bluuebunny/crossref_metadata_2025 huggingface dataset:
Diminished mechanosensitivity and chemosensitivity in patients with achalasia
Authors: Brackbill Stephen , Shi Guoxiang , Hirano Ikuo
Published: March 2015

The pathogenesis of achalasia involves the degeneration of enteric and autonomic nervous systems with resultant effects on esophageal motility. The neural degeneration could affect visceral sensation in achalasia. The aim of this study was to examine mechanosensitivity and chemosensitivity in patients with achalasia. Perceptual responses to esophageal distension and acid perfusion were assessed in nine achalasia patients and nine healthy subjects. Mechanosensitivity was evaluated using a barostat with a double-random staircase distension protocol. Responses were graded as follows: 0, no sensation; 1, initial sensation; 2, mild discomfort; 3, moderate discomfort; and 4, pain. Chemosensitivity was graded along a visual analog scale after perfusion of saline and 0.1 N HCl. Barostat pressure-volume relationships were used to report esophageal body compliance. Barostat pressures for initial sensation and mild discomfort were not significantly different for patients and controls. The pressures for moderate discomfort (37.9 ± 3.5 vs. 25.7 ± 2.4 mmHg; P < 0.05) and pain (47.8 ± 2.3 vs. 32.2 ± 3.5 mmHg; P = 0.002) were significantly higher in achalasics than controls. Seven of the eight achalasia patients never reached pain thresholds at the maximum distension pressure (50 mmHg). Sensation to acid perfusion was significantly lower in achalasics compared with controls (2.2 ± 1.2 vs. 6.7 ± 1.7 cm; P < 0.05). Compliance was significantly increased in patients with achalasia compared with controls. We conclude that both mechanosensitivity and chemosensitivity are significantly diminished in achalasia patients compared with controls. Also, initial sensation and pain sensation are differentially affected in achalasics. These findings suggest that neuropathic defects in achalasia may manifest themselves in visceral sensory and motor dysfunction.
Enhancement of antral contractions and vagal afferent signaling with synchronized electrical stimulation
Authors: Peles Shachar , Petersen Jaime , Aviv Ricardo , Policker Shai , Abu-Hatoum Ossama , Ben-Haim Shlomo A. , Gutterman David D. , Sengupta Jyoti N.
Published: March 2015

Gastric filling activates vagal afferents involved in peripheral signaling to the central nervous system (CNS) for food intake. It is not known whether these afferents linearly encode increasing contractions of the antrum during antral distension (AD). The aim of this study was to investigate effects of AD and electrically enhanced antral contractions on responses of vagal afferents innervating the antrum. Single-fiber recordings were made from the vagal afferents in anesthetized male Long-Evans rats. Antral contractions were measured with a solid-state probe placed in the antrum. A nonexcitatory electrical stimulation (NES) inducing no smooth muscle contractions was applied during the ascending phase of antral contractions to enhance subsequent antral contractions. Fifty-six fibers identified during AD (1 ml for 30 s) were studied through different types of mechanical stimuli. Under normal conditions, one group of fibers exhibited rhythmic firing in phase with antral contractions. Another group of fibers had nonrhythmic spontaneous firing. Responses of 15 fibers were tested with NES during multiple-step distension (MSD). NES produced a mean increase in antral contraction amplitude (177.1 ± 35.3%) and vagal afferent firing (21.6 ± 2.6%). Results show that both passive distension and enhanced antral contractions activate distension-sensitive vagal afferents. Responses of these fibers increase linearly to enhanced antral contraction induced by NES or MSD up to a distending volume of 0.6 ml. However, responses reached a plateau at a distending volume >0.8 ml. We concluded that enhanced contraction of the antrum can activate vagal afferents signaling to the CNS.
Effect of GABABreceptor agonist on distension-sensitive pelvic nerve afferent fibers innervating rat colon
Authors: Sengupta Jyoti N. , Medda Bidyut K. , Shaker Reza
Published: March 2015

Spinal afferents innervating the gastrointestinal tract are the major pathways for visceral nociception. Many centrally acting analgesic drugs attenuate responses of visceral primary afferent fibers by acting at the peripheral site. γ-Amino butyric acid (GABA), a major inhibitory neurotransmitter, acts via metobotropic GABABand ionotropic GABAA/GABACreceptors. The aim of this study was to test the peripheral effect of selective GABABreceptor agonist baclofen on responses of the pelvic nerve afferent fibers innervating the colon of the rat. Distension-sensitive pelvic nerve afferent fibers were recorded from the S1sacral dorsal root in anesthetized rats. The effect of baclofen (1–300 μmol/kg) was tested on responses of these fibers to colorectal distension (CRD; 60 mmHg, 30 s). A total of 21 pelvic nerve afferent fibers was recorded. Mechanosensitive properties of four fibers were also recorded before and after bilateral transections of T12-S3ventral roots (VR). Effect of baclofen was tested on 15 fibers (7 in intact rats, 4 in rats with transected VR, and 4 in rats pretreated with CGP 54626). In nine fibers (5/7 in intact and 4/4 in VR transected rats), baclofen produced dose-dependent inhibition of response to CRD. Pretreatment with selective GABABreceptor antagonist CGP 54626 (1 μmol/kg) reversed the inhibitory effect of baclofen. Results suggest a peripheral role of GABABreceptors in the inhibition of mechanotransduction property of distension-sensitive pelvic nerve afferent fibers.
DLPC decreases TGF-β1-induced collagen mRNA by inhibiting p38 MAPK in hepatic stellate cells
Authors: Cao Qi , Mak Ki M. , Lieber Charles S.
Published: March 2015

