bluuebunny/crossref_metadata_2025 huggingface dataset:
Upregulation of rat intestinal uroguanylin mRNA by dietary zinc restriction
Authors: Blanchard R. K. , Cousins R. J.
Published: December 2017

A cDNA for the rat uroguanylin precursor was identified, by differential display of intestinal mRNA, as upregulated in zinc-deficient rats and subsequently was cloned. The cDNA and deduced amino acid sequences show a high degree of homology to human and opossum preprouroguanylin sequences. When used as a probe for Northern blot analysis of RNA from rat intestinal mucosa, the uroguanylin cDNA hybridized to a single species of mRNA that was 2.5-fold more abundant in zinc deficiency. A tissue distribution survey indicates that although the small intestine expresses a disproportionately high level of preprouroguanylin, this hormone precursor is also expressed in the colon, stomach, kidney, thymus, and testis. The induction by zinc deficiency is the first reported case of gene regulation for this hormone. These results also suggest a potential mechanism to explain, at least in part, the beneficial effects of zinc supplementation for secretory diarrhea prevalent in many areas of the world.
Cholecystokinin depolarizes guinea pig sphincter of Oddi neurons by activating CCK-A receptors
Authors: Gokin A. P. , Hillsley K. , Mawe G. M.
Published: December 2017

Motility studies indicate that cholecystokinin (CCK) acts through a neural mechanism in the sphincter of Oddi (SO) after meals. To evaluate its actions in SO ganglia, CCK was applied by microejection (0.1 mM) or superfusion (0.1 to 300 nM) while recording was carried out intracellularly from intact SO neurons. In tonic cells, microejection and superfusion of CCK caused a prolonged depolarization accompanied by action potentials. In phasic cells, microejection of CCK caused brief and/or prolonged depolarizations, but superfusion caused only prolonged depolarizations. In afterhyperpolarized cells, CCK did not cause a detectable change in the resting membrane potential. In low-Na+ Krebs solution, the prolonged depolarizations in both tonic and phasic cells were significantly reduced. Unsulfated CCK (100 nM) had no effect. CCK-induced depolarization was significantly reduced by a CCK-A, but not a CCK-B, receptor antagonist. It is concluded that CCK can act on CCK-A receptors to depolarize SO neurons. However, it is unlikely that hormonal CCK could mediate such an action because of the discrepancy between the sensitivity of SO neurons for CCK and the peak concentrations of CCK in the serum after a meal.
Asymmetry of lower esophageal sphincter pressure: is it related to the muscle thickness or its shape?
Authors: Liu J. , Parashar V. K. , Mittal R. K.
Published: December 2017

Lower esophageal sphincter (LES) pressure shows axial and circumferential asymmetry, the reasons for which are not clear. Our aim was to determine whether the muscle thickness and shape of the LES were the reasons for the axial and circumferential asymmetry in the LES pressure. High-frequency, catheter-based intraluminal ultrasonography was performed to obtain images of the human LES and esophagus. Station pull-through manometry was performed to record the axial and circumferential asymmetry of LES pressure. Circular and longitudinal muscle were thicker in the LES compared with in the esophagus. There was a close correlation between the axial asymmetry in LES pressure and circular muscle thickness. The thickness of LES muscle increased and decreased with an increase and decrease in the LES pressure, respectively. The circumferential asymmetry in resting LES pressure was related to the noncircular shape of the LES. A swallow-induced LES relaxation was followed by its contraction. During the contraction phase, the LES and esophagus showed relative symmetry in pressure and shape. We conclude that the axial asymmetry of LES pressure is explained by its muscle thickness. On the other hand, circumferential asymmetry is related to the noncircular shape of the LES.
Augmentation of deglutitive upper esophageal sphincter opening in the elderly by exercise
Authors: Shaker R. , Kern M. , Bardan E. , Taylor A. , Stewart E. T. , Hoffmann R. G. , Arndorfer R. C. , Hofmann C. , Bonnevier J.
Published: December 2017

Earlier studies have shown that the cross-sectional area of the deglutitive upper esophageal sphincter (UES) opening in healthy asymptomatic elderly individuals is reduced compared with healthy young volunteers. The aim of this study was to determine the effect of a head-raising exercise on swallow-induced UES opening and hypopharyngeal intrabolus pressure in the elderly. We studied a total of 31 asymptomatic healthy elderly subjects by videofluoroscopy and manometry before and after real (19 subjects) and sham (12 subjects) exercises. A significant increase was found in the magnitude of the anterior excursion of the larynx, the maximum anteroposterior diameter, and the cross-sectional area of the UES opening after the real exercise (P < 0.05). These changes were associated with a significant decrease in the hypopharyngeal intrabolus pressure studied in 12 (real-exercise) and 6 (sham-exercise) subjects (P < 0.05). A similar effect was not found in the sham-exercise group. In normal elderly subjects, deglutitive UES opening is amenable to augmentation by exercise aimed at strengthening the UES opening muscles. This augmentation is accompanied by a significant decrease in hypopharyngeal intrabolus pressure, indicating a decrease in pharyngeal outflow resistance. This approach may be helpful in some patients with dysphagia due to disorders of deglutitive UES opening.
GLUT2 and hexokinase control proximodistal gradient of intestinal glucose metabolism in the newborn pig
Authors: Cherbuy C. , Darcy-Vrillon B. , Posho L. , Vaugelade P. , Morel M. T. , Bernard F. , Leturque A. , Penicaud L. , Duee P. H.
Published: December 2017

