bluuebunny/crossref_metadata_2025 huggingface dataset:
II. Leukocyte-endothelial cell adhesion in the digestive system
Authors: Granger D. Neil
Published: December 2017

The adhesion of leukocytes to vascular endothelial cells is a highly coordinated process that is governed by a number of factors, including the expression of specific adhesion glycoproteins, physical forces generated within the microcirculation, and inflammatory mediators released by a variety of activated cells. The digestive system, with its large resident population of immune cells and its tremendous capacity to generate inflammatory mediators, has proven to be a valuable source of information on the mechanisms involved in the regulation of leukocyte-endothelial cell adhesion. This article considers some of the evolving issues that have surfaced as a consequence of the rapidly growing body of literature on this topic. Particular emphasis is devoted to unresolved issues related to the expression and shedding of endothelial cell adhesion molecules, the contribution of capillaries to the inflammatory response, and the role of mast cells, macrophages, and lymphocytes in the modulation of leukocyte-endothelial cell adhesion.
Evidence for an ATP-independent long-chain phosphatidylcholine translocator in hepatocyte membranes
Authors: Fuchs Michael , Carey Martin C. , Cohen David E.
Published: December 2017

Transport of phosphatidylcholine (PC) molecules across canalicular plasma membranes of the liver is essential for their secretion into bile. To test for evidence of protein-mediated translocation of natural long-chain PCs, we investigated whether hepatocyte membrane subfractions reconstituted into proteoliposomes promoted transmembrane translocation of radiolabeled PCs. Translocation of PC molecules in proteoliposomes was measured by an assay that employed multilamellar acceptor vesicles and the specific PC transfer protein purified from liver. As inferred from the percentage of radiolabel removed from proteoliposomes, facilitated PC translocation occurred in microsomes and canalicular and basolateral plasma membranes from rat liver but not in erythrocyte ghosts, microsomes, homogenates of COS and H35 cells, or Xenopus laevis oocytes. Heat denaturation in the presence of 2-mercaptoethanol and Pronase digestion of solubilized membrane proteins inhibited translocation. In contrast to the mdr2 gene product (Mdr2), which promotes ATP-dependent, verapamil-inhibitable PC translocation, ATP did not enhance and verapamil failed to block PC translocation. These data support the possibility that an ATP-independent PC translocator, possibly distinct from Mdr2, may be present in hepatocyte canalicular plasma membranes.
Opposing effects of l-NAME on capillary filtration rate in the presence or absence of neutrophils
Authors: Harris Norman R.
Published: December 2017

Fluid filtration rate ( J v/ S) from rat mesenteric capillaries was measured using a modified Landis technique before and after superfusion with 100 μM N G-nitro-l-arginine methyl ester (l-NAME), a nitric oxide synthase inhibitor. Three groups were studied: 1) control rats, 2) rats injected with antineutrophil serum (ANS), and 3) rats injected with a monoclonal antibody (CL26) against the leukocyte adhesion molecule CD18. The relative increase in J v/ S(l-NAME/baseline) in control rats averaged 1.66 ± 0.32 ( n = 11), which was significantly higher ( P< 0.05) than in ANS (0.51 ± 0.12; n = 5)- and CL26 (0.45 ± 0.16; n = 6)-injected rats exposed tol-NAME. Thel-NAME-induced changes in J v/ Sin each group were due to altered permeability rather than altered pressure gradients, as determined by measurements of arteriolar hydrostatic pressure (using a servo-null apparatus) and estimates of intravascular oncotic pressure (using plasma protein concentration). These findings indicate that nitric oxide synthase inhibition increases the permeability of mesenteric capillaries to water, a response that is dependent on neutrophil adhesion.
Brain Fos expression and intestinal motor alterations during nematode-induced inflammation in the rat
Authors: Castex Nathalie , Fioramonti Jean , De Lahitte Jacques Ducos , Luffau Gerard , More Jean , Bueno Lionel
Published: December 2017

Brain-gut interactions and intestinal motility were studied during pulmonary and jejunal inflammation induced by Nippostrongylus brasiliensis. Jejunal electromyographic activity was continuously recorded from day 1 before to day 28 after infection. Expression of c- fos was assessed in the brain by immunohistochemistry, and myeloperoxidase (MPO) activity was determined in lung and intestine on days 1, 7, 14, 21, and 28 postinfection. The cyclic intestinal motor pattern was replaced by an irregular activity from day 4, corresponding to larvae migration to the intestine, to day 14. c- fos was expressed in the caudal nucleus of the solitary tract (NTS) and lateral parabrachial nucleus (LPB) on day 1 (lung stage of N. brasiliensis) and in the medial part of the NTS, the LPB, and locus ceruleus on day 7. Pulmonary and intestinal MPO activity was increased from days 1 to 21 postinfection. During N. brasiliensis infection, c- fos expression indicates that specific and different brain nuclei are activated at the onset of pulmonary and intestinal inflammation, which is associated with motor disorders.
Erythromycin gastrokinetic activity is partially vagally mediated
Authors: Mathis C. , Malbert C. H.
Published: December 2017

Erythromycin overcomes postvagotomy gastroparesia in patients without a distal stomach and functional pylorus. We investigate the role of the vagus in gastric emptying increased by erythromycin, using a model that preserves the physiology of the distal stomach and pylorus. The effects of erythromycin lactobionate (10 mg/kg) on transpyloric flow pattern and pyloric resistance were evaluated during repetitive bilateral vagal cooling in anesthetized pigs. Vagal cooling during erythromycin infusion produced a marked decreased of pyloric outflow (23 ± 1.1 vs. 50 ± 2.6 ml/min) related to a reduced stroke volume of the flow pulses (7.8 ± 3.31 vs. 14.1 ± 2.44 ml). The amplitude and frequency of gastric and duodenal pressure events were unchanged or slightly reduced during vagal cooling. The smaller stroke volume of flow pulse was the consequence of increased pyloric resistance (6.2 ± 1.98 vs. 2.3 ± 0.21 mmHg ⋅ ml−1 ⋅ s), which is associated with changes in the temporal relationship between a pyloric pressure event and flow pulse. In conclusion, erythromycin activity on the pylorus requires the integrity of vagal pathways. Enhancement of gastric outflow by erythromycin is also modulated by the vagus, since pyloric resistance was able to overcome increased gastric motility.
II. One size fits all: nonclassical MHC molecules fulfill multiple roles in epithelial cell function
Authors: Blumberg Richard S.
Published: December 2017

