bluuebunny/crossref_metadata_2025 huggingface dataset:
Curcumin prevents alcohol-induced liver disease in rats by inhibiting the expression of NF-κB-dependent genes
Authors: Nanji Amin A. , Jokelainen Kalle , Tipoe George L. , Rahemtulla Amir , Thomas Peter , Dannenberg Andrew J.
Published: March 2015

Induction of NF-κB-mediated gene expression has been implicated in the pathogenesis of alcoholic liver disease (ALD). Curcumin, a phenolic antioxidant, inhibits the activation of NF-κB. We determined whether treatment with curcumin would prevent experimental ALD and elucidated the underlying mechanism. Four groups of rats (6 rats/group) were treated by intragastric infusion for 4 wk. One group received fish oil plus ethanol (FE); a second group received fish oil plus dextrose (FD). The third and fourth groups received FE or FD supplemented with 75 mg · kg−1 · day−1of curcumin. Liver samples were analyzed for histopathology, lipid peroxidation, NF-κB binding, TNF-α, IL-12, monocyte chemotactic protein-1, macrophage inflammatory protein-2, cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and nitrotyrosine. Rats fed FE developed fatty liver, necrosis, and inflammation, which was accompanied by activation of NF-κB and the induction of cytokines, chemokines, COX-2, iNOS, and nitrotyrosine formation. Treatment with curcumin prevented both the pathological and biochemical changes induced by alcohol. Because endotoxin and the Kupffer cell are implicated in the pathogenesis of ALD, we investigated whether curcumin suppressed the stimulatory effects of endotoxin in isolated Kupffer cells. Curcumin blocked endotoxin-mediated activation of NF-κB and suppressed the expression of cytokines, chemokines, COX-2, and iNOS in Kupffer cells. Thus curcumin prevents experimental ALD, in part by suppressing induction of NF-κB-dependent genes.
Glucagon-like peptide-2-enhanced barrier function reduces pathophysiology in a model of food allergy
Authors: Cameron Heather L. , Yang Ping-Chang , Perdue Mary H.
Published: March 2015

Penetration of the gut epithelial barrier by intact luminal antigen is necessary for immunologically mediated pathophysiology in the context of food allergy. We investigated if glucagon-like peptide-2 (GLP-2) could affect immediate hypersensitivity and late-phase allergic inflammation in a murine model. Mice were sensitized to horseradish peroxidase (HRP); studies were conducted 14 days later. Mice were treated with 5 μg GLP-2 subcutaneously 4 h before antigen challenge. For immediate hypersensitivity, jejunal segments in Ussing chambers were challenged by luminal HRP antigen. GLP-2 treatment reduced the uptake of HRP and the antigen-induced secretory response after luminal challenge. GLP-2 appears to reduce macromolecular uptake independent of the CD23-mediated enhanced antigen uptake pathway. For the late phase, mice were gavaged with antigen, and 48 h later the function and histology of the jejunum were examined. GLP-2 prevented the usual prolonged permeability defect and reduced the number of inflammatory cells in the mucosa. Our studies demonstrate that a single treatment of sensitized mice with GLP diminishes both immediate and late-phase hypersensitivity reactions characteristic of food allergy by inhibiting transepithelial uptake of antigen.
Gut-associated lymphoid T cell suppression enhances bacterial translocation in alcohol and burn injury
Authors: Choudhry Mashkoor A. , Fazal Nadeem , Goto Masakatsu , Gamelli Richard L. , Sayeed Mohammed M.
Published: March 2015

The mechanism of alcohol-mediated increased infection in burn patients remains unknown. With the use of a rat model of acute alcohol and burn injury, the present study ascertained whether acute alcohol exposure before thermal injury enhances gut bacterial translocation. On day 2postinjury, we found a severalfold increase in gut bacterial translocation in rats receiving both alcohol and burn injury compared with the animals receiving either injury alone. Whereas there were no demonstrable changes in intestinal morphology in any group of animals, a significant increase in intestinal permeability was observed in ethanol- and burn-injured rats compared with the rats receiving either injury alone. We further examined the role of intestinal immune defense by determining the gut-associated lymphoid (Peyer's patches and mesenteric lymph nodes) T cell effector responses 2 days after alcohol and burn injury. Although there was a decrease in the proliferation and interferon-γ by gut lymphoid T cells after burn injury alone; the suppression was maximum in the group of rats receiving both alcohol and burn injuries. Furthermore, the depletion of CD3+cells in healthy rats resulted in bacterial accumulation in mesenteric lymph nodes; such CD3+cell depletion in alcohol- and burn-injured rats furthered the spread of bacteria to spleen and circulation. In conclusion, our data suggest that the increased intestinal permeability and a suppression of intestinal immune defense in rats receiving alcohol and burn injury may cause an increase in bacterial translocation and their spread to extraintestinal sites.
Neuroimmune interactions in guinea pig stomach and small intestine
Authors: Liu Sumei , Hu Hong-Zhen , Gao Na , Gao Chuanyun , Wang Guodu , Wang Xiyu , Peck Owen C. , Kim Gordon , Gao Xiang , Xia Yun , Wood Jackie D.
Published: March 2015

