Rotaviruses specifically bind to the neutral glycosphingolipid asialo-GM1.

Rotaviruses are the major etiologic agents of severe diarrhea in children. Many rotaviruses encode a hemagglutinin which binds to sialic acids. We report that rotaviruses specifically recognize the neutral glycosphingolipid gangliotetraosylceramide (asialo-GM1 or GA1). GA1 resolved by thin-layer chromatography is bound by rotavirus, and binding is blocked by neutralizing rotavirus antiserum. Similar glycosphingolipid structures, such as globoside, gangliotriaosylceramide, and GA1 oxidized with galactose oxidase are ineffective in binding rotavirus. Other enteric viruses also specifically bind GA1. GA1 adsorbed to polystyrene beads inhibits rotavirus replication in vitro (as do anti-GA1 antibodies). The use of orally administered immobilized GA1 or anti-GA1 antibodies may prove useful in preventing or attenuating rotaviral and other enteric viral infections.

Molecular forms of cholecystokinin in rat intestine.

The predominant immunoreactive cholecystokinin (CCK) forms in acid extracts of rat intestine eluted from reverse-phase high-performance liquid chromatography columns in the positions of CCK-8, CCK-33/39, and CCK-58. Control experiments indicated that smaller CCK forms did not arise from artifactual degradation of large CCK forms. Less than 10% of CCK-8 added to and extracted from intestine was recovered in acid; CCK-8 could not be recovered by subsequent alkaline extraction, but subsequent urea extraction yielded approximately 25% of the added peptide. This suggests that CCK binds to proteins during acid extraction and that the preponderance of large CCK forms in acid extracts is not due to inhibition of CCK degradation but results from poor extraction of small CCK forms. No evidence for a CCK-22-like peptide was found in acid or subsequent urea extracts of rat intestine, suggesting CCK posttranslational processing in adult rats is like that in humans and dogs.

Intestinal metaplasia and dysplasia in gastric ulcer and its tissue repair.

The aim of this work was to study the relationship between intestinal metaplasia and dysplasia in gastric ulcers and their tissue repair in 223 patients with 236 gastric ulcers found endoscopically and treated with H2 blockers. The average duration of follow-up for the men was 32.4 months (range, 12-87 months) and for the women 42.5 months (range, 12-88 months). In 112 patients (50.2%) with 118 gastric ulcers, intestinal metaplasia in the different types was observed. The data obtained allow us to state that severe dysplasia and gastric cancer can occur only in a restricted number of patients with intestinal metaplasia in gastric ulcers and/or gastric ulcer tissue repair (two in our study, more than 60 yr old), and only in the forms with sulphomucins, more precisely type III. In relation to the fact that gastric ulcers rarely become carcinoma, the intestinal metaplasia frequently observed should not be considered "precancerous", as such, but could become so in the presence of several factors which, excluding age, did not emerge from our study.

Developmental changes of Ca2+, PO4, and calcitriol metabolism in spontaneously hypertensive rats.

We have measured Ca and P balance, serum calcitriol, and vitamin D-dependent intestinal calcium-binding protein (CaBP9k) in the spontaneously hypertensive rat (SHR) and its normotensive control, the Wistar-Kyoto rat (WKY) at 4-5 and again at 13-14 wk of age. In rats on a 1% Ca diet, P balance was significantly more positive in the 5-wk-old SHR than WKY (P less than 0.01); Ca balance tended to be greater in the 5-wk-old SHR. In contrast, in the 14-wk-old SHR, P and Ca balance were less positive than in the WKY (P was 5.8 +/- 1.3 vs. 13 +/- 1.7 mg/day, P less than 0.01, and Ca was 53 +/- 4.6 vs. 67 +/- 2.7 mg/day, P less than 0.05). On the 1% Ca diet, plasma calcitriol levels of the 5-wk-old SHR were higher than those of the WKY (58 +/- 3.2 vs. 40 +/- 2.1 pg/ml, P less than 0.002), whereas at 12 wk there was no difference. On Ca-deficient (0.1%) diets, plasma calcitriol was increased in 5-wk-old and 12-wk-old SHR and WKY, compared with the 1% Ca diet (P less than 0.001). Calcitriol metabolic clearance rate was the same in the 13-wk-old SHR and WKY on either Ca diet. Intestinal CaBP9k content of the 5-wk-old (38 +/- 2.6 vs. 40 +/- 3.9 micrograms/mg) and the 12-wk-old (11 +/- 1.6 vs. 14 +/- 1.4 micrograms/mg) SHR and WKY were similar on the 1% Ca diet.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of crystalline rat liver fatty acid binding protein produced in Escherichia coli.

