bluuebunny/crossref_metadata_2025 huggingface dataset:
Part of quercetin absorbed in the small intestine is conjugated and further secreted in the intestinal lumen
Authors: Crespy Vanessa , Morand Christine , Manach Claudine , Besson Catherine , Demigne Christian , Remesy Christian
Published: December 2017

Rutin and quercetin absorption and metabolism were investigated in rats after in situ perfusion of jejunum plus ileum (15 nmol/min). In contrast to rutin, a high proportion of quercetin (two-thirds) disappeared during perfusion, reflecting extensive transfer into the intestinal wall. Net quercetin absorption was not complete (2.1 nmol/min), inasmuch as 52% were reexcreted in the lumen as conjugated derivatives (7.7 nmol/min). Enterohepatic recycling contribution of flavonoids was excluded by catheterization of the biliary duct before perfusion. After a 30-min perfusion period, 0.71 μM of quercetin equivalents were detected in plasma, reflecting a significant absorption from the small intestine. The differential hydrolysis of effluent samples by glucuronidase and/or sulfatase indicates that the conjugated forms released in the lumen were 1) glucuronidated derivatives of quercetin and of its methoxylated forms (64%) and 2) sulfated form of quercetin (36%). In vitro quercetin glucuronides synthetized using jejunal and ileal microsomal fractions were similar to those recovered in the effluent of perfusion. These data suggest that glucuronidation and sulfatation take place in intestinal cells, whereas no glucurono-sulfoconjugates could be detected in the effluent. The present work shows that a rapid quercetin absorption in the small intestine is very effective together with its active conjugation in intestinal cells.
Selective secretion and replenishment of discrete mucin glycoforms from intestinal goblet cells
Authors: Stanley C. Michael , Phillips Thomas E.
Published: December 2017

Antibodies against MUC2, MUC3, and MUC5AC peptide epitopes stained the secretory contents of all goblet cells in the human colon-derived HT29–18N2 cell line. In contrast, four carbohydrate-specific monoclonal antibodies stained mucin glycoforms in consistent subsets of goblet cells. Cholinergic agonist-evoked decreases in total mucin stores were not always mirrored by proportional changes in mucin glycoforms in the same monolayers. Selective secretion of mucin glycoforms did not result from differences in receptor distribution, since cholinergic stimulation was found to increase intracellular free calcium in all cells and selective secretion was also observed when the cells were directly stimulated with the protein kinase C activator phorbol myristate acetate. The results demonstrate that goblet cells cycle through transient periods in which their exocytotic response is unresponsive to cholinergic or protein kinase C-mediated stimuli. Goblet cells replenished intracellular mucin stores to control levels within 1 h, but the relative proportion of mucin glycoforms was not always restored until 24 h after stimulation.
Duodenal neurons provide nicotinic fast synaptic input to sphincter of Oddi neurons in guinea pig
Authors: Mawe Gary M. , Kennedy Audra L.
Published: December 2017

We have investigated the existence of neural connections between the duodenum and the sphincter of Oddi (SO). Stimulation of duodenal myenteric fiber bundles elicited synaptic responses in SO neurons, which included nicotinic fast excitatory postsynaptic potentials (EPSPs), slow EPSPs, and α2-adrenoreceptor-mediated inhibitory postsynaptic potentials. After 48 h in organ culture, when extrinsic fibers had diminished, only the fast EPSPs persisted. Duodenal mucosal stimulation also elicited nicotinic fast EPSPs in SO neurons. There was no association between the SO neurons that received duodenal input and their chemical coding. A reciprocal projection also exists from the SO to the duodenum. In acute and cultured preparations, duodenal myenteric stimulation caused antidromic responses in 20% of SO neurons. Furthermore, 45.6 ± 10.5 neurons in SO ganglia were retrogradely labeled from dye application sites in the duodenum. It is proposed that bidirectional neural communication occurs between the duodenum and the SO and that duodenal neurons provide excitatory fast synaptic input to SO neurons through a reflex that can be activated at the duodenal mucosa.
Expression of somatostatin receptor subtypes on guinea pig gastric and colonic smooth muscle cells
Authors: Corleto Vito D. , Weber H. Christian , Jensen Robert T.
Published: December 2017

In vivo and in vitro studies have demonstrated that somatostatin can influence motility and smooth muscle contractility of the stomach and colon. Recent studies have proposed that some of these effects may be mediated by somatostatin receptors (sst) directly on the smooth muscle cells. If this is correct, the sst receptor subtypes that are present are unknown. This study aimed to resolve these points. Because nucleotide sequences of guinea pig sst genes are unknown, we used sst subtype-specific primers based on comparisons of human and rat sst subtypes and performed RT-PCR of DNase I-treated total RNA from guinea pig total brain. PCR products were cloned in pCR II and sequenced and showed 87% (sst1), 90% (sst2), 90% (sst3), 99% (sst4), and 80% (sst5), respectively, nucleotide homology to the same region (transmembrane 4–6) of the human sst genes. Homology to rat sequences were lower. PCR products were obtained from first-strand cDNA derived from DNase I-treated RNA from dispersed guinea pig gastric and colonic smooth muscle cells. In gastric and colonic smooth muscle cells, we detected sst1–sst3and sst5, and all were confirmed by sequencing. The presence of sst4was shown by Southern blot analysis and hybridization with a guinea pig sst4-specific primer. RT-PCR from cultured colonic and gastric smooth muscle cells devoid of any neural elements gave identical results. These results demonstrate that in the guinea pig all five sst subtypes are present directly on gastric and colonic smooth muscle cells. Previous studies have suggested that a predominant sst3subtype on gastric and a sst5subtype on colonic muscle cells mediated somatostatin’s contractile effects, but the finding here that all five sst subtypes exist on both of these cells suggests that other sst subtypes have only a small or no contractile effect, sst subtypes in guinea pig have a different pharmacological profile from rat or human sst, or these other sst subtypes have some yet undescribed physiological function in muscle cells.
V. Innate mechanisms of mucosal defense and repair: the best offense is a good defense
Authors: Podolsky Daniel K.
Published: December 2017

