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We report a case of intestinal obstruction in a neonate caused by massive muscular hypertrophy of the entire small intestine. Furthermore, the muscularis propria consisted of three layers, in most part. The pathogenesis of this apparently unique lesion is obscure.
BACKGROUND/PURPOSE: The regulatory events that control the adaptive response after massive intestinal resection are poorly understood. An improved understanding could lead to improved clinical therapies. This study tests the hypothesis that massive intestinal resection increases nutrient transport by upregulation of mRNA for the sodium glucose cotransporter (SGLT1). METHOD: Male Lewis rats (225 to 250 g) were studied. Control animals (Con, n = 6) underwent ileal transection 15 cm proximal to the cecum; resected animals (Res, n = 8) underwent resection of all bowel proximal to the terminal 15 cm of ileum. Animals were followed up for 14 days, pair fed, and weighed daily. At the time of death the terminal ileum was studied quantifying SGLT1 mRNA using reverse transcriptase-polymerase chain reaction (RT-PCR) normalized to B-actin, in vitro glucose transport in Ussing Chambers, and histology. RESULTS: All animals survived; animals that were transected regained preoperative weight by day 10, and underwent resection by day 14. Significant adaptation occurred with increases in villus height, (control jejunum 934 +/- 79; control ileum, 811 +/- 64 v resected ileum, 949 +/- 83 [P < .05]). Nutrient flux (J mucosal --> serosa; Jms) was increased in resected ileum (3-0 Methyl-D-Glucose flux: 0.71 +/- 0.2 v 0.56 +/- 0.04 in Con ileum [P < .05; units, micromol/cm2/hr of 3-0 Methyl-D-Glucose]). This was paralleled by an increase in expression of the SGLT1 gene in resected ileum, 104 +/- 14, versus Con ileum, 74 +/- 9 (P < .05; values normalized versus expression of B-actin). CONCLUSION: SGLT1 expression may be useful as a marker of the adaptive process after massive resection and potentially may be used as an endpoint of therapy after resection clinically.
The incidence of polyposis and the number of polyps per mouse were significantly lower in conventionalized (CVz) mice than in germ-free (GF) mice. There was no significant difference in the average number of polyps between GF and gnotobiotic (GB) mice monoassociated with the various strains of intestinal bacteria. However, the incidence of polyposis and the number of polyps per mouse were significantly lower for mice associated with either chloroform-resistant bacteria (CRB) or fusiform bacteria (FB) than for GF mice. This study demonstrated that polyposis was suppressed by FB and CRB in the small intestine of BALB/c mice.
Intestinal transport and metabolism of modified kyotorphin (KTP) were studied in rats. Modified KTPs studied were C-terminally modified KTP with p-aminophenyl-beta-D-glucoside (KTP-pAPbeta glc), N-terminally modified KTP-pAPbeta glc with t-butyloxycarbonyl group (Boc-KTP-pAPbeta glc) and the N- and C-terminally modified KTP by cyclization (cyclic KTP). KTP-pAPbeta glc was metabolized at a similar rate to that of KTP, and did not appear on the serosal side. Although Boc-KTP-pAPbeta glc was also metabolized, it was more stable than KTP and appeared on the serosal side. Cyclic KTP was also quite stable and appeared on the serosal side. The modified KTPs were evaluated kinetically for absorption consisting of membrane transport and metabolism. Absorption clearance (CL(abs)) of cyclic KTP, Boc-KTP-pAPbeta glc and Boc-KTP was higher than that of KTP (0.247 microl/min/cm) (Mizuma et al., Biochim. Biophys. Acta 1335 (1997) 111-119), which is the theoretical maximum by complete inhibition of peptidase activity, indicating that derivatization of KTP increases the membrane permeability. Furthermore, the data clearly showed that the greater the metabolic clearance (CL(met)) of KTP and the KTP derivatives, the lower the absorption clearance (CL(abs)). These results and further simulation study led to the conclusion that metabolic degradation in the intestinal tissues is more critical than membrane permeability (transport) for oral delivery of peptide drugs. Based on the stability of cyclic KTP in serum, this appears to be a good candidate analgesic peptide drug.