The human major histocompatibility complex (MHC) on chromosome 6 encodes three classical class I genes: human leukocyte antigen-A (HLA-A), HLA-B, and HLA-C. These polymorphic genes encode a 43- to 45-kDa cell surface glycoprotein that, in association with the 12-kDa β2-microglobulin molecule, functions in the presentation of nine amino acid peptides to the T cell receptor of CD8-bearing T lymphocytes and killer inhibitory receptors on natural killer cells. In addition to these ubiquitously expressed polymorphic proteins, the human genome also encodes a number of nonclassical MHC class I-like, or class Ib, genes that in general encode nonpolymorphic molecules involved in a variety of specific immunologic functions. Many of these genes, including CD1, the neonatal Fc receptor for immunoglobulin G, HLA-G, the MHC class I chain-related gene A, and Hfe, are prominently displayed on epithelial cells, suggesting an important role in epithelial cell biology.
Shear-induced modulation of vasoconstriction in the hepatic artery and portal vein by nitric oxide
Authors: Macedo M. Paula , Lautt W. Wayne
Published: December 2017

The effect of shear stress on nitric oxide (NO)-mediated suppression of sympathetic nerve (2–6 Hz)- and norepinephrine (0.5 μg ⋅ kg−1 ⋅ min−1)-induced vasoconstriction in the hepatic artery (HA) and portal vein (PV) was studied using a perfusion circuit to regulate blood pressure and flow in the cat liver in situ. Holding flow constant resulted in increased shear stress during constriction; holding pressure steady prevented changes in shear stress. When shear stress was allowed to rise, the vasoconstriction (indicated by elevation in perfusion pressure) in response to nerve stimulation and norepinephrine was significantly potentiated after NO synthase blockade using N G-nitro-l-arginine methyl ester (l-NAME, 2.5 mg/kg iv) in both the HA and PV (response to nerves: HA control 28.8 ± 6.5 mmHg,l-NAME 62.7 ± 14.6 mmHg; PV control 1.5 ± 0.5 mmHg,l-NAME 3.3 ± 0.5 mmHg; response to norepinephrine: HA control 32.4 ± 9.0 mmHg, l-NAME 60.3 ± 8.0 mmHg; PV control 1.3 ± 0.3 mmHg,l-NAME 3.4 ± 0.7 mmHg). The potentiation was reversed byl-arginine (75 mg/kg). When shear stress was held constant by maintaining constant perfusion pressure, l-NAME did not cause potentiation of vasoconstriction. The data are consistent with the hypothesis that elevated shear stress in the hepatic blood vessels leads to NO-dependent postjunctional modulation of vasoconstriction.
GATA-6 stimulates a cell line-specific activation element in the human lactase promoter
Authors: Fitzgerald Kevin , Bazar Leonard , Avigan Mark I.
Published: December 2017

Lactase-phlorizin hydrolase (LPH) synthesis is restricted to differentiated small intestinal enterocytes and is highly regulated during development. Analysis of expression of LPH promoter segments fused with luciferase transfected in Caco-2 cells, a line that uniquely expresses LPH mRNA, mapped an 18-base pair (bp) segment 100 bp upstream of the transcription start site that is required for transactivation. Remarkably, the LPH upstream element (LUE) has no stimulatory activity in both human intestinal and nonintestinal lines in which LPH mRNA is absent. Electrophoretic analysis of sequence-specific DNA-nuclear protein complexes demonstrated the presence of a Caco-2 cell-specific protein(s) (CCP), which is uniformly absent in LPH nonproducer cell lines. Mutational analysis of the LUE demonstrated that bases contained within a GATA consensus motif are critical for both CCP binding and transcription from the LPH promoter. Caco-2 cells express high levels of GATA-6 mRNA in a cell line- specific manner, suggesting that GATA-6 is a CCP that complexes with the LUE. When expressed by a plasmid, GATA-6 transactivated the LPH promoter. The stimulation was abrogated with mutations in the GATA consensus motif as well as mutations in a flanking downstream element. These studies are consistent with an important role of an intestinal GATA binding protein in cell type-specific transactivation of the LPH promoter.
Distribution of heme oxygenase and effects of exogenous carbon monoxide in canine jejunum
Authors: Farrugia G. , Miller S. M. , Rich A. , Liu X. , Maines M. D. , Rae J. L. , Szurszewski J. H.
Published: December 2017

Carbon monoxide (CO) has been postulated to be a messenger in the gastrointestinal tract. The aims of this study were to determine the distribution of heme oxygenase (HO), the source for endogenous CO in the canine jejunum, and to determine the effects of CO on jejunal circular smooth muscle cells. HO-2 isoform was present in a population of myenteric and submucosal neuronal cell bodies, in nerve fibers innervating the muscle layers, and in smooth muscle cells. HO-1 isozyme was not detected in the canine jejunum. Exogenous CO increased whole cell current by 285 ± 86%, hyperpolarized the membrane potential by 8.5 ± 2.9 mV, and increased guanosine 3′,5′-cyclic monophosphate (cGMP) levels in smooth muscle cells. 8-Bromo- cGMP also increased the whole cell current. The data suggest that endogenous activity of HO-2 may be a source of CO in the canine jejunum and that exogenously applied CO can modulate intestinal smooth muscle electrical activity. It is therefore reasonable to suggest a role for endogenously produced CO as a messenger in the canine jejunum.
A 100-year perspective on gastrointestinal motility
Authors: Szurszewski J. H.
Published: December 2017