Enteric neuroimmune interactions in gastrointestinal hypersensitivity responses involve antigen detection by mast cells, mast cell degranulation, release of chemical mediators, and modulatory actions of the mediators on the enteric nervous system (ENS). Electrophysiological methods were used to investigate electrical and synaptic behavior of neurons in the stomach and small intestine during exposure to β-lactoglobulin in guinea pigs sensitized to cow's milk. Application of β-lactoglobulin to sensitized preparations depolarized the membrane potential and increased neuronal excitability in small intestinal neurons but not in gastric neurons. Effects on membrane potential and excitability in the small intestine were suppressed by the mast cell stabilizing drug ketotifen, the histamine H2 receptor antagonist cimetidine, the cyclooxygenase inhibitor piroxicam, and the 5-lipoxygenase inhibitor caffeic acid. Unlike small intestinal ganglion cells, gastric myenteric neurons did not respond to histamine applied exogenously. Antigenic exposure suppressed noradrenergic inhibitory neurotransmission in the small intestinal submucosal plexus. The histamine H3receptor antagonist thioperamide and piroxicam, but not caffeic acid, prevented the allergic suppression of noradrenergic inhibitory neurotransmission. Antigenic stimulation of neuronal excitability and suppression of synaptic transmission occurred only in milk-sensitized animals. Results suggest that signaling between mast cells and the ENS underlies intestinal, but not gastric, anaphylactic responses associated with food allergies. Histamine, prostaglandins, and leukotrienes are paracrine signals in the communication pathway from mast cells to the small intestinal ENS.
V. Confluence of membrane trafficking and motility in epithelial cell models
Authors: Casanova James E.
Published: March 2015

Migration of epithelial cells occurs in a variety of important biological processes including tissue morphogenesis, wound healing, and the metastasis of epithelial tumors. In some instances, the cells remain attached to each other and migrate together as a sheet, maintaining epithelial integrity. In others (e.g., metastasis), junctional complexes are disrupted and cells migrate individually. In both cases, motility involves the extension of membranous protrusions (filopodia and lamellipodia) in the direction of movement and the transient assembly and disassembly of integrin-mediated adhesions with the extracellular matrix. The driving force for these events is provided by regulated changes in the organization of the actin cytoskeleton, which are thought to be coordinated with alterations in intracellular membrane traffic. In this themes article, I review current hypotheses about how these processes are integrated and attempt to identify fruitful areas for future research.
Prepro-endothelin-1 mRNA and its mature peptide in human appendix
Authors: Massai Lauretta , Carbotti Paola , Cambiaggi Caterina , Mencarelli Marzia , Migliaccio Pierluigi , Muscettola Michela , Grasso Giovanni
Published: March 2015

Because the precise immunopathological events occurring in appendicitis are not completely understood, possible local production of endothelin-1 (ET-1) in human appendix was investigated. We used immunohistochemistry and in situ hybridization to detect the presence, distribution, and phenotype of ET-1-positive cells and prepro-ET-1 (pp-ET-1) mRNA-expressing cells. ET-1-positive stromal cells and pp-ET-1 mRNA-expressing cells were detected with different distributions and relative frequencies in normal control appendix, histologically normal appendix, and inflamed appendix. Six of 20 histologically normal appendixes from patients with a clinical diagnosis of acute appendicitis had many ET-1-positive stromal cells and high pp-ET-1 mRNA expression, similar to inflamed appendix. Forty percent of the pp-ET-1 mRNA-expressing cells were neutrophils, and the other positive cells were mast cells and macrophages. We suggest that local production of ET-1 by neutrophils and other inflammatory cells could be a molecular sign of focal inflammation in histologically normal appendixes and that ET-1 could be implicated, with other cytokines, in the pathogenesis of appendicitis by inducing appendiceal ischemia through vasoconstriction.
Transport of ketone bodies and lactate in the sheep ruminal epithelium by monocarboxylate transporter 1
Authors: Müller Frank , Huber Korinna , Pfannkuche Helga , Aschenbach Jörg R. , Breves Gerhard , Gäbel Gotthold
Published: March 2015

Due to intensive intracellular metabolism of short-chain fatty acids, ruminal epithelial cells generate large amounts of d-β-hydroxybutyric acid, acetoacetic acid, and lactic acid. These acids have to be extruded from the cytosol to avoid disturbances of intracellular pH (pHi). To evaluate acid extrusion, pHi was studied in cultured ruminal epithelial cells of sheep using the pH-sensitive fluorescent dye 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Extracellular addition of d-β-hydroxybutyrate, acetoacetate, or lactate (20 mM) resulted in intracellular acidification. Vice versa, removing extracellulard-β-hydroxybutyrate, acetoacetate, or lactate after preincubation with the respective monocarboxylate induced an increase of pHi. Initial rate of pHi decrease as well as of pHi recovery was strongly inhibited by pCMBS (400 μM) and phloretin (20 μM). Both cultured cells and intact ruminal epithelium were tested for the possible presence of proton-linked monocarboxylate transporter (MCT) on both the mRNA and protein levels. With the use of RT-PCR, mRNA encoding for MCT1 isoform was demonstrated in cultured ruminal epithelial cells and the ruminal epithelium. Immunostaining with MCT1 antibodies intensively labeled cultured ruminal epithelial cells and cells located in the stratum basale of the ruminal epithelium. In conclusion, our data indicate that MCT1 is expressed in the stratum basale of the ruminal epithelium and may function as a main mechanism for removing ketone bodies and lactate together with H+ from the cytosol into the blood.
Electroacupuncture accelerates gastric emptying in association with changes in vagal activity
Authors: Ouyang Hui , Yin Jieyun , Wang Zhishun , Pasricha Pankaj J , Chen J. D. Z.
Published: March 2015