The principal absorptive cell of the rat small intestinal epithelium contains two homologous cytosolic proteins that bind long chain fatty acids. These are known as intestinal and liver fatty acid binding proteins (FABP). While their precise physiological roles have not been defined, they are believed to represent a multifunctional cytosolic transport system that is involved in the trafficking of exogenous lipids to sites of metabolic processing. 13C NMR studies have revealed differences in their fatty acid binding stoichiometries, binding mechanisms, and the ionization properties of bound fatty acids. To understand the functional differences, liver FABP has been crystallized for eventual comparison with the known crystal structure of intestinal FABP. The lattice type is trigonal with unit cell dimensions of a = b = 84.1 A and c = 44.2 A. The space group as determined by examination of the Patterson symmetry is either P3(1)21 or P3(2)21.

A review of the effects of glutamine-enriched diets on experimentally induced enterocolitis.

Studies in animal models of enterocolitis have failed to confirm the purported metabolic and functional benefits of bowel rest induced by use of an elemental diet. Recent reports have demonstrated that glutamine-supplemented diets ameliorate or reverse many of the adverse effects of experimentally induced enterocolitis. Human studies are needed to confirm these findings.

Traces of mercury in organs from primates with amalgam fillings.

In order to trace possible accumulations of mercury, three vervet monkeys received occlusal amalgam fillings, three others maxillary bone implants of amalgam, and three untreated monkeys served as controls. One year later all animals were sacrificed by transcardial perfusion with glutaraldehyde. Tissue sections from different organs were subjected to silver amplification by autometallography and analyzed at light and electron microscopical levels. It was found that amalgam fillings (total, 0.7-1.2 g) caused deposition of mercury in the following tissues: spinal ganglia, anterior pituitary, adrenal, medulla, liver, kidneys, lungs, and intestinal lymph glands. In monkeys with maxillary silver amalgam implants (total, 0.1-0.3 g), mercury was found in the same organs except for liver, lungs, and intestinal lymph glands. Organs from the three control animals were devoid of precipitate. To evaluate whether silver released from the corroding amalgam fillings added to the staining pattern, tissue sections were exposed to potassium cyanide prior to being autometallographically developed. This treatment removes all traces of silver, leaving mercury sulfide accumulation untouched. By comparing sections that had been exposed to cyanide with untreated parallels no difference was seen in the pattern confirming that mercury was the only catalyst present in the tissue. These results strongly support what has been suggested previously that dental fillings in primates cause absorption of mercury released from amalgam fillings through lungs and intestinal tract, and that depending on exposure mercury is distributed to most organs and will eventually be found in the central nervous system. The present data also show that silver released from the corroding filling is not absorbed.

Gaucher's disease type I. Report of a case with prominent deposition of ceroid in splenic endothelial cells and intestinal smooth muscle fibers.

A case of type I Gaucher's disease in a 39-year-old male is reported. Autopsy showed marked enlargement of the spleen (3,070 g) and infiltration of typical Gaucher's cells in the spleen, liver, bone, marrow, gastrointestinal tract, lymph nodes and adrenal glands. The diagnosis of Gaucher's disease was ascertained by the very low beta-glucosidase activity of cultured subcutaneous fibroblasts and the high content of glucocerebroside in the spleen tissue. A peculiar finding in this case was prominent deposition of brown pigment in endothelial cells of the spleen and smooth muscles fibers of the gastrointestinal tract, urinary bladder and prostate. Histochemical examination revealed that the granules in endothelial cells and smooth muscle fibers were ceroid. Such deposition of ceroid has never been reported previously in Gaucher's disease. Ceroid deposition in generalized smooth muscle fibers is known as brown bowel syndrome, and is highly suggestive of severe vitamin E deficiency. Although other symptoms of vitamin E deficiency were not noticed in this case, some malnutritional condition might play a role in prominent deposition of ceroid in lysosomes, possibly together with deficient activity of a lysosomal enzyme.

Sexual and age variations of organ iron content in Shaver chickens.

