Nitric oxide production is diminished in colonic circular muscle from acquired megacolon

PURPOSE: Nitric oxide modulates human colonic smooth muscle function. To determine whether nitric oxide production is altered in colon from acquired megacolon, we measured cholinergic nerve-mediated contractionsin vitro before and after inhibition of nitric oxide synthase. METHODS: Intramural nerves in circular smooth muscle from histologically normal colon (n=12) and acquired megacolon (n=3) were activated by electrical field stimulation. RESULTS: In controls blockade of nitric oxide synthase by N^G-Nitro-L-Arginine induced increases (P<0.05) in amplitude of contractions; these increases in amplitudes were blocked by L-Arginine (analysis of variance;P<0.05). By contrast, blockade of nitric oxide synthase did not increase amplitudes of contractions with circular smooth muscle from acquired megacolon. An immediate phasic contraction was blocked by atropine sulfate. CONCLUSIONS: The results support the concept that nitric oxide production modulates cholinergic nerve-mediated contractions in normal colonic circular muscle, whereas acquired megacolon is associated with altered release of this inhibitory neurochemical. Potential explanations include depletion of tissue L-Arginine, decreased capacity to recycle citrulline to arginine, or decreased release of vasoactive intestinal peptide from circular smooth muscle in acquired megacolon.Supported by Veterans Administration Medical Research Funds and by West Virginia University.Presented at Digestive Disease Week, Washington, D.C., May 11 to 14, 1997.     

Decreased colonic peptide histidine-methionine in idiopathic inflammatory bowel diseases

The sequence for peptide histidine-methionine is present within the same preprohormone as vasoactive intestinal polypeptide. Since our previous study using radioimmunoassay had demonstrated significantly decreased colonic concentrations of vasoactive intestinal polypeptide in ulcerative colitis and Crohn's colitis compared to normal colon, we determined the distribution and quantitation of peptide histidine-methionine. Fresh surgical specimens were dissected into mucosal-submucosal and muscularis externa layers prior to acid extraction and specific radioimmunoassay. One immunoreactive species that appeared to coelute with peptide histidine-methionine was separated by reverse-phase high-performance liquid chromatography. Mucosal-submucosal concentrations of peptide histidine-methionine were significantly decreased in ulcerative colitis and Crohn's colitis, compared to those in normal colon. In normal ileum and colon, linear correlation analysis showed no relationship between patient age and tissue concentrations of peptide histidine-methionine. However, a parallel decrease in molar concentrations of peptide histidine-methionine and vasoactive intestinal polypeptide in ulcerative colitis and Crohn's colitis was demonstrated by linear correlation analysis. These results are consistent with the hypothesis that peptide histidine-methionine and vasoactive intestinal polypeptide are colocalized within the same neural structures that have been altered in the idiopathic inflammatory bowel diseases.T.R. Koch is a recipient of a Career Development Award from the National Foundation for Ileitis & Colitis.     

Inhibitory neuropeptides and intrinsic inhibitory innervation of descending human colon

The effects of aging on inhibitory neuropeptide concentrations and intrinsic inhibitory innervation of circular muscle were investigated using normal descending colon obtained at surgery. Immunoreactive vasoactive intestinal peptide, peptide histidine-methionine, met⁵-enkephalin, neuropeptide Y, and somatostatin were extracted from specimens of muscularis externa (patient ages: 19–84 years) and measured by radioimmunoassay. Intracellular electrical activity was recorded from strips of circular muscle (patients ages: 49–84 years) using glass microelectrodes; inhibitory junction potentials were evoked by electrical field stimulation. There were no significant differences (t tests:P>0.05) between neuropeptide concentrations in patients<70 years old (N=28) compared to patients70 years old (N=12). However, the amplitude of inhibitory junction potentials declined with increasing patient age (r=–0.58,P=0.02,N=16), with no change in resting membrane potentials (r=0.22;P>0.05). The decline in amplitude in women (r=–0.68,P=0.03,N=9) preceded the decline in men (r=–0.62,P=0.10,N=7). Age-related decline in inhibitory junction potentials could be related to decreased: density of inhibitory nerves, release of inhibitory neurotransmitter, density of binding sites for inhibitory neurotransmitter on smooth muscle, or a combination thereof. Alternatively, this decline might represent a change in interaction of inhibitory neurotransmitter with the smooth muscle membrane, such as a change in coupling of binding site with the potassium channel, decreased number of potassium channels, or altered permeability of the potassium channel.This study was supported in part by the National Institutes of Health (DK 17238 and DK 34988), and by VA Medical Research funds.     