Dilinoleoylphosphatidylcholine (DLPC), the active component of polyenylphosphatidylcholine extracted from soybeans, decreases collagen accumulation induced by TGF-β1 in cultured hepatic stellate cells (HSCs). Because DLPC exerts antioxidant effects and TGF-β1 generates oxidative stress, we evaluated whether the antifibrogenic effect of DLPC is linked to its antioxidant action. In passage 1 culture of rat HSCs, TGF-β1 induced a concentration-dependent increase in procollagen-α1(I) mRNA levels and enhanced intracellular H2O2and superoxide anion formation and lipid peroxidation but decreased GSH levels. These changes were prevented by DLPC. Upregulation of collagen mRNA by TGF-β1 was likewise inhibited by catalase and p38 MAPK inhibitor SB-203580, suggesting involvement of H2O2and p38 MAPK signaling in this process. TGF-β1 or addition of H2O2to HSCs activated p38 MAPK with a rise in procollagen mRNA level; these changes were blocked by catalase and SB-203580 and likewise by DLPC. α-Smooth muscle actin abundance in HSCs was not altered by TGF-β1 treatment (with or without DLPC), indicating that downregulation of procollagen mRNA by DLPC was not due to alteration in HSC activation. These results demonstrate that DLPC prevents TGF-β1-induced increase in collagen mRNA by inhibiting generation of oxidative stress and associated H2O2-dependent p38 MAPK activation, which explains its antifibrogenic effect. DLPC, an innocuous phospholipid, may be considered for prevention and treatment of liver fibrosis.
Role of passive and adaptive immunity in influencing enterocyte-specific gene expression
Authors: Jenkins Shannon L. , Wang Jiafang , Vazir Mukta , Vela Jose , Sahagun Omar , Gabbay Peter , Hoang Lisa , Diaz Rosa L. , Aranda Richard , Martín Martín G.
Published: March 2015

Numerous genes expressed by intestinal epithelial cells are developmentally regulated, and the influence that adaptive (AI) and passive (PI) immunity have in controlling their expression has not been evaluated. In this study, we tested the hypothesis that both PI and AI influenced enterocyte gene expression by developing a breeding scheme that used T and B cell-deficient recombination-activating gene ( RAG) mice. RNA was isolated from the liver and proximal/distal small intestine at various ages, and the steady-state levels of six different transcripts were evaluated by RNase protection assay. In wild-type (WT) pups, all transcripts [Fc receptor of the neonate ( FcRn), polymeric IgA receptor ( pIgR), GLUT5, lactase-phlorizin hydrolase ( lactase), apical sodium-dependent bile acid transporter ( ASBT), and Na+/glucose cotransporter ( SGLT1)] studied were developmentally regulated at the time of weaning, and all transcripts except ASBT had the highest levels of expression in the proximal small intestine. In WT suckling pups reared in the absence of PI, pIgR mRNA levels were increased 100% during the early phase of development. In mice lacking AI, the expression of pIgR and lactase were significantly attenuated, whereas FcRn and GLUT5 levels were higher compared with WT mice. Finally, in the absence of both passive and active immunity, expression levels of pIgR and lactase were significantly lower than similarly aged WT mice. In summary, we report that the adaptive and passive immune status of mice influences steady-state mRNA levels of several important, developmentally regulated enterocyte genes during the suckling and weaning periods of life.
Longitudinal and circumferential spike patches in the canine small intestine in vivo
Authors: Lammers Wim J. E. P. , Donck Luc Ver , Schuurkes Jan A. J. , Stephen Betty
Published: March 2015

In an open-abdominal anesthetized and fasted canine model of the intact small intestine, the presence, location, shape, and frequency of spike patches were investigated. Recordings were performed with a 240-electrode array (24 × 10, 2-mm interelectrode distance) from several sites sequentially, spanning the whole length of the small intestine. All 240 electrograms were recorded simultaneously during periods of 5 min and were analyzed to reconstruct the origin and propagation of individual spikes. At every level in the small intestine, spikes propagated in all directions before stopping abruptly, thereby activating a circumscribed area termed a “patch.” Two types of spikes were found: longitudinal spikes, which propagated predominantly in the longitudinal direction and occurred most often in the duodenum, and a second type, circumferential spikes, which propagated predominantly in the circular direction and occurred much more frequently in the jejunum and ileum. Circumferential spikes conducted faster than longitudinal spikes (17 ± 6 and 7 ± 2 cm/s, respectively; P < 0.001). Circumferential spikes originated in >90% of all cases from the antimesenteric border, whereas longitudinal spikes were initiated all around the circumference of the intestinal tube. Finally, the spatial sequence of spike patches after the slow wave was very irregular in the upper part of the intestine but much more regular in the lower part. In conclusion, spikes and spike patches occur throughout the small intestine, whereas their type, sites of origin, extent of propagation, and frequencies of occurrence differ along the length of the small intestine, suggesting differences in local patterns of motility.
PKC-ζ prevents oxidant-induced iNOS upregulation and protects the microtubules and gut barrier integrity
Authors: Banan A. , Zhang L. , Fields J. Z. , Farhadi A. , Talmage D. A. , Keshavarzian A.
Published: March 2015