Well-coordinated mechanisms have evolved that provide both innate protection against gastrointestinal mucosal injury and facilitation of rapid mucosal repair following mucosal damage. Generic protection from injury is provided by intrinsic structural features of the epithelium that form a highly competent barrier and a complex formed at the apical surface by trefoil peptides that comprise the interface between mucosa and lumen. When the epithelial barrier has been broken, regardless of the nature of the injury, epithelial surface continuity is rapidly reestablished through restitution as cells migrate and elongate. This process is promoted by trefoil peptides at the apical surface and a large array of cytokines and growth factors acting at the basolateral pole. Many of these regulatory peptides are products of the immune and other lamina propria cell populations, which are activated following disruption of the mucosal barrier. Thus efforts to repair the epithelium follow inherently from inflammatory effects after initial damage; the repair process in turn may allow abrogation of further inflammation. Ultimate repair of injury requires both proliferative replacement of damaged epithelial cells and remodeling of extracellular matrix and deeper cell populations to restore normal architecture and a fully functional mucosa.
III. Lessons learned from gastrin gene deletion in mice
Authors: Hinkle Karen L. , Samuelson Linda C.
Published: December 2017

Gastrin is the principal hormonal inducer of gastric acid secretion. Chronic hypergastrinemia, leading to hypersecretion of gastric acid and increased proliferation of parietal and enterochromaffin-like (ECL) cells, has been well described. In contrast, the physiological consequences of chronic gastrin deficiency had been poorly understood until the recent genetic engineering of mouse mutants containing a gastrin gene deletion by homologous recombination in embryonic stem cells. This themes article describes the consequences of constitutive gastrin deficiency on the development and physiology of the stomach. A lack of gastrin disrupts basal gastric acid secretion and renders the acid secretory system unresponsive to acute histaminergic, cholinergic, and gastrinergic stimulation. The defect in acid secretion is greater than would have been predicted from previous studies in which gastrin action was acutely blocked. Cellular changes include thinning of the gastric mucosa in the gastrin-deficient mice, with a reduction in parietal cells and reduced expression of markers of parietal and ECL cell-differentiated functions. The results suggest that gastrin is required for the functional maturation of the acid-secretory system.
Biosynthesis and secretion of the mannose 6-phosphate receptor and its ligands in polarized Caco-2 cells
Authors: Wick Debra A. , Seetharam Bellur , Dahms Nancy M.
Published: December 2017

We have analyzed the transport of newly synthesized mannose 6-phosphate (Man-6- P)-bearing proteins (i.e., lysosomal enzymes) in the polarized human colon adenocarcinoma cell line, Caco-2, by subjecting filter-grown cells to a pulse-chase labeling protocol using [35S]methionine, and the resulting cell lysate, apical medium, and basolateral medium were immunoprecipitated with insulin-like growth factor II/Man-6- P receptor (IGF-II/MPR)-specific antisera. The results showed that the majority of secreted lysosomal enzymes accumulated in the apical medium at >2 h of chase and that this polarized distribution was facilitated by the IGF-II/MPR selectively endocytosing lysosomal enzymes from the basolateral surface. Treatment with various agents known to affect vesicular transport events demonstrated that incubations at 16°C or incubations with brefeldin A inhibited the secretion of lysosomal enzymes from both the apical and basolateral surface, whereas treatment with nocodazole selectively blocked apical secretion. In contrast, incubation with NH4Cl or nocodazole had a stimulatory effect on basolateral secretion. Taken together, these results demonstrate that the sorting of Man-6- P-containing proteins into the apical and basolateral secretory pathways is regulated by distinct components of the intracellular trafficking machinery.
Estriol sensitizes rat Kupffer cells via gut-derived endotoxin
Authors: Enomoto Nobuyuki , Yamashina Shunhei , Schemmer Peter , Rivera Chantal A. , Bradford Blair U. , Enomoto Ayako , Brenner David A. , Thurman Ronald G.
Published: December 2017

The relationship between gender and alcohol-induced liver disease is complex; however, endotoxin is most likely involved. Recently, it was reported that estriol activated Kupffer cells by upregulation of the endotoxin receptor CD14. Therefore, the purpose of this work was to study how estriol sensitizes Kupffer cells. Rats were given estriol (20 mg/kg ip), and Kupffer cells were isolated 24 h later. After addition of lipopolysaccharide (LPS), intracellular Ca2+concentration was measured using a microspectrofluorometer with the fluorescent indicator fura 2, and tumor necrosis factor-α was measured by ELISA. CD14 was evaluated by Western analysis. One-half of the rats given estriol intraperitoneally 24 h before an injection of a sublethal dose of LPS (5 mg/kg) died within 24 h, whereas none of the control rats died. Mortality was prevented totally by sterilization of the gut with antibiotics. A similar pattern was obtained with liver histology and serum transaminases. Translocation of horseradish peroxidase was increased about threefold in gut segments by treatment with estriol. This increase was not altered by treatment with nonabsorbable antibiotics. On the other hand, endotoxin levels were increased to 60–70 pg/ml in plasma of rats treated with estriol. As expected, this increase was prevented (<20 pg/ml) by antibiotics. In isolated Kupffer cells, LPS-induced increases in intracellular Ca2+concentration, tumor necrosis factor-α production, and CD14 were increased, as previously reported. All these phenomena were blocked by antibiotics. Therefore, it is concluded that estriol treatment in vivo sensitizes Kupffer cells to LPS via mechanisms dependent on increases in CD14. This is most likely due to elevated portal blood endotoxin caused by increased gut permeability.
Responsiveness of β-escin-permeabilized rabbit gastric gland model: effects of functional peptide fragments
Authors: Akagi Keiko , Nagao Taku , Urushidani Tetsuro
Published: December 2017