Gastroparesis is a disorder with a lack of treatment options and this study investigated the effect of electroacupuncture on gastric emptying and involved mechanisms. Dogs implanted with a duodenal cannula and serosal electrodes were fed with Ensure mixed with phenol red, and the gastric effluent was collected. Electroacupuncture was performed from 30 min before until 45 min after the meal. Gastric myoelectrical activity and electrocardiogram were recorded. Gastric emptying was significantly improved with electroacupuncture. Vagal activity assessed from the spectral analysis of heart rate variability was markedly increased with electroacupuncture. Electroacupuncture increased the regularity of gastric slow waves in both the proximal and distal stomach. It also increased the number of spike bursts in the distal but not proximal stomach. Electroacupuncture accelerates gastric emptying of liquid in dogs and its potential for treating gastroparesis may be explored. The effect may be attributed to improvement in gastric slow-wave rhythmicity and antral contractile (spike) activity and may possibly involve the vagal pathway.
Distribution of interstitial cells of Cajal in tunica muscularis of the canine rectoanal region
Authors: Horiguchi Kazuhide , Keef Kathleen D. , Ward Sean M.
Published: March 2015

Electrical and mechanical activity of the circular muscle layer in the rectoanal region of the gastrointestinal tract undergoes considerable changes in the site of dominant pacemaking activity, frequency, and waveform shape. The present study was performed to determine whether changes in the structural organization of the circular layer or in the density, distribution, and ultrastructure of interstitial cells of Cajal (ICC) could account for this heterogeneity in electrical and mechanical activities. Light microscopy revealed that the structural organization of the circular muscle layer underwent dramatic morphological changes, from a tightly packed layer with poorly defined septa in the proximal rectum to one of discrete muscle bundles separated by large septae in the internal anal sphincter. Kit immunohistochemistry revealed a dense network of ICC along the submucosal and myenteric borders in the rectum, whereas in the internal anal sphincter, ICC were located along the periphery of muscle bundles within the circular layer. Changes in electrical activity within the circular muscle layer can be partially explained by changes in the structure of the muscle layer and changes in the distribution of ICC in the rectoanal region of the gastrointestinal tract.
Spatial localization and properties of pacemaker potentials in the canine rectoanal region
Authors: Mutafova-Yambolieva V. N. , O'Driscoll K. , Farrelly A. , Ward S. M. , Keef K. D.
Published: March 2015

The present study investigated the spatial organization of electrical activity in the canine rectoanal region and its relationship to motility patterns. Contraction and resting membrane potential ( E m) were measured from strips of circular muscle isolated 0.5–8 cm from the anal verge. Rapid frequency [25 cycles/min (cpm)] E moscillations (MPOs, 12 mV amplitude) were present across the thickness of the internal anal sphincter (IAS; 0.5 cm) and E m was constant (−52 mV). Between the IAS and the proximal rectum an 18 mV gradient in E mdeveloped across the muscle thickness with the submucosal edge at −70 mV and MPOs were replaced with slow waves (20 mV amplitude, 6 cpm). Slow waves were of greatest amplitude at the submucosal edge. Nifedipine (1 μM) abolished MPOs but not slow waves. Contractile frequency changes were commensurate with the changes in pacemaker frequency. Our results suggest that changing motility patterns in the rectoanal region are associated with differences in the characteristics of pacemaker potentials as well as differences in the sites from which these potentials emanate.
Patterns of esophageal inhibition during swallowing, pharyngeal stimulation, and transient LES relaxation
Authors: Pouderoux Philippe , Verdier Eric , Kahrilas Peter J.
Published: March 2015

Lower esophageal sphincter (LES) relaxation and esophageal body inhibition co-occur during esophageal peristalsis but not necessarily during pharyngeal stimulation or transient LES relaxation (tLESR). This study examined these relationships and the impact on reflux. Nine young volunteers were studied. An artificial high-pressure zone (HPZ) was established, and pH was recorded 8 and 5 cm proximal to the LES. Pharyngeal stimulation was by water injection and gastric distension with liquid or gas. Peristalsis, pharyngeal stimulation, and spontaneous events were recorded. Swallowing relaxed the LES in 100% of trials (the HPZ in 80%) and caused no reflux. Pharyngeal stimulation relaxed the LES in two-thirds of trials, had no effect on the HPZ, and caused no reflux. Gastric distension was associated with 117 tLESRs, 48% with acid reflux, and 32% with gas reflux; there was no effect on the HPZ. We conclude that LES relaxation is a necessary but not sufficient condition for reflux. LES relaxation and esophageal body inhibition are independent events that may be concurrent (swallowing) or dissociated (tLESR).
85-kDa cPLA2plays a critical role in PPAR-mediated gene transcription in human hepatoma cells
Authors: Han Chang , Demetris A. Jake , Michalopoulos George , Shelhamer James H. , Wu Tong
Published: March 2015

In an effort to understand the role of key eicosanoid-forming enzymes in the activation of peroxisome proliferator-activated receptor (PPAR), this study was designed to evaluate the possible contributions of cytosolic phospholipase A2(cPLA2) and group IIA secretory phospholipase A2(sPLA2) in the regulation of PPAR-mediated gene transcription in a human hepatoma cell line (HepG2). The HepG2 cells express both PPAR-α and -γ but not PPAR-β. Overexpression of cPLA2, but not group IIA sPLA2in the HepG2 cells, caused a significantly increased PPAR-α/γ-mediated reporter activity. Antisense inhibition of cPLA2resulted in a significantly decreased PPAR-α/γ activity. The PPAR-α/γ-induced gene transcription in the HepG2 cells was inhibited by the cPLA2inhibitors methyl arachidonyl fluorophosphonate and arachidonyltrifluoromethyl ketone, but not by the sPLA2inhibitor LY311727. The expression of PPAR-α-mediated endogenous gene apolipoprotein A-II was increased in cells with overexpression of cPLA2, decreased in cells with antisense inhibition of cPLA2, but unaltered in cells with overexpression of group IIA sPLA2. The above results demonstrated an important role of cPLA2, but not group IIA sPLA2in the control of PPAR activation. The cPLA2-mediated PPAR activation was likely mediated by arachidonic acid and prostaglandin E2. This study reveals a novel intracellular function of cPLA2in PPAR activation in HepG2 cells. The cPLA2thus may represent a potential therapeutic target for the control of PPAR-related liver and metabolic disorders such as obesity, lipid metabolic disorders, diabetes mellitus, and atherosclerosis.
Increased expression of ileal apical sodium-dependent bile acid transporter in postpartum rats
Authors: Mottino Aldo D. , Hoffman Tim , Dawson Paul A. , Luquita Marcelo G. , Monti Juan A. , Sánchez Pozzi Enrique J. , Catania Viviana A. , Cao Jingsong , Vore Mary
Published: March 2015

