Expression of nitric oxide synthases and effects of L-arginine and L-NMMA on nitric oxide production and fluid transport in collagenous colitis

BACKGROUND AND AIMS: 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. PATIENTS: 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.     

n-3 fatty acids in adipose tissue and coronary artery disease are inversely related.

The incidence of coronary heart disease is supposedly influenced by dietary fish-oil intake. The purpose of this study was to investigate the possible beneficial effect of fish consumption on coronary atherosclerosis. Adipose-tissue biopsies and segments of coronary arteries were sampled from 40 consecutive autopsies. Degree of coronary artery disease was estimated quantitatively by morphometry, and the fatty acid composition of adipose tissue was analyzed with gas-liquid chromatography. The group with the highest degree (68-92% stenosis, n = 13) of coronary artery disease had lower concentrations of docosahexaenoic acid in adipose tissue than did the group with the lowest degree (23-53% stenosis, n = 13) of coronary artery disease (P less than 0.05). Multiple-regression analysis showed that the degree of coronary artery disease was dependent on docosahexaenoic acid in adipose tissue (P less than 0.01) and body weight (P less than 0.05). The study supports the hypothesis that fish consumption is associated with a reduced risk of coronary heart disease.     

The impact of an end-point committee in a large multicentre, randomized, placebo-controlled clinical trial: results with and without the end-point committee's final decision on end-points.

BACKGROUND: A multicentre study permits rapid recruitment of a large number of patients. However, there is a risk of heterogeneities in end-point evaluations, as complex definitions of criteria are interpreted by several local investigators from different hospitals. Reports discussing end-point evaluation are sparse. The TRIM trial was a multicentre trial of a thrombin inhibitor in patients with unstable angina or non-Q myocardial infarction. In this study, an independent end-point committee evaluated all the reported events of death, acute myocardial infarction and refractory angina pectoris in order to obtain uniform judgements of major end-points. STUDY AIMS: To describe the work of the end-point committee, to analyse its possible effect on the final study results and to discuss the impact on the design of future trials. METHOD: The end-point committee consisted of four members, one from each participating country. After the data were processed by the study monitors, completed case record forms and patient files for patients with reported end-points were mailed to the national member of the end-point committee for judgement. The end-point committee met regularly and made final decisions about the end-points. The work of the end-point committee was documented on a separate case record form. RESULTS: The end-point committee assessed 246 events of death, acute myocardial infarction and refractory angina pectoris in 187 of the 1209 patients (15.5%) in the TRIM trial. Misinterpretation of the index event, an inclusion myocardial infarction, as an early cardiac event was found in 12 patients. After end-point committee judgements, the number of patients with acute myocardial infarction or refractory angina pectoris during 30 days of follow-up was reduced from 177 to 153 (13. 6% reduction). The classification of events was changed in 53 of 187 patients (28.3%) with death, acute myocardial infarction or refractory angina pectoris. The data assessed by the safety committee was significantly different from the final database after end-point committee judgements. CONCLUSION: The end-point committee corrected misinterpretations in such a high proportion of cases that the final results differed significantly from the preliminary results delivered to the safety committee. End-point judgements by an end-point committee should be performed in multicentre clinical trials to improve the quality and reliability of study results.     

Eaton-Lambert syndrome and malignant thymoma

Eaton-Lambert syndrome is most frequently associated with small-cell carcinoma of the lung. We describe the first case of this disorder in a patient with malignant thymoma. Guanidine treatment had no clinical or electrophysiologic effects.     

Expression of inducible nitric oxide synthase and effects of L-arginine on colonic nitric oxide production and fluid transport in patients with "minimal colitis"