Adult neurons possess the ability to adapt to a changing environment. Loss of target-derived neurotrophic factors due to axotomy or isolation by culturing is known to induce changes in neuropeptide expression in several types of peripheral neurons. The aim of the present study was to investigate changes in the expression of vasoactive intestinal polypeptide (VIP) and nitric oxide synthase (NOS) in cultured myenteric ganglia and dissociated neurons. Myenteric ganglia and neurons from rat small intestine were dissociated and cultured for up to 21 days. Immunocytochemistry was used to determine the total number of neurons and the proportions of subpopulations containing VIP or NOS or both in preparations of whole mounts (controls used to determine the conditions in vivo), myenteric ganglion culture and dissociated myenteric neuronal culture. In situ hybridization was used to determine changes in the expressions of NOS and VIP mRNA. The relative number of VIP-expressing neurons increased significantly during culturing. The percentage of all neurons expressing VIP was 3.6+/-0.3% in whole mounts, 22-24% in cultured myenteric ganglia, and up to 35% in cultured dissociated neurons. NOS-expressing neurons constituted approximately 30-40% of all neurons in whole mounts as well as in cultured ganglia or dissociated neurons. A dramatic increase in NOS/VIP-containing neurons were detected in cultured neurons irrespective of whether they were arranged in ganglia or dissociated, as compared to whole mount preparations. This suggests that the NOS-containing neurons are the ones that increase their VIP expression. The induced expression of VIP in cultured adult myenteric neurons indicates that VIP is important for neuronal adaptation, maintenance and survival.
The purpose of this work was to solubilize vasoactive intestinal peptide (VIP) receptors from rat small intestinal plasma membranes and to analyze the nature and function of its molecular form(s) in a nondenaturing environment. Membranes were incubated with 3 nM 125I-VIP, washed, and treated with 1% Triton X-100. Chromatography on Sephadex G-50 showed that 60% of the extractable radioactivity was eluted with macromolecular components in the void volume. This radioactive material was dramatically reduced when 1 microM unlabeled VIP was present in the incubation medium or when membranes were pretreated with trypsin or dithiothreitol. Macromolecular components that had bound 125I-VIP were further chromatographed on Sephacryl S-300. Two peaks were observed: a major one (80%) and a minor one (20%) with Stokes radii of 5.2 and 3.1 nm, respectively. The labeling of both components was inhibited by unlabeled VIP or peptide with NH2-terminal histidine and COOH-terminal isoleucine amide (a VIP agonist). The presence of GTP (0.1 mM) in the incubation medium of membranes completely abolished the labeling of the 5.2-nm component but did not affect that of the 3.1-nm one. Moreover, GTP induced dissociation of 125I-VIP from the 5.2-nm component isolated by Sephacryl S-300 chromatography. This effect was time dependent and nucleotide specific. In contrast, GTP did not affect the stability of the 3.1-nm component. After cholera toxin catalyzed [32P]ADP-ribosylation of membranes, chromatography of solubilized material on Sephacryl S-300 showed that a peak of 32P radioactivity was coeluted with the 5.2-nm component.(ABSTRACT TRUNCATED AT 250 WORDS)
In epithelial cells isolated from rat small intestine, we have studied the influence of vasoactive intestinal peptide (VIP), a neurotransmitter which markedly increases enterocyte cyclic AMP, and of two cyclic AMP analogues (8-bromo cyclic AMP and N6,2'-O-dibutyryl cyclic AMP) on the rate of glycolysis, fructose 2,6-bisphosphate concentration and 6-phosphofructo-2-kinase activity, as well as on the rate of 3-O-methyl-D-[14C]glucose uptake. Our results show that, without affecting the rate of 3-O-methyl-D-[14C]glucose accumulation, VIP and cyclic AMP analogues were able to inhibit glucose consumption and L-lactate formation by isolated rat enterocytes. These effects occurred parallel to a significant decrease in the cellular concentration of fructose 2,6-bisphosphate and to a partial inactivation of 6-phosphofructo-2-kinase. These findings support the hypothesis that VIP inhibits glycolysis in rat enterocytes through a cyclic AMP-dependent mechanism.