Idiopathic chronic constipation is associated with decreased colonic vasoactive intestinal peptide

To investigate the reported association between idiopathic chronic constipation and morphologic abnormalities of enteric nerves, we measured the concentrations of six neuropeptides, vasoactive intestinal peptide, peptide histidine-methionine, substance P, methionine5-enkephalin, neuropeptide Y, and the bombesinlike intestinal peptides, in descending colon from 4 patients with idiopathic chronic constipation. Decreased concentrations of vasoactive intestinal peptide (707 +/- 112 ng/g wet tissue) and peptide histidine-methionine (543 +/- 58 ng/g) were found in the muscularis externa obtained from constipated patients compared with normal concentrations (40 patients) of vasoactive intestinal peptide (1199 +/- 47 ng/g) and peptide histidine-methionine (815 +/- 45 ng/g). Vasoactive intestinal peptide was identified by immunocytochemistry in nerve fibers within the circular smooth muscle layer of descending colon obtained from 6 control patients, but not in nerve fibers within the circular smooth muscle of descending colon obtained from 3 patients with idiopathic chronic constipation. By contrast, the distribution of immunoreactive met5-enkephalin was similar in normal descending colon and in descending colon obtained from patients with idiopathic chronic constipation. Decreased colonic concentrations of vasoactive intestinal peptide (a candidate nonadrenergic, noncholinergic inhibitory neurotransmitter) may be associated with diminution of inhibitory innervation of colonic circular smooth muscle in some patients with idiopathic chronic constipation.     

Enteric innervation in idiopathic megarectum and megacolon

We have studied the resection specimens from 5 patients with idiopathic megarectum and megacolon and 10 control subjects with non-obstructing colonic cancer. Histological staining with haematoxylin and eosin, and immunocytochemical staining for protein gene product 9.5 (PGP 9.5), S100 protein, vasoactive intestinal polypeptide (VIP) and calcitonin gene-related peptide (CGRP), and histochemical localization of NADPH diaphorase was performed. The amount of VIP and CGRP present in samples was measured using an enzyme-linked immunosorbent assay. Patients with idiopathic megarectum and megacolon showed hypertrophy of the muscularis mucosae and muscularis externa. The architecture of the innervation as assessed by immunoreactivity for PGP 9.5 and S100 protein appeared normal. There was a decrease in the density of innervation of the longitudinal muscle in rectal tissue from patients with idiopathic megarectum, with fewer VIP- and NADPH-diaphorase-containing nerves. In the muscularis mucosae and lamina propria of the rectal samples of patients with idiopathic megarectum, VIP immunoreactivity was higher and more NADPH-diaphorase-containing nerves were seen. CGRP-immunoreactive nerve fibres were only seen in the myenteric plexus. No CGRP-immunoreactive cell bodies were seen. In summary, there is an increase in VIP and nitric oxide containing fibres in the muscularis mucosae and lamina propria and a decrease in the longitudinal muscle in rectal tissue of patients with idiopathic megarectum. Both are NANC (nonadrenergic noncholinergic) inhibitory transmitters in the gut and the possible relationship of the changes in their density with gut function is discussed.