The expression and activity of the apical ileal sodium-dependent bile acid transporter (asbt) was examined in the small intestine of control, pregnant, and lactating postpartum rats 2, 12, and 21 days after delivery. Western blot analysis of brush border membrane vesicles (BBMV) prepared from different regions of the small intestine demonstrated that expression of asbt was maximal in the most distal segments for all experimental groups, was not substantially affected in pregnant and 2-day postpartum rats, and was significantly increased in 12- and 21-day postpartum rats. Analysis of mRNA suggested that asbt protein was regulated at the posttranscriptional level in postpartum rats. Increased expression of asbt protein postpartum was maximal (∼2-fold) in the proximal region of the ileum, consistent with a 60% increase in taurocholate (TC) transport in BBMV from the proximal ileum in 14- to 21-day postpartum rats relative to control rats. Absorption of TC, determined from the intact proximal ileum using an intestinal loop model, demonstrated a 30% increase in TC uptake per unit weight of tissue in 14- to 21-day postpartum rats relative to control rats. Together with the marked increase in intestinal mass observed at peak lactation, these data indicate a significant increase in asbt-mediated reclamation of bile acids in the intestine of lactating rats.
Regulation and role of p21 and p27 cyclin-dependent kinase inhibitors during hepatocyte differentiation and growth
Authors: Ilyin Gennady P. , Glaise Denise , Gilot David , Baffet Georges , Guguen-Guillouzo Christiane
Published: March 2015

Unlike a large number of cell types that undergo terminal differentiation associated with permanent withdrawal from the cell cycle, mature quiescent hepatocytes retain high proliferative potential. We report here a specific behavior of members of the Cip/Kip family of cyclin-dependent kinase (Cdk) inhibitors during development of the rat liver and proliferation of normal hepatocytes. Expression of p21, p27, and p57 transcripts and proteins was downregulated during the differentiation process to low or undetectable levels in adult liver. In contrast to p27, p21 protein increased in a mitogen-dependent manner in isolated hepatocytes and its expression pattern correlated with that of cyclin D1. In proliferating hepatocytes, p21 was predominantly associated with cyclin D1, these proteins were colocalized in the nucleus and p21-associated retinoblastoma protein (pRb) kinase activity increased in parallel with that of cyclin D1. Overexpression of p21 in mitogen-stimulated hepatocytes reduced DNA synthesis. In contrast, inhibition of p21 expression by antisense or small interfering RNAs oligonucleotides accelerated S phase entry. Finally, expression of p21 and cyclin D1, but not p27 proteins was regulated by MAPK kinase/extracellular signal-regulated kinase and phosphatidylinositol 3-kinase-ferric-reducing ability power/mammalian target of rapamycin signal transduction pathways. In conclusion, these results demonstrate a specific and differential regulation of p21 and p27 during hepatocyte differentiation and proliferation that may contribute to the control of quiescent differentiated hepatic cell proliferating activity.
Differential mechanism and site of action of CCK on the pancreatic secretion and growth in rats
Authors: Yamamoto Mitsuyoshi , Otani Munenori , Jia Dong-Mei , Fukumitsu Ken-Ichiro , Yoshikawa Hiroyuki , Akiyama Toshiharu , Otsuki Makoto
Published: March 2015

Recent studies demonstrated that cholecystokinin (CCK) at physiological levels stimulates pancreatic enzyme secretion via a capsaicin-sensitive afferent vagal pathway. This study examined whether chemical ablation of afferent vagal fibers influences pancreatic growth and secretion in rats. Bilateral subdiaphragmatic vagal trunks were exposed, and capsaicin solution was applied. Pancreatic wet weight and pancreatic secretion and growth in response to endogenous and exogenous CCK were examined 7 days after capsaicin treatment. Perivagal application of capsaicin increased plasma CCK levels and significantly increased pancreatic wet weight compared with those in the control rats. Oral administration of CCK-1 receptor antagonist loxiglumide prevented the increase in pancreatic wet weight after capsaicin treatment. In addition, continuous intraduodenal infusion of trypsin prevented the increase in plasma CCK levels and pancreatic wet weight after capsaicin treatment. There were no significant differences in the expression levels of CCK-1 receptor mRNA and protein in the pancreas in capsaicin-treated and control rats. Intraduodenal administration of camostat or intravenous infusion of CCK-8 stimulated pancreatic secretion in control rats but not in capsaicin-treated rats. In contrast, repeated oral administrations of camostat or intraperitoneal injections of CCK-8 significantly increased pancreatic wet weight in both capsaicin-treated and control rats. Present results suggest that perivagal application of capsaicin stimulates pancreatic growth via an increase in endogenous CCK and that exogenous and endogenous CCK stimulate pancreatic growth not via vagal afferent fibers but directly in rats.
Characterization of CCKAreceptor affinity states and Ca2+signal transduction in vagal nodose ganglia
Authors: Lankisch Tim O. , Tsunoda Yasuhiro , Lu Yuanxu , Owyang Chung
Published: March 2015