Human angiotensin I-converting enzyme (ACE) exists in intestinal epithelial cells as a membrane-bound (ACEm) and secretory glycoprotein (ACEsec). The electrophoretic mobilities of ACEsec and ACEm on SDS/polyacrylamide gels are similar; the N-deglycosylated ACEsec and ACEm, in contrast, display slight differences in their apparent molecular masses, indicating that the carbohydrate contents of ACEsec and ACEm are different. Moreover, ACEsec is solely N-glycosylated whereas ACEm is N- and O-glycosylated, assessed by lectin binding studies. Spontaneous release of ACEsec is achieved by incubation of brush border membranes at 37 degrees C. Aprotinin, leupeptin and EDTA partly inhibit the generation of ACEsec, strongly suggesting that ACEsec is generated from ACEm by proteolytic cleavage. The expression levels of ACEsec in the intestine may be associated with the differentiation state of mucosal cells. Thus ACEsec is more abundant than ACEm in immature non-epithelial crypt cells of patients with coeliac disease. Well-differentiated epithelial cells, by contrast, contain predominantly ACEm. The variations in the proportions of cleaved ACEsec in differentiated and non-differentiated cells may be due to varying levels of the cleaving protease. Alternatively, because epithelial cell differentiation is accompanied by alterations in the levels of oligosaccharyltransferases, the results suggest a critical role for the glycosylation pattern of ACEm in its susceptibility to the putative cleaving protease.
A fatal case of the megacystis-microcolon syndrome in a newborn girl is presented and two previously unreported findings are discussed. The umbilical hernia is thought to represent a mechanically induced embryological defect, secondary to the expanding nonfunctional megacystis which appears to be the dominant component from an early stage. Hypertrophic changes of the large intestinal wall musculature, adjacent to dilated segments, may indicate an impaired propulsive activity in utero, as a result of defective neurotransmission. The latter is proposed as the common pathophysiological mechanism for both urinary and intestinal malfunction.
Spontaneous perforation of the small intestine in the newborn is rare. In most cases the etiology is unknown. Three cases of perforation in preterm newborn infants are described. The clinical diagnosis was necrotizing enterocolitis (NEC), and an urgent laparotomy was performed. The histologic examination was compatible with the diagnosis of a congenital defect of the intestinal musculature. All three infants survived.
Intestinal mucosal uptake of protein complexed-fatty acids was studied in ligated duodenal loops in the chick. Increasing the concentration of an albumin oleic acid-complex resulted in a linear increase in uptake of oleic acid. Varying the albumin-to-oleic-acid ratio with constant albumin concentration resulted in depressed oleic acid uptake when the ratio was below 1:3. Uptake of oleic acid complexed to albumin was increased by some 60% on addition of taurocholic acid above its critical micellar concentration. In the absence of albumin, oleic acid uptake was some 60% high from a micellar solution. Uptake of lauric acid from aqueous solution was linear with concentration until its maximum solubility was reached, whereas uptake from albumin complexes at varying lauric acid concentrations was not linear with increasing concentration. Stearic acid exhibited lowest uptake and linoleic and linolenic acid highest uptake both when complexed to albumin or from micellar solution, although albumin-complexed fatty acids were transported at about half the rate of micellar fatty acids. We concluded that some proportion of fatty acids complexed to lipophilic proteins can be absorbed in the intestine in the absence of bile acids. When oleic acid was complexed to casein, bovine serum albumin or beta-lactoglobulin at protein:oleic acid ratio of 1:10 serosal transport was 40 to 50% of mucosal uptake.