This contribution to the centennial commemorative issue of the American Journal of Physiology: Gastrointestinal and Liver Physiology identifies some of the important studies of spontaneous electrical and motor activity in the gastrointestinal tract published in the Journal between 1898 and 1996. Emphasis is given to the contributions made by Walter B. Cannon, Walter C. Alvarez, Emil Bozler, C. Ladd Prosser, and James Christensen.
Nonionic diffusion of short-chain fatty acids across rat colon
Authors: Charney Alan N. , Micic Ljubisa , Egnor Richard W.
Published: December 2017

Short-chain fatty acid (SCFA) transport across the colon may occur by nonionic diffusion and/or via apical membrane SCFA−/[Formula: see text]exchange. To examine the relative importance of these processes, stripped segments of rat ( Ratus ratus) proximal and distal colon were studied in Ussing chambers, and the unidirectional fluxes of radiolabeled SCFA butyrate, propionate, or weakly metabolized isobutyrate were measured. In N-2-hydroxyethylpiperazine- N′-2-ethanesulfonic acid (HEPES) or 1 or 5 mM [Formula: see text] Ringer, decreases in mucosal pH stimulated mucosal-to-serosal flux ( J m→s) of all SCFA, decreases in serosal pH stimulated serosal-to-mucosal flux ( J s→m), and bilateral pH decreases stimulated both fluxes equally. These effects were observed whether the SCFA was present on one or both sides of the tissue, in both proximal and distal colon, in the absence of luminal Na+, and in the presence of either luminal or serosal ouabain. Changes in intracellular pH or intracellular [[Formula: see text]] did not account for the effects of extracellular pH. Luminal Cl− removal, to evaluate the role of apical membrane Cl−/SCFA−exchange, had no effect on J m→s but decreased J s→m 32% at pH 6.5 and 22% at 7.2. Increasing SCFA concentration from 1 to 100 mM, at pH 6.4 or 7.4, caused a linear increase in J m→s. We conclude that SCFA are mainly transported across the rat colon by nonionic diffusion.
Differential susceptibility of inbred mouse strains to dextran sulfate sodium-induced colitis
Authors: Mähler Michael , Bristol Ian J. , Leiter Edward H. , Workman Aletha E. , Birkenmeier Edward H. , Elson Charles O. , Sundberg John P.
Published: December 2017

Dextran sulfate sodium (DSS)-induced murine colitis represents an experimental model for human inflammatory bowel disease. The aim of this study was to screen various inbred strains of mice for genetically determined differences in susceptibility to DSS-induced colitis. Mice of strains C3H/HeJ, C3H/HeJBir, C57BL/6J, DBA/2J, NOD/LtJ, NOD/LtSz- Prkdcscid/Prkdcscid, 129/SvPas, NON/LtJ, and NON.NOD- H2g7were fed 3.5% DSS in drinking water for 5 days and necropsied 16 days later. Ceca and colons were scored for histological lesions based on severity, ulceration, hyperplasia, and area involved. Image analysis was used to quantitate the proportion of cecum ulcerated. Histological examination revealed significant differences among inbred strains for all parameters scored. In both cecum and colon, C3H/HeJ and a recently selected substrain, C3H/HeJBir, were highly DSS susceptible. NOD/LtJ, an autoimmune-prone strain, and NOD/LtSz- Prkdcscid/Prkdcscid, a stock with multiple defects in innate and adoptive immunity, were also highly DSS susceptible. NON/LtJ, a strain closely related to NOD, was quite DSS resistant. The major histocompatibility (MHC) haplotype of NOD mice ( H2g7), a major component of the NOD autoimmune susceptibility, was not crucial in determining DSS susceptibility, since NON mice congenic for this MHC haplotype retained resistance. C57BL/6J, 129/SvPas, and DBA/2J mice showed various degrees of susceptibility, depending upon the anatomical site. A greater male susceptibility to DSS-induced colonic but not cecal lesions was observed. In summary, this study demonstrates major differences in genetic susceptibility to DSS-induced colitis among inbred strains of mice. Knowledge of these strain differences in genetic responsiveness to acute inflammatory stress in the large intestine will permit design of genetic crosses to elucidate the genes involved.
Involvement of the 5-HT3 receptor in CRH-induced defecation in rats
Authors: Miyata Keiji , Ito Hiroyuki , Fukudo Shin
Published: December 2017

We evaluated the possibility that serotonin (5-HT) mediates defecation induced by corticotropin-releasing hormone (CRH) exogenously administered or released from the central nervous system by stress via the 5-HT3 receptor in rats. Intracerebroventricular (ICV) injection of CRH (1, 3, and 10 μg/rat) dose dependently increased the number of stools excreted in rats, whereas intravenous (IV) injection of up to 100 μg/kg CRH did not affect defecation. α-Helical CRH-(9—41) and 5-HT3 receptor antagonists ramosetron and azasetron inhibited CRH (10 μg icv)-induced defecation in a dose-dependent manner with ED50 values of 4.3 μg/kg iv, 3.8 μg/kg po, and 70.4 μg/kg po, respectively. α-Helical CRH-(9—41) also inhibited CRH-induced defecation by ICV injection with an ED50 value of 0.078 μg/rat. In contrast, ramosetron and azasetron injectied ICV had no effect on CRH-induced defecation. α-Helical CRH-(9—41), ramosetron, and azasetron reduced defecation caused by restraint stress with ED50 values of 0.32, 3.6, and 19.7 μg/kg iv, respectively. These results indicate that CRH exogenously administered or released from the central nervous system by stress peripherally promotes the release of 5-HT, which in turn stimulates defecation through the 5-HT3 receptor.
Effect of nitric oxide on calcium-activated potassium channels in colonic smooth muscle of rabbits
Authors: Lu Gang , Mazet Bruno , Sarr Michael G. , Szurszewski Joseph H.
Published: December 2017