OBJECTIVE: Some patients with idiopathic, chronic diarrhoea have minimal, non-specific colonic inflammation. As nitric oxide (NO) acts as a secretagogue in the colon, we studied the expression of inducible NO synthase (iNOS) in mucosal biopsies and the effects of NOS stimulation on colonic transfer of fluid and output of NO in patients with "minimal colitis". MATERIAL AND METHODS: Twelve patients with idiopathic, chronic diarrhoea and "minimal colitis" and 6 healthy volunteers were included in the study. Expression of iNOS in colonic mucosal biopsies was quantified by Western blot analysis and localized by immunohistochemistry. The effects of topical L-arginine or placebo on colonic net fluid transfer and nitrite/nitrate (NOx) output were assessed during "steady state" perfusion of the whole colon. Concentrations of NOx were measured by Griess' assay. RESULTS: The expression of iNOS was increased 10-fold (p<0.01) in patients with "minimal colitis" compared with that in healthy volunteers and localized to the colonic epithelium. Colonic absorption of fluid was impaired (mean (SEM) 1.5 (0.2) versus 3.0 (0.2) ml/min, p<0.001) and the output of NOx was increased (47 (4) nmol/min versus <37 nmol/min, p<0.05) in patients with "minimal colitis" compared with that in healthy volunteers. Luminal L-arginine (20 mM) reduced colonic absorption of fluid in both groups (95% confidence intervals (CIs) 21-50% in patients with "minimal colitis" versus 4-18% in healthy volunteers), but an increase in NOx output was detectable only in the group of patients (8-106%). In time control experiments, colonic net transfer rates of fluid and outputs of NOx were unaffected by placebo. CONCLUSIONS: In patients with idiopathic, chronic diarrhoea and histopathological evidence of "minimal colitis", colonic absorption of fluid is impaired, while epithelial expression of iNOS and mucosal production of NO is enhanced. It could be speculated that NO in excess contributes to the diarrhoea observed in "minimal colitis".     

Craniofacial tissues including tooth buds in fetal hypohidrotic ectodermal dysplasia

OBJECTIVE: Hypohidrotic ectodermal dysplasia (HED) comprises defects in hair, teeth, and sweat glands. Disturbances in other ectodermal tissues have been associated with the condition. Our objective was to examine ectodermal craniofacial structures histologically in a fetus with HED and to compare the findings to similar structures in normal control fetuses. MATERIALS AND METHODS: A male fetus diagnosed with HED was therapeutically aborted in the 15th week of gestation. One male and two female healthy fetuses were used as normal controls. All fetuses were examined with parental consent, and had comparable sizes. Their bone maturation stage in the hand was identical. Tissue blocks from the craniofacial region were excised from all fetuses and prepared for histological analysis (formalin fixed, stained with toluidine blue or Alcian blue). The tissues examined were: tooth buds, skin and skin appendages, oral mucosa including minor salivary glands, major salivary glands, lacrimal glands, and adenohypophysis. RESULTS: Fewer tooth buds, minor salivary glands, and hair follicles were observed in the HED fetus as compared to controls. The structures of the epidermal components in the developing HED organs were loose and disorganised. The adhesion between the ectodermal and mesenchymal organ components in the HED fetus seemed to be disturbed.     

Subclassification of Anencephalic Human Fetuses According to Morphology of the Posterior Cranial Fossa

Anencephaly is a designation for congenital absence of the cranial vault with cerebral hemispheres completely missing or decreased to small masses attached to the base of the skull. The etiology is unknown. Whether the bony tissue or soft brain tissue is a primary factor is also unknown. The present study has focused on the posterior cranial fossa in anencephaly. The goal is to determine whether differences in the posterior cranial fossa could provide a basis for subclassification of anencephalic fetal skeletons. Twenty-three human anencephalic fetuses, at gestational ages 13 to 22 weeks, were studied. Radiologic and cephalometric analyses, including measurements of bone sizes and different angles, were performed. Permission for autopsy of the central nervous system was not available. For comparison of anencephalic findings with normal conditions, standards from a recent publication were used. Foot length served as a parameter for age comparison. The study showed 2 morphologic types of the posterior cranial fossa. One type had a fossa cranial morphology close to normal morphology, whereas the other had a malformed and much smaller posterior cranial fossa. The latter condition was presumed to be due to a primary error in chondral and cranial development. The current skeletal subgrouping might be essential for clinicians’ or pathologists’ future assessment of the autopsy results. The skeletal subgrouping should, if possible, be associated with karyotyping and analysis of the central nervous system. The goal is to distinguish between congenital conditions resulting in anencephaly and acquired conditions resulting in anencephaly. published online November 17, 2004.