CCKAreceptors are present on vagal afferent fibers. The objectives of this study were to identify the presence of high- and low-affinity CCKAreceptors on nodose ganglia and to characterize the intracellular calcium signal transduction activated by CCK. Stimulation of acutely isolated nodose ganglion cells from rats with 1 nM CCK-8 primarily evoked a Ca2+transient followed by a sustained Ca2+plateau (45% of cells responded), whereas 10 pM CCK-8 evoked Ca2+oscillations (37% of cells responded). CCK-OPE, a high-affinity agonist and low-affinity antagonist of CCKAreceptors, primarily elicited Ca2+oscillations (29% of cells responded). CCK-OPE inhibited the Ca2+transient induced by 1 nM CCK-8 but not by carbachol and high K+. This result suggests the presence of high- and low-affinity states of CCKAreceptors on nodose ganglia. We further demonstrated that nicardipine (10 μM) but not ω-conotoxins GVIA and MVIIC (10–100 nM) abolished Ca2+signaling induced by CCK-8, indicating that an L-type voltage-dependent Ca2+channel and not an N- or Q-type Ca2+channel is coupled to CCKAreceptors. In a separate study, we showed that the G protein activator NaF (10 mM) elicited a Ca2+transient and inhibited CCK-8-evoked Ca2+signaling, indicative of G protein(s) involvement in CCKAreceptor activity. The Gqprotein antagonist Gp antagonist-2A (10 μM) also abolished the action of CCK-8. This study indicates that CCKAreceptors exist in both high- and low-affinity states in the nodose ganglia. Activation of high-affinity CCKAreceptors elicits Ca2+oscillations, whereas stimulation of low-affinity CCKAreceptors evokes a sustained Ca2+plateau. These Ca2+-signaling modes are mediated through the L-type Ca2+channel and involve the participation of Gqprotein.
Trypsin activity is not involved in premature, intrapancreatic trypsinogen activation
Authors: Halangk Walter , Krüger Burkhard , Ruthenbürger Manuel , Stürzebecher Jörg , Albrecht Elke , Lippert Hans , Lerch Markus M.
Published: March 2015

A premature and intracellular activation of digestive zymogens is thought to be responsible for the onset of pancreatitis. Because trypsin has a critical role in initiating the activation cascade of digestive enzymes in the gut, it has been assumed that trypsin also initiates intracellular zymogen activation in the pancreas. We have tested this hypothesis in isolated acini and lobules from rat pancreas. Intracellular trypsinogen activation was induced by supramaximal secretagogue stimulation and measured using either specific trypsin substrates or immunoreactivity of the trypsinogen activation peptide (TAP). To prevent a trypsin-induced trypsinogen activation, we used the cell-permeant, highly specific, and reversible inhibitor Nα-(2-naphthylsulfonyl)-3-amidinophenylalanine-carboxymethylpiperazide (S124), and to prevent cathepsin-induced trypsinogen activation, we used the cysteine protease inhibitor E-64d. Incubation of acini or lobules in the presence of S124 completely prevented the generation of trypsin activity in response to supramaximal caerulein but had no effect whatsoever on the generation of TAP. Conversely, when trypsin activity was recovered at the end of the experiment by either washout of S124 from acini or extensive dilution of lobule homogenates, it was up to 400% higher than after caerulein alone and corresponded, in molar terms, to the generation of TAP. Both trypsin activity and TAP release were inhibited in parallel by E-64d. We conclude that caerulein-induced trypsinogen activation in the pancreas is caused by an E-64d-inhibitable mechanism such as cathepsin-induced trypsinogen activation, and neither involves nor requires intracellular trypsin activity. Specific trypsin inhibition, on the other hand, prevents 80% of trypsin inactivation or autodegradation in the pancreas.
ERG K+ channels modulate the electrical and contractile activities of gallbladder smooth muscle
Authors: Parr Edward , Pozo Maria J. , Horowitz Burton , Nelson Mark T. , Mawe Gary M.
Published: March 2015

The current study was undertaken to test the existence and possible role of ether-a-go-go-related gene 1 (ERG1) protein K+ channels in gallbladder smooth muscle (GBSM). Transcripts encoding ERG1 were detected in human, mouse, and guinea pig GBSM, and ERG1 immunoreactivity was observed in GBSM cells. In intracellular voltage recordings, addition of E-4031 (100 nM–1 μM) or cisapride (100 nM–2 μM) caused concentration-dependent excitation of guinea pig GBSM that was not affected by 500 nM TTX + 5 μM atropine, and E-4031 also depolarized the resting membrane potential. In muscle strip studies, E-4031 either induced phasic contractions or significantly increased the amplitude of phasic contractions in spontaneously active tissues ( P = 0.001). E-4031 also potentiated bethanechol-induced contractions. In conclusion, ERG1 channels are expressed in the GBSM, where they play a role in excitation-contraction coupling probably by contributing to repolarization of the plateau phase of the action potential and to the resting membrane potential.
Fatty acid binding protein is a major determinant of hepatic pharmacokinetics of palmitate and its metabolites
Authors: Hung Daniel Y. , Burczynski Frank J. , Chang Ping , Lewis Andrew , Masci Paul P. , Siebert Gerhard A. , Anissimov Yuri G. , Roberts Michael S.
Published: March 2015