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Résumé. Nous avons étudié les pièces opératoires de cinq patients traités pour un mégarectum idiopathique et un mégac?lon et celles de dix sujets témoins traités pour un cancer colique non obstructif. Les coupes histologiques ont été colorées à l'hématoxine-éosine et avec des colorants immuno-histo-chimiques pour la protéine génique 9.5 (PGP 9.5), la protéine S100, les polypeptides intestinaux vaso-actifs (VIP) et le peptide de la calcitonine (CGRP) ainsi que pour la localisation histo-chimique de la diaphorase NADPH. Les concentrations de VIP et de CGRP présents dans les échantillons ont été mesurées au moyen d'un essai immuno-absorbant enzymatiquement lié. Les patients avec un mégarectum idiopathique et un mégac?lon présentaient une hypertrophie de la muscularis mucosae et de la musculature externe. L'architecture de l'innervation déterminée par l'immuno-réactivité pour le PGP 9.5 et la protéine S100 appara?t comme normale. Il y a une diminution de la densité de l'innervation de la musculature longitudinale du rectum chez les patients avec un mégarectum idiopathique, ainsi qu'une diminution des nerfs contenant de la VIP et de la diaphorase-NADH. Dans la muscularis mucosae et dans la lamina propria, des échantillons de rectum des patients avec un mégarectum idiopathique, l'immuno-réactivité pour la VIP était plus haute et les nerfs contiennent davantage de diaphorase NADPH. L'immuno-réactivité pour le CGRP et fibres nerveuses n'a été retrouvée que dans le plexus myentérique. Aucune inclusion celulaire immuno-réactive pour CGRP n'a été observée. En résumé, il y a une augmentation des fibres contenant VIP et des oxydes nitriques. Dans la muscularis mucosae et la lamina propria et une diminution pour ces substances dans la musculature longitudinale des patients avec un mégarectum idiopathique. Tous deux sont des transmetteurs inhibiteurs NANC de l'intestin (non adrénergiques et non cholénergiques) et une relation possible dans les changements de leur densité avec la fonction intestinale est discutée.

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Accepted: 18 August 1996     

Immunohistochemical study of the muscularis externa of the esophagus in achalasia patients

The etiology of achalasia is believed to be the neuropathy associated with chronic inflammation of the nerve plexus, but the cause of plexus inflammation is unknown. The purpose of this study was to evaluate the pathophysiology of achalasia by examining the muscularis externa of the esophagus. We used the muscularis externa of the esophagus of 62 patients with achalasia (median 44 years, male : female 32:30) who underwent surgical treatment (achalasia group) and of 10 patients (median 65.5 years, male : female 9:1) who underwent esophagectomy for thoracic esophageal cancer (control group) to perform immunohistochemical staining with S‐100, CD43, c‐kit (CD117), n‐NOS, vasoactive intestinal polypeptide (VIP), and ubiquitin. The cell counts that were positive for S‐100, n‐NOS, VIP, and ubiquitin were significantly lower in the achalasia group compared with the control group (P < 0.001, P= 0.001, P < 0.001, and P= 0.001, respectively). There were no statistically significant differences with respect to CD43 and c‐kit staining (P= 0.586 and P= 0.209, respectively). In conclusion, the pathophysiology of achalasia is therefore considered to be an impaired production of NO and VIP, which both affect interstitial cell of Cajal and smooth muscles, and this impairment is therefore considered to play a role in the pathophysiology of achalasia.     

Induction of Enlarged Intestinal Lymphoid Aggregates During Acute Glutathione Depletion in a Murine Model

Glutathione is a nonenzymatic antioxidant synthesized by most animal cells and is depleted in inflammatory bowel disease. The effects of glutathione depletion on intestinal histology and inhibitory neurochemicals was examined in a mouse model. Glutathione depletion in A/J mice involved inhibition of -glutamylcysteine synthetase using l-buthionine-(S,R)-sulfoximine (BSO) for 10 days. Ileum and colon were obtained from saline-control mice, BSO-treated mice, and BSO-treated mice receiving ascorbate or glutathione monoethylester. Glutathione, lipid peroxides, and nicotineamide adenine dinucleotide phosphate diaphorase activity were measured by colorimetric assays. Vasoactive intestinal peptide was measured by radioimmunoassay. Glutathione depletion induced enlargement of mucosal–submucosal lymphoid aggregates without germinal centers in ileum and colon. These aggregates were prevented by supplementation with glutathione monoethylester but not ascorbate. Tissue levels of inhibitory neurochemicals were unchanged. Depletion of glutathione appears to induce enlarged lymphoid aggregates by recruitment of lymphocytes from the peripheral circulation. A component of the inflammation that develops in inflammatory bowel disease could be related to depletion of tissue levels of glutathione.     