Nitric oxide (NO) hyperpolarizes intestinal smooth muscle cells. This study was designed to determine the mechanism whereby NO activates KCa channels of circular smooth muscle of the rabbit colon. Transmural biopsies of the rabbit colon were stained for NADPH-diaphorase. Freshly dispersed circular smooth muscle cells were studied in the whole cell configuration, as well as in on-cell and excised inside-out patch recording configurations, while KCa current and the activity of KCa channels, respectively, were monitored. NADPH-diaphorase-positive nerve fibers were found in both muscle layers. NO (1%) increased whole cell net outward current by 79% and hyperpolarized resting membrane voltage from −59 to −73 mV ( n = 8 cells, P < 0.01). In the on-cell patch recording configuration, NO (0.5% or 1%) in the bath increased NP o of KCa channels; charybdotoxin (125 nM) in the pipette solution blocked this effect. In the excised inside-out patch recording configuration, NO (1%) had no effect on NP o of KCa channels. In the on-cell patch recording configuration, methylene blue (1 μM) or cystamine (5 mM) in the bath solution decreased the effect of NO (1%) on NP o of KCa channels. NP o was increased by 8-bromo-cGMP (8-BrcGMP; 1 mM), a cGMP analog, and zaprinast (100 μM), an inhibitor of cGMP phosphodiesterase. These data suggest that NO increased whole cell outward K+current by activating KCa channels through a cGMP pathway.
Lack of defect in insulin action on hepatic glycogen synthase and phosphorylase in insulin-resistant monkeys
Authors: Ortmeyer Heidi K. , Bodkin Noni L.
Published: December 2017

It is well known that an alteration in insulin activation of skeletal muscle glycogen synthase is associated with insulin resistance. To determine whether this defect in insulin action is specific to skeletal muscle, or also present in liver, simultaneous biopsies of these tissues were obtained before and during a euglycemic hyperinsulinemic clamp in spontaneously obese insulin-resistant male rhesus monkeys. The activities of glycogen synthase and glycogen phosphorylase and the concentrations of glucose 6-phosphate and glycogen were measured. There were no differences between basal and insulin-stimulated glycogen synthase and glycogen phosphorylase activities or in glucose 6-phosphate and glycogen contents in muscle. Insulin increased the activities of liver glycogen synthase ( P < 0.05) and decreased the activities of liver glycogen phosphorylase ( P ≤ 0.001). Insulin also caused a reduction in liver glucose 6-phosphate ( P = 0.05). We conclude that insulin-resistant monkeys do not have a defect in insulin action on liver glycogen synthase, although a defect in insulin action on muscle glycogen synthase is present. Therefore, tissue-specific alterations in insulin action on glycogen synthase are present in the development of insulin resistance in rhesus monkeys.
Cyclooxygenase-2 inhibitors suppress the growth of gastric cancer xenografts via induction of apoptosis in nude mice
Authors: Sawaoka Hitoshi , Kawano Sunao , Tsuji Shingo , Tsujii Masahiko , Gunawan Edhi S. , Takei Yoshiyuki , Nagano Kouichi , Hori Masatsugu
Published: December 2017

To clarify the role of mitogen-inducible cyclooxygenase (COX-2) in the development of malignant tumors, we investigated the effects of COX-2 inhibitors on the growth of gastric cancer xenografts in nude mice in vivo. MKN45 gastric cancer cells (5 × 106cells/animal) that overexpress COX-2 were inoculated subcutaneously into athymic mice. NS-398, a specific COX-2 inhibitor, or indomethacin, a nonspecific COX-2 inhibitor, was administered orally to animals every day for 20 days. These drugs reduced the tumor volume significantly. Immunohistochemistry using bromodeoxyuridine, nick end labeling, and electron microscopy showed that NS-398 induced apoptosis in cancer cells in a dose-dependent manner and inhibited cancer cell replication slightly. Indomethacin also induced apoptosis and suppressed replication of tumor cells. There was a significant negative correlation between tumor volume and apoptotic cell number within the tumor. These results are consistent with the hypothesis that COX-2 inhibitors suppress growth of gastric cancer xenografts mainly by inducing apoptosis and suppressing replication of the neoplastic cells. It follows that COX-2 plays an important role in the development of gastric cancer.
Involvement of intracellular Ca2+stores in inhibitory effects of NO donor SIN-1 and cGMP
Authors: Franck Hartmut , Storr Martin , Puschmann Andreas , Schusdziarra Volker , Allescher Hans-Dieter
Published: December 2017

We investigated the role of K+channels and intracellular Ca2+stores in the relaxations induced by the NO donor 3-morpholinosydnonimine (SIN-1) and 8-bromo-cGMP (8-BrcGMP), 8-(4-chlorophenylthio)-cGMP (pCPT-cGMP), and α,β-methylene-ATP in isolated segments of rat ileum. The inhibitory responses to SIN-1 and the cGMP analogs were not influenced by the K+blockers apamin, charybdotoxin, iberiotoxin, or glibenclamide, whereas relaxations induced by α,β-methylene-ATP were abolished by apamin and tetraethylammonium. The NO-donor SIN-1 and the cGMP analogs were able to inhibit contractions induced by activation of L-type Ca2+channels (BAY-K-8644), by carbachol (CCh), and by cyclopiazonic acid (CPA), a blocker of sarcoplasmic Ca2+-ATPase. However, the inhibition of the combined CPA and CCh response was reduced and the dose-response curve of SIN-1 shifted to the right. Intracellular Ca2+stores were emptied by incubation in Ca2+-free buffer and repetitive stimulation with CCh or BAY-K-8644. After restoration of extracellular Ca2+, the inhibitory effect of SIN-1 and pCPT-cGMP was only attenuated, whereas in the additional presence of CPA, the inhibitory effect of SIN-1 was blocked and the effect of 8-BrcGMP reduced. Thus depleting intracellular Ca2+stores attenuated the effect of SIN-1 and 8-BrcGMP, suggesting an involvement of functional Ca2+stores.
Sterol carrier protein-2 expression increases NBD-stearate uptake and cytoplasmic diffusion in L cells
Authors: Murphy Eric J.
Published: December 2017