We have reported previously that a high glycolytic capacity develops soon after birth in enterocytes isolated from suckling newborn pigs. In the present work, we investigated whether such metabolic changes could affect intestinal glucose utilization in vivo and examined possible variations in glucose metabolism along the small intestine. Glucose utilization by individual tissues was assessed using the 2-deoxyglucose technique. The overall glucose utilization rate was doubled in suckling vs. fasting 2-day-old pigs because of significantly higher rates in all tissues studied, except for the brain. In parallel, enterocytes were isolated from the proximal, medium, or distal jejunoileum of newborn vs. 2-day-old pigs and assessed for their capacity to utilize, transport, and phosphorylate glucose. Intestinal glucose consumption accounted for approximately 15% of glucose turnover rate in suckling vs. 8% in fasting pigs. Moreover, there was a proximal-to-distal gradient of glucose utilization in the intestinal mucosa of suckling pigs. Such a gradient was also evidenced on isolated enterocytes. The stimulation of both hexokinase activity (HK2 isoform) and basolateral glucose transporter (GLUT2), as observed in the proximal jejunum, could account for such a site-specific effect of suckling.
Effects of telenzepine and L-364,718 on canine pancreatic secretion before and after vagotomy
Authors: Niebergall-Roth E. , Teyssen S. , Wetzel D. , Hartel M. , Beglinger C. , Riepl R. L. , Singer M. V.
Published: December 2017

In six conscious dogs we compared the action of the M1-receptor antagonist telenzepine (20.25-81.0 nmol.kg-1.h-1), the cholecystokinin (CCK) antagonist L-364,718 (0.025-0.1 mg.kg-1.h-1), and combinations of both on the pancreatic secretory response to intraduodenal tryptophan, given against a secretin background before and after truncal vagotomy. Before vagotomy, the higher doses of telenzepine and of L-364,718 significantly (P < 0.05) decreased the protein response to tryptophan by up to 97%. After vagotomy, all doses of L-364,718 abolished the protein response, whereas telenzepine had no further effect. Before and after vagotomy, all combinations abolished the protein response. The plasma CCK-like immunoreactivity basally, during secretin, and in response to tryptophan was not altered by vagotomy, telenzepine, and/or L-364,718. These findings indicate that in dogs 1) potentiation exists between M1 receptors and CCK for stimulation of the pancreatic enzyme response to intraduodenal tryptophan, 2) the cholinergic fibers of the enteropancreatic reflex activated by tryptophan run within the vagus nerves and end at least in part on M1 receptors, 3) CCK acts in part independently of the vagal nerves, and 4) the CCK release by intestinal tryptophan is not influenced by vagotomy, telenzepine, and/or L-364,718.
Hypertrophy of colonic smooth muscle: contractile proteins, shortening velocity, and regulation
Authors: Siegman M. J. , Butler T. M. , Mooers S. U. , Trinkle-Mulcahy L. , Narayan S. , Adam L. , Chacko S. , Haase H. , Morano I.
Published: December 2017

Smooth muscle in megacolon was studied in the lethal spotted mouse and its normal sibling. In megacolon, structural remodeling and a very large increase in total protein content are associated with some changes in the contractile protein isoform composition. 1) There is a higher actin concentration in megacolon, primarily caused by a larger proportion of gamma-isoforms. 2) There was no difference in myosin concentration or in SM1/SM2 heavy chains in megacolon and normal muscle contractile proteins. 3) Only LC17a essential light chain is present in both normal and megacolon. 4) The 25- to 50-kDa 5'-insert occurs in 15-20% of the myosin in normal colon, compared with 5- to 10-fold lower amounts in megacolon. In permeabilized muscles there was no significant difference in unloaded shortening velocity (Vo) with maximal thiophosphorylation of the 20-kDa light chains, nor was there significant difference in the force vs. Ca2+ and force vs. myosin light chain phosphorylation relationships. At approximately 60% myosin light chain phosphorylation after Ca2+ activation, Vo of megacolon was approximately two times faster than Vo of normal muscle. These cellular changes largely account for the higher propulsive velocity of the colon in situ. The distribution of myosin and actin isoforms and the lack of a simple relationship between myosin light chain phosphorylation and Vo point to the operation of additional regulatory processes.
Gut ischemia and mesenteric synthesis of inflammatory cytokines after hemorrhagic or endotoxic shock
Authors: Tamion F. , Richard V. , Lyoumi S. , Daveau M. , Bonmarchand G. , Leroy J. , Thuillez C. , Lebreton J. P.
Published: December 2017

The intestine plays a major role in the pathophysiology of multiorgan failure. Although the systemic inflammatory response might be induced by endotoxin released through bacterial translocation, other factors such as intestinal ischemia might be implicated. We investigated the relationship between intestinal ischemia-reperfusion and cytokine release in rat models of hemorrhagic or endotoxic shock. Plasma levels of tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), lactate, and endotoxin, as well as macrophage TNF-alpha and IL-6 mRNA expression, were assessed at the end of shock and resuscitation. Hemodynamic changes and lactate levels suggested the presence of intestinal ischemia in both models. Mesenteric levels of TNF-alpha and IL-6 were increased by hemorrhage and further increased after saline resuscitation. Similar results were obtained with mRNA cytokine gene expression in macrophages. Endotoxin was not detectable in the hemorrhagic group. Endotoxic shock also increased production of cytokines, which, in contrast to hemorrhage, was not further increased by resuscitation. These results suggest that intestinal ischemia-reperfusion upon hemorrhage and resuscitation may be a major trigger for cytokine gene expression in the absence of endotoxin.
Chronic rejection causes early destruction of the intrinsic nervous system in rat intestinal transplants
Authors: Heeckt P. F. , Halfter W. , Schraut W. H. , Bauer A. J.
Published: December 2017