Total Antioxidant Capacity of Colon in Patients with Chronic Ulcerative Colitis

It has been proposed that oxidative stress is involved in the pathophysiology of ulcerative colitis. We have reported the depletion of the nonenzymatic antioxidant, glutathione, in colon from active and inactive ulcerative colitis. The colon contains several biochemically linked antioxidant systems. We hypothesized that diminished total antioxidant capacity in active ulcerative colitis would be associated with increased colonic lipid peroxidation. This study was designed to determine total antioxidant capacity and lipid hydroperoxide levels using colon obtained at surgery from controls (N = 16; 4 females, 12 males; mean age 70 years), and active and inactive ulcerative colitis (N = 15; 3 females, 12 males; mean age 39). Total antioxidant capacity of control colon was higher in muscularis externa compared to the mucosal–submucosal layer (P < 0.05). There were no differences in colonic total antioxidant capacity or lipid hydroperoxide levels comparing control colon to inactive and active ulcerative colitis. The results did not support depletion of tissue total antioxidant capacity by free radicals. Depletion of glutathione in ulcerative colitis may be a specific disorder rather than a secondary defect attributable to global oxidative stress. Nonspecific antioxidant supplements appear unlikely to be beneficial in the treatment of ulcerative colitis.     

Distributions of neuropeptides in the human esophagus

The distributions of nerve cells and fibers with immunoreactivity for the peptides substance P, somatostatin, enkephalin, vasoactive intestinal peptide, gastrin-releasing peptide, and neuropeptide Y and the enzyme tyrosine hydroxylase were examined in 25 samples of human esophagus. These were compared with samples of stomach and intestine. In the smooth muscle of the muscularis externa, the muscularis mucosae, and beneath the epithelium, the most abundant nerve fibers contained vasoactive intestinal peptide and neuropeptide Y, in contrast to the scarcity of substance P, enkephalin, somatostatin, and gastrin-releasing peptide. Gastric and intestinal samples contained dense populations of fibers containing vasoactive intestinal peptide, neuropeptide Y, substance P, and enkephalin in the equivalent layers, but somatostatin- and gastrin-releasing peptide-immunoreactive fibers were scarce. Complete coexistence of vasoactive intestinal peptide and neuropeptide Y in nerve fibers within the muscle layers was demonstrated in the esophagus, but not in gastric and intestinal samples. The myenteric plexus along the length of the esophagus contained cell bodies and fibers reactive for vasoactive intestinal peptide, neuropeptide Y, enkephalin, and substance P. Somatostatin-immunoreactive cell bodies were very rare in the myenteric plexus, no gastrin-releasing peptide-immunoreactive cell bodies were seen, and both somatostatin and gastrin-releasing peptide-immunoreactive fibers were rare. In the upper esophagus, striated muscle bundles did not contain nerve fibers reactive for these peptides but immunoreactive fibers were seen in the muscularis mucosae and subepithelium. It is concluded that the esophagus has a different pattern of innervation by peptide-containing neurons than the stomach and intestines. Esophageal neurons can be classified into separate classes on the basis of their peptide content.     

Distribution and quantitation of gut neuropeptides in normal intestine and inflammatory bowel diseases

To study hyperplasia of peptidergic nerves purported to be diagnostic of Crohn's disease, we determined the distribution and concentrations of gut neuropeptides in specimens of normal intestine, ulcerative colitis, and Crohn's disease. Tissue specimens obtained at surgery were dissected into the mucosal-submucosal and muscularis externa layers, and immunoreactive gut neuropeptides were acid-extracted for measurement by radioimmunoassay. The immunoreactive species were characterized by column chromatography. Mucosal-submucosal layer concentrations of vasoactive intestinal peptide were significantly decreased in Crohn's colitis and ulcerative colitis, while mucosal-submucosal layer concentrations of substance P were significantly increased in left-sided ulcerative colitis. Muscularis externa layer concentrations of vasoactive intestinal peptide and met5-enkephalin were decreased in left-sided Crohn's colitis. These neuropeptide concentration abnormalities did not clearly differentiate between Crohn's colitis and ulcerative colitis, and no increase in concentration of a neuropeptide diagnostic of Crohn's disease was identified.     