We established a β-escin-permeabilized gland model with the use of rabbit isolated gastric glands. The glands retained an ability to secrete acid, monitored by [14C]aminopyrine accumulation, in response to cAMP, forskolin, and histamine. These responses were all inhibited by cAMP-dependent protein kinase inhibitory peptide. Myosin light-chain kinase inhibitory peptide also suppressed aminopyrine accumulation, whereas the inhibitory peptide of protein kinase C or that of calmodulin kinase II was without effect. Guanosine-5′- O-(3-thiotriphosphate) (GTPγS) abolished cAMP-stimulated acid secretion concomitantly, interfering with the redistribution of H+-K+-ATPase from tubulovesicles to the apical membrane. To identify the targets of GTPγS, effects of peptide fragments of certain GTP-binding proteins were examined. Although none of the peptides related to Rab proteins showed any effect, the inhibitory peptide of Arf protein inhibited cAMP-stimulated secretion. These results demonstrate that our new model, the β-escin-permeabilized gland, allows the introduction of relatively large molecules, e.g., peptides, into the cell, and will be quite useful for analyzing signal transduction of parietal cell function.
Stimulation of jejunal synthesis of apolipoprotein A-IV by ileal lipid infusion is blocked by vagotomy
Authors: Kalogeris Theodore J. , Holden V. Roger , Tso Patrick
Published: December 2017

We examined the role of vagal innervation in lipid-stimulated increases in expression and synthesis of intestinal apolipoprotein A-IV (apoA-IV). In rats with duodenal cannulas and superior mesenteric lymph fistulas given duodenal infusions of lipid emulsion, vagotomy had no effect on either intestinal lipid transport, lymphatic apoA-IV output, or jejunal mucosal apoA-IV synthesis. In rats with jejunal Thiry-Vella fistulas, ileal lipid infusion elicited a twofold stimulation of apoA-IV synthesis without affecting apoA-IV mRNA levels; vagotomy blocked this increase in apoA-IV synthesis. Direct perfusion of jejunal Thiry-Vella fistulas produced 2- to 2.5-fold increases in both apoA-IV synthesis and mRNA levels in the Thiry-Vella segment; these effects were not influenced by vagal denervation. These results suggest two mechanisms whereby lipid stimulates intestinal apoA-IV production: 1) a vagal-dependent stimulation of jejunal apoA-IV synthesis by distal gut lipid that is independent of changes in apoA-IV mRNA levels and 2) a direct stimulatory effect of proximal gut lipid on both synthesis and mRNA levels of jejunal apoA-IV that is independent of vagal innervation.
Amount and fate of egg protein escaping assimilation in the small intestine of humans
Authors: Evenepoel Pieter , Claus Dirk , Geypens Benny , Hiele Martin , Geboes Karen , Rutgeerts Paul , Ghoos Yvo
Published: December 2017

Studies attempting to evaluate protein assimilation in humans have hitherto relied on either ileostomy subjects or intubation techniques. The availability of stable isotope-labeled protein allowed us to determine the amount and fate of dietary protein escaping digestion and absorption in the small intestine of healthy volunteers using noninvasive tracer techniques. Ten healthy volunteers were studied once after ingestion of a cooked test meal, consisting of 25 g of13C-,15N-, and2H-labeled egg protein, and once after ingestion of the same but raw meal. Amounts of 5.73% and 35.10% ( P < 0.005) of cooked and raw test meal, respectively, escaped digestion and absorption in the small intestine. A significantly higher percentage of the malabsorbed raw egg protein was recovered in urine as fermentation metabolites. These results 1) confirm that substantial amounts of even easily digestible proteins may escape assimilation in healthy volunteers and 2) further support the hypothesis that the metabolic fate of protein in the colon is affected by the amount of protein made available.
Differentiated intestinal epithelial cells exhibit increased migration through polyamines and myosin II
Authors: Rao Jaladanki N. , Li Ji , Li Li , Bass Barbara L. , Wang Jian-Ying
Published: December 2017

Early mucosal restitution is a rapid process by which differentiated intestinal epithelial cells migrate to reseal superficial wounds. However, most of the in vitro studies for restitution employ undifferentiated intestinal crypt cells as a model. The transcription factor, Cdx2, plays an important role in the regulation of intestinal epithelial differentiation. Forced expression of the Cdx2 gene in undifferentiated intestinal crypt cells induces the development of a differentiated phenotype. The current study was designed to determine changes in differentiated intestinal epithelial cell migration after wounding in the stable Cdx2-transfected IEC-6 cells and then to examine involvement of polyamines and nonmuscle myosin II in the process of cell motility. Cdx2-transfected IEC-6 cells were associated with a highly differentiated phenotype and exhibited increased cell migration after wounding. Migration of Cdx2-transfected IEC-6 cells were approximately four times that of nontransfected IEC-6 cells. Migration after wounding was associated with significant increases in polyamine synthesis. Depletion of cellular polyamines by 5 mM α-difluoromethylornithine (DFMO), a specific inhibitor of polyamine biosynthesis, inhibited cell migration without affecting the differentiated phenotype. DFMO also decreased levels of nonmuscle myosin II mRNA and protein and resulted in reorganization of myosin II, along with a marked reduction in stress fibers. Exogenous spermidine given together with DFMO not only returned nonmuscle myosin II levels and cellular distribution toward normal but also restored cell migration to control levels. These results indicate that 1) Cdx2-transfected IEC-6 cells exhibit increased cell migration after wounding and 2) cellular polyamines are absolutely required for stimulation of cell migration in association with their ability to modulate the structural organization of nonmuscle myosin II.
CYP2E1 is not involved in early alcohol-induced liver injury
Authors: Kono Hiroshi , Bradford Blair U. , Yin Ming , Sulik Kathleen K. , Koop Dennis R. , Peters Jeffrey M. , Gonzalez Frank J. , McDonald Tasha , Dikalova Anna , Kadiiska Maria B. , Mason Ronald P. , Thurman Ronald G.
Published: December 2017