Using intestinal (Caco-2) monolayers, we reported that inducible nitric oxide synthase (iNOS) activation is key to oxidant-induced barrier disruption and that EGF protects against this injury. PKC-ζ was required for protection. We thus hypothesized that PKC-ζ activation and iNOS inactivation are key in EGF protection. Wild-type (WT) Caco-2 cells were exposed to H2O2(0.5 mM) ± EGF or PKC modulators. Other cells were transfected to overexpress PKC-ζ or to inhibit it and then pretreated with EGF or a PKC activator (OAG) before oxidant. Relative to WT cells exposed to oxidant, pretreatment with EGF protected monolayers by 1) increasing PKC-ζ activity; 2) decreasing iNOS activity and protein, NO levels, oxidative stress, tubulin oxidation, and nitration); 3) increasing polymerized tubulin; 4) maintaining the cytoarchitecture of microtubules; and 5) enhancing barrier integrity. Relative to WT cells exposed to oxidant, transfected cells overexpressing PKC-ζ (+2.9-fold) were protected as indicated by decreases in all measures of iNOS-driven pathways and enhanced stability of microtubules and barrier function. Overexpression-induced inhibition of iNOS was OAG independent, but EGF potentiated this protection. Antisense inhibition of PKC-ζ (−95%) prevented all measures of EGF protection against iNOS upregulation. Thus EGF protects against oxidative disruption of the intestinal barrier by stabilizing the cytoskeleton in large part through the activation of PKC-ζ and downregulation of iNOS. Activation of PKC-ζ is by itself required for cellular protection against oxidative stress of iNOS. We have thus discovered novel biologic functions, suppression of the iNOS-driven reactions and cytoskeletal oxidation, among the atypical PKC isoforms.
Influence of caveolin on constitutively activated recombinant eNOS: insights into eNOS dysfunction in BDL rat liver
Authors: Hendrickson H. , Chatterjee S. , Cao S. , Ruiz M. Morales , Sessa W. C. , Shah V.
Published: March 2015

Diminished endothelial nitric oxide (NO) synthase (eNOS)-derived NO production from the hepatic vascular endothelium contributes to hepatic vasoconstriction in portal hypertension. The aim of this study was to examine the mechanism of this process by testing the influence of a constitutively active form of eNOS (S1179DeNOS) in both primary and propagated liver cells in vitro and in the sham and bile duct ligated (BDL) rat liver in vivo, using an adenoviral vector encoding green fluorescent protein (AdGFP) and S1179DeNOS (AdS1179DeNOS). AdS1179DeNOS transduction augmented basal and agonist-stimulated NO generation in nonparenchymal liver cells. Sham rats transduced in vivo with AdS1179DeNOS evidenced a decreased pressor response to incremental doses of the vasoconstrictor methoxamine compared with sham rats transduced with AdGFP. However, BDL rats transduced with AdS1179DeNOS did not display improved vasodilatory responses as evidenced by similar flow-dependent pressure increases to that observed in BDL rats transduced with AdGFP, despite similar levels of viral transgene expression. We next examined the influence of the eNOS inhibitory protein caveolin on S1179DeNOS dysfunction in cirrhotic liver. Immunogold electron microscopic analysis of caveolin in BDL liver demonstrated prominent expression not only in liver endothelial cells, but also in hepatic stellate cells. In vitro studies in the LX2 hepatic stellate cell line demonstrate that caveolin precipitates recombinant S1179DeNOS in LX2 cells, that recombinant S1179DeNOS coprecipitates caveolin, and that binding is enhanced in the presence of overexpression of caveolin. Furthermore, caveolin overexpression inhibits recombinant S1179DeNOS activity. These studies indicate that recombinant S1179DeNOS protein functions appropriately in normal liver cells and tissue but evidences dysfunction in the cirrhotic rat liver and that caveolin expression and inhibition in BDL nonparenchymal cells, including hepatic stellate cells, may account for this dysfunction.
Cryptdin 3 forms anion selective channels in cytoplasmic membranes of human embryonic kidney cells
Authors: Yue Gang , Merlin Didier , Selsted Michael E. , Lencer Wayne I. , Madara James L. , Eaton Douglas C.
Published: March 2015

Cryptdins are antimicrobial peptides secreted by Paneth cells located at the base of intestinal crypts. In addition to their antimicrobial function, cryptdins may also regulate salt and water secretion by intestinal epithelial cells. Recent work with short-circuit current measurements indicated that at least one cryptdin peptide, cryptdin 3, induces apical conductance(s) in Cl−secretory, including cystic fibrosis, epithelia. In the present study, we characterized the cryptdin 3-induced anion channel activity in human embryonic kidney (HEK) cells with single-channel patch-clamp techniques. The patch pipette was filled with solution containing different concentrations of cryptdin 3, and, after gigaseal formation, the channel activity was recorded with either cell-attached or inside-out patch modes. We found an anion selective channel with a conductance of 15 pS and open probability of 0.19, regardless of cryptdin 3 concentration. The mean open and closed times varied with the cryptdin 3 concentration. For cryptdin 3 concentrations of 10, 4, 1, and 0.5 μg/ml in the pipette, the corresponding mean open times were 1.2, 7.0, 9.0, and 17.4 ms and the corresponding mean closed times were 1.1, 1.6, 4.2, and 12.5 ms. These results suggest that cryptdin 3 forms anion-selective channels on the cytoplasmic membrane of HEK cells and that the kinetics of one such channel are affected by its interaction with other such channels.
Gastric protective effect of peripheral PYY through PYY preferring receptors in anesthetized rats
Authors: Kawakubo Keishi , Yang Hong , Taché Yvette
Published: March 2015

The influence of intravenous peptide YY (PYY) on the gastric injury induced by 45% ethanol was investigated in urethane-anesthetized rats. PYY (25, 75, 125, and 250 pmol · kg−1 · h−1) significantly reduced gastric lesions by 36, 59, 40, and 38%, respectively. Antibody against ratPYY (2 mg/rat) injected intravenously completely prevented the gastroprotective effect of intravenous PYY (75 pmol · kg−1 · h−1), whereas injected intracisternally (460 μg/20 μl), it significantly prevented intracisternal PYY (24 pmol/rat)-induced 58% reduction of ethanol lesions but not that induced by intravenous PYY. Vagotomy did not influence the gastroprotective effect of intravenous PYY. The Y1/“PYY-preferring” receptor agonist [Pro34]PYY (75 pmol · kg−1 ·h−1iv) significantly decreased ethanol-induced gastric lesions by 82%, whereas [Leu31, Pro34]NPY, a Y1/Y3 agonist, and PYY-(3–36), a Y2 agonist, had no effect. These data indicate that PYY-infused intravenously at doses reported to mimic postprandial peak blood levels prevents ethanol-induced gastric injury through vagal independent pathways and PYY-preferring receptors.
Mechanism of n-butyrate uptake in the human proximal colonic basolateral membranes
Authors: Tyagi S. , Venugopalakrishnan J. , Ramaswamy K. , Dudeja P. K.
Published: March 2015