Enteric neural modulation of slow-wave activity in cat colon

These studies test the hypothesis that the generation of colonic slow waves can be modulated by stimulation of intrinsic enteric nerves and attempt to identify a neurotransmitter that may be responsible for this change in slow-wave activity. Isolated segments from the mid-colon of the cat generated regular, continuous slow waves at 6.5±1.1 cpm. Activation of the intrinsic nerves by electrical field stimulation transiently reduced the rate of slow-wave generation to 4.7±0.7 cpm (P<0.001). The response to electrical stimulation was blocked by tetrodotoxin and -chymotrypsin. The following antagonists were not effective in blocking the response: atropine, hexamethonium, phenoxybenzamine, propranolol, methysergide, naloxone, or imidazole. Vasoactive intestinal polypeptide (5×10^–7 M) decreased slow wave frequency to 4.5±0.4 cpm. Vasoactive intestinal polypeptide (VIP) fragment 10-28 inhibited the effect of electrical field stimulation but also decreased the slow-wave frequency. VIP-immunoreactive nerves were much more abundant in the plexus submucosus extremus than in the circular muscle of the muscularis externa. Thus, pacemakers for colonic slow waves may be modulated by intrinsic colonic nerves, and vasoactive intestinal polypeptide may be the neurotransmitter responsible for this modulation.This work was supported by NIH grants DK 1750 and Digestive Diseases Core Center grant AM 34986 and a grant from the Veterans Administration.     

Vasoactive intestinal peptide

Dose—response characteristics of feline corpus circular muscle were studiedin vitro for three neuropeptides individually and with vasoactive intestinal peptide. Bombesin, substance P, and cholecystokinin-octapeptide each elicited concentration-dependent isometric contractions that were reduced by 10^–8 M or 10^–7 M vasoactive intestinal peptide (P<0.01). The concentration of each neuropeptide producing a half-maximal response was increased more than one logfold to 10^–6 M by vasoactive intestinal peptide. Tetrodotoxin blocked responses to bombesin (P<0.001) and reduced responses to substance P (P<0.05), but had no effect on responses to cholecystokinin-octapeptide (P>0.1). These results demonstrate inhibition of neuropeptide responses of gastric smooth muscle and support vasoactive intestinal peptide as an inhibitory regulator of gastric motor function.     

Altered inhibitory innervation of circular smooth muscle in Crohn's colitis. Association with decreased vasoactive intestinal polypeptide levels

To determine whether decreased tissue vasoactive intestinal polypeptide levels might affect inhibitory neural input, fresh colonic specimens were obtained from patients with Crohn's colitis (n = 7) and normal subjects (n = 13). Immunoreactive vasoactive intestinal polypeptide levels were measured in the muscularis externa by radioimmunoassay and localized in tissue sections by immunostaining. Circular muscle strips were maintained in an organ bath; inhibitory junction potentials evoked by short- and long-duration field stimulation and resting membrane potentials were recorded using intracellular impalements. In Crohn's colitis, vasoactive intestinal polypeptide levels displayed a bimodal distribution in which 3 specimens had vasoactive intestinal polypeptide levels greater than or equal to 4 SE lower than the mean in normal specimens. In 3 specimens from Crohn's colitis with decreased vasoactive intestinal polypeptide levels, immunoreactive material was absent from the circular muscle layer and the myenteric plexus. Mean resting membrane potentials, mean amplitude of inhibitory junction potentials evoked by short-duration stimulation, and mean amplitude of initial inhibitory junction potentials evoked by long-duration stimulation were not different between the two groups. However, the mean amplitude of the 60th inhibitory junction potential during prolonged stimulation was decreased (p less than 0.01) in Crohn's colitis (6 mV) compared with normal specimens (11 mV). These results show that diminished neural input to circular muscle in Crohn's colitis was associated with decreased extractable vasoactive intestinal polypeptide levels and decreased staining of nerve fibers containing vasoactive intestinal polypeptide.     