Chronic rejection is the major cause of late intestinal allograft dysfunction. We have previously shown that chronic rejection alters the muscularis externa of the graft. This study determined structural and functional changes to the enteric nerves during chronic rejection. Chronic rejection was achieved in orthotopic intestinal transplants (ACI to Lewis) by limited immunosuppression. Syngeneic transplants (ACI to ACI) and unoperated ACI rats served as controls. Animals were clinically healthy and showed no significant alterations in the mucosal architecture on postoperative day 90. Staining for NADPH diaphorase activity (nitric oxide synthase-containing neurons) and with neurofilament antibody RT-97 revealed that chronic rejection decreased the number of jejunal myenteric ganglia by approximately 50%. Inhibitory junction potentials (IJPs) to circular muscle cells were determined by electrical field stimulation (EFS). In controls and syngeneic grafts, EFS caused a stimulus-dependent increase in IJP amplitude, with a maximal amplitude of 9 +/- 0.4 and 10 +/- 0.8 mV, respectively. Chronic rejection in allografts markedly increased the threshold for IJP initiation and decreased the maximal IJP amplitude (5 +/- 0.8 mV). Our data indicate that chronic rejection severely damages the muscularis and the enteric nervous system before mucosal changes become evident.
Oxidant-induced disruption of intestinal epithelial barrier function: role of protein tyrosine phosphorylation
Authors: Rao R. K. , Baker R. D. , Baker S. S. , Gupta A. , Holycross M.
Published: December 2017

The effect of hydrogen peroxide (H2O2) on intestinal epithelial barrier function was examined in Caco-2 and T84 cell monolayers. H2O2reduced transepithelial electrical resistance (TER) of Caco-2 and T84 cell monolayers. This decrease in TER was associated with a decrease in dilution potential and an increase in [3H]mannitol permeability, suggesting an H2O2-induced disruption of the paracellular junctional complexes. H2O2administration also induced tyrosine phosphorylation of several proteins (at the molecular mass ranges of 50–90, 100–130, and 150–180 kDa) in Caco-2 cell monolayers. Phenylarsine oxide and sodium orthovanadate, inhibitors of protein tyrosine phosphatase, decreased TER and increased mannitol permeability and protein tyrosine phosphorylation (PTP). A low concentration of sodium orthovanadate also potentiated the effect of H2O2on TER, dilution potential, mannitol permeability, and PTP. Pretreatment with genistein (30–300 μM) and tyrphostin (100 μM) inhibited the effect of H2O2on TER, dilution potential, mannitol permeability, and PTP. These studies show that H2O2increases the epithelial permeability by disrupting paracellular junctional complexes, most likely by a PTP-dependent mechanism.
Dynamics of esophageal bolus transport in healthy subjects studied using multiple intraluminal impedancometry
Authors: Nguyen H. N. , Silny J. , Albers D. , Roeb E. , Gartung C. , Rau G. , Matern S.
Published: December 2017

The dynamics of a bolus transport through the esophagus are largely unexplored. To study this physiological process, we applied multiple intraluminal impedancometry in 10 healthy subjects. Three different protocols were used: 1) liquid bolus administered with subject supine, 2) liquid bolus with subject upright, or 3) semisolid bolus with subject supine. Transit of different parts of a bolus (bolus head, body, and tail) was analyzed at different anatomic segments, namely the pharynx and the proximal, middle, and distal thirds of the esophagus. A characteristic pattern of bolus transport was seen in all subjects. Impedance changes related to air were observed preceding the bolus head. The bolus head propelled significantly faster than did the bolus body and tail. Pharyngeal bolus transit was significantly faster than esophageal bolus transit. Within the esophagus, bolus propulsion velocity gradually decreased. Bolus transport was significantly accelerated in the upright position and delayed with increase of bolus viscosity. In conclusion, the dynamics of a bolus transport from the pharynx into the stomach are complex. It varies within both different anatomic segments and different parts of the bolus and depends on bolus characteristics and test conditions. The spatial and temporal resolution of a bolus transport can be obtained by the impedance technique.
Gastrin inhibits secretin-induced ductal secretion by interaction with specific receptors on rat cholangiocytes
Authors: Glaser Shannon S. , Rodgers Rebecca E. D. , Phinizy Jo Lynne , Robertson Willie E. , Lasater John , Caligiuri Alessandra , Tretjak Ziga , Lesage Gene D. , Alpini Gianfranco
Published: December 2017

We assessed the effect of gastrin on ductal secretion in normal and bile duct-ligated (BDL) rats. The effect of gastrin on ductal secretion was examined in the presence of proglumide, a specific antagonist for gastrin receptor (GR). We isolated pure cholangiocytes from normal and BDL rats and assessed gastrin effects on secretin receptor (SR) gene expression and intracellular adenosine 3′,5′-cyclic monophosphate (cAMP) levels. We examined the presence of GR mRNA in cholangiocytes by reverse transcription polymerase chain reaction (RT-PCR). In normal or BDL rats, gastrin produced no changes in spontaneous bile secretion. Simultaneous infusion of gastrin inhibited secretin-induced choleresis and bicarbonate output in BDL rats. In the presence of proglumide gastrin did not inhibit secretin-induced choleresis in BDL rats. Gastrin decreased in cholangiocytes from BDL rats 1) SR gene expression and 2) secretin-induced cAMP levels. With the use of RT-PCR, GR mRNA was detected in cholangiocytes. Similar to what is shown for secretin and somatostatin, we propose that the opposing effects of secretin and gastrin on cholangiocyte secretory activity regulate ductal secretion in rats.
Induction of nitric oxide synthase in rat gastric smooth muscle preparations
Authors: Zheng Xi-Long , Sharkey Keith A. , Hollenberg Morley D.
Published: December 2017