1. Haematological values and iron content in liver, spleen, kidneys and intestine were determined in Shaver chickens of both sexes at 4, 8, 13 and 18 weeks and in females at 24 weeks (the beginning of the laying period). 2. The haematocrit decreased significantly in laying compared with non-laying females and the haemoglobin concentration was similar to that in the prelaying state. Plasma iron in laying females increased to four times the basal value at 13 weeks. 3. Females of 13 and 18 weeks (prelaying state) stored more iron than males at the same age. A simultaneous liver and spleen mobilisation of stored iron and increased intestinal iron accumulation took place in the laying process. The haematological variables examined were minimally altered. 4. The iron contents of both heart and kidneys were influenced by age and followed a linear trend, except that in the heart of females where a quadratic response was observed.

Identification of chicken calbindin D28K pre-messenger RNA sequences by polymerase chain reaction.

A transcribed RNA sequence encompassing the junction between the first intron and the second exon of the chicken calbindin D28K gene was copied in a cDNA fragment and subsequently amplified by polymerase chain reaction. When intestinal RNA is used as template, the appearance of the 161 bp amplified fragment is strictly dependent on the vitamin D status of the animal. In fact no amplified fragment is obtained when the RNA is extracted from the intestine of vitamin D-deficient chickens, while it is easily detected when the RNA is extracted only 30 min after injection with 1,25-dihydroxycholecalciferol. Conversely, the amplified fragment is obtained, irrespectively of the vitamin D status of the animal, when the RNA template is extracted from the brain. The appearance of unspliced RNA sequences upon vitamin D induction is followed, after a 30 min lag, by the appearance of the corresponding mature mRNA sequences.

Rat heparan sulphates. A study of the antithrombin-binding properties of heparan sulphate chains from rat adipose tissue, brain, carcase, heart, intestine, kidneys, liver, lungs, skin and spleen.

Adult male rats were given [35S]sulphate intraperitoneally. Heparan [35S]sulphate (HS) chains were recovered from adipose tissue, brain, carcase, heart, intestine, kidneys, liver, lungs, skin and spleen by digestion with Pronase, precipitation with cetylpyridinium chloride, digestion with chondroitin ABC lyase and DNAase and gradient elution from DEAE-Sephacel. Purity was confirmed by agarose-gel electrophoresis and degradation with HNO2. Fractionation by gradient elution from antithrombin-agarose indicated that the proportion of HS with high binding affinity for antithrombin (HA-HS) ranged from 4.7% (kidneys) to 21.5% (brain). On a mass basis the major sources of HA-HS were carcase, skin and intestine. HA-HS from intestine was arbitrarily divided into subfractions I-VI, with anticoagulant activities ranging from 1 to 60 units/mg [by amidolytic anti-(Factor IIa) assay] and from 4 to 98 units/mg [by amidolytic anti-(Factor Xa) assay], indicating that the antithrombin-binding-site densities of HA-HS chains covered a wide range, as shown previously for rat HA-heparin chains [Horner, Kusche, Lindahl & Peterson (1988) Biochem. J. 251, 141-145]. HA-HS subfractions II, IV and VI were mixed with samples of HA-[3H]heparin chains and rechromatographed on antithrombin-agarose. Affinity for matrix-bound antithrombin did not correlate with anticoagulant activity, e.g. HA-HS subfraction IV [38 anti-(Factor Xa) units/mg] was co-eluted with HA-heparin chains [127 anti-(Factor Xa) units/mg].

Presence and binding characteristics of calcitriol receptors in human fetal gut.

In the present study, we show for the first time the presence of calcitriol-specific binding sites in hypertonic extracts of cells isolated from human fetal small intestine and colon from 13-21 weeks of gestation. Woolf plot analysis of the binding characteristics revealed the presence of a single class of high affinity receptors. The presence of specific receptors for calcitriol in fetal intestine and colon opens interesting possibilities as to the role of this hormone in human gut development.

Characterization of an endogenous quail intestinal lectin.

Soluble extracts of quail intestine scrapings contain a lectin activity specific for chicken and rabbit trypsinized, glutaraldehyde-fixed erythrocytes. The lectin displayed a specificity for the simple sugar haptens lactose and galactose and for mucin. Quail lectin was purified by affinity chromatography on either asialofetuin- or mucin-Sepharose, followed by DEAE-Sepharose chromatography, and demonstrated an apparent molecular weight of 14,500 on sodium dodecyl sulfate - polyacrylamide gel electrophoresis and a pI of 6.2 upon isoelectric focusing. Immunohistochemical localization of this lectin in the intestine was carried out using polyclonal antibody raised in rabbits and tested for specificity in Western blots. Immunoperoxidase staining for quail lectin showed the lectin to be prominent in secretions at the mucosal surface and in goblet cells.