Peptidergic innervation of human sphincter of Oddi

The innervation of the sphincter of Oddi (SO) has been extensively studied experimentally, but human studies have not been published, which is why this study was undertaken. Biopsies, taken by gastroscopy-biopsy forceps from duodenal epithelium of the papilla of Vater and from ampullary epithelium after sphincterotomy, did not demonstrate nerves and could not be used for studying SO innervation. Therefore SO specimens were obtained from brain-dead organ donors (N=5) and from autopsies (N=14). By staining with a myelin marker S-100, a rich network of nerves was demonstrated in SO. The occurrence of vasoactive intestinal polypeptide (VIP), peptide histidine-isoleucine (PHI) (or its immunologically similar human equivalent peptide histidine methioninamide, PHM), neuropeptide Y, calcitonin gene-related peptide (CGRP), galanin, substance P, enkephalin, bombesin, and somatostatin were studied by immunohistochemical technique. SO demonstrated immunoreactivity for VIP, PHI (PHM), neuropeptide Y, CGRP, galanin, somatostatin, substance P, and enkephalin, but no immunoreactivity was observed for bombesin. The SO immunoreactivity was similar in specimens from organ donors and from autopsies of victims of violence without pancreatobiliary diseases (N=3) when the specimens were taken within 48 hr of death. Autopsy specimens of SO from subjects with gallstone disease (N=5), recurrent pancreatitis (N=3) or periampullary carcinoma (N=3) also demonstrated similar immunoreactivity. We conclude that VIP-, PHI- (PHM-), neuropeptide Y-, CGRP-, galanin-, substance P-, somatostatin-, and enkephalin-like immunoreactivity occur in human SO. These neuropeptides may have role in the neural control of human SO function.     

Urinary organ specific neoantigen

Urinary organ-specific neoantigen from colorectal cancer patients has been used to make a monoclonal antibody, BAC 18.1. In this study we assessed the potential of this antibody for the diagnosis of colorectal cancer. We evaluated binding in both urine and effluent samples and compared it with effluent carcinoembryonic antigen standardized for both volume (nanograms per milliliter) and protein. Urinary organ-specific antigen as detected by BAC 18.1 was significantly greater in 29 cancer patients (A₄₀₅:0.717±0.500) vs 27 controls [0.121 ±0.273 (P<0.05)]. Considerable overlap of binding of BAC 18.1 was observed in the colonic effluent of patients with CRC (N=13), adenomas (N=26), inflammatory bowel disease (N=8), or having a normal colonoscopic examination (N=24). CEA levels (nanograms per milliliter) were significantly elevated in the effluent samples of patients with a past history of colorectal cancer, as compared to that of normal individuals (P<0.05). The presence of the Mr 30,000 organ-specific neoantigen in colonic effluent was also demonstrated by western blot. Organ-specific neoantigen originates in the colon and is excreted into the urine, so the BAC 18.1 binding levels in the urine may be a diagnostic aid for CRC.The work reported in this paper was supported in part by a grant from the Israeli Cancer Association and Tel Aviv University, and in part by grants from the Israel Cancer Association and the Sackler School of Medicine, Tel Aviv, Israel.     

