bluuebunny/crossref_metadata_2025 huggingface dataset:
Ethanol modulation of intestinal epithelial tight junction barrier
Authors: Ma Thomas Y. , Nguyen Don , Bui Vuong , Nguyen Hanh , Hoa Neil
Published: December 2017

Previous studies have shown that high concentrations of ethanol (≥40%) cause functional damage of the gastrointestinal epithelial barrier by direct cytotoxic effect on the epithelial cells. The effects of lower noncytotoxic doses of ethanol on epithelial barrier function are unknown. A major function of gastrointestinal epithelial cells is to provide a barrier against the hostile substances in the gastrointestinal lumen. The apicolaterally located tight junctions (TJs) form a paracellular seal between the lateral membranes of adjacent cells and act as a paracellular barrier. In this study, we investigated the effects of lower doses of ethanol on intestinal epithelial TJ barrier function using filter-grown Caco-2 intestinal epithelial monolayers. The Caco-2 TJ barrier function was assessed by measuring epithelial resistance or paracellular permeability of the filter-grown monolayers. Ethanol (0, 1, 2.5, 5, 7.5, and 10%) produced a dose-related drop in Caco-2 epithelial resistance and increase in paracellular permeability. Ethanol also produced a progressive disruption of TJ protein (ZO-1) with separation of ZO-1 proteins from the cellular junctions and formation of large gaps between the adjacent cells. Ethanol, at the doses used (≤10%), did not cause cytotoxicity (lactate dehydrogenase release) to the Caco-2 cells. Ethanol produced a disassembly and displacement of perijunctional actin and myosin filaments from the perijunctional areas. On ethanol removal, actin and myosin filaments rapidly reassembled at the cellular borders. Ethanol stimulated the Caco-2 myosin light chain kinase (MLCK) activity but did not affect the MLCK protein levels. Specific MLCK inhibitor ML-7 inhibited both ethanol increases in MLCK activity and TJ permeability without affecting the MLCK protein levels. Consistent with these findings, metabolic inhibitors sodium azide and 2,4-dinitrophenol significantly prevented ethanol-induced increase in Caco-2 TJ permeability, whereas cycloheximide or actinomycin D had no effect. The results of this study indicate that ethanol at low noncytotoxic doses causes a functional and structural opening of the Caco-2 intestinal epithelial TJ barrier by activating MLCK.
V. Therapeutic potential of nitric oxide donors and inhibitors
Authors: Muscará Marcelo N. , Wallace John L.
Published: December 2017

Nitric oxide is a crucial mediator of gastrointestinal mucosal defense, but, paradoxically, it also contributes to mucosal injury in several situations. Inhibitors of nitric oxide synthesis and compounds that release nitric oxide have been useful pharmacological tools for evaluating the role of nitric oxide in gastrointestinal physiology and pathophysiology. Newer inhibitors with selectivity for one of the isoforms of nitric oxide synthase are even more powerful tools and may have utility as therapeutic agents. Also, agents that can scavenge nitric oxide or peroxynitrite are promising as drugs to prevent nitric oxide-associated tissue injury. Compounds that release nitric oxide in small amounts over a prolonged period of time may also be very useful for prevention of gastrointestinal injury associated with shock and with the use of drugs that have ulcerogenic effects. Indeed, the coupling of a nitric oxide-releasing moiety to nonsteroidal anti-inflammatory drugs has proven to be a valid means of substantially reducing the gastrointestinal toxicity of these drugs without decreasing their efficacy.
IL-2-deficient mice raised under germfree conditions develop delayed mild focal intestinal inflammation
Authors: Schultz Michael , Tonkonogy Susan L. , Sellon Rance K. , Veltkamp Claudia , Godfrey Virginia L. , Kwon Julie , Grenther Wetonia B. , Balish Edward , Horak Ivan , Sartor R. Balfour
Published: December 2017

Interleukin-2 (IL-2) amplifies immune stimuli and influences B cell differentiation. IL-2-deficient mice spontaneously develop intestinal inflammation if raised under specific pathogen-free (SPF) conditions. We quantitatively determined the aggressiveness and kinetics of gastrointestinal and hepatic inflammation in the presence or absence of viable bacteria in IL-2-deficient mice. Breeding colonies were maintained under SPF and germfree (GF) conditions. Intestinal tissues, serum, and mesenteric lymph nodes were obtained from mice at different ages for blind histological scoring, immunoglobulin measurements, mucosal T cell infiltration, and cytokine secretion. GF IL-2 −/− mice developed mild, focal, and nonlethal intestinal inflammation with delayed onset, whereas the more aggressive inflammation in SPF IL-2 −/− mice led to their death between 28 and 32 wk. Periportal hepatic inflammation was equal in the presence or absence of bacterial colonization. Intestinal immunoglobulin secretion decreased significantly by 13 wk of age in IL-2 −/− mice in both GF and SPF environments. In contrast to other genetically engineered rodents, IL-2 −/− mice develop mild focal gastrointestinal and active portal tract inflammation in the absence of viable bacteria.
Po2measurements in the rat intestinal microcirculation
Authors: Sinaasappel M. , Donkersloot C. , van Bommel J. , Ince C.
Published: December 2017