The continuous intragastric enteral feeding protocol in the rat was a major development in alcohol-induced liver injury (ALI) research. Much of what has been learned to date involves inhibitors or nutritional manipulations that may not be specific. Knockout technology avoids these potential problems. Therefore, we used long-term intragastric cannulation in mice to study early ALI. Reactive oxygen species are involved in mechanisms of early ALI; however, their key source remains unclear. Cytochrome P-450 (CYP)2E1 is induced predominantly in hepatocytes by ethanol and could be one source of reactive oxygen species leading to liver injury. We aimed to determine if CYP2E1 was involved in ALI by adapting the enteral alcohol (EA) feeding model to CYP2E1 knockout (−/−) mice. Female CYP2E1 wild-type (+/+) or −/− mice were given a high-fat liquid diet with either ethanol or isocaloric maltose-dextrin as control continuously for 4 wk. All mice gained weight steadily over 4 wk, and there were no significant differences between groups. There were also no differences in ethanol elimination rates between CYP2E1 +/+ and −/− mice after acute ethanol administration to naive mice or mice receiving EA for 4 wk. However, EA stimulated rates 1.4-fold in both groups. EA elevated serum aspartate aminotransferase levels threefold to similar levels over control in both CYP2E1 +/+ and −/− mice. Liver histology was normal in control groups. In contrast, mice given ethanol developed mild steatosis, slight inflammation, and necrosis; however, there were no differences between the CYP2E1 +/+ and −/− groups. Chronic EA induced other CYP families (CYP3A, CYP2A12, CYP1A, and CYP2B) to the same extent in CYP2E1 +/+ and −/− mice. Furthermore, POBN radical adducts were also similar in both groups. Data presented here are consistent with the hypothesis that oxidants from CYP2E1 play only a small role in mechanisms of early ALI in mice. Moreover, this new mouse model illustrates the utility of knockout technology in ALI research.
Inhibition of gastric acid secretion in rat stomach by PACAP is mediated by secretin, somatostatin, and PGE2
Authors: Li P. , Chang T.-M. , Coy D. , Chey W. Y.
Published: December 2017

Pituitary adenylate cyclase-activating polypeptide (PACAP), existing in two variants, PACAP-27 and PACAP-38, is found in the enteric nervous system and regulates function of the digestive system. However, the regulatory mechanism of PACAP on gastric acid secretion has not been well elucidated. We investigated the inhibitory action of PACAP-27 on acid secretion and its mechanism in isolated vascularly perfused rat stomach. PACAP-27 in four graded doses (5, 10, 20, and 50 μg/h) was vascularly infused to determine its effect on basal and pentagastrin (50 ng/h)-stimulated acid secretion. To study the inhibitory mechanism of PACAP-27 on acid secretion, a rabbit antisecretin serum, antisomatostatin serum, or indomethacin was administered. Concentrations of secretin, somatostatin, PGE2, and histamine in portal venous effluent were measured by RIA. PACAP-27 dose-dependently inhibited both basal and pentagastrin-stimulated acid secretion. PACAP-27 at 10 μg/h significantly increased concentrations of secretin, somatostatin, and PGE2in basal or pentagastrin-stimulated state. The inhibitory effect of PACAP-27 on pentagastrin-stimulated acid secretion was reversed 33% by an antisecretin serum, 80.0% by an antisomatostatin serum, and 46.1% by indomethacin. The antisecretin serum partially reduced PACAP-27-induced local release of somatostatin and PGE2. PACAP-27 at 10 μg/h elevated histamine level in portal venous effluent, which was further increased by antisomatostatin serum. However, antisomatostatin serum did not significantly increase acid secretion. It is concluded that PACAP-27 inhibits both basal and pentagastrin-stimulated gastric acid secretion. The effect of PACAP-27 is mediated by local release of secretin, somatostatin, and PGE2in isolated perfused rat stomach. The increase in somatostatin and PGE2levels in portal venous effluent is, in part, attributable to local action of the endogenous secretin.
Bile salt excretion in skate liver is mediated by a functional analog of Bsep/Spgp, the bile salt export pump
Authors: Ballatori Nazzareno , Rebbeor James F. , Connolly Gregory C. , Seward David J. , Lenth Benjamin E. , Henson John H. , Sundaram Pazhani , Boyer James L.
Published: December 2017