The induction in vitro of inducible nitric oxide synthase (iNOS) in intact gastric circular (CM) and longitudinal (LM) smooth muscle preparations was evaluated 1) pharmacologically, by the appearance of 1 mM l-arginine (l-Arg)-induced relaxation in a precontracted tissue; 2) biochemically, according to the appearance of iNOS mRNA using a reverse transcriptase-polymerase chain reaction; and 3) immunohistochemically, using an iNOS-specific antibody. Functionally, iNOS induction affected the contractile properties of the CM but not the LM preparation. The time course of iNOS induction monitored pharmacologically paralleled exactly the appearance of iNOS mRNA. The relaxant response to l-Arg in iNOS-induced CM tissues was blocked by the iNOS inhibitor aminoguanidine and by the guanylyl cyclase inhibitor LY-83583. The addition of oxyhemoglobin to the organ bath also attenuated the relaxant response, but tetrodotoxin had no effect. The transcriptional inhibitor actinomycin D completely blocked iNOS induction as assessed by both pharmacological and biochemical criteria. In iNOS-induced preparations the iNOS immunoreactivity was not detected in the smooth muscle elements but was localized in a layer of macrophage-related cells that were in apposition to the CM smooth muscle elements. We conclude that the spontaneous induction of iNOS in rat gastric tissue can affect the pharmacomechanical reactivity of the CM elements and that this regulation of the CM contractility is due to the induction of iNOS in a set of macrophage-related cells that are closely apposed to the CM elements so that they selectively affect only the contractility of the CM preparation.
III. Leukocyte adhesion and transmigration in the liver vasculature*
Authors: Jaeschke Hartmut , Smith C. Wayne
Published: December 2017

Leukocytes, i.e., neutrophils, monocytes, and lymphocytes, can accumulate in the hepatic vasculature and contribute to the pathophysiology of various liver diseases. Recently, significant progress has been made in the understanding of the basic mechanisms of neutrophil infiltration and cytotoxicity in the liver. However, there are a substantial number of unresolved issues. This article describes the current knowledge and the gaps in our understanding of mechanisms of neutrophil sequestration in sinusoids and venules, adhesion to endothelial cells, and transmigration and adherence to parenchymal cells. From these data, it is clear that assumptions regarding the roles of adhesion molecules in the liver may be misleading if drawn from studies of peripheral vascular beds. Greater insight into these mechanisms is critical for the development of selective therapeutic strategies that attenuate excessive inflammatory responses without compromising the vital host defense.
IV. Lymphocyte trafficking in the intestine and liver
Authors: Salmi M. , Adams D. , Jalkanen S.
Published: December 2017

Naive lymphocytes patrol continuously between the blood and different lymphatic tissues to sample the whole body for foreign antigens. During inflammation, leukocyte recruitment into tissue is enhanced to promote the recruitment of a range of effector cells into the affected area. The complex recirculatory pathways that underlie this process are governed by adhesion receptors on blood-borne leukocytes and by their specific ligands expressed on the luminal aspect of endothelial cells lining the vessels. Gut-associated lymphatic tissues are positioned strategically at the major port of entry for foreign antigens. They form a functionally unified entity that utilizes both the afferent and efferent arms of the immune response to respond to the large array of antigens entering via the gut under normal conditions as well as during inflammation. Once antigens have been absorbed from the gut, they may enter the portal vein and the liver where the immune response can be further regulated by the resident immune cells of the liver. Thus the gut and liver form an important barrier to enteral antigens, and leukocyte recruitment to these sites will need to be carefully regulated to ensure effective immune surveillance. In this article, we describe the current concepts of lymphocyte adhesion in these two organs as revealed by animal models. Subsequently, we discuss how well these principles apply to the lymphocyte-endothelial cell interactions in humans and what additional insights can be obtained from human studies.
Nucleotide metabolism by gastric glands and H+-K+-ATPase-enriched membranes
Authors: Rong Qinfen , Utevskaya Olga , Ramilo Marlon , Chow Dar C. , Forte John G.
Published: December 2017

α-Toxin-permeabilized gastric glands represent a functional model in which acid secretion can be elicited by either adenosine 3′,5′-cyclic monophosphate (cAMP) or ATP, with proven morphological and functional transition between resting and secretory states [X. Yao, S. M. Karam, M. Ramilo, Q. Rong, A. Thibodeau, and J. G. Forte. Am. J. Physiol. 271 ( Cell Physiol. 40): C61–C73, 1996.] In this study we use α-toxin-permeabilized rabbit gastric glands to study energy metabolism and the interplay between nucleotides to support acid secretion, as indicated by the accumulation of aminopyrine (AP). When permeabilized glands were treated with a phosphodiesterase inhibitor, the secretory response to cAMP was inhibited, whereas the secretory response to ATP was potentiated. This implied that 1) ATP provided support not only as an energy source but also as substrate for adenylate cyclase, 2) activation of acid secretion by cAMP needed ATP, and 3) ATP and cAMP exchanged rapidly inside parietal cells. To address these issues, we tested the action of adenine nucleotides in the presence and absence of oxidizable substrates. All adenine nucleotides, including AMP, ADP, ATP, and cAMP, could individually enhance the glandular AP accumulation in the presence of substrates, whereas only a high concentration of ATP (5 mM) was able to support secretory activity in substrate-free buffer. Moreover, ATP could maintain 75–80% of maximal secretory activity in phosphate-free buffer; cAMP alone could not support secretion in phosphate-free buffer. In glands and in H+-K+-adenosinetriphosphatase-rich gastric microsomes, we showed the operation of adenylate kinase, creatine kinase, and ATP/ADP exchange activities. These enzymes, together with endogenous adenylate cyclase and phosphodiesterase, provide the recycling of nucleotides essential for the viability of α-toxin-permeabilized gastric glands and imply the importance of nucleotide recycling for energy metabolism in intact parietal cells.
Peripheral mediators involved in gastric hyperemia to vagal activation by central TRH analog in rats
Authors: Király Ágnes , Sütö Gábor , Guth Paul H. , Taché Yvette
Published: December 2017