Microvascular partial oxygen pressure (Po2) data measured with the quenched phosphorescence of palladium-porphyrin (Pd-porphyrin) with the use of optical fibers have provided new insight into the behavior of the microvascular oxygenation in models of shock. However, the actual microcirculatory compartment measured by this fiber technique has not yet been demonstrated. The purpose of this study was to investigate whether the Po2of the intestines, as measured using a fiber phosphorometer, reflects the microvascular compartment. To this end, a new intravital phosphorometer with an improved sensitivity to high-Po2levels (up to 180 mmHg) was developed and validated. With this setup, Po2values were measured at different inspired oxygen fractions (15, 25, and 50% oxygen) in first-order arterioles, capillaries, and venules of the ileum of rats. Simultaneously, the Po2was measured with an optical fiber attached to another phosphorometer. Po2measurements with the fiber phosphorometer show excellent correlation with the Po2in the capillaries and first-order venule vessels ( r2= 0.94, slope 0.99). We therefore conclude that values measured with the fiber phosphorometer correlate with the capillary and venular Po2.
Modulation of host cell membrane fluidity: a novel mechanism for preventing bacterial adhesion
Authors: Ismaili Arif , Meddings Jonathan B. , Ratnam Samuel , Sherman Philip M.
Published: December 2017

Adhesion of bacterial enteropathogens to host mucosal surfaces is a critical primary step in the pathogenesis of diarrheal disease. We investigated the effects of altering the physical properties of eukaryotic cells on bacterial adhesion with the use of a series of three structurally dissimilar membrane fluidizers and several Escherichia coli as test strains. Lipid fluidity of the cell plasma membrane was measured by steady-state fluorescence anisotropy employing the probe 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene. There was a dose-dependent and reversible inhibition of bacterial adhesion with increasing membrane fluidity. Time course experiments indicated that increasing membrane fluidity during the early stages of bacterial adhesion was essential for inhibition of attachment. None of the fluidizers affected the viability of either eukaryotic or prokaryotic cells. These findings demonstrate, for the first time, that changes in plasma membrane physical properties of epithelial cells can prevent microbial adhesion. This also suggests that altering the membrane properties of host cells could form a basis for novel strategies to prevent bacterial adhesion during infection in vivo.
Generation of slow waves in membrane potential is an intrinsic property of interstitial cells of Cajal
Authors: Lee Jonathan C. F. , Thuneberg Lars , Berezin Irene , Huizinga Jan D.
Published: December 2017

To reveal the unique intrinsic properties of interstitial cells of Cajal (ICC), morphological and electrophysiological characteristics of isolated ICC from the adult mouse small intestine were investigated and compared with those of smooth muscle cells. All typical ultrastructural features of in situ ICC were evident in isolated ICC throughout the isolation procedure and short-term culture. With the use of the nystatin perforated patch-clamp technique, ICC demonstrated spontaneous voltage oscillations that were not abolished by hyperpolarization nor by L-type calcium channel blockers. This rhythmic activity occurred at room temperature at a frequency of 13.9 ± 11.2 cycles/min, with an amplitude of 13.4 ± 11.2 mV at membrane potentials from −20 to −70 mV. Smooth muscle cells from the same culture only generated voltage-sensitive action potentials above the threshold potential of −35 mV. Hyperpolarization as well as the addition of L-type calcium channel blockers abolished the action potentials. In whole cell voltage-clamp recordings from ICC, a large noninactivating outward current was observed to be activated (5% threshold) at −49.6 mV with a half-activation voltage of −18.7 mV and slope factor of 9.9 mV. In contrast, in smooth muscle cells, smaller outward currents with distinctive transient outward currents were present. In conclusion, the generation of L-type calcium channel blocker-insensitive slow waves in membrane potential is a unique intrinsic property of ICC.
Detection of gastric slow wave propagation from the cutaneous electrogastrogram
Authors: Chen J. D. Z. , Zou Xiaoping , Lin Xuemei , Ouyang Shou , Liang Jie
Published: December 2017

The gastric slow wave is originated in the proximal stomach and propagates distally toward the pylorus. It determines the maximum frequency and propagation of gastric contractions. The aim of this study was to detect the propagation of the gastric slow wave from the surface electrogastrogram (EGG). The study was performed in 11 healthy subjects of normal weight. Gastric myoelectrical activity was recorded for 1 h in the fasting state with the use of a specially designed multichannel recording device that was composed of four identical amplifiers with cutoff frequencies of 1.8 and 16.0 cycles/min. Four active electrodes were placed on the abdomen along the gastric axis and were connected to a common reference electrode placed near the xiphoid process, yielding four-channel bipolar EGG signals. Cross-covariance analysis was performed to compute the time lag among the four channels. There was a time lag in EGG waveform between channels 1 and 4 (9.6 ± 1.1 s); the average time during which the time lag was observed (≥3 s) was 89.9 ± 9.0%. There was a significant difference in the time lag among different adjacent channels ( P< 0.04); the time lag observed between channels 3 and 4 was significantly smaller than that between channels 1 and 2( P < 0.03). No correlation was found between the body mass index and the time lag between channels 1 and 4 ( r= −0.31, P = 0.3). It was concluded that, with a multichannel recording device with identical multiamplifiers and an appropriate arrangement of abdominal electrodes, the propagation of the gastric slow wave could be identified from the EGG in healthy subjects. This method may be used to detect the coupling of the gastric slow wave noninvasively.
Electrical physiological evidence for highand low-affinity vagal CCK-A receptors
Authors: Li Ying , Zhu Jinxia , Owyang Chung
Published: December 2017