Biliary secretion of bile salts in mammals is mediated in part by the liver-specific ATP-dependent canalicular membrane protein Bsep/Spgp, a member of the ATP-binding cassette superfamily. We examined whether a similar transport activity exists in the liver of the evolutionarily primitive marine fish Raja erinacea, the little skate, which synthesizes mainly sulfated bile alcohols rather than bile salts. Western blot analysis of skate liver plasma membranes using antiserum raised against rat liver Bsep/Spgp demonstrated a dominant protein band with an apparent molecular mass of 210 kDa, a size larger than that in rat liver canalicular membranes, ∼160 kDa. Immunofluorescent localization with anti-Bsep/Spgp in isolated, polarized skate hepatocyte clusters revealed positive staining of the bile canaliculi, consistent with its selective apical localization in mammalian liver. Functional characterization of putative ATP-dependent canalicular bile salt transport activity was assessed in skate liver plasma membrane vesicles, with [3H]taurocholate as the substrate. [3H]taurocholate uptake into the vesicles was mediated by ATP-dependent and -independent mechanisms. The ATP-dependent component was saturable, with a Michaelis-Menten constant ( Km) for taurocholate of 40 ± 7 μM and a Kmfor ATP of 0.6 ± 0.1 mM, and was competitively inhibited by scymnol sulfate (inhibition constant of 23 μM), the major bile salt in skate bile. ATP-dependent uptake of taurocholate into vesicles was inhibited by known substrates and inhibitors of Bsep/Spgp, including other bile salts and bile salt derivatives, but not by inhibitors of the multidrug resistance protein-1 or the canalicular multidrug resistance-associated protein, indicating a distinct transport mechanism. These findings provide functional and structural evidence for a Bsep/Spgp-like protein in the canalicular membrane of the skate liver. This transporter is expressed early in vertebrate evolution and transports both bile salts and bile alcohols.
Defect of receptor-G protein coupling in human gallbladder with cholesterol stones
Authors: Xiao Zuo-Liang , Chen Qian , Amaral Joseph , Biancani Piero , Behar Jose
Published: December 2017

Human gallbladders with cholesterol stones (ChS) exhibit an impaired muscle contraction and relaxation and a lower CCK receptor-binding capacity compared with those with pigment stones (PS). This study was designed to determine whether there is an abnormal receptor-G protein coupling in human gallbladders with ChS using35S-labeled guanosine 5′- O-(3-thiotriphosphate) ([35S]GTPγS) binding,125I-labeled CCK-8 autoradiography, immunoblotting, and G protein quantitation. CCK and vasoactive intestinal peptide caused significant increases in [35S]GTPγS binding to Gαi-3and Gsα, respectively. The binding was lower in ChS than in PS ( P < 0.01). The reduced [35S]GTPγS binding in ChS was normalized after the muscles were treated with cholesterol-free liposomes ( P < 0.01). Autoradiography and immunoblots showed a decreased optical density (OD) for CCK receptors, an even lower OD value for receptor-G protein coupling, and a higher OD for uncoupled receptors or Gαi-3protein in ChS compared with PS ( P < 0.001). G protein quantitation also showed that there were no significant differences in the Gαi-3and Gsα content in ChS and PS. We conclude that, in addition to an impaired CCK receptor-binding capacity, there is a defect in receptor-G protein coupling in muscle cells from gallbladder with ChS. These changes may be normalized after removal of excess cholesterol from the plasma membrane.
IX. Mast cell-deficient mice and intestinal biology
Authors: Wershil Barry K.
Published: December 2017

Mutant mice that express abnormalities in mast cell development represent a powerful tool for the investigation of multiple aspects of mast cell biology. In addition, the identification of the genes affected by these mutations has not only increased our knowledge about the mast cell but has opened new areas of investigation as to the role of these gene products in gastrointestinal pathology, immunology, and physiology.
Complete lower esophageal sphincter relaxation observed in some achalasia patients is functionally inadequate
Authors: Mearin Fermín , Malagelada Juan-R.
Published: December 2017

Generally accepted manometric criteria for the diagnosis of achalasia are absent peristalsis and incomplete lower esophageal sphincter (LES) relaxation. However, in some patients with otherwise typical features of achalasia, esophageal manometry shows complete LES relaxation during swallowing. To establish whether such apparently complete LES relaxations are functionally adequate, we quantified changes in resistance to flow at the esophagogastric junction (EGJ) during wet swallowing. We studied seven achalasia patients with manometrically complete (>80%) LES relaxation, eight achalasia patients with incomplete (<40%) LES relaxation, and eight healthy volunteers. Complete LES relaxation on standard manometry (open-tip catheters) was confirmed in five of the seven achalasia patients by a Dentsleeve. Changes in EGJ resistance to flow were quantified using a pneumatic resistometer. Manometrically, the relaxation time span was significantly longer in patients with complete LES relaxation than in those with incomplete relaxation (7.3 ± 0.5 vs. 4.4 ± 0.7 s; P < 0.05). The fall in EGJ resistance from basal values during swallowing was markedly reduced in both achalasia groups (21 ± 8% in those with manometrically complete relaxation and 4 ± 2% in those with incomplete relaxation) by comparison with healthy individuals, in whom resistance fell by 90 ± 3% ( P< 0.05 vs. both achalasia groups). The duration of EGJ resistance drop was also much shorter in achalasia with (0.7 ± 0.2 s) and without (0.2 ± 0.1 s) complete LES relaxation compared with healthy control values (6.6 ± 1.2 s). Our results reveal that the apparently complete LES relaxation observed manometrically in some patients with achalasia is functionally inadequate since it is not associated with the normal profound fall in EGJ resistance to flow.
Increased insulin-like growth factor binding protein-4 expression after partial hepatectomy in the rat
Authors: Demori Ilaria , Balocco Sara , Voci Adriana , Fugassa Emilia
Published: December 2017