Irritable bowel syndrome (IBS), which affects up to 25% of the population in western countries and accounts for up to 50% of referrals to gastroenterologist, remains mainly a diagnosis of exclusion. We have studied, for the first time, prolonged ambulatory motility recordings from the oesophagus, small intestine and colon of a patient who was referred to us with a 12-year history of abdominal pain and a presumptive diagnosis of IBS. The results indicated that the diagnosis was pseudo-obstruction syndrome rather than IBS. Wider clinical use of the new techniques of prolonged ambulatory intestinal motility monitoring in IBS would offer considerable potential not only in achieving a greater understanding of its pathophysiology but also in providing a more precise definition of clinical and therapeutic subgroups.
Laser Doppler fluxmetry (LDF) has been used to assess mucosal blood-flux in the intestine of normal CD1 mice and groups of animals with altered iron metabolism (i.e. iron-deficient, hypoxic and hypotransferrinaemic [hpx/hpx]). All experimental animals showed a 2-3-fold increase in duodenal iron absorption, mainly due to changes in the 'mucosal transfer' phase. However, only the hypoxic mice exhibited any increases in duodenal blood-flux. In the hpx/hpx group, blood flux was maintained at control levels despite the anaemia and without any significant alterations in red blood cell velocity. Moreover, these two groups demonstrated a further capacity to increase blood-flux above basal levels during onset of acute hypoxia (inhalation of 10% O2). Changes in duodenal blood-flux were apparent on both the mucosal and serosal surfaces, suggesting that the LDF signal reflects flux throughout the thickness of the mouse intestine. Iron absorption is likely to be more related to plasma flux changes than to blood-flux changes. Deduction of plasma flux changes from blood-flux changes is complicated by vasodilation effects on capillary haematocrits. It is clear from the data, however, that plasma flux changes do not parallel changes in iron absorption in the experimental models. The lack of correlation between the duodenal blood-flux and iron absorption values in the experimental models argues against the possible involvement of blood-flux in the control of duodenal iron absorption.
(1) A method is described for counting nuclei in wax-embedded histological sections of epithelium. The counts are independent of section thickness over a wide range (4-10 mum). (2) counts of nuclei were made in villi and crypts of rat small intestine fixed either collapsed or distended by a hydrostatic pressure equivalent to 350 mm of water. There was no significant difference between the counts of nuclei in collapsed or distended gut. (3) Distension reduced the absolute height of villi, and increased their absolute width at the base. (4) The implications of these findings for studies of the morphological basis of intestinal adaptation are discussed.
The cleavable cross-linking reagent dithiobis (succinimidyl propionate) or DTSP was shown to link 125I-labeled vasoactive intestinal peptide (125I-VIP) covalently to its receptors in rat intestinal epithelial membranes. DTSP treatment of 125I-VIP-labeled membranes inhibited the dissociation of VIP-receptor complexes in a way which was dependent on both time and concentration (ED50 = 200 microM). Polyacrylamide gel electrophoresis of membrane proteins revealed three 125I-VIP-protein complexes of Mr 76 000, 36 000 and 17 000. The labeling of those compounds was not observed when: (a) treatment of membranes by DTSP was omitted; (b) the reagent quench, ammonium acetate, was added together with DTSP; (c) DTSP-treated membranes were incubated with 2-mercaptoethanol which reduces the disulfide bond present within DTSP. Labeling of Mr-76 000 and Mr-36 000 complexes was specific in that it could be abolished by native VIP, while the labeling of the Mr-17 000 was not. Densitometric scanning of autoradiographs indicated that: (a) labeling of the Mr-76 000 complex was abolished by low VIP concentrations (0.03--10 nM), by VIP agonists with the relative potency VIP greater than a peptide having N-terminal histidine and C-terminal isoleucine amide greater than secretin, and by GTP (10(-5)--1 mM) but was unaffected by various other peptide hormones; (b) labeling of the Mr-36 000 complex was inhibited by high VIP concentrations (1--300 nM), by VIP agonists at high concentrations but was not affected by GTP and various peptide hormones. Assuming one molecule of 125I-VIP was bound per molecule of protein, two proteins with Mr-73 000 and 33 000 were identified as VIP binding sites. The Mr-73 000 protein displays many characteristics (affinity, specificity, discriminating power toward agonists, sensitivity to GTP regulation) of the high-affinity VIP receptors mediating adenylate cyclase activation. The Mr-33 000 protein displays the characteristics (affinity, specificity) of a low-affinity VIP binding site. This study thus shows the molecular characteristics of the VIP receptor and further argues for the molecular heterogeneity of VIP binding sites.