Improved survival in intestinal ischemia by allopurinol not related to xanthine-oxidase inhibition.

Allopurinol, a xanthine-oxidase (XO) inhibitor, has been used to improve the resistance to ischemia with disappointing results that have been attributed to administration regimen of the drug. Our aim was to investigate the effect of different administration schedules of allopurinol on the survival in rats undergoing intestinal ischemia testing the blockade of XO. Intestinal ischemia was achieved by 90 min of clamping the superior mesenteric artery (SMA) close to its origin from the aorta. Three groups of animals were evaluated: A-group: only the allopurinol solvent was given; B-group: the full dose of allopurinol (100 mg/k b.w.) was given iv and C-group: the 75% dose was administered orally 24 hr before and the remaining 25% was administered 30 min before. Survival was evaluated at 48 hr and the blockade of XO was assayed by High Efficacy Liquid Chromatography (HELC) in homogenate of intestinal wall. Survival was only improved in the C-group (P = 0.02). Levels of hypoxanthine were significantly increased both in B-group and C-group (P = 0.003) when compared with the A-group. Levels of uric acid in B-group (P = 0.0003) and C-group (P = 0.0009) were significantly decreased with respect to A-group. That means that an effective blockade of XO is achieved whichever the regimen of administration. Allopurinol and oxypurinol levels were significantly increased (P = 0.05 and P = 0.008) in C-group when compared with B-group. We conclude that the protective effect of allopurinol on survival in intestinal ischemia in rats is not related to the blockade of XO but rather to the allopurinol and oxypurinol levels in intestinal wall.

Distribution of calcitonin gene-related peptide and calcitonin-like immunoreactivity in trout.

Radioimmunoassay and chromatography were used to study the occurrence of calcitonin gene-related peptide in various tissues of the rainbow trout, Salmo gairdnerii. The highest concentrations of the peptide were found in gill (1.68 +/- 0.09 ng/mg protein) and in intestine (1.06 +/- 0.4 ng/mg protein). Significant concentrations were also found in heart and stomach. The level in brain was very low. In trout, the plasma concentration accounted for 283 +/- 82 pg/ml. Chromatographic analysis of the calcitonin gene-related peptide (CGRP)-like immunoreactivity occurring in gills showed that two molecular forms cross-reacted with the anti-human CGRP antibody, one co-eluting with the synthetic human CGRP. In addition, calcitonin in fish is not confined to the ultimobranchial organ but is also present in organs as heart, intestine, kidney, spleen and stomach. The evidence of CGRP in fish emphasizes the role of this hormone in evolution and leads us to investigate its physiological role in this species.

Comparative distribution of carbonic anhydrase isozymes III and II in rodent tissues.

Carbonic anhydrase (CA) III was demonstrated immunocytochemically in epithelium in some regions of salivary gland ducts, colon, bronchi, and male genital tract and in adipocytes, in addition to skeletal muscle and liver where the isozyme was previously localized. Basal cells beneath the submandibular gland's excretory ducts in guinea pig stained for CA III. Carbonic anhydrase III occurred alone in some and with CA II in other sites but was often absent from CA-II-containing types of cells. This was exemplified by CA III's abundance in CA-II-positive proximal colon and its sparsity in the CA-II-rich distal colon of the mouse. Striated ducts in guinea pig, but not mouse salivary glands, stained darker for CA and appeared accordingly to function more actively in ion transport compared with excretory ducts. Carbonic anhydrase content varied among genera in liver and pancreas and between mouse species and strains in salivary glands and kidney. Newly observed murine sites of CA II activity included Auerbach's plexus and a population of leukocytes infiltrating the lamina propria in small intestine, and several types of cells in the male genital tract. In immunoblot tests, antisera to CA III showed no cross reactivity with antisera to CA II, but those to CA II disclosed weak cross reactivity with CA III.

The role of intestinal microflora in the metabolic activation of 6-nitrochrysene to DNA-binding derivatives in mice.