The insulin-like growth factor (IGF) binding proteins (IGFBPs) are important regulators of cell growth produced by different tissues. The IGFBPs regulate cell growth by modulating the activity and bioavailability of IGFs. The evidence that IGFBP-1 is a liver-specific immediate-early gene highly induced after 70% partial hepatectomy (PHx) suggests a role for the IGF-IGFBP system in hepatic regeneration. In this work we analyzed the effect of PHx on the expression of IGFBP-4, which is highly produced by the liver and very abundant in rat serum. Our results show a marked increase in hepatic IGFBP-4 mRNA levels 6–12 h after PHx and no significant change in sham-operated control animals. A parallel rise in IGFBP-4 transcript abundance was observed in the kidneys of PHx rats but not in sham-operated animals. Moreover, ligand blot analysis demonstrated that serum IGFBP-4 levels began to increase 12–24 h after surgery, consistent with the rise in the corresponding mRNA. This enhancement in IGFBP-4 production after PHx could be part of a fine regulatory mechanism to modulate IGF activity during liver regeneration.
Minor involvement of nitric oxide during chronic endotoxemia in anesthetized pigs
Authors: Pastor Catherine M. , Hadengue Antoine , Nussler Andreas K.
Published: December 2017

To study the modifications of hepatic blood flow and hepatic function over time during endotoxemia, 10 pigs received a continuous intravenous infusion of endotoxin (Endo, 160 ng ⋅ kg−1⋅ h−1) over 18 h and 7 control (Ctrl) animals received a saline infusion. The involvement of nitric oxide (NO) in this endotoxic model was assessed by measuring plasma concentrations of[Formula: see text],[Formula: see text], and cGMP, by testing vascular reactivity to ACh, and by evaluating inducible NO synthase (NOS 2) expression in hepatic biopsies. Endotoxin induced hypotensive and normokinetic shock in association with few modifications of hepatic blood flow, and hepatic injury was observed in both groups. Endotoxin did not increase plasma concentrations of[Formula: see text],[Formula: see text], and cGMP. The ACh-dependent decrease of mean arterial pressure was reduced in Endo pigs, whereas a minor difference was observed between Ctrl and Endo pigs for ACh-dependent modification of hepatic perfusion. Hepatic NOS 2 mRNA was not detected in Ctrl pigs. In Endo pigs, NOS 2 protein expression was detected only in tissues surrounding the portal vein and the inferior vena cava, whereas NOS 2 mRNA was expressed in all hepatic biopsies. Thus, although endotoxemia induces NOS 2 expression in the liver, our findings show that NO involvement is lower in pigs than in rodents during endotoxemia.
Upregulation of HGF activator inhibitor type 1 but not type 2 along with regeneration of intestinal mucosa
Authors: Itoh Hiroshi , Kataoka Hiroaki , Tomita Masaki , Hamasuna Ryouichi , Nawa Yukifumi , Kitamura Naomi , Koono Masashi
Published: December 2017

Hepatocyte growth factor (HGF) activator inhibitor type 1 (HAI-1) and type 2 (HAI-2) are new Kunitz-type serine protease inhibitors that were recently purified and cloned from the human stomach cancer cell line MKN45 as specific inhibitors against HGF activator. Both proteins contain two Kunitz inhibitor domains and are expressed abundantly throughout the gastrointestinal tract, in addition to the placenta, pancreas, and kidney. In this study, to assess the possible roles of HAI-1 and HAI-2 in the intestinal mucosa, we examined the expression of HAI-1 and HAI-2 during regeneration of the intestinal mucosa. Immunohistochemical studies revealed that HAI-1 but not HAI-2 was detected more strongly in regenerative epithelium than in normal epithelium, although both proteins were detected throughout the human gastrointestinal tract. During the course of acetic acid-induced experimental colitis in an in vivo mouse model, HAI-1 but not HAI-2 was upregulated in the recovery phase, suggesting that HAI-1 but not HAI-2 is associated with the regeneration of damaged colonic mucosa. Upregulation of HAI-1 may serve to downregulate the proliferative response after initial activation of MET receptor by HGF/scatter factor after an injury.
Uptake of lactate by the liver: effect of red blood cell carriage
Authors: Goresky Carl A. , Bach Glen G. , Simard André , Schwab Andreas J. , Bracht Adelar
Published: December 2017

Multiple-indicator dilution experiments with labeled lactate were performed in the livers of anesthetized dogs. A mixture of51Cr-labeled erythrocytes, [3H]sucrose, andl-[1-14C]lactate or a mixture of51Cr-labeled erythrocytes, [14C]sucrose, andl-[2-3H]lactate was injected into the portal vein, and samples were obtained from the hepatic vein. Data were evaluated using a model comprising flow along sinusoids, exchange of lactate between plasma and erythrocytes and between plasma and hepatocytes, and, in the case ofl-[1-14C]lactate, metabolism to H[14C][Formula: see text]within hepatocytes. The coefficient for lactate efflux from erythrocytes was 0.62 ± 0.24 s−1, and those for influx into and efflux from hepatocytes were 0.44 ± 0.13 and 0.14 ± 0.07 s−1, respectively. The influx permeability-surface area product of the hepatocyte membrane for lactate ( PinS, in ml ⋅ s−1⋅ g−1) varied with total flow rate ( F, in ml s−1⋅ g−1) according to PinS = (3.1 ± 0.5) F+ (0.021 ± 0.014). Lactate in plasma, erythrocytes, and hepatocytes was close to equilibrium, whereas lactate metabolism was rate limiting.
XII. IL-10-deficient (IL-10−/−) mice and intestinal inflammation
Authors: Rennick Donna M. , Fort Madeline M.
Published: December 2017