The effects of sterol carrier protein-2 (SCP-2) expression on fatty acid uptake and cytoplasmic diffusion were determined using L cell fibroblasts transfected with cDNA encoding either the 15- or 13.2-kDa form of SCP-2. Cis-parinarate and 12- N-methyl-(7-nitrobenz-2-oxa-1,3-diazol)aminostearate (NBD-stearate) were used as nonesterifiable fluorescent fatty acid probes. NBD-stearate and cis-parinarate uptake was rapid and saturable. In 15-kDa SCP-2-expressing cells, the extent of NBD-stearate and cis-parinarate uptake was increased 1.4- and 1.2-fold, respectively, compared with control. In the 13.2-kDa SCP-2-expressing cells, the extent of NBD-stearate and cis-parinarate uptake was increased 1.3- and 1.1-fold, respectively, compared with control cells. NBD-stearate cytoplasmic diffusion was increased 1.5-fold in 15-kDa SCP-2-expressing cells, but not in 13.2-kDa SCP-2-expressing cells, compared with control cells. After incubation with NBD-stearate for 30 min at 37°C, fluorescence imaging indicated that NBD-stearate was localized primarily in lipid droplets in all cell lines. These results suggest that SCP-2 may be involved not only in fatty acid uptake but also in intracellular fatty acid trafficking.
Rat serum induces a differentiated phenotype in a rat parotid acinar cell line
Authors: Zhu Yingting , Aletta John M. , Wen Jiayu , Zhang Xuejun , Higgins Dennis , Rubin Ronald P.
Published: December 2017

To establish a continuous cell line, freshly prepared rat parotid acinar cells were stably transfected with a plasmid vector containing the SV40 large T antigen. The acinar origin of these cells was confirmed by Western blotting, enzyme analysis, and morphological analysis. Transformed cells grown in 10% rat serum showed a modest reduction in cell number after 7 days and a concentration- and time-dependent increase in amylase levels ∼16 times greater than those observed in fetal bovine serum-treated cells. Ultrastructural analysis revealed that cells grown in rat serum harbored protein-filled secretory granules localized adjacent to the endoplasmic reticulum, and punctate amylase-specific immunofluorescence distributed throughout the cytoplasm was consistent with the presence of amylase in secretory organelles. Clonal cells express tissue-specific proline-rich proteins and the four protein kinase C isozymes present in primary culture. Carbachol and isoproterenol stimulated [3H]protein secretion and isoproterenol enhanced amylase secretion from cells grown in rat serum. Moreover, norepinephrine, carbachol, and substance P produced a time- and concentration-dependent rise in cytoplasmic Ca2+. This continuous cell line of parotid acinar cells, which after treatment with rat serum retains the basic structural and functional properties of primary culture cells, will be utilized as a model system for studying long-term biological processes that regulate parotid cell function.
Effects of rectal distensions on nociceptive flexion reflexes in humans
Authors: Bouhassira Didier , Sabaté Jean-Marc , Coffin Benoit , Le Bars Daniel , Willer Jean-Claude , Jian Raymond
Published: December 2017

We previously showed that gastric distension inhibits the somatic nociceptive flexion RIII reflex. To explore further the viscerosomatic interactions, we tested in the present study the effects of rectal distensions on RIII reflexes. Rapid and slow-ramp rectal distensions were performed in 10 healthy volunteers with an electronic barostat. The RIII reflex was continuously recorded from the lower limb during both types of distension and from the upper limb during rapid distensions. The visceral sensations were scored on a graded questionnaire. Rapid distensions facilitated the RIII reflex recorded from the lower limb, but at the highest distension level, facilitation was followed by inhibition. Slow-ramp distension induced gradual inhibition of the RIII reflex, which correlated with both distension volume and visceral sensation. RIII reflex recorded from the upper limb was also inhibited by rapid rectal distensions. Reflex inhibitions were probably related to the activation of pain modulation systems. One plausible explanation for the facilitatory effects, observed only at the lower limb, is the convergence of rectal and reflex afferents at the same levels of the spinal cord. The differential effects of rapid and slow-ramp distensions suggest the activation of two distinct populations of mechanoreceptors by these two modes of distension.
Capacitative Ca2+entry in enteric glia induced by thapsigargin and extracellular ATP
Authors: Sarosi George A. , Barnhart Douglas C. , Turner Douglas J. , Mulholland Michael W.
Published: December 2017