We have demonstrated that under physiological conditions CCK acts through vagal high-affinity CCK-A receptors to mediate pancreatic secretion. In this study, we evaluated the vagal afferent response to endogenous CCK in rats and defined the CCK-receptor affinity states and the vagal-receptive field responsive to CCK stimulation using electrophysiological studies. Experiments were performed on anesthetized rats prepared with bile-pancreatic fistula. Plasma CCK levels were elevated by diverting bile-pancreatic juice (BPJ). The single-unit discharge of sensory neurons supplying the gastrointestinal tract was recorded from the nodose ganglia. All units studied were either silent or they had a very low resting discharge frequency. Thirty-two single units were studied extensively; seven were shown to be stimulated by diversion of BPJ (2.6 ± 2 impulses/min at basal to 40 ± 12 impulses/min after diversion). Acute subdiaphragmatic vagotomy or perivagal capsaicin treatment abolished the response. The CCK-A-receptor antagonist CR-1409, but not the CCK-B antagonist L-365260, blocked the vagal response to endogenous CCK stimulation. Infusion of the low-affinity CCK-receptor antagonist CCK-JMV-180 completely blocked the vagal afferent response to the diversion of BPJ in three of seven rats tested but had no effect on the response in the remaining four. In a separate study, we demonstrated that gastric, celiac, or hepatic branch vagotomy abolished the response in different subgroups of neurons. In conclusion, under physiological conditions, CCK acts on both high- and low-affinity CCK-A receptors present on distinct vagal afferent fibers. The vagal CCK-receptor field includes the regions innervated by the gastric, celiac, and hepatic vagal branches. This study provides electrophysiological evidence that vagal CCK receptors are present on the vagal gastric, celiac, and hepatic branches and may occur in high- and low-affinity states.
LPS-induced muscularis macrophage nitric oxide suppresses rat jejunal circular muscle activity
Authors: Eskandari Mark K. , Kalff Jörg C. , Billiar Timothy R. , Lee Kenneth K. W. , Bauer Anthony J.
Published: December 2017

Cellular mechanisms of sepsis-induced ileus remain an enigma. The study aim was to determine the role of nitric oxide (NO) in mediating the suppression of rat jejunal circular smooth muscle activity during endotoxemia. Isolated muscularis inducible NO synthase (iNOS) mRNA was measured by RT-PCR, immunohistochemistry was employed to localize iNOS protein, and contractile activity was measured in an organ bath. The low basal expression of muscularis iNOS mRNA expression was increased in a time-dependent fashion after lipopolysaccharide (LPS), resulting in a 20-fold increase over controls 3 h after injection. Immunohistochemistry of muscularis whole mounts and dissociated muscularis cells for iNOS revealed staining only in the muscularis macrophages 12 h after LPS. LPS caused a 68% reduction in spontaneous muscle activity 12 h after injection, which improved by 53% after the in vitro application of the selective iNOS inhibitorl- N6-(1-iminoethyl)lysine. Similar results were obtained in C57BL/6 mice but not in iNOS knockout mice. These data demonstrate that macrophage iNOS plays an important role in mediating LPS-induced intestinal circular muscle suppression.
Stroking human jejunal mucosa induces 5-HT release and Cl− secretion via afferent neurons and 5-HT4receptors
Authors: Kellum John M. , Albuquerque Francisco C. , Stoner Michael C. , Harris R. Paul
Published: December 2017

5-Hydroxytryptamine (5-HT) release and neural reflex pathways activated in response to mucosal stroking were investigated in muscle-stripped human jejunum mounted in modified Ussing chambers. The mucosa was stroked with a brush at 1/s for 1–10 s. Mucosal stroking resulted in a significant increase in the concentration of 5-HT in the mucosal bath within 5 min. It also was associated with a reproducible positive change (Δ) in short-circuit current ( I sc), which was abolished by inhibitors of chloride secretion. Capsaicin and hexamethonium significantly inhibited the Δ I sc but not the release of 5-HT. The Δ I sc was inhibited by TTX but not by atropine. It was also inhibited by the 5-HT3,4 receptor antagonist tropisetron (10 μM) and the 5-HT4,3 receptor antagonist SDZ-205-557 (10 μM) but not by preferential antagonists of 5-HT1P, 5-HT2A, or 5-HT3 receptors. These results suggest that mucosal stroking induces release of mucosal 5-HT, which activates a 5-HT4 receptor on enteric sensory neurons, evoking a neuronal reflex that stimulates chloride secretion.
Identification of a functional Ca2+-sensing receptor in normal human gastric mucous epithelial cells
Authors: Rutten Michael J. , Bacon Kathy D. , Marlink Katie L. , Stoney Mark , Meichsner Camie L. , Lee Fred P. , Hobson Susan A. , Rodland Karin D. , Sheppard Brett C. , Trunkey Donald D. , Deveney Karen E. , Deveney Clifford W.
Published: December 2017

The purpose of the present study was to determine whether human gastric mucous epithelial cells express a functional Ca2+-sensing receptor (CaR). Human gastric mucous epithelial cells were isolated from surgical tissues and cultured on glass coverslips, plastic dishes, or porous membrane filters. Cell growth was assessed by the MTT assay, CaR localization was detected by immunohistochemistry and confocal microscopy, CaR protein expression was assessed by Western immunoblotting, and intracellular Ca2+concentration ([Ca2+]i) was determined by fura 2 spectrofluorometry. In paraffin sections of whole stomach, we found strong CaR immunohistochemical staining at the basolateral membrane, with weak CaR-staining at the apical membrane in mucous epithelial cells. Confocal microscopy of human gastric mucous epithelial cell cultures showed abundant CaR immunofluorescence at the basolateral membrane and little to no CaR immunoreactivity at the apical membrane. Western immunoblot detection of CaR protein in cell culture lysates showed two significant immunoreactive bands of 140 and 120 kDa. Addition of extracellular Ca2+to preconfluent cultures of human gastric mucous epithelial cells produced a significant proliferative response. Changes in [Ca2+]iwere also observed in response to graded doses of extracellular Ca2+and Gd3+. The phospholipase C inhibitor U-73122 specifically inhibited Gd3+-induced changes in [Ca2+]iin the gastric mucous epithelial cell cultures. In conclusion, we have identified the localization of a functional CaR in human gastric mucous epithelial cells.
Biomechanical basis for lingual muscular deformation during swallowing
Authors: Napadow Vitaly J. , Chen Qun , Wedeen Van J. , Gilbert Richard J.
Published: December 2017