Interleukin (IL)-10−/−mice spontaneously develop intestinal inflammation characterized by discontinuous transmural lesions affecting the small and large intestine and by dysregulated production of proinflammatory cytokines. The uncontrolled generation of IFN-γ-producing CD4+T cells (Th1 type) has been shown to play a causal role in the development of enterocolitis affecting these mutants. This article discusses studies of IL-10−/−mice that have investigated the role of enteric organisms in triggering intestinal disease, the mediators responsible for initiating and maintaining intestinal disease, the role IL-10 plays in the generation and/or function of regulatory cells, and the results of IL-10 therapy in experimental animal models of inflammatory bowel disease (IBD) and human patients with IBD.
Mechanism of the alcohol cyclic pattern: role of the hypothalamic-pituitary-thyroid axis
Authors: Li J. , Nguyen V. , French B. A. , Parlow A. F. , Su G. L. , Fu P. , Yuan Q. X. , French S. W.
Published: December 2017

The cause of the cycle of urinary alcohol levels (UALs) in rats fed ethanol continually at a fixed rate is unknown. Rats were fed ethanol intragastrically at a constant dose for 2 mo, and daily body temperatures and UALs were recorded. Body temperature cycled inversely to UAL, suggesting that the rate of metabolism could be mechanistically involved in the rate of ethanol elimination during the cycle. To document this, whole body O2consumption rate was monitored daily during the cycle. The rate of O2consumption correlated positively with the change in body temperature and negatively with the change in UAL. Since the metabolic rate responds to changes in body temperature, thyroid hormone levels were measured during the UAL cycle. T4levels correlated positively with the O2consumption rate and negatively with the UALs. In a second experiment using propylthiouracil treatment, UALs did not cycle and a fall in body temperature failed to stimulate an increase in the rate of ethanol elimination. Consequently, rats died of overdose. Likewise, in a third experiment using rats with severed pituitary stalks, UALs failed to cycle and rats died of overdose. From these observations it was concluded that the UAL cycle depends on an intact hypothalamic-pituitary-thyroid axis response to the ethanol-induced drop in body temperature by increasing the rate of ethanol elimination.
Primary structures of PYY, [Pro34]PYY, and PYY-(3–36) confer different conformations and receptor selectivity
Authors: Keire David A. , Mannon Peter , Kobayashi Mitsuo , Walsh John H. , Solomon Travis E. , Reeve Joseph R.
Published: December 2017

We synthesized PYY-(1–36) (nonselective between Y1and Y2receptor subtype agonists), [Pro34]PYY (selective for Y1), and PYY-(3–36) (selective for Y2) to determine whether solution conformation plays a role in receptor subtype selectivity. The three peptides exhibited the expected specificities in displacing labeled PYY-(1–36) from cells transfected with Y1receptors (dissociation constants = 0.42, 0.21, and 1,050 nM, respectively) and from cells transfected with Y2receptors (dissociation constants = 0.03, 710, and 0.11 nM, respectively) for PYY-(1–36), [Pro34]PYY, and PYY-(3–36). Sedimentation equilibrium analyses revealed that the three PYY analogs were 80–90% monomer at the concentrations used for the subsequent circular dichroism (CD) and1H-nuclear magnetic resonance (NMR) studies. CD analysis measured helicities for PYY-(1–36), [Pro34]PYY, and PYY-(3–36) of 42%, 31%, and 24%, suggesting distinct differences in secondary structure. The backbone1H-NMR resonances of the three peptides further substantiated marked conformational differences. These patterns support the hypothesis that Y1and Y2receptor subtype binding affinities depend on the secondary and tertiary solution state structures of PYY and its analogs.
A cortical evoked potential study of afferents mediating human esophageal sensation
Authors: Hobson Anthony R. , Sarkar Sanchoy , Furlong Paul L. , Thompson David G. , Aziz Qasim
Published: December 2017

The aim of this study was to compare the characteristics of esophageal cortical evoked potentials (CEP) following electrical and mechanical stimulation in healthy subjects to evaluate the afferents involved in mediating esophageal sensation. Similarities in morphology and interpeak latencies of the CEP to electrical and mechanical stimulation suggest that they are mediated via similar pathways. Conduction velocity of CEP to either electrical or mechanical stimulation was 7.9–8.6 m/s, suggesting mediation via thinly myelinated Aδ-fibers. Amplitudes of CEP components to mechanical stimulation were significantly smaller than to electrical stimulation at the same levels of perception, implying that electrical stimulation activates a larger number of afferents. The latency delay of ∼50 ms for each mechanical CEP component compared with the corresponding electrical CEP component is consistent with the time delay for the mechanical stimulus to distend the esophageal wall sufficiently to trigger the afferent volley. In conclusion, because the mechanical and electrical stimulation intensities needed to obtain esophageal CEP are similar and clearly perceived, it is likely that both spinal and vagal pathways mediate esophageal CEP. Esophageal CEP to both modalities of stimulation are mediated by myelinated Aδ-fibers and produce equally robust CEP responses. Both techniques may have important roles in the assessment of esophageal sensory processing in health and disease.
Biological activities of novel lipid mediator sphingosine 1-phosphate in rat hepatic stellate cells
Authors: Ikeda Hitoshi , Yatomi Yutaka , Yanase Mikio , Satoh Hiroaki , Maekawa Hisato , Ogata Itsuro , Ozaki Yukio , Takuwa Yoh , Mochida Satoshi , Fujiwara Kenji
Published: December 2017