Mechanisms mediating the increase in gastric mucosal blood flow (GMBF) induced by the stable thyrotropin-releasing hormone (TRH) analog RX-77368 injected intracisternally at a gastric acid secretory dose (30 ng) were investigated using hydrogen gas clearance in urethan-anesthetized rats. The histamine H1 receptor antagonist pyrilamine (intravenously), capsaicin (subcutaneously, −10 days), and N G-nitro-l-arginine methyl ester (l-NAME, intracisternally) failed to impair the 150% rise in GMBF induced by intracisternal injection of RX-77368. By contrast, atropine (subcutaneously) and N G-monomethyl-l-arginine (intravenously) completely inhibited the increase in GMBF evoked by intracisternal RX-77368. l-NAME (intravenously) blocked the intracisternal RX-77368-induced increase in GMBF in capsaicin-pretreated rats, and thel-NAME effect was reversed by intravenous l- but notd-arginine. These findings indicate that vagal efferent activation induced by TRH analog injected intracisternally at a gastric acid secretory dose increases GMBF through atropine-sensitive mechanisms stimulatingl-arginine-nitric oxide pathways, whereas H1 receptors and capsaicin-sensitive afferent fibers do not play a role.
Protamine enhances the proliferative activity of hepatocyte growth factor in rats
Authors: Liu Ke-Xin , Kato Yukio , Kaku Tai-Ichi , Matsumoto Kunio , Nakamura Toshikazu , Sugiyama Yuichi
Published: December 2017

The effect of protamine on the proliferative activity of hepatocyte growth factor (HGF) was examined in α-naphthyl isothiocyanate-intoxicated rats. Protamine preinjection increased the hepatocyte labeling index induced by HGF four- to fivefold. A similar effect was also observed in partially hepatectomized rats. Because a cell surface heparin-like substance can bind to HGF and protamine has an affinity for heparin, protamine may affect HGF pharmacokinetics. In fact, protamine injection caused a transient increase in plasma HGF concentrations after administration of HGF and, in vitro, protamine eluted HGF prebound to heparin-Sepharose. Protamine also reduced the plasma clearance of HGF and increased 2.5-fold the exposure of hepatocytes to HGF in vivo. The enhancing effect of protamine on the mitogenic response of hepatocytes to HGF was also observed in vitro (∼2-fold after protamine pretreatment compared with HGF alone), suggesting that the enhancing effect of protamine on HGF-induced liver regeneration results from dual effects exerted by protamine 1) lowering the overall elimination of HGF and 2) directly stimulating hepatocyte mitosis induced by HGF.
Locations and molecular forms of PACAP and sites and characteristics of PACAP receptors in canine ileum
Authors: Mao Y. K. , Wang Y. F. , Moogk C. , Fox-Threlkeld J. E. T. , Xiao Q. , McDonald T. J. , Daniel E. E.
Published: December 2017

In canine ileum we investigated the distribution of pituitary adenylate cyclase-activating peptide (PACAP), using immunofluorescence and radioimmunoassay and the binding of125I-PACAP-27 to membranes. Nerve profiles immunoreactive to PACAP-27, and often to vasoactive intestinal polypeptide (VIP) as well, were found in all plexi, but PACAP was present in ∼100-fold lesser amounts than VIP. High-performance liquid chromatography analysis of deep muscular plexus (DMP) synaptosomes suggested the presence of PACAP-38, PACAP-27, and a third unidentified molecular form. High- and low-affinity125I-PACAP-27 binding sites were found in DMP synaptosomes and circular smooth muscle (CM) plasma membranes. In competition studies with DMP membranes, high (H)- and low (L)-affinity dissociation constants ( Kd) and maximal binding capacities (Bmax) were as follows: Kd H= 66.9 pM, Bmax H= 101 fmol/mg; Kd L= 2.18 nM, Bmax L= 580 fmol/mg protein. The binding of125I-PACAP-27 was fast. Dissociation was slow and incomplete in the presence of unlabeled PACAP-27 but accelerated by pretreatment with guanosine 5′- O-(3-thiotriphosphate) (GTPγS). GTPγS or cholera toxin treatment eliminated high-affinity binding in both membranes. VIP had ∼100-fold lower affinity than PACAP-27 in both membranes. Cross-linking studies identified an ∼70-kDa PACAP receptor in each membrane. Thus PACAP coexists with VIP in ileal enteric nerves and acts on PACAP-preferring, possibly Gs-coupled, receptors in DMP synaptosomes and CM membranes.
ICAM-1 and P-selectin expression in a model of NSAID-induced gastropathy
Authors: Morise Z. , Komatsu S. , Fuseler J. W. , Granger D. N. , Perry M. , Issekutz A. C. , Grisham M. B.
Published: December 2017