6-Nitrochrysene has previously been shown to be a potent lung and liver carcinogen following i.p. administration to newborn mice and to be metabolically activated to DNA-binding derivatives by nitro-reduction or a combination of nitro-reduction and ring oxidation. In this study, we have examined fecal metabolites and DNA-carcinogen adducts in 5-week-old conventional and germfree Balb/c mice treated with [3H]6-nitrochrysene in order to determine if the metabolic activation pathway(s) for this compound in these mice differs from that observed in preweanling mice. We further evaluated the role of the intestinal microflora on the metabolism and generation of DNA-reactive metabolites in this system. The amount of 6-aminochrysene excreted in the feces of germfree mice within 48 h after treatment with a single i.p. dose of [3H]6-nitrochrysene (0.03 mumol/5 microliters/g body wt) was approximately 25% of that excreted in identically treated conventional mice. However, the levels of carcinogen-DNA adducts in the lungs and livers of conventional and germfree Balb/c mice were similar at the 24 and 48 h time points examined. HPLC analysis of hydrolysates of liver and lung DNA indicated that adducts derived from both N-hydroxy-6-aminochrysene and trans-1,2-dihydroxy-1,2-dihydro-6-aminochrysene metabolites were formed in the liver whereas only the latter adduct was detected in the lung. This contrasts with previous findings in preweanling mice where the adduct derived from the trans-1,2-dihydroxy-1,2-dihydro-6-aminochrysene metabolite was the single major adduct detected in both liver and lung DNA. The proportion of adducts derived from N-hydroxy-6-aminochrysene was significantly greater in the liver DNA of germfree mice than in the liver DNA of conventional mice.

Extrahepatic replication of duck hepatitis B virus: more than expected.

Replication of duck hepatitis B virus in extrahepatic tissue such as pancreas, kidney and spleen has been well documented. To assess whether there is more widespread extrahepatic virus replication, we assayed brain, heart, lung, thymus, pancreas, kidney, spleen and intestine of 1- to 16-wk-old ducklings for the presence of duck hepatitis B virus DNA and mRNA by blotting and in situ methods. Replicative intermediates and single-stranded duck hepatitis B virus DNA and RNA transcripts were detected in the brain, lung, heart, intestine, kidney, pancreas and spleen. In situ hybridization showed evidence of viral replication in the lung epithelium, germinal center of spleen, acinar cell of pancreas and tubular epithelium of kidney. These data suggest that extrahepatic duck hepatitis B virus replication is more widespread than previously thought. It is yet to be determined whether widespread extrahepatic replication is unique to duck hepatitis B virus infection or is a common feature of other mammalian hepatitis B-like viruses.

Isolation and partial characterization of a lactotransferrin receptor from mouse intestinal brush border.

Several lines of evidence have recently suggested the occurrence of a specific lactotransferrin receptor in the small intestinal brush-border membrane in several animal species, which is thought to be involved in lactotransferrin-mediated intestinal iron absorption. We report here for the first time the isolation and partial characterization of this receptor from mouse intestinal brush border. The receptor has been purified to homogeneity by affinity chromatography on an immobilized human lactotransferrin column. The purified receptor was found to be active in that it binds iron-free and iron-saturated lactotransferrin with a Kd of 0.1 microM. Anti-receptor antibodies were prepared, and the receptor was further isolated by immunoaffinity chromatography in higher yield but in a denatured form. The purified receptor was revealed by sodium dodecyl sulfate-polyacrylamide electrophoresis to be a protein of about Mr = 130,000, consisting of a single polypeptide chain. The isoelectric point was determined to be 5.8. The receptor was further shown to bear concanavalin A and phytohemagglutinin L binding glycans. Digestion by N-glycanase and endo-N-acetyl-beta-D-glucosaminidase B led to a decrease of Mr = 25,000, while the endo-N-acetyl-beta-D-glucosaminidase H was uneffective, suggesting that the lactotransferrin receptor is mainly glycosylated by bi- and triantennary glycans. To gain further insight into the interaction of the receptor with lactotransferrin, namely, the number of ligand molecules bound per molecule of receptor, mouse lactotransferrin was cross-linked to its membrane-bound enterocyte receptor by use of radiolabeled sulfosuccinimidyl 3-[[2-(p-azidosalicylamido)ethyl]dithio]propionate (SASD).(ABSTRACT TRUNCATED AT 250 WORDS)