Disposition kinetics of [3H]palmitate and its low-molecular-weight metabolites in perfused rat livers were studied using the multiple-indicator dilution technique, a selective assay for [3H]palmitate and its low-molecular-weight metabolites, and several physiologically based pharmacokinetic models. The level of liver fatty acid binding protein (L-FABP), other intrahepatic binding proteins (microsomal protein, albumin, and glutathione S-transferase) and the outflow profiles of [3H]palmitate and metabolites were measured in four experimental groups of rats: 1) males; 2) clofibrate-treated males; 3) females; and 4) pregnant females. A slow-diffusion/bound model was found to better describe the hepatic disposition of unchanged [3H]palmitate than other pharmacokinetic models. The L-FABP levels followed the order: pregnant female > clofibrate-treated male > female > male. Levels of other intrahepatic proteins did not differ significantly. The hepatic extraction ratio and mean transit time for unchanged palmitate, as well as the production of low-molecular-weight metabolites of palmitate and their retention in the liver, increased with increasing L-FABP levels. Palmitate metabolic clearance, permeability-surface area product, retention of palmitate by the liver, and cytoplasmic diffusion constant for unchanged [3H]palmitate also increased with increasing L-FABP levels. It is concluded that the variability in hepatic pharmacokinetics of unchanged [3H]palmitate and its low-molecular-weight metabolites in perfused rat livers is related to levels of L-FABP and not those of other intrahepatic proteins.
Characterization of vesicular glutamate transporter in pancreatic α- and β-cells and its regulation by glucose
Authors: Bai Liqun , Zhang Xiaohong , Ghishan Fayez K.
Published: March 2015

Glutamate has been suggested to play an important role in the release of insulin and glucagon from pancreatic cells via exocytosis. Vesicular glutamate transporter is a rate-limiting step for glutamate release and is involved in the glutamate-evoked exocytosis. Two vesicular glutamate transporters (VGLUT1 and -2) have recently been cloned from the brain. In this report, we first functionally characterized vesicular glutamate transporter in cultured pancreatic α- and β-cells, and then detected mRNA expression of VGLUT1 and -2 in these cells. We also investigated the effect of high or low level of glucose on vesicular glutamate transport in cultured pancreas cells. Our results suggest that both α- and β-cells contain functional vesicular glutamate transporter. The transport characteristics are similar to the cloned neuronal VGLUT1 and -2 in regard to ATP dependence, substrate specificity, kinetics, and chloride dependence. VGLUT2 mRNA is expressed in both α- and β-cells, whereas VGLUT1 is only expressed in β-cells. High (12.8 mM) and low (2.8 mM) concentrations of glucose increased vesicular glutamate transport in β- and α-cells, respectively. VGLUT2 mRNA was significantly increased in β- and α-cells by high and low glucose concentration, respectively. This increase in VGLUT2 mRNA was suppressed by actinomycin D. We conclude that both α- and β-cells possess functional vesicular glutamate transporters regulated by alteration in glucose concentration, partly via the transcriptional mechanism.
Patterns of excitability in human esophageal sensorimotor cortex to painful and nonpainful visceral stimulation
Authors: Hamdy Shaheen , Rothwell John C. , Fraser Chris , Power Maxine , Gow David , Thompson David G.
Published: March 2015

To better understand the relationship between cortical plasticity and visceral pain, we developed a pain-induced model of altered esophageal corticobulbar excitability. In eight healthy volunteers, corticoesophageal electromyographic responses were recorded via an intraluminal catheter, following magnetic stimulation of the right sensorimotor cortex using perithreshold intensities. Corticothenar responses were used as control. Responses were assessed both before and for up to 1 h after either painful or nonpainful balloon distension of the esophagus (frequency = 1 Hz, dwell time = 200 ms, duration = 10 min), each being delivered to each subject in random order. Painful esophageal distension (mean volume = 11 ± 3 ml) induced a profound increase in esophageal responses compared with baseline levels (at 30 min: 141 ± 12 vs. 101 ± 9 μV, P < 0.01), whereas nonpainful esophageal distension (mean volume = 4 ± 2 ml) showed a decrease (at 30 min: 72 ± 8 vs. 88 ± 12 μV, P < 0.03). Thenar responses were unaffected. The results show that painful and nonpainful stimuli induce different patterns of esophageal corticobulbar excitability, suggesting a physiological link between cortical plasticity and visceral pain.
Molecular mechanisms of hepatic ischemia-reperfusion injury and preconditioning
Authors: Jaeschke Hartmut
Published: March 2015

Ischemia-reperfusion injury is, at least in part, responsible for the morbidity associated with liver surgery under total vascular exclusion or after liver transplantation. The pathophysiology of hepatic ischemia-reperfusion includes a number of mechanisms that contribute to various degrees in the overall injury. Some of the topics discussed in this review include cellular mechanisms of injury, formation of pro- and anti-inflammatory mediators, expression of adhesion molecules, and the role of oxidant stress during the inflammatory response. Furthermore, the roles of nitric oxide in preventing microcirculatory disturbances and as a substrate for peroxynitrite formation are reviewed. In addition, emerging mechanisms of protection by ischemic preconditioning are discussed. On the basis of current knowledge, preconditioning or pharmacological interventions that mimic these effects have the greatest potential to improve clinical outcome in liver surgery involving ischemic stress and reperfusion.
Mesenteric afferent nerves are sensitive to vascular perfusion in a novel preparation of rat ileum in vitro
Authors: Brunsden A. M. , Jacob S. , Bardhan K. D. , Grundy D.
Published: March 2015