The H(+)-dependent uptake system responsible for the enteral absorption of oligopeptides and orally active beta-lactam antibiotics was functionally reconstituted into liposomes. Membrane proteins from rabbit small intestinal brush border membrane vesicles were solubilized with n-octyl glucoside and incorporated into liposomes using a gel filtration method. At protein/lipid ratios of 1:10 and 1:40, the uptake of the orally active alpha-amino-cephalosporin, D-cephalexin into proteoliposomes was stimulated by an inwardly directed H+ gradient and was protein-dependent. In these proteoliposomes the binding protein for oligopeptides and beta-lactam antibiotics of Mr 127,000 could be labeled by direct photoaffinity labeling with [3H]benzylpenicillin revealing an identical binding specificity as in the original brush border membrane vesicles. The uptake system for beta-lactam antibiotics and oligopeptides showed a remarkable stereospecificity; only D-cephalexin was taken up by intact brush border membrane vesicles, whereas the L-enantiomer was not taken up to a significant extent. This stereospecificity for uptake was also seen after reconstitution of solubilized brush border membrane proteins into liposomes demonstrating a functional reconstitution of the peptide transporter. Both enantiomers however, bound to the 127-kDa binding protein as was shown by a decrease in the extent of photoaffinity labeling of the 127-kDa protein in the presence of both enantiomers. After reconstitution of subfractions of brush border membrane proteins obtained by wheat germ lectin affinity chromatography into proteoliposomes, only liposomes containing the 127-kDa binding protein showed a significant uptake of D-cephalexin whereas the L-enantiomer was not transported. The uptake rates for D-cephalexin into proteoliposomes correlated with the content of 127-kDa binding protein in these liposomes as was determined by specific photoaffinity labeling with [3H]benzylpenicillin. The purified 127-kDa binding protein was also reconstituted into liposomes and its ability for specific binding of substrates as well as stereospecific uptake of cephalexin could be restored. These results indicate that the binding protein for oligopeptides and beta-lactam antibiotics of Mr 127,000 mediates the stereospecific and H(+)-dependent transport of orally active beta-lactam antibiotics across the enterocyte brush border membrane. We therefore suggest that this 127-kDa binding protein is the intestinal peptide transport system (or a component thereof).
Peritonitis induced by serosal application of 0.1 M hydrochloric acid causes net fluid secretion via the enteric nervous system. The aim of the present study was to investigate the roles of vasoactive intestinal peptide (VIP) and tachykinins in this reflex(es). The release of tachykinins (substance P [SP], neurokinin A [NKA], neuropeptide K [NPK]) and VIP into the mesenteric circulation, net fluid transport, intestinal blood flow and sometimes motility were recorded simultaneously in extrinsically denervated jejunal segments of the cat in vivo. The release of both VIP and NKA was increased upon application of HCl to the cat jejunal serosa. Tetrodotoxin, hexamethonium and methionine enkephalin inhibited both the induced VIP release and the secretory response. The increased release of NKA was unaffected by hexamethonium. We propose that the intramural secretory reflex evoked by acid application of the serosa consists of an 'afferent' tachykinin neuron, a cholinergic interneuron and an 'efferent' VIPergic neuron innervating the secretory enterocytes.
Cysteine-rich intestinal protein (CRIP) is a newly identified peptide that occurs primarily in the intestine and binds zinc in the mucosa during absorption. High dietary zinc does not affect CRIP concentration, but it greatly increases metallothionein, which may compete with CRIP to decrease zinc absorption.