A growing body of experimental evidence suggests that neutrophilic polymorphonuclear leukocyte (PMN)-endothelial cell interactions play a critical role in the pathophysiology of nonsteroidal anti-inflammatory drug (NSAID)-induced gastropathy. The objective of this study was to directly determine whether the expression of endothelial cell adhesion molecules is enhanced in a model of NSAID-induced gastropathy. Gastropathy was induced in male Sprague-Dawley rats via oral administration of indomethacin (Indo, 20 mg/kg). Lesion scores, blood-to-lumen clearance of51Cr-EDTA (mucosal permeability), and histological analysis (epithelial necrosis) were used as indexes of gastric mucosal injury. Gastric mucosal vascular expression of intercellular adhesion molecule 1 (ICAM-1) or P-selectin were determined at 1 and 3 h after Indo administration using the dual radiolabeled monoclonal antibody (MAb) technique. For some experiments, a blocking MAb directed at either ICAM-1 (1A29) or P-selectin (RMP-1) or their isotype-matched controls was injected intravenously 10 min before Indo administration. We found that P-selectin expression was significantly increased at 1 h but not 3 h after Indo administration, whereas ICAM-1 expression was significantly increased at both 1 and 3 h after Indo treatment. The blocking ICAM-1 and P-selectin MAbs both inhibited Indo-induced increases in lesion score, mucosal permeability, and epithelial cell necrosis. However, the Indo-induced gastropathy was not associated with significant PMN infiltration into the gastric mucosal interstitium, nor did Indo reduce gastric mucosal blood flow. We propose that NSAID-induced gastric mucosal injury may be related to the expression of P-selectin and ICAM-1; however, this mucosal injury does not appear to be dependent on the extravasation of inflammatory cells or mucosal ischemia.
Differential localization of colonic H+-K+-ATPase isoforms in surface and crypt cells
Authors: Rajendran Vazhaikkurichi M. , Singh Satish K. , Geibel John , Binder Henry J.
Published: December 2017

Two distinct colonic H+-K+-adenosinetriphosphatase (H+-K+-ATPase) isoforms can be identified in part on the basis of their sensitivity to ouabain. The colonic H+-K+-ATPase α-subunit (HKcα) was recently cloned, and its message and protein are present in surface (and the upper 20% of crypt) cells in the rat distal colon. These studies were performed to establish the spatial distribution of the ouabain-sensitive and ouabain-insensitive components of both H+-K+-ATPase activity in apical membranes prepared from surface and crypt cells and K+-dependent intracellular pH (pHi) recovery from an acid load both in isolated perfused colonic crypts and in surface epithelial cells. Whereas H+-K+-ATPase activity in apical membranes from surface cells was 46% ouabain sensitive, its activity in crypt apical membranes was 96% ouabain sensitive. Similarly, K+-dependent pHirecovery in isolated crypts was completely ouabain sensitive, whereas in surface cells K+-dependent pHirecovery was insensitive to ouabain. These studies provide compelling evidence that HKcα encodes the colonic ouabain-insensitive H+-K+-ATPase and that a colonic ouabain-sensitive H+-K+-ATPase isoform is present in colonic crypts and remains to be cloned and identified.
Downregulation of a human colonic sialyltransferase by a secondary bile acid and a phorbol ester
Authors: Li Ming , Vemulapalli Ravi , Ullah Asad , Izu Leighton , Duffey Michael E. , Lance Peter
Published: December 2017

Fecal constituents such as bile acids and increased sialylation of membrane glycoproteins by α-2,6-sialyltransferase (HST6N-1) may contribute to colorectal tumorigenesis. We hypothesized that bile acids and phorbol ester [12- O-tetradecanoylphorbol-13-acetate (TPA)] would upregulate HST6N-1 in colonic cells. However, deoxycholate (DOC) (300 μmol/l), a secondary bile acid, and TPA (20 ng/ml) decreased expression of an ∼100-kDa glycoprotein bearing α-2,6-linked sialic acid in a colon cancer cell line (T84) in vitro. HST6N-1 mRNA levels were reduced ∼80% by treatment (≤24 h) with DOC or TPA but not by cholate, a primary bile acid. Treatment (24 h) with DOC or TPA decreased activity of this enzyme to 30% and 13% of control, respectively. These effects of DOC and TPA were transcriptional and were mediated by Ca2+and protein kinase C, respectively. Thus DOC and TPA both downregulated, and did not upregulate, α-2,6-sialyltransferase expression in vitro, but by different transduction pathways. As colorectal tumors grow, their progressive removal from the fecal milieu that normally downregulates this enzyme may favor invasion and metastasis.
II. Regulation of neuropeptide receptors in enteric neurons
Authors: McConalogue Karen , Bunnett Nigel W.
Published: December 2017

Neuropeptides exert their diverse biological effects by interacting with G protein-coupled receptors (GPCRs). In this review we address the question, What regulates the ability of a target cell, in particular a neuron, to respond to a neuropeptide? Available evidence from studies of many GPCRs in reconstituted systems and transfected cell lines indicates that much of this regulation occurs at the level of the receptor and serves to alter the capacity of the receptor to bind ligands with high affinity and to couple to heterotrimeric G proteins. Although some of the knowledge gained from these studies is applicable to the regulation of neuropeptide receptors on neurons, at present there are far more questions than answers.
Secretory PAF-acetylhydrolase of the rat hepatobiliary system: characterization and partial purification
Authors: Svetlov Stanislav I. , Howard Katherine M. , Debuysere Michael S. , Olson Merle S.
Published: December 2017