Using novel in vitro preparations of vascularly perfused rat ileum, we investigated mesenteric afferent sensitivity to vascular perfusion. Gut (GPP) and vascular (VPP) perfusion pressures were recorded simultaneously with afferent discharge (AD). After preconstriction (l-phenylephrine), capsaicin (100 μM, gut lumen) caused a transient increase in AD and a sustained fall in VPP, supporting afferent modulation of vascular tone. In turn, AD was affected by vascular perfusion rate (VPR). Increasing VPR step-wise (0.6 to 1.0, 1.4 and 1.8 ml/min) caused concomitant falls in AD, returning at 0.6 ml/min. Terminating flow (5 min) increased AD. Afferent responses were independent of changes in GPP, vascular O2, or the gut “tube” (“gut-off”). In gut-off studies, where capsaicin (100 nM ia) still reduced VPP, flow-associated falls in AD were abolished by the enzyme neuraminidase (0.2 U/ml ia or extravascularly over 20 min). In contrast, increased AD after stopped flow was unaffected. We propose that mesenteric afferents “sense” changes in vascular perfusion. The precise stimuli (pressure and/or flow) and the physiological relevance to control of local circulation remain to be determined.
Effect of iron status on DMT1 expression in duodenal enterocytes from β2-microglobulin knockout mice
Authors: Moos Torben , Trinder Debbie , Morgan Evan H.
Published: March 2015

Divalent metal transporter I (DMT1) is thought to be involved in transport of iron across the apical cell membrane of villus duodenal cells. To determine its role in hereditary hemochromatosis (HH), we used β2-microglobulin knockout ( B2M−/−) mice that accumulate iron as in HH. The B2M−/− and control C57BL/6 ( B2M+/+) mice were fed diets with different iron contents. Increasing the iron availability increased plasma iron levels in both B2M+/+ and B2M−/−mice. Reducing the iron availability decreased the plasma iron concentration in B2M+/+ mice but was without effect on plasma iron in B2M−/− mice. DMT1 was not detectable in mice fed normal or iron-loaded diets when using immunohistochemistry. In Western blots, however, the protein was consistently observed regardless of the dietary regimen. DMT1 expression was increased to the same extent in B2M+/+ and B2M−/− mice when fed an iron-poor diet. In both strains of mice fed an iron-poor diet, DMT1 was evenly distributed in the differentiated enterocytes from the base to the tip of the villi but was absent from the crypts of Lieberkühn. These data suggest that the observed effects were due to the state of iron deficiency in mucosal cells rather than genetic defect.
Intestinal preconditioning prevents systemic inflammatory response in hemorrhagic shock. Role of HO-1
Authors: Tamion Fabienne , Richard Vincent , Lacoume Yann , Thuillez Christian
Published: March 2015

Intestinal ischemia-reperfusion has been implicated in the systemic inflammatory response and organ injury in hemorrhagic shock, but the exact role of the intestine has never been directly demonstrated. Preconditioning (PC) with brief periods of intermittent ischemia is a known potent anti-ischemic intervention and thus can be used as a tool to assess the role of local intestinal ischemia-reperfusion injury in systemic inflammatory response. Thus rats were first subjected to sham surgery or intestinal preconditioning with four cycles of 1-min ischemia and 10 min of reperfusion 24 h before hemorrhagic shock followed by resuscitation. PC reduced fluid requirements, lung edema, and lactate and tumor necrosis factor-α production. These effects were abolished by the heme-oxygenase-1 (HO-1) inhibitor tin protoporphyrin (Sn-PP). PC induced more than fivefold in intestinal HO-1 expression. These results suggest that intestinal ischemia-reperfusion is a major trigger for inflammatory response and organ injury in nonseptic shock. HO-1 appears to play an important role in the protective effect of intestinal preconditioning.
Progastrin1–80stimulates growth of intestinal epithelial cells in vitro via high-affinity binding sites
Authors: Singh P. , Lu X. , Cobb S. , Miller B. T. , Tarasova N. , Varro A. , Owlia A.
Published: March 2015

Proliferation and carcinogenesis of the large intestinal epithelial cells (IEC) cells is significantly increased in transgenic mice that overexpress the precursor progastrin (PG) peptide. It is not known if the in vivo growth effects of PG on IEC cells are mediated directly or indirectly. Full-length recombinant human PG (rhPG1–80) was generated to examine possible direct effects of PG on IEC cells. Surprisingly, rhPG (0.1–1.0 nM) was more effective than the completely processed gastrin 17 (G17) peptide as a growth factor. Even though IEC cells did not express CCK1and CCK2receptors (-R), fluorescently labeled G17 and Gly-extended G17 (G-Gly) were specifically bound to the cells, suggesting the presence of binding proteins other than CCK1-R and CCK2-R on IEC cells. High-affinity ( Kd= 0.5–1.0 nM) binding sites for125I-rhPG were discovered on IEC cells that demonstrated relative binding affinity for gastrin-like peptides in the order PG ≥ COOH-terminally extended G17 ≥ G-Gly > G17 > *CCK-8 (* significant difference; P< 0.05). In conclusion, our studies demonstrate for the first time direct growth effects of the full-length precursor peptide on IEC cells in vitro that are apparently mediated by the high-affinity PG binding sites that were discovered on these cells.
Role of nerve growth factor in modulation of gastric afferent neurons in the rat
Authors: Bielefeldt K. , Ozaki N. , Gebhart G. F.
Published: March 2015

Recent studies demonstrated that experimental ulcers are associated with changes in the properties of voltage-sensitive sodium currents in sensory neurons. We hypothesized that nerve growth factor (NGF) contributes to these changes. Gastric ulcers were induced by acetic acid injection into the wall of the rat stomach. NGF expression was determined by ELISA and immunohistochemically. Sensory neurons were labeled by injection of a retrograde tracer into the gastric wall. Sodium currents were recorded in gastric sensory neurons from nodose and dorsal root ganglia cultured for 24 h in the presence of NGF or a neutralizing NGF antibody, respectively. Gastric ulcer formation caused a rise in NGF concentration within the gastric wall and an increase in NGF immunoreactivity. Exposure to NGF caused a significant increase in the TTX-resistant sodium current, whereas the TTX-sensitive sodium current remained unchanged. This was associated with an acceleration of the recovery from inactivation in spinal sensory neurons. Production and release of NGF in the gastric wall may contribute to sensitization of primary afferent neurons during gastric inflammation.
Electrical charge on protein regulates its absorption from the rat small intestine
Authors: Nishikawa Makiya , Hasegawa Susumu , Yamashita Fumiyoshi , Takakura Yoshinobu , Hashida Mitsuru
Published: March 2015