Detoxifying enzymes in human ovarian tissues: comparison of normal and tumor tissues and effects of chemotherapy

Summary Many anticancer drugs exert their cytotoxic effects via formation of oxygen free radicals. Cellular thiols, glutathione (GSH)-dependent enzymes and other redox enzymes are involved in the metabolism of these anticancer drugs and of the oxygen free radicals that may be generated during their metabolism. We quantified these biochemical parameters in cytosol from human ovarian tissues. We compared non-protein thiol levels, GSH transferase, GSH peroxidase, Superoxide dismutase, catalase, DT diaphorase and aldehyde dehydrogenase activity in serous ovarian tumors (n=15), other malignant ovarian tumors (n=12), benign ovarian tissue (n=10) and histologically normal ovarian tissue (n=12). Mean GSH transferase and DT diaphorase activitie were similar in serous and other malignant ovarian tumors. GSH transferase activity was decreased in malignant tissues relative to normal and benign tissues. Mean DT diaphorase and Superoxide dismutase activities were increased in the malignant tissues, although this was not statistically significant. The mean levels of all anzymes except Superoxide dismutase and aldehyde dehydrogenase in benign tissues were fairly similar to the mean levels found in normal tissue samples. Tissues from patients with serous ovarian tumors, who had received cyclophosphamide and cisplatin prior to surgery, also were analyzed (n=7). Except for aldehyde dehydrogenase, all the parameters measured were decreased in these samples relative to serous tissue from untreated patients. These biochemical analyses may be useful in understanding the mechanisms involved in the response to chemotherapy.Abbreviations ANOVA analysis of variance - GSH glutathione Partial support for this study was obtained from the Comprehensive Cancer Center of Metropolitan Detroit, core grant CA-22453, National Institutes of Health.     

Indium scanning in assessment of acute Crohn's disease

The aim of this study was to determine whether the sensitivity of indium-111 (¹¹¹In) scanning in the assessment of the activity and extent of Crohn's disease correlates with the severity of intestinal lesions as measured by the newly validated Crohn's disease endoscopic index of severity (CDEIS). Nineteen patients with active (CDAI>200) colonic (N=11) or ileocolonic (N=8) Crohn's disease were assessed by colonoscopy and indium scanning. The intestine was divided into five segments in both studies (rectum, sigmoid and left colon, transverse colon, right colon, and ileum). Seventy of the 86 intestinal segments seen at colonoscopy presented macroscopic lesions of Crohn's disease. On third-hour scintigrams¹¹¹In uptake was observed in 52 segments, 51 of which were found to be abnormal at colonoscopy. Predictive positive and negative values of scanning with respect to disease extent assessment were equal to 98% and 44%, respectively. Complete agreement between endoscopic and scintigraphic findings was observed in only six of the 19 patients (32%). Segmental endoscopic indexes of severity (SEIS) were significantly (P<0.001) lower in false negative (7.9±4.2) (mean ±sd) than in true positive (18.0±9.7) segments as defined by scintigraphy. SEIS values above which¹¹¹In uptake was constantly observed did not differ in the different disease locations. When comparing macroscopically abnormal intestinal segments according to their¹¹¹In uptake grade, the corresponding mean SEIS values increased significantly as the grade increased. Scintigraphic activity, as assessed by the fall in splenic activity, was equal to 23±11% (N=19). It correlated significantly with CDEIS (r=0.63,P<0.005), but even more so when the highest SEIS of each patient had been taken into account (r=0.75,P<0.0005). In conclusion, when considering disease extent and activity, scanning results correlate well with the endoscopic severity of intestinal lesions in active Crohn's disease. Nevertheless, minor endoscopic lesions can be missed by¹¹¹In scanning.     

Effects of a transplantable insulinoma upon regulatory peptide concentrations in the gastrointestinal tract of the rat

