Luminal nitric oxide : marker of intestinal inflammation

Abstract

Nitric oxide (NO) is a pluripotent molecule with several important physiological functions in the human body. Large amounts of NO are released by various cell types during inflammation, although the role of NO in the inflammatory process remains unclear. Being a free radical, NO is very short-lived in biological systems and consequently, indirect methods are often used to evaluate the production of this gas in vivo. In contrast, NO in the gaseous phase is rather stable, allowing for direct measurement in air-filled luminal structures such as the airways and the intestines. NO is increased in exhaled air of asthmatics and in gas sampled from the large intestine of patients with colitis. The aim of this thesis was to develop a feasible method for measurement of intestinal NO in man, and to use this method for further investigations of NO in relation to inflammatory responses in the human gastrointestinal tract.

We propose a novel concept for sampling of luminal NO. This method uses a balloon catheter made of thin silicone, which is inserted in the rectum. The balloon is inflated with NO-free air and left for incubation, during which NO will diffuse into the balloon until equilibrium is reached. The gas sample is su sequent y asp rate and immediately analysed for NO content with chemiluminescence technique.

Two different catheters were evaluated in vitro regarding recovery rate for NO. Healthy volunteers, patients with irritable bowel syndrome (IBS) and hospitalised patients without gastrointestinal disease were studied for assessment of NO levels in the non-inflamed intestine. Furthermore, patients with gastrointestinal diseases of different immunological background were investigated. These included infective gastroenteritis, coeliac disease, microscopic colitis and IBD (i.e. ulcerative colitis and Crohn's disease). Due to the special considerations regarding diagnostic procedure in paediatric gastroenterology, we also evaluated the method in children with IBD. In order to determine the time-course of rectal NO levels, we additionally performed repeated measurements in healthy controls and patients treated for active ulcerative colitis.

The in vitro recovery rate of NO for the first catheter used was rather low and varying, but ensuing research granted a catheter with improved recovery rate (above 70%). The latter was used in the majority of the studies. Variation was minor in the recovery rate when compared between individual catheters and between different concentrations. The method was well tolerated and without complications in the some three hundred individuals tested, including very young children.

Using the high recovery catheter, healthy controls displayed rectal NO levels of 89±1 8 parts per billion, (adults and children) and minor individual variation over time. Patients with IBI) in remission displayed similar levels as did patients with IBS and hospitalised patients without gastrointestinal disease.

In sharp contrast, patients with active IBD, microscopic colitis or infective gastroenteritis all displayed greatly increased luminal NO levels (10 to 100-fold increase) as compared to controls or subjects with inactive disease. In ulcerative colitis, rectal NO correlated to disease activity as measured by the Powell-Tuck index (r=0.41, p<0.001).

Patients with treated coeliac disease had basal levels not different from healthy controls. However, 24 hours after local challenge with gluten, the patients, but not the controls, displayed a 20-fold increase of rectal NO. Despite the elevated NO levels none of the patients had any gastrointestinal symptoms after rectal gluten challenge.

As a further evidence of increased NO production, plasma nitrite was increased in infective gastroenteritis and plasma nitrite + nitrate was increased in microscopic colitis.

We conclude that luminal NO is greatly increased in intestinal inflammation, seemingly regardless of aetiology of the inflammatory response, and that luminal NO levels easily can be estimated with rectal sampling. This novel method may be a means for the continued investigation of the role of NO in the intestines in health and disease. Furthermore, rectal NO seems to be a sensitive marker of intestinal inflammation in adults and children, and might thus constitute a rapid, safe and minimally invasive tool for the diagnosing and monitoring of certain intestinal disorders.