Current studies were undertaken to characterize the mechanism of short-chain fatty acid (SCFA) transport in isolated human proximal colonic basolateral membrane vesicles (BLMV) utilizing a rapid-filtration n-[14C]butyrate uptake technique. Human colonic tissues were obtained from mucosal scrapings from organ donor proximal colons. Our results, consistent with the existence of a HCO[Formula: see text]/SCFA exchanger in these membranes, are summarized as follows: 1) n-[14C]butyrate influx was significantly stimulated into the vesicles in the presence of an outwardly directed HCO[Formula: see text] and an inwardly directed pH gradient; 2) n-[14C]butyrate uptake was markedly inhibited (∼40%) by anion exchange inhibitor niflumic acid (1 mM), but SITS and DIDS (5 mM) had no effect; 3) structural analogs e.g., acetate and propionate, significantly inhibited uptake of HCO[Formula: see text] and pH-gradient-driven n-[14C]butyrate; 4) n-[14C]butyrate uptake was saturable with a Kmfor butyrate of 17.5 ± 4.5 mM and a Vmaxof 20.9 ± 1.2 nmol · mg protein−1· 5 s−1; 5) n-[14C]butyrate influx into the vesicles demonstrated a transstimulation phenomenon; and 6) intravesicular or extravesicular Cl−did not alter the anion-stimulated n-[14C]butyrate uptake. Our results indicate the presence of a carrier-mediated HCO[Formula: see text]/SCFA exchanger on the human colonic basolateral membrane, which appears to be distinct from the previously described anion exchangers in the membranes of colonic epithelia.
Interleukin-6 inhibits hepatic growth hormone signaling via upregulation of Cis and Socs-3
Authors: Denson Lee A. , Held Matthew A. , Menon Ram K. , Frank Stuart J. , Parlow Albert F. , Arnold Dodie L.
Published: March 2015

Cytokines may cause an acquired growth hormone (GH) resistance in patients with inflammatory diseases. Anabolic effects of GH are mediated through activation of STAT5 transcription factors. We have reported that TNF-α suppresses hepatic GH receptor (GHR) gene expression, whereas the cytokine-inducible SH2-containing protein 1 (Cis)/suppressors of cytokine signaling ( Socs) genes are upregulated by TNF-α and IL-6 and inhibit GH activation of STAT5. However, the relative importance of these mechanisms in inflammatory GH resistance was not known. We hypothesized that IL-6 would prevent GH activation of STAT5 and that this would involve Cis/Socs protein upregulation. GH ± LPS was administered to TNF receptor 1 (TNFR1) or IL-6 null mice and wild-type (WT) controls. STAT5, STAT3, GHR, Socs 1–3, and Cis phosphorylation and abundance were assessed by using immunoblots, EMSA, and/or real time RT-PCR. TNF-α and IL-6 abundance were assessed by using ELISA. GH activated STAT5 in WT and TNFR1 or IL-6 null mice. LPS pretreatment prevented STAT5 activation in WT and TNFR1 null mice; however, STAT5 activation was preserved in IL-6 null mice. GHR abundance did not change with LPS administration. Inhibition of STAT5 activation by LPS was temporally associated with phosphorylation of STAT3 and upregulation of Cis and Socs-3 protein in WT and TNFR1 null mice; STAT3, Cis, and Socs-3 were not induced in IL-6 null mice. IL-6 inhibits hepatic GH signaling by upregulating Cis and Socs-3, which may involve activation of STAT3. Therapies that block IL-6 may enhance GH signaling in inflammatory diseases.
Luminal nutrient signals for intestinal adaptation in pythons
Authors: Secor Stephen M. , Lane John S. , Whang Edward E. , Ashley Stanley W. , Diamond Jared
Published: March 2015

Python intestine responds rapidly to luminal nutrients by increasing mass and upregulating nutrient transport. Candidates for luminal signals triggering those responses include mechanical stimulation, single or several dietary nutrients, and endogenous secretions. To identify signals, we infused into the python's small intestine either a nonnutrient solution (saline) or a single- or multinutrient solution. Python intestine failed to respond trophically or functionally to luminal infusions of saline, glucose, lipid, or bile. Infusion of amino acids and peptides, with or without glucose, induced an intermediate response. Infusion of nutritionally complete liquid formula or natural diet induced full intestinal response. Intact meals triggered full intestinal responses without pancreatic or biliary secretions, whereas direct cephalic and gastric stimulation failed to elicit any response. Hence neither physical stimulation (cephalic, gastric, or intestinal) nor the luminal presence of glucose, lipids, or bile can induce intestinal response; instead, a combination of nutrients is required (even without pancreaticobiliary secretions), the most important being amino acids and peptides. This is understandable because pythons, as carnivores, have a high-protein diet.
Expression of the lung resistance-related protein in human and rat hepatocarcinogenesis
Authors: Raidl Maria , Berger Walter , Schulte-Hermann Rolf , Kandioler-Eckersberger Daniela , Kappel Sonja , Wrba Fritz , Micksche Michael , Grasl-Kraupp Bettina
Published: March 2015

