compared to exhalation. We were not able to show a relation between NO formation from nitrite in saliva and the low levels of exhaled NO seen in smokers. However, we do find indications for a significant iNOSmediated NO formation in the squamous epithelium of the oropharyngeal tract. Thus, the reduced exhaled NO levels observed in smokers could still be due to an effect in the oropharyngeal tract. A major part of the nitrite measured in exhaled breath condensate originates above the upper trachea, indicating a proximal generation of respiratory droplets, and is primarily a product of the bacterial reduction of nitrate. Additionally, we show that plasma nitrate is taken up from the circulation by the lower airways, but not by the nasal airways. Moreover, we found that pH measured in exhaled breath condensate is strongly affected by oral ammonia formed by urea-splitting bacteria, thus refuting that pH measurements in exhaled breath condensate would reflect pH homeostasis in the lower airways. In conclusion, the oropharyngeal tract is a major site of origin for many biological markers found in exhaled breath, in the gaseous as well as in the aqueous phase. Some of these exhaled markers (nitrite and ammonia) are formed by bacteria, and since the levels of these markers are altered in asthma, the collected data indicate that patients with airway disease have abnormal bacterial activity in the oropharyngeal tract.