Mobilization of intracellular Ca2+stores is coupled to Ca2+influx across the plasma membrane, a process termed capacitative Ca2+entry. Capacitative Ca2+entry was examined in cultured guinea pig enteric glia exposed to 100 μM ATP, an inositol trisphosphate-mediated Ca2+-mobilizing agonist, and to 1 μM thapsigargin, an inhibitor of microsomal Ca2+ATPase. Both agents caused mobilization of intracellular Ca2+stores followed by influx of extracellular Ca2+. This capacitative Ca2+influx was inhibited by Ni2+(88 ± 1%) and by La3+(87 ± 1%) but was not affected by L- or N-type Ca2+channel blockers. Pretreatment of glia with 100 nM phorbol 12-myristate 13-acetate for 24 h decreased capacitative Ca2+entry by 48 ± 2%. Chelerythrine (0.1–10 μM), a specific antagonist of protein kinase C (PKC), dose dependently inhibited capacitative Ca2+entry. The nitric oxide synthase inhibitor NG-nitro-l-arginine (1 mM) decreased Ca2+influx by 42 ± 1%. Capacitative Ca2+entry was inhibited to a similar degree by the guanylate cyclase inhibitor (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one). Capacitative Ca2+entry occurs in enteric glial cells via lanthanum-inhibitable channels through a process regulated by PKC and nitric oxide.
Enteroinvasive bacteria directly activate expression of iNOS and NO production in human colon epithelial cells
Authors: Witthöft Thomas , Eckmann Lars , Kim Jung Mogg , Kagnoff Martin F.
Published: December 2017

In these studies, we investigated whether bacterial infection of human colon epithelial cells is a sufficient stimulus to upregulate epithelial cell expression of inducible nitric oxide synthase (iNOS) and nitric oxide (NO) production. Human colon epithelial cells (Caco-2 and HT-29) rapidly upregulated iNOS mRNA and protein expression and NO production after infection with enteroinvasive Escherichia coli, Salmonella dublin, or Shigella flexneri but not after infection with noninvasive E. coli or an invasion-deficient mutant of S. dublin. Bacterial infection in the absence of added cytokines was as potent or more potent a stimulus of iNOS expression and NO production as stimulation of cells with combinations of cytokines known to strongly upregulate this epithelial cell response. Enteroinvasive E. coli increased epithelial NO production to a greater extent than S. dublin, although S. dublin was a stronger stimulus of epithelial cell interleukin-8 (IL-8) production. After enteroinvasive E. coli infection of polarized epithelial cell monolayers, nitrite, a stable NO end product, was released predominately into the apical compartment early after infection, whereas IL-8 was released in parallel into the basolateral compartment. These studies suggest NO and/or its redox products are an important component of the intestinal epithelial cell response to microbial infection.
II. Alcoholic liver injury involves activation of Kupffer cells by endotoxin
Authors: Thurman R. G.
Published: December 2017

It is well known that females show a greater susceptibility to alcohol-induced liver injury than males. Additionally, females who consume alcohol regularly and have been obese for 10 years or more are at greater risk for both hepatitis and cirrhosis. Female rats on an enteral alcohol protocol exhibit injury more quickly than males, with widespread fatty changes over a larger portion of the liver lobule. Levels of plasma endotoxin, intercellular adhesion molecule-1, free radical adducts, infiltrating neutrophils, and nuclear factor-κB are increased about twofold more in livers from female than male rats after enteral alcohol treatment. Estrogen treatment in vivo increases the sensitivity of Kupffer cells to endotoxin. Evidence has been presented that Kupffer cells are pivotal in the development of alcohol-induced liver injury. Destruction of Kupffer cells with gadolinium chloride (GdCl3) or reduction of bacterial endotoxin by sterilization of the gut with antibiotics blocks early inflammation due to alcohol. Similar results have been obtained with anti-tumor necrosis factor-α antibody. These findings led to the hypothesis that alcohol-induced liver injury involves increases in circulating endotoxin, leading to activation of Kupffer cells, which causes a hypoxia-reoxygenation injury. This idea has been tested using pimonidazole, a nitroimidazole marker, to quantitate hypoxia in downstream pericentral regions of the liver lobule. After chronic enteral alcohol, pimonidazole binding increases twofold. Enteral alcohol also increases free radicals detected with electron spin resonance. Importantly, hepatic hypoxia and radical production detected in bile are decreased by destruction of Kupffer cells with GdCl3. These data are consistent with the hypothesis that Kupffer cells participate in important gender differences in liver injury caused by alcohol.
PYY stimulates synthesis and secretion of intestinal apolipoprotein AIV without affecting mRNA expression
Authors: Kalogeris Theodore J. , Qin Xiaofa , Chey William Y. , Tso Patrick
Published: December 2017

We tested whether exogenous peptide YY (PYY) can stimulate synthesis and lymphatic secretion of intestinal apolipoprotein AIV (apo AIV). Rats with mesenteric lymph fistulas and right atrial cannulas were given continuous intravenous infusions of control vehicle or PYY at 25, 50, 75, 100, or 200 pmol ⋅ kg−1⋅ h−1. PYY (75–200 pmol ⋅ kg−1⋅ h−1) stimulated lymphatic apo AIV output from 1.5- to 3.5-fold higher than basal output. In separate experiments, PYY (100 pmol ⋅ kg−1⋅ h−1) produced a 60% increase in jejunal mucosal apo AIV synthesis but had no effect on mucosal apo AIV mRNA levels at doses up to 200 pmol ⋅ kg−1⋅ h−1. Finally, exogenous PYY infusion (100 pmol ⋅ kg−1⋅ h−1) produced a plasma PYY increment of 30 pM compared with an increment of 18.7 pM in response to ileal infusion of lipid. These results support the hypothesis that PYY may be an endocrine mediator of the effects of distal gut lipid on production and release of intestinal apo AIV, likely via a posttranscriptional mechanism of action.
Characterization of the 5′-flanking region of the murine polymeric IgA receptor gene
Authors: Martín Martín G. , Wang Jiafang , Li Tony W. H. , Lam Jason T. , Gutierrez Edgar M. , Solorzano-Vargas R. Sergio , Tsai And Hugh V.
Published: December 2017