Our goal was to quantify intramural mechanics in the tongue through an assessment of local strain during the physiological phases of swallowing. Subjects were imaged with an ultrafast gradient echo magnetic resonance imaging (MRI) pulse sequence after the application of supersaturated magnetized bands in the x and y directions. Local strain was defined through deformation of discrete triangular elements defined by these bands and was depicted graphically either as color-coded two-dimensional strain maps or as three-dimensional octahedra whose axes correspond to the principal strains for each element. During early accommodation, the anterior tongue showed positive strain (expansive) in the anterior-posterior direction ( x), whereas the middle tongue showed negative strain (contractile) in the superior-inferior direction ( y). During late accommodation, the anterior tongue displayed increased positive x-direction and y-direction strain, whereas the posterior tongue displayed increased negative y-direction strain. These findings were consistent with contraction of the anterior-located intrinsic muscles and the posterior-located genioglossus and hyoglossus muscles. During propulsion, posterior displacement of the tongue was principally associated with positive strain directed in the x and y directions. These findings were consistent with posterior passive stretch in the midline due to contraction of the laterally inserted styloglossus muscle, as well as contraction of the posterior located transversus muscle. We conclude that MRI of lingual deformation during swallowing resolves the synergistic contractions of the intrinsic and extrinsic muscle groups.
CFTR-mediated inhibition of epithelial Na+ conductance in human colon is defective in cystic fibrosis
Authors: Mall M. , Bleich M. , Kuehr J. , Brandis M. , Greger R. , Kunzelmann K.
Published: December 2017

Cystic fibrosis (CF) patients show characteristic defects in epithelial ion transport, such as failure in cAMP-dependent Cl−secretion. Because the cystic fibrosis transmembrane conductance regulator (CFTR) also functions as a downregulator of epithelial Na+ channels (ENaC), enhanced Na+ conductance was found in the airways of CF patients. Here, we examined whether enhanced epithelial Na+ conductance is also present in the colonic epithelium of CF patients and examined the underlying mechanisms. Thus transepithelial voltages were measured, and equivalent short-circuit currents ( I sc-eq) were determined by means of a novel type of Ussing chamber. Non-CF tissues demonstrated cAMP-dependent Cl− secretion that was absent in biopsies of CF patients. Correspondingly, Isc-eq was inhibited in non-CF but not in CF epithelia when synthesis of endogenous prostaglandins was blocked by indomethacin. In the presence of indomethacin, a larger portion of amiloride-sensitive Isc-eq was detected in CF tissues, suggesting enhanced ENaC conductance in colonic mucosa of CF patients. Increase of intracellular cAMP by forskolin and IBMX inhibited amiloride-sensitive ENaC currents in non-CF tissues but not in CF biopsies. Therefore, enhanced epithelial Na+ conductance is present in the CF colon and is probably due to missing downregulation by CFTR.
Intestinal tachyarrhythmias during small bowel ischemia
Authors: Seidel Scott A. , Hegde Sanjay S. , Bradshaw L. Alan , Ladipo J. K. , Richards William O.
Published: December 2017

The electrical control activity (ECA) of the bowel is the omnipresent slow electrical wave of the intestinal tract. Characterization of small bowel electrical activity during ischemia may be used as a measure of intestinal viability. With the use of an animal model of mesenteric ischemia, serosal electrodes and a digital recording apparatus utilizing autoregressive spectral analysis were used to monitor the ECA of 20 New Zealand White rabbits during various lengths of ischemia. ECA frequency fell from 18.2 ± 0.5 cycles per minute (cpm) at baseline to 12.2 ± 0.9 cpm ( P < 0.05) after 30 min of ischemia and was undetectable by 90 min of ischemia in all animals. Tachyarrhythmias of the ECA were recorded in 55% of the animals as early as 25 min after ischemia was induced and lasted from 1 to 48 min. Frequencies ranged from 25 to 50 cpm. These tachyarrhythmias were seen only during ischemia, suggesting that they are pathognomonic for intestinal ischemia. The use of the detection of ECA changes during intestinal ischemia may allow earlier diagnosis of mesenteric ischemia.
VII. Apoptosis in intestinal epithelium: lessons from transgenic and knockout mice
Authors: Watson Alastair J. M. , Pritchard D. Mark
Published: December 2017

Apoptosis plays an important role in homeostasis of intestinal epithelia and is also a stress response to toxic stimuli. Transgenic and knockout mice have provided insights into the regulation of intestinal epithelial apoptosis that could not have been obtained by cell culture techniques. Two broad types of apoptosis have been characterized: spontaneous apoptosis, which occurs continuously at low levels in the normal, unstressed intestine, and stress-induced apoptosis, which occurs after genotoxic insult such as exposure to gamma radiation or DNA-damaging drugs. Spontaneous apoptosis occurs at the base of the crypt at or near the position of epithelial stem cells. Knockout studies have shown that spontaneous apoptosis is independent of p53 and Bax in both small and large intestine, whereas Bcl2 only regulates spontaneous apoptosis in the colon. Little is known about the regulation of the specialized form of cell death at the villus tip. In contrast, knockout studies have demonstrated that both p53 and Bcl2 are important regulators of stress-induced apoptosis but that there are significant differences between early and late time points. Bax plays only a minor role in the regulation of stress-induced apoptosis. The cumulative effect of stress-induced apoptosis on tissue architecture is not straightforward, and cell cycle arrest also plays a critical role. Nevertheless, p53 is an important determinant of the histopathological damage induced by 5-fluorouracil in murine intestinal epithelium. These studies have important implications for the development of more effective treatment for inflammatory bowel disease and cancer.
Reduction of EGF is associated with the delay of ulcer healing by cigarette smoking
Authors: Ma L. , Wang W. P. , Chow J. Y. C. , Yuen S. T. , Cho C. H.
Published: December 2017