Sphingosine 1-phosphate (S-1-P), a lipid mediator shown to be a ligand for G protein-coupled receptors (AGRs), endothelial differentiation gene (EDG)1, EDG3, and AGR16/EDG5, is stored in platelets and released on their activation. Platelet consumption occurs in acute liver injury. Hepatic stellate cells (HSCs) play an important role in wound healing. Effects of S-1-P on HSCs were investigated. S-1-P enhanced proliferation of culture-activated HSCs. The mitogenic effect was pertussis toxin sensitive, mitogen-activated protein kinase dependent, and more prominent at lower cell density. S-1-P increased contraction of collagen lattices containing HSCs, irrespective of activation state, in a C3 exotoxin-sensitive manner. mRNAs of EDG1 and AGR16, but not of EDG3, were detected in HSCs. In HSC activation, EDG1 mRNA levels were downregulated, whereas AGR16 mRNA levels were unchanged. Considering that HSCs are capable of production of extracellular matrices and modulation of blood flow in sinusoids, our results suggest that S-1-P may play a role in wound healing process in the liver.
Altered intestinal transport of amino acids in cirrhotic rats: the effect of insulin-like growth factor-I
Authors: Pascual Maria , Castilla-Cortazar Inma , Urdaneta Elena , Quiroga Jorge , Garcia Maria , Picardi Antonio , Prieto Jesus
Published: December 2017

The intestine is an important target organ for insulin-like growth factor-I (IGF-I), an anabolic hormone synthesized in the liver upon growth hormone (GH) stimulation. Levels of IGF-I are reduced in cirrhosis, and altered GH/IGF-I axis may contribute to malnutrition in cirrhotic patients. Our aim was to study Na+-dependent jejunal transport of amino acids (l-leucine, l-proline,l-glutamic acid, and l-cysteine) in cirrhotic rats and to analyze the effect of IGF-I on this function. IGF-I or saline was administered for 2 wk to rats with CCl4-induced cirrhosis and saline was administered to healthy control rats. Transport of amino acids was assessed in brush-border membrane vesicles (BBMV) using14C- or35S-labeled amino acids, and the kinetic constants Vmaxand Ktwere determined. Na+-independent uptake of l-leucine, l-proline,l-glutamic acid, and l-cysteine by BBMV was similar in all groups. Na+-dependent uptake of all four amino acids was significantly diminished in cirrhotic rats compared with both controls and IGF-I-treated cirrhotic rats. The latter two groups exhibited similar Vmaxand Kt, whereas untreated cirrhotic rats had reduced Vmaxand increased Ktcompared with normal controls and IGF-I-treated cirrhotic animals. In conclusion, the transport of all four tested amino acids by BBMV is impaired in cirrhotic rats, and low doses of IGF-I can correct this defect.
Glutathione oxidation and PTPase inhibition by hydrogen peroxide in Caco-2 cell monolayer
Authors: Rao R. K. , Li L. , Baker R. D. , Baker S. S. , Gupta A.
Published: December 2017

The role of H2O2and protein thiol oxidation in oxidative stress-induced epithelial paracellular permeability was investigated in Caco-2 cell monolayers. Treatment with a H2O2generating system (xanthine oxidase + xanthine) or H2O2(20 μM) increased the paracellular permeability. Xanthine oxidase-induced permeability was potentiated by superoxide dismutase and prevented by catalase. H2O2-induced permeability was prevented by ferrous sulfate and potentiated by deferoxamine and 1,10-phenanthroline. GSH, N-acetyl-l-cysteine, dithiothreitol, mercaptosuccinate, and diethylmaleate inhibited H2O2-induced permeability, but it was potentiated by 1,3-bis(2-chloroethyl)-1-nitrosourea. H2O2reduced cellular GSH and protein thiols and increased GSSG. H2O2-mediated reduction of GSH-to-GSSG ratio was prevented by ferrous sulfate, GSH, N-acetyl-l-cysteine, diethylmaleate, and mercaptosuccinate and potentiated by 1,10-phenanthroline and 1,3-bis(2-chloroethyl)-1-nitrosourea. Incubation of soluble fraction of cells with GSSG reduced protein tyrosine phosphatase (PTPase) activity, which was prevented by coincubation with GSH. PTPase activity was also lower in H2O2-treated cells. This study indicates that H2O2, but not O2−· or ·OH, increases paracellular permeability of Caco-2 cell monolayer by a mechanism that involves oxidation of GSH and inhibition of PTPases.
Divergent homeobox geneHexregulates promoter of the Na+-dependent bile acid cotransporter
Authors: Denson Lee A. , Karpen Saul J. , Bogue Clifford W. , Jacobs Harris C.
Published: December 2017

The divergent homeobox gene Hex is expressed in both developing and mature liver. A putative Hex binding site was identified in the promoter region of the liver-specific Na+-bile acid cotransporter gene ( ntcp), and we hypothesized that Hex regulates the ntcp promoter through this site. Successive 5′-deletions of the ntcp promoter in a luciferase reporter construct transfected into Hep G2 cells confirmed a Hex response element (HRE) within the ntcppromoter (nt −733/−714). Moreover, p-CMHex transactivated a heterologous promoter construct containing HRE multimers (p4xHRELUC), whereas a 5-bp mutation of the core HRE eliminated transactivation. A dominant negative form of Hex (p- Hex-DN) suppressed basal luciferase activity of p-4xHRELUC and inhibited activation of this construct by p-CMHex. Interestingly, p-CMHex transactivated the HRE in Hep G2 cells but not in fibroblast-derived COS cells, suggesting the possibility that Hex protein requires an additional liver cell-specific factor(s) for full activity. Electrophoretic mobility shift assays confirmed that liver and Hep G2 cells contain a specific nuclear protein that binds the native HRE. We have demonstrated that the liver-specific ntcp gene promoter is the first known target of Hex and is a useful tool for evaluating function of the Hex protein.