The regulatory elements that control basal and activated transcriptional expression of the polymeric IgA receptor gene ( pIgR) have not been defined. In this study, we performed functional analysis of the murine pIgR 5′-upstream region. Transient transfection studies identified the gene’s minimal promoter to reside within 110 nucleotides upstream from the start of transcription. Substitution mutations of this region identified both a putative activator (−78 to −70) and a repressor (−66 to −52) element. DNase I footprint analysis confirmed an area of protection that spans from nucleotides −85 to −62. Mobility shift assays of the putative region confirmed binding of upstream stimulatory factor 1 (USF1) to an E box element at positions −75 and −70, representing the putative enhancer. Overexpression studies using various forms of USF suggest that both USF1 and USF2 enhance activity of the pIgR minimal promoter. We report the identification and characterization of the murine pIgR minimal promoter, as well as the critical role of USF in enhancing its basal level of transcription in Caco-2 cells.
Sorting of rat liver and ileal sodium-dependent bile acid transporters in polarized epithelial cells
Authors: Sun An-Qiang , Ananthanarayanan Meenakshisundaram , Soroka Carol J. , Thevananther Sundararajah , Shneider Benjamin L. , Suchy Frederick J.
Published: December 2017

The rat ileal apical Na+-dependent bile acid transporter (ASBT) and the liver Na+-taurocholate cotransporting polypeptide (Ntcp) are members of a new family of anion transporters. These transport proteins share limited sequence homology and almost identical predicted secondary structures but are localized to the apical surface of ileal enterocytes and the sinusoidal surface of hepatocytes, respectively. Stably transfected Madin-Darby canine kidney (MDCK) cells appropriately localized wild-type ASBT and Ntcp apically and basolaterally as assessed by functional activity and immunocytochemical localization studies. Truncated and chimeric transporters were used to determine the functional importance of the cytoplasmic tail in bile acid transport activity and membrane localization. Two cDNAs were created encoding a truncated transporter in which the 56-amino-acid COOH-terminal tail of Ntcp was removed or substituted with an eight-amino-acid epitope FLAG. For both mutants there was some loss of fidelity in basolateral sorting in that ∼75% of each protein was delivered to the basolateral surface compared with ∼90% of the wild-type Ntcp protein. In contrast, deletion of the cytoplasmic tail of ASBT led to complete loss of transport activity and sorting to the apical membrane. An Ntcp chimera in which the 56-amino-acid COOH-terminal tail of Ntcp was replaced with the 40-amino-acid cytoplasmic tail of ASBT was largely redirected (82.4 ± 3.9%) to the apical domain of stably transfected MDCK cells, based on polarity of bile acid transport activity and localization by confocal immunofluorescence microscopy. These results indicate that a predominant signal for sorting of the Ntcp protein to the basolateral domain is located in a region outside of the cytoplasmic tail. These studies have further shown that a novel apical sorting signal is localized to the cytoplasmic tail of ASBT and that it is transferable and capable of redirecting a protein normally sorted to the basolateral surface to the apical domain of MDCK cells.
Acute effect of EtOH on Mg2+homeostasis in liver cells: evidence for the activation of an Na+/Mg2+exchanger
Authors: Tessman Patrick A. , Romani Andrea
Published: December 2017

The acute administration of ethanol mobilizes a considerable amount of Mg2+from perfused rat livers and isolated hepatocytes in a dose-dependent fashion in the absence of release of cellular K+or lactate dehydrogenase (LDH) in the extracellular medium. Mg2+extrusion becomes detectable within 2 min and reaches the maximum within 8 min after ethanol addition, declining toward the basal value thereafter irrespective of the persistence of alcohol in the perfusion system and the dose of ethanol administered. The effect is the result of a specific impairment of Mg2+transport and/or regulatory mechanisms. In fact, Mg2+extrusion does not occur under conditions in which 1) ethanol is replaced by an equivalent dose of DMSO, 2) amiloride or imipramine are used as inhibitors of the Na+/Mg2+exchanger, 3) extracellular Na+is replaced by an equimolar concentration of choline chloride, and 4) 4-methylpyrazole is used to specifically inhibit alcohol dehydrogenase and cytochrome P-4502E1. Finally, the observation that the cellular level of ATP is markedly reduced after acute ethanol administration would suggest that Mg2+extrusion results from a decreased buffering capacity of cytosolic Mg-ATP complex.
Estradiol alters cholecystokinin stimulus-response coupling in rat pancreatic acini
Authors: Blevins George. T. , McCullough Sandra S. , Wilbert Tasha N. , Isom Rémelle M. , Chowdhury Parimal , Miller Stephania T.
Published: December 2017

We have previously demonstrated that altered exocrine pancreatic stimulus-secretion coupling is associated with ovariectomy and chronic estradiol administration. To elucidate possible mechanisms underlying those effects we examined the ability of chronic administration of different doses of estradiol to regulate the CCK signal transduction pathway in isolated rat pancreatic acini. Doses of estradiol ranging from 0.5 to 119 μg/day were administered to ovariectomized rats for 18 days. Ovariectomy was associated with enhanced CCK-stimulated pancreatic amylase release, whereas estradiol dose dependently decreased the magnitude of CCK-stimulated amylase release. Ovariectomy was also associated with enhanced CCK receptor numbers on acinar cell membranes. Estradiol administration was associated with dose-dependent decreases in CCK receptor numbers. Neither ovariectomy nor estradiol administration affected CCK receptor affinity. Moreover, estradiol administration was associated with increased expression of the α-subunit of the heterotrimeric G protein Gq/11(Gαq/11). Recent findings (H. Ohnishi, S. A. Ernst, D. I. Yule, C. W. Baker, and J. A. Williams. J. Biol. Chem. 272: 16056–16061, 1997) demonstrate that Gαq/11may exert a tonic inhibitory effect on pancreatic enzyme release. In view of these findings, the increased expression of Gαq/11 induced by estradiol likely contributes to the inhibition of pancreatic enzyme release. We conclude that the effect of estradiol to decrease pancreatic secretion is mediated through regulation of CCK receptor density and Gαq/11 expression.
Mechanism of inhibition of Na+-bile acid cotransport during chronic ileal inflammation in rabbits
Authors: Sundaram U. , Wisel S. , Stengelin S. , Kramer W. , Rajendran V.
Published: December 2017