Cigarette smoking is associated with peptic ulcer diseases. Smokers have lower levels of salivary epidermal growth factor (EGF) than nonsmokers. We investigated whether reduction of EGF is involved in the delay of gastric ulcer healing by cigarette smoking. Rats with acetic acid-induced ulcers were exposed to cigarette smoke (0, 2, or 4% vol/vol) 1 day after ulcer induction. EGF level was elevated 1 day after ulcer induction in salivary glands and serum, and 4 days after ulcer induction in the gastric mucosa. However, cigarette smoke depressed these beneficial effects and EGF mRNA expression in salivary glands and gastric mucosa. Cigarette smoke delayed gastric ulcer healing and reduced cell proliferation, angiogenesis, and mucus synthesis. Exogenous EGF (10 and 20 μg/kg iv) before smoke exposure reversed the adverse effects of cigarette smoke, whereas vascular endothelial growth factor level and nitric oxide synthase activity were unaffected. It is concluded that the detrimental effect of cigarette smoke on ulcer healing is a consequence of reduction of angiogenesis, cell proliferation, and mucus secretion through the depressive action on EGF biosynthesis and its mRNA expression in salivary glands and gastric mucosa.
NF-κB determines between apoptosis and proliferation in hepatocytes during liver regeneration
Authors: Plümpe Jörg , Malek Nisar P. , Bock C.-Thomas , Rakemann Tim , Manns Michael P. , Trautwein Christian
Published: December 2017

Tumor necrosis factor (TNF)-α is a potent inducer of apoptotic cell death in various tissues, whereas the transcription factor nuclear factor (NF)-κB is essential to protect against TNF-α-induced apoptosis. Human hepatoma cell lines were used to investigate the effectiveness and specificity of the fungal metabolite gliotoxin in inhibiting TNF-α-induced NF-κB activation in transformed cells. Gliotoxin-TNF-α cotreatment induced massive apoptosis in these otherwise TNF-α-resistant cell lines. With the use of the mouse partial hepatectomy model, we were also able to demonstrate in vivo the capacity of gliotoxin to act as inhibitor of NF-κB activation. Bromodeoxyuridine staining of liver sections showed that the lack of NF-κB activation correlated with 80% reduction of DNA synthesis 48 h after hepatectomy compared with untreated controls. Additionally, animals treated with gliotoxin showed nuclear condensation and DNA laddering of hepatocytes indicative of apoptosis 24 h after hepatectomy. In summary, our results demonstrate that NF-κB is essential in defining the fate of liver cells in response to TNF-α in vivo and furthermore implicate gliotoxin as a potential new response modifier for TNF-α-based therapy.
Genistein augments prostaglandin-induced recovery of barrier function in ischemia-injured porcine ileum
Authors: Blikslager Anthony T. , Roberts Malcolm C. , Young Karen M. , Rhoads J. Marc , Argenzio Robert A.
Published: December 2017

We have previously shown that PGE2enhances recovery of transmucosal resistance ( R) in ischemia-injured porcine ileum via a mechanism involving chloride secretion. Because the tyrosine kinase inhibitor genistein amplifies cAMP-induced Cl−secretion, we postulated that genistein would augment PGE2-induced recovery of R. Porcine ileum subjected to 45 min of ischemia was mounted in Ussing chambers, and R and mucosal-to-serosal fluxes of [3H] N-formyl-methionyl-leucyl phenylalanine (FMLP) and [3H]mannitol were monitored as indicators of recovery of barrier function. Treatment with genistein (10−4M) and PGE2(10−6M) resulted in synergistic elevations in R and additive reductions in mucosal-to-serosal fluxes of [3H]FMLP and [3H]mannitol, whereas treatment with genistein alone had no effect. Treatment of injured tissues with genistein and either 8-bromo-cAMP (10−4M) or cGMP (10−4M) resulted in synergistic increases in R. However, treatment of tissues with genistein and the protein kinase C (PKC) agonist phorbol myristate acetate (10−5–10−6M) had no effect on R. Genistein augments recovery of Rin the presence of cAMP or cGMP but not in the presence of PKC agonists.
Impairment of Ca2+mobilization in circular muscle cells of the inflamed colon
Authors: Shi Xuan-Zheng , Sarna Sushil K.
Published: December 2017

This study investigated whether inflammation modulates the mobilization of Ca2+in canine colonic circular muscle cells. The contractile response of single cells from the inflamed colon was significantly suppressed in response to ACh, KCl, and BAY K8644. Methoxyverapamil and reduction in extracellular Ca2+concentration dose-dependently blocked the response in both normal and inflamed cells. The increase in intracellular Ca2+concentration in response to ACh and KCl was significantly reduced in the inflamed cells. However, Ca2+efflux from the ryanodine- and inositol 1,4,5-trisphosphate (IP3)-sensitive stores, as well as the decrease of cell length in response to ryanodine and IP3, were not affected. Heparin significantly blocked Ca2+efflux and contraction in response to ACh in both conditions. ACh-stimulated accumulation of IP3and the binding of [3H]ryanodine to its receptors were not altered by inflammation. Ruthenium red partially inhibited the response to ACh in normal and inflamed states. We conclude that the canine colonic circular muscle cells utilize Ca2+influx through L-type channels as well as Ca2+release from the ryanodine- and IP3-sensitive stores to contract. Inflammation impairs Ca2+influx through L-type channels, but it may not affect intracellular Ca2+release. The impairment of Ca2+influx may contribute to the suppression of circular muscle contractility in the inflamed state.
Fas and Fas ligand in gut and liver
Authors: Pinkoski Michael J. , Brunner Thomas , Green Douglas R. , Lin Tesu
Published: December 2017