Hepatocytes and Kupffer cells in primary culture both secrete plasma-type platelet-activating factor-acetylhydrolase (pPAF-AH) into serum-free culture medium. The rate of secretion of pPAF-AH by Kupffer cells was 20 to 25 times higher than from hepatocytes, and Kupffer cells expressed a higher level of pPAF-AH mRNA than did hepatocytes. Purified liver cell-secreted pPAF-AH exhibited a major protein band of 65–67 kDa on SDS-PAGE; this was the band predominantly labeled when the enzyme catalytic center was reacted with [3H]diisopropylfluorophosphate ([3H]DFP). Rat bile collected from cannulated bile ducts contained significant PAF-AH activity, and bile samples possessed a prominent band at 30–32 kDa, which was the exclusive target for [3H]DFP. Experiments using tunicamycin, an inhibitor of N-linked glycosylation, and endoglycosidase H suggested that pPAF-AH secreted constitutively by cultured hepatocytes and Kupffer cells is glycosylated. The present study supports the notion that hepatic secretion of pPAF-AH into the blood contributes to the regulation of PAF and oxidized phospholipid levels in the circulation, whereas secretion of PAF-AH into the bile may allow hepatic control of these phospholipid signaling molecules in the gastrointestinal tract.
Molecular identification of a component of delayed rectifier current in gastrointestinal smooth muscles
Authors: Schmalz Felicitas , Kinsella Jacqueline , Koh Sang Don , Vogalis Fivos , Schneider Anne , Flynn Elaine R. M. , Kenyon James L. , Horowitz Burton
Published: December 2017

Kv2.2, homologous to the shab family of Drosophila voltage-gated K+channels, was isolated from human and canine colonic circular smooth muscle-derived mRNA. Northern hybridization analysis performed on RNA prepared from tissues and RT-PCR performed on RNA isolated from dispersed and selected smooth muscle cells demonstrate that Kv2.2 is expressed in smooth muscle cells found in all regions of the canine gastrointestinal (GI) tract and in several vascular tissues. Injection of Kv2.2 mRNA into Xenopus oocytes resulted in the expression of a slowly activating K+current (time to half maximum current, 97 ± 8.6 ms) mediated by 15 pS (symmetrical K+) single channels. The current was inhibited by tetraethylammonium (IC50= 2.6 mM), 4-aminopyridine (IC50= 1.5 mM at +20 mV), and quinine (IC50= 13.7 μM) and was insensitive to charybdotoxin. Low concentrations of quinine (1 μM) were used to preferentially block the slow component of the delayed rectifier current in native colonic myocytes. These data suggest that Kv2.2 may contribute to this current in native GI smooth muscle cells.
Novel properties of hepatic canalicular reduced glutathione transport revealed by radiation inactivation
Authors: Mittur Aravind V. , Kaplowitz Neil , Kempner Ellis S. , Ookhtens Murad
Published: December 2017

Transport of GSH at the canalicular pole of hepatocytes occurs by a facilitative carrier and can account for ∼50% of total hepatocyte GSH efflux. A low-affinity unit with sigmoidal kinetics accounts for 90% of canalicular transport at physiological GSH concentrations. A low-capacity transporter with high affinity for GSH has also been reported. It is not known whether the same or different proteins mediate low- and high-affinity GSH transport, although they do differ in inhibitor specificity. The bile of rats with a mutation in the canalicular multispecific organic anion transporter (cMOAT or MRP-2, a 170-kDa protein) is deficient in GSH, implying that cMOAT may transport GSH. However, transport of GSH in canalicular membrane vesicles (CMV) from these mutant rats remains intact. We examined the functional size of the two kinetic components of GSH transport by radiation inactivation of GSH uptake in rat hepatic CMV. High-affinity transport of GSH was inactivated as a single exponential function of radiation dose, yielding a functional size of ∼70 kDa. In contrast, low-affinity canalicular GSH transport exhibited a complex biexponential response to irradiation, characterized by an initial increase followed by a decrease in GSH transport. Inactivation analysis yielded a ∼76-kDa size for the low-affinity transporter. The complex inactivation indicated that the low-affinity transporter is associated with a larger protein of ∼141 kDa, which masked ∼80% of the potential transport activity in CMV. Additional studies, using inactivation of leukotriene C4transport, yielded a functional size of ∼302 kDa for cMOAT, indicating that it functions as a dimer.
Effects of Helicobacter pylorigastritis on gastric secretion in healthy human beings
Authors: Feldman Mark , Cryer Byron , Lee Edward
Published: December 2017

Helicobacter pylori gastritis is common, but effects on gastric secretion are not well understood. We measured basal and pentagastrin-stimulated gastric acidity, pepsin activity, and fluid output, as well as serum gastrin concentrations and H. pylori antibody levels, before and after treatment of H. pylori gastritis in 28 men and women. Subjects were studied before and 1 and 3 mo after a course of bismuth, metronidazole, and tetracycline. Elimination of H. pylori gastritis, accomplished in 14 subjects, increased basal and pentagastrin-stimulated gastric acidity (by 15 meq/l) and basal acid output significantly (by 2.1 meq/h 1 mo after therapy). Elimination of H. pylori had an opposite effect on pepsin secretion, significantly decreasing pepsin output by 30%. Elimination of H. pylori significantly reduced nonparietal fluid output by 35%, without affecting fluid output from parietal cells. Serum gastrin and H. pylori antibody levels declined significantly after elimination of H. pylori. None of these changes was observed in 14 subjects whose H. pylori gastritis was resistant to antimicrobial therapy. In summary, eradication of H. pylori infection increases gastric acidity by reducing nonparietal gastric secretion from peptic and other cells.
Pancreatic trypsinogen I expression during cell growth and differentiation of two human colon carcinoma cells
Authors: Bernard-Perrone Françoise , Carrere Jacqueline , Renaud Wanda , Moriscot Christine , Thoreux Karine , Bernard Patrice , Servin Alain , Balas Daniel , Senegas-Balas Françoise
Published: December 2017