Summary The rapid growth (0.8±0.3 g/day) of a transplantable insulinoma, which also contained substance P (2.9±2.3 pmol/g) and gastrin-releasing peptide (3.2± 2.1 pmoll/g), resulted in the development of hyperphagia, hyperinsulinaeinia and hypoglycaemia in rats (n=8). After a 14-day growth period, the insulinoma-bearing rats showed an increase (49%; p<0.01) in the weight of the small intestine but no significant change in stomach weight compared with control animals. The content (pmol/organ) of somatostatin, substance P, neurokinin A and vasoactive intestinal peptide in the stomachs of the tumour rats was unchanged. A depletion in the content (53% p<0.01) and concentration (57%; p<0.01) of gastrin-releasing peptide, however, suggested either hypersecretion, possibly mediated through hypoglycaemia-induced vagal stimulation, or inhibition of synthesis. The concentration and content of glucagon-like immunoreactivity (enteroglucagon) in the small intestine of the insulinoma rats increased markedly (47%; p<0.01 and 120%; p<0.01). This increase is consistent with a proposed role of this peptide as a factor trophic to the intestinal mucosa. No significant changes in the concentrations of somatostatin, substance P, neurokinin A, vasoactive intestinal peptide and gastrin-releasing peptide in the small intestine were observed. However, the increase in gut weight resulted in a greater content of vasoactive intestinal peptide (40%; p<0.01) and substance P (37%; p<0.05) in the insulinoma rats.     

Mechanisms and sites of mannitol permeability of small and large intestine in the rat

Mannitol is commonly used as an intestinal permeability probe, yet the mechanisms of its penetration of the intestinal barrier are not entirely clear. Therefore, we studied mannitol's permeability of different segments of the intestine and studied the kinetics and influence of intraluminal factors on mannitol permeabilityin vivo in perfused intestinal segments of rats. There was linear relationship between permeability rate of mannitol and its luminal concentration (y=7.2x+1.7;r=0.98), indicating that passive diffusion is involved in mannitol's permeability. Increased luminal fluid osmolarity from 0.3 to 0.6 osmol/liter resulted in decreased net water flux with a corresponding decrease in mannitol permeability in both jejunum and colon (P<0.01), indicating the prominent influence of solvent drag on net mannitol permeability. The relationship between mannitol permeability and water absorption at different osmolarities was linear in the jejunum and colon. At luminal osmolarity of 0.3 osmol/liter, 34.6% of mannitol permeability was mediated by passive diffusion and 65.4% was mediated by solvent drag in the jejunum. Mannitol permeability was much more dependent on solvent drag in the colon (88.9%) than in the small intestine (65.4%). The net permeability rate of mannitol was similar in the jejunum and ileum but was much higher in the colon (P<0.01). Addition of chenodeoxycholate (5 mM) to the perfusate resulted in a significant decrease in absorption of water (P<0.01) with a corresponding decrease in mannitol permeability (P<0.01). These studies indicate that mannitol permeability of the intestinal barrier is mediated by passive diffusion and solvent drag, with the latter accounting for a greater fraction of the total permeability.This study was supported by the Goldsmith Family Foundation.     

Effects of metronidazole and misoprostol on indomethacin-induced changes in intestinal permeability

In previous open studies, misoprostol and metronidazole reduced nonsteroidal anti-inflammatory drug-induced intestinal permeability changes and inflammation respectively. We assessed the effects of indomethacin treatment (50 mg three times a day) for one week with either coadministered metronidazole (400 mg twice a day, group 1,N=9) or misoprostol (200 g four times a day, group 2,N=7) on intestinal permeability to [⁵¹Cr]EDTA and mannitol in healthy volunteers, using double-blind, placebo-controlled, randomized techniques. Given alone, neither metronidazole nor misoprostol affected [⁵¹Cr]EDTA permeation, whereas indomethacin alone increased it from 1.20 (0.40) [mean percent urinary recovery (sd) groups 1 and 2] to 2.43 (0.72),P<0.002. Coadministered metronidazole (group 1) prevented this increase [1.10 (0.39) before, 1.55 (0.54) after,P>0.05], whereas misoprostol (group 2) did not [1.31 (0.51) before, 3.26 (1.10) after,P=0.005]. No drug regimen altered mannitol permeation. Indomethacin and misoprostol did not affect urinary recovery of intravenously administered probes. The results with metronidazole, if related to its antibacterial effects, support evidence from animal models that bacteria contribute to NSAID-induced intestinal damage. The previously reported reduction of indomethacin-induced increased permeability by misoprostol during a one-day study is not seen when the drugs are used in standard clinical doses for one week.Dr. Davies is partly supported by a grant from Searle and Dr. Wilkie by a grant from Bayer.