Apoptosis (programmed cell death) has been shown to play a major role in development and in the pathogenesis of numerous diseases. A principal mechanism of apoptosis is molecular interaction between surface molecules known as the “death receptors” and their ligands. Perhaps the best-studied death receptor and ligand system is the Fas/Fas ligand (FasL) system, in which FasL, a member of the tumor necrosis factor (TNF) family of death-inducing ligands, signals death through the death receptor Fas, thereby resulting in the apoptotic death of the cell. Numerous cells in the liver and gastrointestinal tract have been shown to express Fas/FasL, and there is a growing body of evidence that the Fas/FasL system plays a major role in the pathogenesis of many liver and gastrointestinal diseases, such as inflammatory bowel disease, graft vs. host disease, and hepatitis. Here we review the Fas/FasL system and the evidence that it is involved in the pathogenesis of liver and gastrointestinal diseases.
Mechanism of glucocorticoid-mediated reversal of inhibition of Cl−/ HCO 3 − exchange during chronic ileitis
Authors: Coon Steve , Sundaram Uma
Published: December 2017

In the normal ileum, coupled NaCl absorption occurs via the dual operation of Na+/H+and Cl−/[Formula: see text] exchange on the brush-border membrane (BBM) of villus cells. In a rabbit model of chronic small intestinal inflammation we determined the cellular mechanism of inhibition of NaCl absorption and the effect of steroids on this inhibition. Cl−/[Formula: see text] but not Na+/H+exchange was reduced in the BBM of villus cells during chronic ileitis. Cl−/[Formula: see text] exchange was inhibited secondary to a decrease in the affinity for Cl−rather than an alteration in the maximal rate of uptake of Cl−(Vmax). Methylprednisolone (MP) stimulated Cl−/[Formula: see text] exchange in the normal ileum by increasing the Vmaxof Cl−uptake rather than altering affinity for Cl−. MP reversed the inhibition of Cl−/[Formula: see text] exchange in rabbits with chronic ileitis. However, MP alleviated the Cl−/[Formula: see text] exchange inhibition by restoring the affinity for Cl−rather than altering the Vmaxof Cl−uptake. These data suggest that glucocorticoids mediate the alleviation of Cl−/[Formula: see text] exchange inhibition in chronically inflamed ileum by reversing the same mechanism that was responsible for inhibition of this transporter rather than exerting a direct effect on the transporter itself, as was the case in normal ileum.
Regulation of the human Na+-glucose cotransporter gene,SGLT1, by HNF-1 and Sp1
Authors: Martín Martín G. , Wang Jiafang , Solorzano-Vargas R. Sergio , Lam Jason T. , Turk Eric , Wright Ernest M.
Published: December 2017

The Na+-glucose cotransporter ( SGLT1) is expressed primarily by small intestinal epithelial cells and transports the monosaccharides glucose and galactose across the apical membrane. Here we describe the isolation and characterization of 5.3 kb of the 5′-flanking region of the SGLT1 gene by transiently transfecting reporter constructs into a variety of epithelial cell lines. A fragment (nt −235 to +22) of the promoter showed strong activity in the intestinal cell line Caco-2 but was inactive in a nonintestinal epithelial cell line (Chinese hamster ovary). Within this region, three cis-elements, a hepatocyte nuclear factor-1 (HNF-1) and two GC box sites are critical for maintaining the gene's basal level of expression. The two GC boxes bind to several members of the Sp1 family of transcription factors and, in the presence of HNF-1, synergistically upregulate transactivation of the promoter. A novel 16-bp element just downstream of one GC box was also shown to influence the interaction of Sp1 to its binding site. In summary, we report the identification and characterization of the human SGLT1 minimal promoter and the critical role that HNF-1 and Sp1-multigene members have in enhancing the basal level of its transcription in Caco-2 cells.
Gender differences in early alcohol-induced liver injury: role of CD14, NF-κB, and TNF-α
Authors: Kono Hiroshi , Wheeler Michael D. , Rusyn Ivan , Lin Min , Seabra Vitor , Rivera Chantal A. , Bradford Blair U. , Forman Donald T. , Thurman Ronald G.
Published: December 2017

The purpose of this study was to determine whether early alcohol-induced liver injury (ALI) in females is associated with changes in CD14 on Kupffer cells, activation of hepatic nuclear factor (NF)-κB, and expression of tumor necrosis factor (TNF)-α mRNA. Male and female rats were given high-fat control or ethanol-containing diets for 4 wk using the intragastric enteral protocol. Physiological parameters were similar in both genders. Ethanol was increased as tolerance developed with higher blood levels than previously observed, resulting in a fourfold increase in aspartate aminotransferase (males 389 ± 47 IU/l vs. females 727 ± 66 IU/l). Hepatic pathology developed more rapidly and was nearly twofold greater and endotoxin levels were significantly higher in females after ethanol. Also, expression of CD14 on Kupffer cells was 1.5-fold greater and binding of transcription factor NF-κB in hepatic nuclear extracts and TNF-α mRNA expression were threefold greater in females. These data are consistent with the hypothesis that elevated endotoxin after ethanol triggers more activation of Kupffer cells via enhanced CD14 expression in females. NF-κB is activated in this process, leading to increases in TNF-α mRNA expression in the liver and more severe liver injury in females. It is concluded that gender differences in ALI are dependent on endotoxin and a signaling cascade leading to TNF-α.
Early aldosterone effect in distal colon by transcriptional regulation of ENaC subunits
Authors: Epple H. J. , Amasheh S. , Mankertz J. , Goltz M. , Schulzke J. D. , Fromm M.
Published: December 2017