Lung resistance-related protein (LRP) plays an important role in chemoresistance of tumor cells probably by altering nuclear-cytoplasmic transport processes. We analyzed the association between LRP expression and hepatocarcinogenesis in humans and rats by RT-PCR, immunoblotting, and immunohistochemistry. LRP was found in hepatocytes and bile epithelia of normal human and rat liver showing distinct interindividual variations. In human tissues, the LRP expression levels of dysplastic liver nodules, hepatocellular adenomas, and carcinomas were highly variable, including decreased but also distinctly increased staining intensities. Mean expression levels, however, were comparable to the surrounding tissue. Considerable levels of LRP mRNA and protein were also found in human hepatoma cell lines. To study LRP expression from the beginning of hepatocarcinogenesis onward, rats were subjected to a tumor initiation/promotion protocol leading to preneoplastic hepatocytes present as single cells or multicellular clones, followed by adenoma and carcinoma. All of the (pre)neoplastic rat liver lesions expressed, comparable to the surrounding tissue, considerable amounts of LRP. We conclude that LRP might be one mechanism involved in the intrinsically high but variable chemoresistance of normal and (pre)neoplastic hepatocytes.
Aquaporins in rat pancreatic interlobular ducts
Authors: Ko Shigeru B. H. , Naruse Satoru , Kitagawa Motoji , Ishiguro Hiroshi , Furuya Sonoko , Mizuno Nobumasa , Wang Youxue , Yoshikawa Toshiyuki , Suzuki Atsushi , Shimano Shoko , Hayakawa Tetsuo
Published: March 2015

The aquaporin (AQP) family of water channels is distributed ubiquitously in many epithelia and plays a fundamental role in transmembrane water transport. The aim of this study is to identify the water transport pathway in pancreatic duct cells where most of the HCO[Formula: see text]-rich fluid originates. Using digital videomicroscopy, we measured the osmotic water permeability ( Pf) of pancreatic duct epithelium by exposing isolated rat interlobular ducts to the hypotonic solution (145 mosM). To identify mRNA and protein of AQPs expressed in duct cells, we conducted RT-PCR analysis and immunohistochemistry of the isolated duct and pancreas. The calculated Pf(160–230 μm/s) of the isolated ducts was significantly reduced to 16–35 μm/s by 80–90% with either basolateral or luminal applications of HgCl2. Fluid secretion evoked by secretin was almost completely abolished by a basolateral or luminal application of HgCl2. A large amount of AQP1 and small amounts of AQP5 transcripts were detected in the isolated duct cells by RT-PCR. AQP1, but not AQP5, immunoreactivity was present in both luminal and basolateral membranes of the interlobular duct cells. Mercury-sensitive water channels are present in both luminal and basolateral membranes of rat pancreatic ducts. AQP1 of the known AQPs appears to be the main water pathway in interlobular ducts.
Expression of enteropeptidase in differentiated enterocytes, goblet cells, and the tumor cells in human duodenum
Authors: Imamura Takahisa , Kitamoto Yasunori
Published: March 2015

Enteropeptidase (EP) is a serine proteinase and activates trypsinogen to trypsin, thus playing an important role in food digestion. Nevertheless, the localization of EP is still controversial, likely due to a lack of studies using specific antibodies against EP. The aim of this study was to define cellular localization of EP in human duodenum and expression in tumor cells at the duodenal region. Immunohistochemical staining for resected tissues was performed with two antibodies against recombinant EP light and heavy chains, respectively. In situ hybridization was done with two RNA probes that include either the light or the heavy chain sequences of proEP, respectively. The two antibodies reacted with enterocytes, accentuated on the brush border, and goblet cells, with increasing intensity from the bottom of crypts to the top of villi. Paneth cells, neuroendocrine cells, Brunner's glands, lymphocytes, smooth muscle, or connective tissue did not react with the antibodies. The two RNA probes detected EP mRNA expression only in enterocytes and goblet cells. EP is produced in enterocytes and goblet cells, and the localization on the brush border of the cells is reasonable for the physiological activation of digestive enzymes. Interestingly, the antibodies reacted with tumor cells in duodenal polyps and adenocarcinoma at the duodenum but not in Brunner's gland adenoma. EP seems to be a marker of differentiated enterocytes and goblet cells, which suggests the existence of a common progenitor of these cells. Furthermore, EP may be a useful marker of tumor cells originating from these cells.
Determinants of transpyloric fluid transport: a study using combined real-time ultrasound, manometry, and impedance recording
Authors: Savoye-Collet Céline , Savoye Guillaume , Smout André
Published: March 2015

Intraluminal impedance recording has made it possible to record fluid transport across the pylorus during the interdigestive state without filling the stomach. During antral phase II, fluid transport occurs with and without manometrically detectable antral contraction. Our aim was to investigate the relationships between ultrasonographic patterns of antral contraction, manometric pressure waves, and transpyloric fluid transport during antral phase II. Antral wall movements were recorded by real-time ultrasound (US) in eight healthy volunteers (mean age 24 ± 7 yr) during 17 ± 5 min of antral phase II. Concomitantly, a catheter positioned across the pylorus, monitored by transmucosal potential difference measurement, recorded five impedance signals (1 antral, 1 pyloric, and 3 duodenal) and six manometric signals (2 antral, 1 pyloric, and 3 duodenal). Antral contractions detected by US at the level of the two antral impedance electrodes were classified according to their association with a pyloric opening or a duodenal contraction. Transpyloric liquid transport events (impedance drop of >40% of the baseline with an antegrade or retrograde propagation) and manometric pressure waves (amplitude and duration) were identified during the whole study and especially during each period of US antral contraction. A total of 110 antral contractions was detected by US. Of these, 79 were also recorded by manometry. Fluid transport across the pylorus was observed in 70.9% of the US-detected antral contractions. Pyloric opening was observed in 98.6% of the contractions associated with fluid transport compared with 50% in the absence of fluid transport ( P < 0.05). Antral contractions associated with fluid transport were significantly ( P < 0.05) more often propagated to the duodenum (92%) than those without fluid transport (53%). Pressure waves associated with fluid transport were of higher amplitude (208 mmHg, range 22-493) and longer duration (7 s, range 2.5-13.5 s) than those not associated with fluid transport (102 mmHg, range 18-329 mmHg, and 4.1 s, range 2-8.5 s; P < 0.05). The propagation of the antral contractions in the duodenum in US was always associated with a pyloric opening, whereas only 8 of the 25 contractions without duodenal propagation were associated with a pyloric opening ( P < 0.05). The presence of duodenal contractile activity before the onset of an antral contraction in US was always accompanied by pyloric opening and with fluid transport in 93.3%, compared with 56.8% in its absence ( P < 0.05). In antral phase II, US is the most sensitive technique to detect antral contractions. Transpyloric fluid transport observed in relation to antral contractions occurs mainly in association with contractions of high amplitude and long duration and is associated with pyloric opening and/or duodenal propagation.
Localization and effects of orexin on fasting motility in the rat duodenum
Authors: Näslund E. , Ehrström M. , Ma J. , Hellström P. M. , Kirchgessner A. L.
Published: March 2015

