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Abstract:

We have used a chemiluminescence technique to measure gaseous nitric oxide (NO) directly in hollow organs of humans. NO production was studied in the airways of healthy controls and patients with chronic inflammatory airway disease, in the stomach of healthy subjects, in the colon of patients with inflammatory bowel disease, and in the urinary bladder of patients with cystitis. We have found that almost all NO found in exhaled air is derived from the upper airways, with only a minor contribution from the lower airways and the lungs. Thus, in subjects with a permanent tracheostomy, exhaled NO levels were much higher during nasal or oral breathing than when the subjects breathed through the tracheostomy. The major NO source in the upper airways seems to be situated in the paranasal sinuses, whereas the nasal mucosa seems to release much less NO. Sinus derived NO enters the nasal cavity via the sinus ostia. The paranasal sinuses are quantitatively the most important contributors to the high levels of NO found in nasally exhaled air. In immunohistochemical and mRNA in situ hybridization studies we show that an enzyme most closely resembling the inducible NO synthase (NOS) is constitutively expressed apically in sinus epithelium. Furthermore, NO synthase activity in sinus mucosa is mostly Ca2+-independent, which is characteristic of the inducible NOS. The regulation of sinus NOS, however, differs profoundly for what has earlier been described for inducible NOS. Thus, sinus NOS is continuously expressed and seems resistant to glucocorticoids. Sinus NO concentrations (1000-30000 parts per billon) greatly exceed those that are bacteriostatic indicating a role for NO in host-defence. It is an intriguing possibility that the sterility of the paranasal sinuses is maintained by means of the profuse epithelial NO production. NO derived from the upper airways will normally reach the lungs with each breath, especially during nasal breathing. We here show that arterial oxygenation increases during nasal breathing compared to oral breathing. Furthermore, in intubated patients, who are deprived of autogenous, nasally produced NO, arterial oxygenation increases and pulmonary vascular resistance may decrease when NO-containing air, derived from the patients' own nose, is added to the inspired air. This action of NO in the airways might be termed "aerocrine", and may represent a novel physiological principle, namely that of an enzymatically produced airborne messenger. In fact, these findings may help to explain one biological role of the paranasal sinuses, the major sources of NO in the airways.

We have also found that a high production of NO takes place normally in the stomach lumen. We analysed NO levels in air that was expelled from the stomach after ingestion of carbonated water. Interestingly, stomach NO production is not catalysed by NO synthases. Instead, intragastric NO is derived from nitrite present in swallowed saliva. When acidified in the stomach, nitrite is reduced to NO and accordingly, NO production is blocked if gastric acid secretion is prevented by the proton pump inhibitor omeprazole. This is the first evidence of non- enzymatic NO production in humans. NO produced in the stomach may be involved in host-defence against swallowed pathogens and in the regulation of mucosal blood flow and mucus secretion.

We also show that chronic inflammatory diseases of the airways are associated with alterations of NO excretion at different levels of the repiratory tract. Thus, asthmatic children show increased orally exhaled NO levels whereas children with Kartagener's syndrome or cystic fibrosis show markedly reduced nasal NO levels. In the asthmatics, there was an inverse relationship between orally exhaled NO levels and daily doses of inhaled corticosteroids. This supports the idea that an inducible NOS is expressed in the lower airways of asthmatics, since this NOS isoform is known to be down regulated by steroids. Administration of L-arginine i.v. resulted in increased NO excretion in the upper airways, which is interesting considering the possible involvement of NO in airway host defence. Thus, L-arginine supplementation might increase resistance to airway infections. Luminal levels of NO are greatly increased in the colon of patients with ulcerative colitis or Crohn's colitis, and inthe urinary bladder of patients with cystitis as compared to controls.

In conclusion: Large amounts of NO are excreted in the upper airways of humans. This NO originates mainly in the paranasal sinuses, where an "inducible-like" steroid-resistant NO synthase is constitutively expressed in healthy subjects. The high local concentrations of NO in the sinuses may contribute to the sterility of these cavities. When NO is diluted in inhaled air, this vasodilator gas may serve to modulate pulmonary function. This novel action of NO in the airways may help to explain one biological function of the enigmatic human paranasal sinuses. A non-enzymatic production of NO takes place in the acidic stomach through the reduction of nitrite present in swallowed saliva. Stomach NO may be involved in local host defence and in regulation of mucosal blood flow and mucus secretion.

Inflammation is associated with altered mucosal production of NO. Measurements of airborne NO may be used as a simple test to detect mucosal inflammation in hollow organs. This may be of clinical value in the diagnosis and monitoring of inflammation e.g. in the airways, gastrointestinal tract and in the urinary bladder.

Keywords: ARDS, asthma, cystic fibrosis, cystitis, exhaled air, inflammatory bowel disease, inhaled nitric oxide, Kartagener's syndrome, nitric oxide synthase, sinusitis,