Aldosterone-induced sodium absorption is mediated by the epithelial Na+channel (ENaC). It is thought that the “early effect” is not based on genomic regulation of ENaC expression, because ENaC subunit transcription was reported to start later than Na+transport. We investigated electrogenic Na+absorption ( JNa) and, in identical tissues, mRNA expression of ENaC subunits in early (EDC) and late (LDC) distal colon of the rat. In both segments, 8-h in vitro incubation with 3 nM aldosterone enhanced expression of β- and γ-ENaC mRNA and induced JNa. JNawas 10 times higher in LDC than in EDC. α-ENaC mRNA was unchanged in EDC, whereas it decreased in LDC. In LDC, β- and γ-ENaC mRNA was induced 1 h after aldosterone addition, whereas JNabecame apparent >1 h later. Downregulation of α-ENaC mRNA did not take part in acute regulation because it started after a lag time of 3 h. Time correlation of β- and γ-ENaC induction and JNastimulation suggests that the early aldosterone effect on Na+absorption in distal colon is caused by transcriptional upregulation of β- and γ-ENaC expression.
Cyclooxygenase-1 and an alternatively spliced mRNA in the rat stomach: effects of aging and ulcers
Authors: Vogiagis Daphne , Glare Eric M. , Misajon Aileen , Brown Wendy , O'Brien Paul E.
Published: December 2017

Prostaglandins play a critical role in gastric mucosal cytoprotection and decrease progressively with age. Cyclooxygenase (COX), the rate-limiting enzyme for prostaglandin synthesis, exists in two isoforms, COX-1 and COX-2. The rat COX-1 gene expresses an alternatively spliced mRNA COX-1 splice variant (SV) that may, at best, code for a truncated COX-1 protein. With the use of competitive PCR, we determined whether COX gene expression was altered in the stomach with increasing age and after gastric ulcer induction. COX-1 mRNA was significantly reduced in the aged, and COX-1SV mRNA was significantly higher in the adults compared with the young and aged stomach. Levels of COX-1 and COX-2 were similarly expressed in the normal stomach. In acute gastric ulcers, only COX-2 mRNA levels were significantly elevated. When ulcers were undergoing healing and repair, COX-1 and COX-2 mRNA levels were significantly elevated. Age-related changes in COX-1 and COX-1SV but not COX-2 mRNA may alter gastric mucosal cytoprotection. Furthermore, COX-1 and COX-2 may both contribute to the healing of a gastric ulcer.
Chronic stress impairs rat growth and jejunal epithelial barrier function: role of mast cells
Authors: Santos Javier , Benjamin Michelle , Yang Ping-Chang , Prior T. , Perdue Mary H.
Published: December 2017

We examined the impact of chronic stress on rat growth rate and intestinal epithelial physiology and the role of mast cells in these responses. Mast cell-deficient (Ws/Ws) rats and +/+ littermate controls were submitted to water avoidance stress or sham stress, 1 h/day, for 5 days. Seven hours after the last sham or stress session, jejunal segments were mounted in Ussing chambers, in which secretion and permeability were measured. Body weight (as a growth index) and food intake were determined daily. Stress increased baseline jejunal epithelial ion secretion (indicated by short-circuit current), ionic permeability (conductance), and macromolecular permeability (horseradish peroxidase flux) in +/+ rats, but not in Ws/Ws rats, compared with nonstressed controls. Stress induced weight loss and reduced food intake similarly in the groups. In +/+ rats, these parameters remained altered 24–72 h after the cessation of stress. Modulation of stress-induced mucosal mast cell activation may help in the management of certain intestinal conditions involving epithelial pathophysiology.
Intraepithelial lymphocytes coinduce nitric oxide synthase in intestinal epithelial cells
Authors: Hoffman Rosemary A.
Published: December 2017

The study of mucosal immunity has revealed that complex reciprocal interactions occur between intestinal intraepithelial lymphocytes (IEL) and intestinal epithelial cells (IEC). The present study focuses on the induction of inducible nitric oxide (NO) synthase in cocultures of freshly isolated rat IEL and the rat epithelial cell line IEC-18 after the addition of interleukin-1β (IL-1β), tumor necrosis factor-α, or lipopolysaccharide. When IEL and IEC were separated using Transwell chambers, NO synthesis was not induced, indicating that cell-cell contact was required. Culture of IEC-18 with IEL, even in the absence of inflammatory stimuli such as IL-1β, resulted in upregulation of class I and II antigens on IEC-18, due to the interferon-γ (IFN-γ) that is constitutively produced by IEL. Addition of anti-IFN-γ antibody to the NO-producing cocultures resulted in inhibition of NO synthesis as well as the upregulation of class I and II antigen expression. These data indicate that IFN-γ production by IEL conditions IEC for the expression of other components of the inflammatory process.
Pramlintide, an amylin analog, selectively delays gastric emptying: potential role of vagal inhibition
Authors: Samsom Melvin , Szarka Lawrence A. , Camilleri Michael , Vella Adrian , Zinsmeister Alan R. , Rizza Robert A.
Published: December 2017