In the chronically inflamed ileum, unique mechanisms of alteration of similar transport processes suggest regulation by different immune-inflammatory mediator pathways. In a rabbit model of chronic ileitis, we previously demonstrated that Na+-glucose cotransport was inhibited by a decrease in the cotransporter numbers, whereas Na+-amino acid cotransport was inhibited by a decrease in the affinity for the amino acid. In this study, we demonstrated that Na+-bile acid cotransport was reduced in villus cells from the chronically inflamed ileum. In villus cell brush-border membrane vesicles from the chronically inflamed ileum, Na+-bile acid cotransport was reduced as well, suggesting a direct effect at the cotransporter itself. Kinetic studies demonstrated that Na+-bile acid cotransport was inhibited by both a decrease in the affinity as well as a decrease in the maximal rate of uptake of the bile acid. Analysis of steady-state mRNA and immunoreactive protein levels of the Na+-bile acid cotransporter also demonstrated some reduction during chronic ileitis. Thus, in the chronically inflamed ileum, the mechanisms of inhibition of Na+-glucose, Na+-amino acid, and Na+-bile acid cotransport are different. These data suggest that different cotransporters are uniquely altered either secondary to their intrinsic differences or by different immune-inflammatory mediators during chronic ileitis.
Regional and transmural density of interstitial cells of Cajal in human colon and rectum
Authors: Hagger Robert , Gharaie Sussan , Finlayson Caroline , Kumar Devinder
Published: December 2017

The interstitial cells of Cajal (ICC) are thought to play an important role in the control of gut motility. The regional and transmural pattern of distribution of ICC in the normal human colon and rectum was evaluated with immunohistochemistry using an anti-c- kit antibody. The transmural distribution of ICC was constant throughout the whole colon, the density of ICC was significantly greater at the myenteric plexus than at either the longitudinal or circular muscle layers, and in the rectum the transmural distribution was more even. Regionally, at the myenteric plexus, the transverse colon had a significantly greater density of ICC compared with the right colon ( P = 0.038), left colon ( P = 0.006), and rectum ( P = 0.008). The pattern of distribution of ICC identified in this study is consistent with the proposed roles of ICC as colorectal pacemakers, intermediaries of the neural control of muscle activity, and coordinators of colorectal muscle activity. The highest density of ICC was at the myenteric plexus of the transverse colon, which is the proposed region of pacemaking activity.
Modulation of cholinergic neuromuscular transmission by nitric oxide in canine colonic circular smooth muscle
Authors: Rae M. G. , Khoyi M. A. , Keef K. D.
Published: December 2017

This study examines the effect of nitric oxide (NO) on cholinergic transmission in strips of canine colonic circular muscle in which neural plexus-pacemaker regions had been removed. Electrical field stimulation gave rise to atropine- and TTX-sensitive excitatory junction potentials (EJPs), the amplitude of which were frequency dependent. In 47% of control muscles, the EJP was followed by an inhibitory junction potential (IJP), whereas in the presence of atropine all preparations exhibited only IJPs. The NO synthase inhibitor N ω-nitro-l-arginine (l-NNA), the guanylyl cyclase inhibitor 1H-[1,2,4]-oxadiazolo-[4,3-a]-quinoxaline-1-one (ODQ), and the protein kinase G (PKG) antagonist Rp-8-bromo-PET-cGMPS all significantly increased EJP amplitude and reduced or abolished IJPs. The potentiation of EJPs by l-NNA was reversed by the NO donors sodium nitroprusside (SNP) and S-nitroso- N-acetylpenicillamine in a manner blocked by ODQ. [14C]ACh overflow was also measured to evaluate the possible prejunctional effects of NO. Both norepinephrine and TTX significantly decreased [14C]ACh overflow; however, l-NNA, ODQ, and SNP were without effect. These data suggest that both cholinergic and nitrergic motoneurons functionally innervate the interior of the circular muscle layer. The inhibitory actions of NO on cholinergic transmission appear to be post- rather than prejunctional and to involve guanylyl cyclase as well as possibly PKG.
Role of nitric oxide in inflammation-induced suppression of secretion in a mouse model of acute colitis
Authors: MacNaughton Wallace K. , Lowe Sonya S. , Cushing Kelly
Published: December 2017

The role of nitric oxide (NO) derived from the inducible isoform of NO synthase (iNOS) in epithelial transport dysfunction was studied in a model of colitis induced in mice by intrarectal 2,4,6-trinitrobenzenesulfonic acid in 30% ethanol. Expression of iNOS mRNA was determined by RT-PCR. Electrolyte transport studies were conducted in Ussing chambers in which segments of inflamed colon were incubated with or without the selective iNOS inhibitorl- N 6-(1-iminoethyl)lysine (l-NIL). Seven days after the induction of colitis, colonic tissue exhibited increased myeloperoxidase activity compared with saline controls. There was a detectable basal expression of iNOS mRNA, but expression was increased 3.7-fold in inflamed colons. Inflammation also caused an increase in iNOS activity and a concomitant decrease in constitutive NOS activity. In Ussing chamber experiments, there was a decreased response to electrical field stimulation in inflamed tissue, which was partially reversed by pretreatment of the tissue withl-NIL. The short-circuit current response to the muscarinic agonist carbachol was also reduced in inflammation, but this was not reversed byl-NIL. In summary, NO derived from iNOS mediates, in part, inflammation-induced suppression of neurally evoked electrolyte transport.