Pancreatic trypsin has been found to induce tight junction or dome formation in some colon cancer cell lines (HT-29, Caco-2), and a tumor-associated trypsinogen, trypsinogen type II, has been isolated from another colon cancer cell line (COLO 205). We have tried to determine if trypsinogen is present and how its expression varies during cell culture in HT-29 Glc+/− and Caco-2 cells, which exhibit enterocytic differentiation, and in HT-29 Glc+ cells, which never differentiate. Trypsinogen mRNA presence and expression were demonstrated in these cells by mRNA hybridization, RT-PCR, cytoimmunofluorescence, Western immunoblot analysis, and gel filtration. Trypsinogen was found to be trypsinogen type I and was mainly in zymogen form in culture media. Differentiating cells exhibited variations in trypsinogen I expression, but cells that remained undifferentiated did not. In the differentiated cells, a high and transient peak in trypsinogen I expression was observed during the first steps of differentiation.
Origin of molecular species of diacylglycerol induced by bombesin in smooth muscle cells from rabbit rectosigmoid
Authors: Sbrissa Diego , Hajra Amyia , Bitar Khalil N.
Published: December 2017

The source of early production of sn-1,2-diacylglycerol (DAG) has for a long time been exclusively linked to hydrolysis of phosphatidylinositol 4,5-diphosphate, which on receptor activation is hydrolyzed into DAG and inositol 1,4,5-trisphosphate. We have investigated the origin of lipid sources of DAG production in smooth muscle cells, in response to contraction induced by peptide agonists. We have performed a quantitative analysis of the molecular species of DAG formed in relation to the known molecular composition of parent phospholipids. The molecular species of phospholipids are sufficiently unique that the phospholipid origin of DAGs and its quantitative contribution to their formation can be measured by HPLC. Cell suspensions (10–15 × 106cells/ml) from the circular muscle of rabbit rectosigmoid were incubated in the presence of the contractile peptide agonist bombesin (BB) at 10−6M. Reactions were stopped at different time intervals from 30 s to 4 min. DAGs were extracted, purified by TLC, and benzoylated with benzoic anhydride. The benzoylated DAGs were first purified by TLC and then by normal phase HPLC before they were injected onto a reverse-phase column and eluted isocratically. Furthermore, phospholipids in the lipid extract [phosphatidylinositol (PI), phosphatidylcholine (PC), phosphatidylserine (PS), and phosphatidylethanolamine (PE)] were purified by TLC and similarly analyzed after hydrolysis to DAGs with phospholipase C (PLC). The DAG molecular species profiles for PI, PC, PS, and PE were all unique. Contraction of cells with BB gave noticeable increases (17–55%) in newly formed DAGs. The major phospholipid source of the newly formed DAGs at 30 s was only ∼30% from PI, and the remainder was from PC. In contrast, after 4 min of BB stimulation, a decrease was seen in newly formed DAGs in the peak specific for PI hydrolysis. The data suggest that BB-induced contraction by activation of PLCs results in hydrolysis of different phospholipids. The DAGs formed as a result are qualitatively and quantitatively distinct. This could be the basis for the kinetically different pattern of sustained contraction observed with BB.
Substance P receptor expression in intestinal epithelium inClostridium difficiletoxin A enteritis in rats
Authors: Pothoulakis Charalabos , Castagliuolo Ignazio , Leeman S. E. , Wang Chi-Chung , Li Hanzong , Hoffman Beth J. , Mezey Eva
Published: December 2017

We previously reported that the inflammatory effects of Clostridium difficile toxin A on rat intestine can be significantly inhibited with a specific neurokinin-1 receptor (NK-1R) antagonist. In this study we investigated the localization and expression of NK-1R mRNA and protein in rat intestine by in situ hybridization, Northern blot analysis, and immunohistochemistry, respectively, after exposure to toxin A. Northern blot analysis showed increased mucosal levels of NK-1R mRNA starting 30 min after toxin A administration. In situ hybridization showed that toxin A increased NK-1R mRNA expression in intestinal epithelial cells after 30, 120, and 180 min. In rats pretreated with the NK-1R antagonist CP-96345 the increase in NK-1R mRNA levels after exposure to toxin A was inhibited, indicating that NK-1R upregulation is substance P (SP) dependent. One hour after exposure to toxin A many of the intestinal epithelial cells showed staining for NK-1R compared with controls. Specific125I-SP binding to purified epithelial cell membranes obtained from ileum exposed to toxin A for 15 min was increased twofold over control and persisted for 4 h. This report provides evidence that NK-1R expression is increased in the intestinal epithelium shortly after exposure to toxin A and may be important in toxin A-induced inflammation.
Role of calcium in adaptive cytoprotection and cell injury induced by deoxycholate in human gastric cells
Authors: Kokoska Evan R. , Smith Gregory S. , Wolff Andrew B. , Deshpande Yashwant , Rieckenberg Christopher L. , Banan Ali , Miller Thomas A.
Published: December 2017

We have developed an in vitro model of adaptive cytoprotection induced by deoxycholate (DC) in human gastric cells and have shown that pretreatment with a low concentration of DC (mild irritant, 50 μM) significantly attenuates injury induced by a damaging concentration of DC (250 μM). This study was undertaken to assess the effect of the mild irritant on changes in intracellular Ca2+and to determine if these perturbations account for its protective action. Protection conferred by the mild irritant was lost when any of its effects on intracellular Ca2+were prevented: internal Ca2+store release via phospholipase C and inositol 1,4,5-trisphosphate sustained Ca2+influx through store-operated Ca2+channels or eventual Ca2+efflux. We also investigated the relationship between Ca2+accumulation and cellular injury induced by damaging concentrations of DC. In cells exposed to high concentrations of DC, sustained Ca2+accumulation as a result of extracellular Ca2+influx, but not transient changes in intracellular Ca2+content, appeared to precede and induce cellular injury. We propose that the mild irritant disrupts normal Ca2+homeostasis and that this perturbation elicits a cellular response (involving active Ca2+efflux) that subsequently provides a protective action by limiting the magnitude of intracellular Ca2+accumulation.