The amylin analog pramlintide delays gastric emptying in type I diabetics. The effects of multiple doses of pramlintide and the mechanism of action in non-amylin-deficient humans are unknown. We investigated the effects of pramlintide on gastrointestinal and colonic transit and on the plasma pancreatic polypeptide response to the meal in a parallel-group dose-response study with subjects randomized to placebo, or 30 or 60 μg (tid, sc) of pramlintide. Pramlintide delayed gastric emptying [half-time ( t½): 112 min (SE 8.7 min), 169 min (SE 12 min), or 177 min (SE 25 min) after placebo or 30- or 60-μg pramlintide treatment, respectively; P = 0.033]. Pramlintide did not significantly affect small bowel or colonic transit. Pancreatic polypeptide concentrations in the first postprandial hour were lower with pramlintide than with placebo ( P < 0.01 for drug effect). An inverse correlation was observed between mean pancreatic polypeptide concentrations in the first postprandial hour and gastric emptying t½[Spearman correlation coefficient ( Rs) = 0.48; P = 0.044]. Pramlintide at 30 and 60 μg delays gastric emptying in healthy humans without affecting small bowel or colonic transit. Vagal inhibition is a potential mechanism of the effects of pramlintide on gastric emptying.
Lipolysis and lipid oxidation in cirrhosis and after liver transplantation
Authors: Shangraw Robert E. , Jahoor Farook
Published: December 2017

On the basis of the finding that plasma glycerol concentration is not controlled by clearance in healthy humans, it has been proposed that elevated plasma free fatty acid (FFA) and glycerol concentrations in cirrhotic subjects are caused by accelerated lipolysis. This proposal has not been validated. We infused 10 volunteers, 10 cirrhotic subjects, and 10 patients after orthotopic liver transplantation (OLT) with [1-13C]palmitate and [2H5]glycerol to compare fluxes (Ra) and FFA oxidation. Cirrhotic subjects had higher plasma palmitate (52%) and glycerol (33%) concentrations than controls. Palmitate Ra was faster (1.45 ± 0.18 vs. 0.85 ± 0.17 μmol ⋅ kg−1 ⋅ min−1) but glycerol Ra and clearance slower (1.20 ± 0.09 vs. 1.90 ± 0.24 μmol ⋅ kg−1 ⋅ min−1and 21.2 ± 1.2 vs. 44.7 ± 4.9 ml ⋅ kg− ⋅ h−1, respectively) than in controls. After OLT, plasma palmitate and glycerol concentrations and palmitate Ra did not differ, but glycerol Ra (1.16 ± 0.11 μmol ⋅ kg−1 ⋅ min−1) and clearance (26.7 ± 2.4 ml ⋅ kg− ⋅ h−1) were slower than in controls. We conclude that 1) impaired reesterification, not accelerated lipolysis, elevates FFA in cirrhotic subjects; 2) normalized FFA after OLT masks impaired reesterification; and 3) plasma glycerol concentration poorly reflects lipolytic rate in cirrhosis and after OLT.
Gi-1/Gi-2-dependent signaling by single-transmembrane natriuretic peptide clearance receptor
Authors: Murthy K. S. , Teng B.-Q. , Zhou H. , Jin J.-G. , Grider J. R. , Makhlouf G. M.
Published: December 2017

Single-transmembrane natriuretic peptide clearance receptor (NPR-C), which is devoid of a cytoplasmic guanylyl cyclase domain, interacts with pertussis toxin (PTx)-sensitive G proteins to activate endothelial nitric oxide synthase (eNOS) expressed in gastrointestinal smooth muscle cells. We examined the ability of NPR-C to activate other effector enzymes in eNOS-deficient tenia coli smooth muscle cells; these cells expressed NPR-C and NPR-B but not NPR-A. Atrial natriuretic peptide (ANP), the selective NPR-C ligand cANP-(4–23), and vasoactive intestinal peptide (VIP) inhibited125I-ANP and125I-VIP binding to muscle membranes in a pattern indicating high-affinity binding to NPR-C. Interaction of VIP with NPR-C was confirmed by its ability to inhibit125I-ANP binding to membranes of NPR-C-transfected COS-1 cells. In tenia muscle cells, all ligands selectively activated Gi-1and Gi-2; VIP also activated Gsvia VIP2receptors. All ligands stimulated phosphoinositide hydrolysis, which was inhibited by ANP-(1–11), PTx, and antibodies to phospholipase C-β3 (PLC-β3) and Gβ. cANP-(4–23) contracted tenia muscle cells; contraction was blocked by U-73122 and PTx and by antibodies to PLC-β3 and Gβ in intact and permeabilized muscle cells, respectively. VIP and ANP contracted muscle cells only after inhibition of cAMP- and cGMP-dependent protein kinases. ANP and cANP-(4–23) inhibited forskolin-stimulated cAMP in a PTx-sensitive fashion. We conclude that NPR-C is coupled to activation of PLC-β3 via βγ-subunits of Gi-1and Gi-2and to inhibition of adenylyl cyclase via α-subunits.
Bile salts mediate hepatocyte apoptosis by increasing cell surface trafficking of Fas
Authors: Sodeman Thomas , Bronk Steven F. , Roberts Patricia J. , Miyoshi Hideyuki , Gores Gregory J.
Published: December 2017

Toxic bile salts induce hepatocyte apoptosis by a Fas-dependent, Fas ligand-independent mechanism. To account for this observation, we formulated the hypothesis that toxic bile salts induce apoptosis by effecting translocation of cytoplasmic Fas to the cell surface, resulting in transduction of Fas death signals. In McNtcp.24 cells the majority of Fas was cytoplasmic, as assessed by cell fractionation and immunofluorescence studies. However, cell surface Fas increased sixfold after treatment with the toxic bile salt glycochenodeoxycholate (GCDC) in the absence of increased Fas protein expression. Moreover, in cells transfected with Fas-green fluorescence protein, cell surface fluorescence also increased in GCDC-treated cells, directly demonstrating Fas translocation to the plasma membrane. Both brefeldin A, a Golgi-disrupting agent, and nocodazole, a microtubule inhibitor, prevented the GCDC-induced increase in cell surface Fas and apoptosis. In conclusion, toxic bile salts appear to induce apoptosis by promoting cytoplasmic transport of Fas to the cell surface by a Golgi- and microtubule-dependent pathway.