Expression of nitric oxide synthases and effects of L-arginine and L-NMMA on nitric oxide production and fluid transport in collagenous colitis.
OBJECTIVE
Luminal nitric oxide (NO) is greatly increased in the colon of patients with collagenous and ulcerative colitis. To define the source and consequence of enhanced NO production we have studied expression of NO synthase (NOS) isoforms and nitrotyrosine in mucosal biopsies from these patients. In addition, effects on colonic fluid transfer caused by manipulating the substrate of NOS were studied in patients with collagenous colitis.


METHODS
Eight patients with collagenous colitis, nine with active ulcerative colitis, and 10 with uninflamed bowel were included.


METHODS
Expression of NOS isoforms was quantified by western blotting. Inducible NOS (iNOS) and nitrotyrosine were localised by immunohistochemistry. Modulation of NOS activity by topical N(G)-monomethyl-L-arginine (L-NMMA) or L-arginine was assessed during perfusion of whole colon. Plasma and perfusate nitrite/nitrate (NOx) was measured by Griess' reaction.


RESULTS
Both in collagenous and ulcerative colitis, expression of iNOS was 10(2)-10(3) higher (p<0.001) than in uninflamed bowel and localised primarily to the epithelium. Endothelial NOS was evenly expressed in all groups while neuronal NOS was undetectable. Nitrotyrosine was markedly expressed in active ulcerative colitis but rarely detected in collagenous colitis and never in uninflamed bowel. In collagenous colitis, the output of NOx was markedly increased compared with uninflamed bowel (283 (58) v <37 nmol/min; p<0.01) and fluid was net secreted. L-NMMA reduced the output of NOx by 13-66% (95% confidence intervals) and secretion of fluid by 25-109% whereas L-arginine increased the output of NOx by 3-39% and secretion of fluid by 15-93%.


CONCLUSIONS
In collagenous colitis, as opposed to ulcerative colitis, upregulation of iNOS occurs in the absence of nitrotyrosine formation and mucosal damage. Excess generation of NO may be the primary cause of diarrhoea in this condition.
Perner
A
A
Department of Gastroenterology, Herlev Hospital, DK-2730 Herlev, University of Copenhagen, Denmark.
Andresen
L
L
Normark
M
M
Fischer-Hansen
B
B
Sørensen
S
S
Eugen-Olsen
J
J
Rask-Madsen
J
J
D000328:Adult; D000368:Aged; D001120:Arginine / Q000502:physiology*; D015153:Blotting, Western; D001826:Body Fluids / Q000378:metabolism*; D016022:Case-Control Studies; D003093:Colitis, Ulcerative / Q000378:metabolism*; D016001:Confidence Intervals; D005260:Female; D006801:Humans; D007408:Intestinal Absorption / Q000502:physiology; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D018709:Statistics, Nonparametric; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism; D015854:Up-Regulation; D019323:omega-N-Methylarginine / Q000502:physiology*
D019323:omega-N-Methylarginine; D009569:Nitric Oxide; C002744:3-nitrotyrosine; D014443:Tyrosine; D001120:Arginine; C496320:NOS1 protein, human; C496319:NOS2 protein, human; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III