The orexins [orexin A (OXA) and orexin B (OXB)] are novel neuropeptides that increase food intake in rodents. The aim of this study was to determine the distribution of orexin and orexin receptors (OX1R and OX2R) in the rat duodenum and examine the effects of intravenous orexin on fasting gut motility. OXA-like immunoreactivity was found in varicose nerve fibers in myenteric and submucosal ganglia, the circular muscle, the mucosa, submucosal and myenteric neurons, and numerous endocrine cells of the mucosa. OXA neurons displayed choline acetyltransferase immunoreactivity, and a subset contained vasoactive intestinal peptide. OXA-containing endocrine cells were identified as enterochromaffin (EC) cells based on the presence of 5-hydroxytryptamine immunoreactivity. OX1R was expressed by neural elements of the gut, and EC cells expressed OX2R. OXA at 100 and 500 pmol · kg−1 · min−1significantly increased the myoelectric motor complex (MMC) cycle length compared with saline. Similarly, OXB increased the MMC cycle length at 100 pmol · kg−1 · min−1, but there was no further effect at 500 pmol · kg−1 · min−1. We postulate that orexins may affect the MMC through actions on enteric neurotransmission after being released from EC cells and/or enteric neurons.
Cephalic phase of acid secretion involves activation of medullary TRH receptor subtype 1 in rats
Authors: Martínez Vicente , Barrachina María Dolores , Ohning Gordon , Taché Yvette
Published: March 2015

Mechanisms involved in the cephalic phase of gastric acid secretion were studied in awake fasted rats with chronic gastric fistula and exposed to the sight and smell of chow for 30 min. Acid secretion was monitored using constant intragastric perfusion and automatic titration. Sham feeding induced a peak acid response reaching 82 ± 7 μmol/10 min within 20 min compared with the average 22 ± 2 μmol/10 min in controls. The sham-feeding response was abolished by intracisternal pretreatment with the TRH1-receptor antisense oligodeoxynucleotides or subcutaneous injection of atropine, whereas TRH1 mismatch oligodeoxynucleotides had no effect. Serum gastrin was not altered by the sham feeding and increased by refeeding. Gastrin antibody did not block the rise in acid during sham feeding, although the net acid response was reduced by 47% compared with the control group. Glycine-gastrin antibody, indomethacin and nitro-l-arginine methyl ester had no effect. Atropine and gastrin antibody decreased basal acid secretion by 98 and 75%, respectively, whereas all other pretreatments did not. These results indicate that the cholinergic-dependent acid response to sham feeding is mediated by brain medullary TRH1 receptors in rats.
T cell receptor δ repertoire in inflamed and noninflamed colon of patients with IBD analyzed by CDR3 spectratyping
Authors: Holtmeier Wolfgang , Hennemann Andreas , May Ekkehard , Duchmann Rainer , Caspary Wolfgang F.
Published: March 2015

γ/δ T cells might play an important role in autoimmune conditions like inflammatory bowel disease (IBD). In the present study, we characterized the T cell receptor (TCR)-δ repertoire by complementarity determining region 3 (CDR3) spectratyping in the inflamed and noninflamed mucosa and in the peripheral blood of subjects with Crohn's disease and ulcerative colitis. In contrast to previously published data about α/β T cells, we rarely found oligoclonal expansions of γ/δ T cells specific only for the inflamed mucosa. The same dominant γ/δ T cell expansions were also present in the noninflamed colon. Furthermore, the peripheral γ/δ TCR repertoire was oligoclonal but clearly distinct from that in the inflamed intestine. Thus our results do not support a role for antigen-specific γ/δ T cells in IBD, and dominant γ/δ T cells of the peripheral blood are not likely to be derived from the inflamed gut. However, in several patients, the TCR-δ-repertoire was highly diversified, whereas in others we observed a loss of dominant γ/δ T cell clones when inflamed and noninflamed mucosa were compared. In conclusion, those changes indicate that γ/δ T cells might play an important role in a subset of patients with IBD.
III. Senescent enteric nervous system: lessons from extraintestinal sites and nonmammalian species
Authors: Wiley John W.
Published: March 2015