The effect of the electrical charge on the intestinal absorption of a protein was studied in normal adult rats. Chicken egg lysozyme (Lyz), a basic protein with a molecular weight of 14,300, was selected and several techniques for chemical modification were applied. Then the intestinal absorption of Lyz derivatives was evaluated by measuring the radioactivity in plasma and tissues, after the administration of an111In-labeled derivative to an in situ closed loop of the jejunum. After the administration of111In-Lyz, the level of radioactivity in plasma was comparable with the lytic activity of Lyz, supporting the fact that the radioactivity represents intact Lyz.111In-cationized Lyz showed a 2–3 times higher level of radioactivity in plasma, whereas the radioactivity of111In-anionized Lyz was much lower. The absorption rate of111In-Lyz derivatives calculated by a deconvolution method was correlated for the strength of their positive net charge. A similar relationship was observed using superoxide dismutase. These findings indicate that the intestinal absorption of a protein is, at least partially, determined by its electrical charge.
Rectal and colonic distension elicit viscerovisceral reflexes in humans
Authors: Law Ngai-Moh , Bharucha Adil E. , Zinsmeister Alan R.
Published: March 2015

Colonic transit is slowed in patients with disordered rectal evacuation, but the mechanism of this phenomenon is unclear. Our objective was to investigate rectocolonic inhibitory reflexes in humans to provide potential insight into patients with obstructed defecation. In 30 healthy subjects, a barostat-manometric assembly recorded colonic tone and phasic activity in the descending colon during rectal distension and recorded rectal tone during colonic distension. Phasic distensions were 8, 16, and 32 mmHg above balloon operating pressure, and staircase inflations were comprised of balloon inflation then deflation in 2-mmHg increments at 30-s intervals from 0 to 36 mmHg. Colonic balloon volumes increased to a similar extent during phasic rectal distensions 8, 16, and 32 mmHg above operating pressure, reflecting reduced colonic tone; balloon volumes also increased and phasic pressure activity decreased during staircase rectal distensions. In contrast, rectal balloon volume declined, reflecting increased tone during phasic and staircase colonic distensions. Thus rectal distension inhibited colonic motor activity, indicative of a viscerovisceral inhibitory reflex.
A common Dubin-Johnson syndrome mutation impairs protein maturation and transport activity of MRP2 (ABCC2)
Authors: Keitel Verena , Nies Anne T. , Brom Manuela , Hummel-Eisenbeiss Johanna , Spring Herbert , Keppler Dietrich
Published: March 2015

Absence of a functional multidrug resistance protein 2 (MRP2; symbol ABCC2) from the hepatocyte canalicular membrane is the molecular basis of Dubin- Johnson syndrome, an inherited disorder associated with conjugated hyperbilirubinemia in humans. In this work, we analyzed a relatively frequent Dubin-Johnson syndrome mutation that leads to an exchange of two hydrophobic amino acids, isoleucine 1173 to phenylalanine (MRP2I1173F), in a predicted extracellular loop of MRP2. HEK-293 cells stably transfected with MRP2I1173F cDNA synthesized a mutant protein that was mainly core-glycosylated, predominantly retained in the endoplasmic reticulum, and degraded by proteasomes. MRP2I1173F did not mediate ATP-dependent transport of leukotriene C4(LTC4) into vesicles from plasma membrane and endoplasmic reticulum preparations while normal MRP2 was functionally active. Human HepG2 cells were used to study localization of MRP2I1173F in a polarized cell system. Quantitative analysis showed that GFP-tagged MRP2I1173F was localized to the apical membrane in only 5% of transfected, polarized HepG2 cells compared with 80% for normal MRP2-GFP. Impaired protein maturation followed by proteasomal degradation of inactive MRP2I1173F explain the deficient hepatobiliary elimination observed in this group of Dubin-Johnson syndrome patients.
High-fat diet effects on gut motility, hormone, and appetite responses to duodenal lipid in healthy men
Authors: Boyd K. A. , O'Donovan D. G. , Doran S. , Wishart J. , Chapman I. M. , Horowitz M. , Feinle C.
Published: March 2015

There is evidence that gastrointestinal function adapts in response to a high-fat (HF) diet. This study investigated the hypothesis that an HF diet modifies the acute effects of duodenal lipid on appetite, antropyloroduodenal pressures, plasma CCK and plasma glucagon-like peptide-1 (GLP-1) levels in humans. Twelve healthy men were studied twice in randomized, crossover fashion. The effects of a 90-min duodenal lipid infusion (6.3 kJ/min) on the above parameters were assessed immediately following 14-day periods on either an HF or a low-fat (LF) diet. After the HF diet, pyloric tonic and phasic pressures were attenuated, and the number of antropyloroduodenal pressure-wave sequences was increased when compared with the LF diet. Plasma CCK and GLP-1 levels did not differ between the two diets. Hunger was greater during the lipid infusion following the HF diet, but there was no difference in food intake. Therefore, exposure to an HF diet for 14 days attenuates the effects of duodenal lipid on antropyloroduodenal pressures and hunger without affecting food intake or plasma hormone levels.