Functional changes in GI motility associated with advanced age include slowing of gastric emptying, decreased peristalsis, and slowing of colonic transit. These changes appear to be associated with region-specific loss of neurons and impaired function. The mechanism(s) underlying physiological aging are likely to be multifactorial. Alterations in specific signal transduction pathways have been reported at the level of the receptor and postreceptor events including kinase expression and function, mitochondrial function, and activation of the apoptosis cascade. Advanced age is associated with increased oxidative stress and its concomitant effects on cellular function. Whereas no specific genes have been causally linked to life span in mammals, studies involving nonmammalian species suggest that specific genes are involved in determining life span and age-related changes in cellular function. Caloric restriction is the only intervention shown to slow aging in a variety of species. Recent studies implicate a possible role for an insulin/IGF-I cascade in the region- and tissue-specific changes associated with physiological aging.
Differential effects of polyunsaturated fatty acids on sterol synthesis rates in adult and fetal tissues of the hamster: consequence of altered sterol balance
Authors: Schmid Kara E. , Woollett Laura A.
Published: March 2015

Cholesterol is necessary for the proper growth and development of the fetus. Consequently, disruptions in cholesterol biosynthesis lead to abnormal fetal development. It has been shown that in cells exposed to polyunsaturated fatty acids (PUFA), the expressions of genes and activities of enzymes involved in cholesterol synthesis are reduced. Similarly, we found that adult male hamsters fed PUFA-enriched diets had an ≈60% reduction in in vivo hepatic sterol synthesis rates. If fetal tissues respond to PUFA in the same manner as do adult livers, then maternal dietary PUFA could lead to a reduction in fetal sterol synthesis rates and possibly abnormal development. To investigate the impact of maternal dietary fatty acids on fetal sterol synthesis rates, female hamsters were fed diets enriched in various fatty acids before and throughout gestation. In vivo sterol synthesis rates were measured in fetuses at mid- and late gestation. At both gestational stages, dietary PUFA had no effect on fetal sterol synthesis rates. This lack of effect was not a consequence of a lack of PUFA enrichment in fetal fatty acids or the lack of PUFA receptor expression in the fetus. We hypothesize that the fetus may experience a dysregulation of sterol synthesis as the result of the fetus being in a negative sterol balance; the PUFA-induced suppression of sterol synthesis in the adult male hamster liver was ablated by creating a net negative sterol balance across the adult hepatocyte.
Prosurvival and antiapoptotic effects of PGE2in radiation injury are mediated by EP2receptor in intestine
Authors: Houchen Courtney W. , Sturmoski Mark A. , Anant Shrikant , Breyer Richard M. , Stenson William F.
Published: March 2015

The biological activities of PGE2are mediated through EP receptors (EP1–EP4), plasma membrane G protein-coupled receptors that differ in ligand binding and signal-transduction pathways. We investigated gastrointestinal EP2receptor expression in adult mice before and after radiation injury and evaluated intestinal stem cell survival and crypt epithelial apoptosis after radiation injury in EP2null mice. EP2was expressed throughout the gut. Intestinal EP2mRNA increased fivefold after γ-irradiation. Crypt survival was diminished in EP2−/−mice (4.06 crypts/cross section) compared with wild-type littermates (8.15 crypts/cross section). Radiation-induced apoptosis was significantly increased in EP2−/−mice compared with wild-type littermates. Apoptosis was 1.6-fold higher in EP2−/−mice (5.9 apoptotic cells/crypt) than in wild-type mice (3.5 apoptotic cells/crypt). The EP2receptor is expressed in mouse gastrointestinal epithelial cells and is upregulated following radiation injury. The effects of PGE2on both crypt epithelial apoptosis and intestinal crypt stem cell survival are mediated through the EP2receptor.
Heat shock induces intestinal-type alkaline phosphatase in rat IEC-18 cells
Authors: Harada Tsuyoshi , Koyama Iwao , Kasahara Toshihiko , Alpers David H. , Komoda Tsugikazu
Published: March 2015

We demonstrate a previously unknown regulation for intestinal-type alkaline phosphatase (IAP) as a heat shock protein (HSP). Heat shock to rat intestinal epithelial cells (IEC)-18 at 43°C induced the expression of IAP-I and HSP72 mRNAs time dependently (<60 min) but did not induce expression of IAP-II, tissue nonspecific-type alkaline phosphatase (TNAP), or HSP90 as determined by the RT-PCR method. To confirm the identity of the IAP-I gene, we sequenced the amplification product of IAP-I and found the gene to have 99% homology with the sequence of the IAP-I gene in rat intestine. Under the subculture conditions used, no IAP protein was detected in IEC-18 cells, but it became detectable as a 62-kDa band on a Western blot after heat shock. IAP-I was also induced by sodium arsenite, which generates reactive oxygen species and is an inducer of members of the HSP family. Glutathione suppressed activating protein-1 and cAMP response element-binding protein activation caused by heat shock but did not suppress the expression of IAP-I. These results suggest that cellular stress induces the elevation of IAP-I mRNA and protein synthesis. IAP-I may play an important role as a dephosphorylating enzyme under stress conditions.
Taurocholate feeding prevents CCl4-induced damage of large cholangiocytes through PI3-kinase-dependent mechanism
Authors: Marucci Luca , Alpini Gianfranco , Glaser Shannon S. , Alvaro Domenico , Benedetti Antonio , Francis Heather , Phinizy Jo Lynne , Marzioni Marco , Mauldin Jeremy , Venter Julie , Baumann Brandy , Ugili Laura , LeSage Gene
Published: March 2015

Bile acids are cytoprotective in hepatocytes by activating phosphatidylinositol-3-kinase (PI3-K) and its downstream signal AKT. Our aim was to determine whether feeding taurocholate to CCl4-treated rats reduces cholangiocyte apoptosis and whether this cytoprotective effect is dependent on PI3-K. Cholangiocyte proliferation, secretion, and apoptosis were determined in cholangiocytes from bile duct ligation (BDL), CCl4-treated BDL rats, and CCl4-treated taurocholate-fed rats. In vitro, we tested whether CCl4induces apoptosis and whether loss of cholangiocyte proliferation and secretion is dependent on PI3-K. The CCl4-induced cholangiocyte apoptosis and loss of cholangiocyte proliferation and secretion were reduced in CCl4-treated rats fed taurocholate. CCl4-induced cholangiocyte apoptosis, loss of cholangiocytes secretion, and proliferation were prevented by preincubation with taurocholate. Taurocholate cytoprotective effects were ablated by wortmannin. Taurocholate prevented, in vitro, CCl4-induced decrease of phosphorylated AKT protein expression in cholangiocytes. The cytoprotective effects of taurocholate on CCl4effects on cholangiocyte proliferation and secretion were abolished by wortmannin. Taurocholate protects cholangiocytes from CCl4-induced apoptosis by a PI3-K-dependent mechanism. Bile acids are important in the prevention of drug-induced ductopenia in cholangiopathies.
Cloning, expression, and localization of a rat hepatocyte inwardly rectifying potassium channel
Authors: Hill Ceredwyn E. , Briggs M. Martha , Liu Junjun , Magtanong Leslie
Published: March 2015

Bile formation involves anion accumulation within the apical lumen of hepatocytes. Potassium flux through hepatocellular basolateral membrane channels may provide the counterion for apical anion efflux. Here we cloned a molecular candidate for maintaining charge balance during bile secretion. Two transcripts resembling the Kir4.2 subclass of inwardly rectifying potassium channels were found. The longer deduced isoform (4.2a) has 30 additional NH3-terminal amino acids, which identifies this as a new isoform. The short-form isoform shared 86–91% identity with the mouse, human, and guinea pig channels. Whole cell currents of either rat isoform expressed in HEK293T cells demonstrated time independence and inward rectification. Antibodies against a COOH-terminal fragment recognized bands between 40 and 45 kDa and at 90 kDa and recognized a high molecular mass band around 200 kDa in overexpressing HEK cells. Immunohistology of liver tissue shows hepatocellular plasma membrane localization. In hepatocyte couplets, Kir4.2 was predominantly localized to the basolateral membrane. Results demonstrate expression of a new Kir4.2 isoform in the rat hepatocyte whose functional properties are compatible with a role in maintaining electrical integrity of bile-generating hepatocytes.
Effects ofHelicobacter pylorion intracellular Ca2+signaling in normal human gastric mucous epithelial cells
Authors: Marlink Katie L. , Bacon Kathy D. , Sheppard Brett C. , Ashktorab Hassan , Smoot Duane T. , Cover Timothy L. , Deveney Clifford W. , Rutten Michael J.
Published: March 2015

In stomach, Helicobacter pylori ( Hp) adheres to gastric mucous epithelial cells (GMEC) and initiates several different signal transduction events. Alteration of intracellular Ca2+concentration ([Ca2+]i) is an important signaling mechanism in numerous bacteria-host model systems. Changes in [Ca2+]iinduced by Hp in normal human GMEC have not yet been described; therefore, we examined effects of Hp on [Ca2+]iin normal human GMEC and a nontransformed GMEC line (HFE-145). Cultured cells were grown on glass slides, porous filters, or 96-well plates and loaded with fura 2 or fluo 4. Hp wild-type strain 60190 and vacA–, cagA–, and picB–/cagE–isogenic mutants were incubated with cells. Changes in [Ca2+]iwere recorded with a fluorimeter or fluorescence plate reader. Wild-type Hp produced dose-dependent biphasic transient [Ca2+]ipeak and plateau changes in both cell lines. Hp vacA–isogenic mutant produced changes in [Ca2+]isimilar to those produced by wild type. Compared with wild type, cagA–and picB–/cagE–isogenic mutants produced lower peak changes and did not generate a plateau change. Preloading cultures with intracellular Ca2+chelator BAPTA blocked all Hp-induced [Ca2+]ichanges. Thapsigargin pretreatment of cultures to release Ca2+from internal stores reduced peak change. Extracellular Ca2+removal reduced plateau response. Hp-induced peak response was sensitive to G proteins and PLC inhibitors. Hp-induced plateau change was sensitive to G protein inhibitors, src kinases, and PLA2. These findings are the first to show that H. pylori alters [Ca2+]iin normal GMEC through a Ca2+release/influx mechanism that depends on expression of cagA and picB/cagE genes.
Development and characterization of secretin-stimulated secretion of cultured rat cholangiocytes
Authors: Alpini Gianfranco , Phinizy Jo Lynne , Glaser Shannon , Francis Heather , Benedetti Antonio , Marucci Luca , LeSage Gene
Published: March 2015

We sought to develop a cholangiocyte cell culture system that has preservation of receptors, transporters, and channels involved in secretin-induced secretion. Isolated bile duct fragments, obtained by enzyme perfusion of normal rat liver, were seeded on collagen and maintained in culture up to 18 wk. Cholangiocyte purity was assessed by staining for γ-glutamyl transpeptidase (γ-GT) and cytokeratin-19 (CK-19). We determined gene expression for secretin receptor (SR), cystic fibrosis transmembrane conductance regulator, Cl−/HCO[Formula: see text] exchanger, secretin-stimulated cAMP synthesis, Cl−/HCO3exchanger activity, secretin-stimulated Cl−efflux, and apical membrane-directed secretion in polarized cells grown on tissue culture inserts. Cultured cholangiocytes were all γ-GT and CK-19 positive. The cells expressed SR and Cl−/HCO[Formula: see text] exchanger, and secretin-stimulated cAMP synthesis, Cl−/HCO[Formula: see text] exchanger activity, and Cl−efflux were similar to freshly isolated cholangiocytes. Forskolin (10−4M) induced fluid accumulation in the apical chamber of tissue culture inserts. In conclusion, we have developed a novel cholangiocyte line that has persistent HCO[Formula: see text], Cl−, and fluid transport functions. This cell system should be useful to investigators who study cholangiocyte secretion.