bluuebunny/crossref_metadata_2025 huggingface dataset:
Canine corticosteroid-induced alkaline phosphatase in serum was solubilized by phospholipase activity in vivo
Authors: Solter P. F. , Hoffmann W. E.
Published: December 2017

In this study, gas chromatography/mass spectrometry revealed the presence of stoichiometric amounts of myo-inositol in association with serum corticosteroid-induced isozyme of alkaline phosphatase (CALP) in canine serum. Such remnants are consistent with prior membrane attachment of serum CALP and its release into serum by endogenous phospholipase activity. Serum CALP was further shown to behave similarly to CALP released from hepatocyte membranes by glycosyl phosphatidylinositol phospholipase D (GPI-PLD) and differently from CALP solubilized by GPI-phospholipase C (PLC) on both native polyacrylamide gel electrophoresis and Western blot analysis using anti-cross-reacting determinant antibody. In addition to bile canalicular surfaces, CALP activity was found over hepatocyte sinusoidal surfaces by histochemical staining of canine liver sections. A significantly higher ratio of CALP to total alkaline phosphatase activity was observed in serum as opposed to bile in 10 of 11 paired serum and bile samples from dogs. This suggested that bile is not likely to be the source of serum CALP and is consistent with the release of CALP from hepatocyte basolateral surfaces directly into serum. It was concluded that serum CALP was once membrane bound and was released by phospholipase activity into serum. Our findings are consistent with release of CALP from the sinusoidal surfaces of hepatocytes into serum either by endogenous GPI-PLD activity or release by GPI-PLC followed by modification of the phosphatidylinositol remnant in vivo.
Effect of intestinal resection and arginine-free diet on rat physiology
Authors: Wakabayashi Y. , Yamada E. , Yoshida T. , Takahashi N.
Published: December 2017

The small intestine has been presumed to release citrulline as a precursor for the endogenous arginine synthesis. We studied the effect of intestinal resection and arginine-free diet on rat physiology. We maintained rats with massively resected small intestine (R rats) and those with transected intestines (T rats) on either control or an arginine-free diet. After 4 wk, R rats fed deficient diet [R(-)] lost weight by a mean of 46 g, whereas R rats fed control diet [R(+)] and T rats fed control [T(+)] and deficient diet [T(-)] gained 30-96 g. Average nitrogen balance was 150, 60, 110, and -33 mg/day for T(+), T(-), R(+), and R(-), respectively. The concentrations of arginine in skeletal muscle were 654, 163, 230, and 84 nmol/g, respectively, and those in plasma were 133, 50, 103, and 54 microM, respectively. The concentrations of citrulline in R rats were halved compared with T rats irrespective of diet. We conclude that arginine is synthesized in a small intestine-dependent manner in the rat.
Novel model of integration of signaling pathways in rat pancreatic acinar cells
Authors: Rivard N. , Rydzewska G. , Lods J. S. , Morisset J.
Published: December 2017

Cholecystokinin (CCK) is the major pancreatic secretagogue and acinar cell mitogen. This study was performed to determine by which effector systems CCK regulates tyrosine kinases, phosphatidylinositol (PtdIns) 3-kinase, and phospholipase D (PLD) activities. Pancreatic acini loaded with [3H]myristic acid or [3H]inositol were used to assay PLD and PtdIns 3-kinase. G protein activation with NaF increased particulate and crude cytosolic tyrosine kinase and PLD activities. PLD activation was pertussis toxin sensitive. Inhibition of phospholipase C (PLC) slightly reduced caerulein-stimulated particulate tyrosine kinase and blocked crude cytosolic tyrosine kinase activity without affecting caerulein-induced PLD activity. Ca2+ is an important factor in caerulein stimulation of tyrosine kinase and PLD activities. Protein kinase C and tyrosine kinase inhibition abolished caerulein-activated particulate and crude cytosolic tyrosine kinase and PtdIns 3-kinase activities without any effect on PLD. Wortmannin inhibited PLD and PtdIns 3-kinase activation. Caerulein-induced amylase secretion was partially reduced by tyrosine kinase inhibition, with no effect from wortmannin. Caerulein can stimulate a pertussis toxin-insensitive G protein, leading to particulate tyrosine kinase activation and a Ca(2+)-sensitive cytosolic tyrosine kinase through PLC activation. However, PLD activation by caerulein is pertussis toxin sensitive, cytosolic Ca2+ sensitive, and independent of previous PLC and tyrosine kinase activation.
Bicarbonate diffusion through mucus
Authors: Livingston E. H. , Miller J. , Engel E.
Published: December 2017

The mucus layer overlying duodenal epithelium maintains a pH gradient against high luminal acid concentrations. Despite these adverse conditions, epithelial surface pH remains close to neutrality. The exact nature of the gradient-forming barrier remains unknown. The barrier consists of mucus into which HCO3- is secreted. Quantification of the ability of HCO3- to establish and maintain the gradient depends on accurate measurement of this ion's diffusion coefficient through mucus. We describe new experimental and mathematical methods for diffusion measurement and report diffusion coefficients for HCO3- diffusion through saline, 5% mucin solutions, and rat duodenal mucus. The diffusion coefficients were 20.2 +/- 0.10, 3.02 +/- 0.31, and 1.81 +/- 0.12 x 10(-6) cm2/s, respectively. Modeling of the mucobicarbonate layer with this latter value suggests that for conditions of high luminal acid strength the neutralization of acid by HCO3- occurs just above the epithelial surface. Under these conditions the model predicts that fluid convection toward the lumen could be important in maintaining the pH gradient. In support of this hypothesis we were able to demonstrate a net luminal fluid flux of 5 microliters.min-1.cm-2 after perfusion of 0.15 N HCl in the rat duodenum.
Tight junctions and the molecular basis for regulation of paracellular permeability
Authors: Anderson J. M. , Van Itallie C. M.
Published: December 2017

Tight junctions create a regulated paracellular barrier to the movement of water, solutes, and immune cells between both epithelial and endothelial cells. Recent progress has been made in identifying the proteins that create this barrier. The transmembrane protein occludin is an excellent candidate for the sealing protein and is bound on the cytoplasmic membrane surface to the proteins ZO-1 and ZO-2. Functions for ZO-1 and ZO-2 are suggested by their invertebrate homologues, one of which is a tumor suppressor and another is required in epidermal growth factor receptor signaling. Multiple cellular signaling pathways affect assembly and sealing of junctions. Dynamic regulation of perijunctional actin has emerged as a unifying hypothesis for controlling paracellular permeability. Understanding and manipulating permeability will require a more detailed molecular characterization of tight junction proteins and in particular a characterization of how cell signaling regulates their attachment to the perijunctional cytoskeleton.
Gender influences sphincter of Oddi response to cholecystokinin in the prairie dog
Authors: Tierney S. , Qian Z. , Yung B. , Lipsett P. A. , Pitt H. A. , Sostre S. , Lillemoe K. D.
Published: December 2017

Although gallstones and disorders of biliary tract motility are both more common in women than men, sphincter of Oddi motility has not previously been compared between the sexes. In this study, cholescintigraphy (under ketamine and diazepam anesthesia) was used to determine gallbladder emptying rate and ejection fraction in response to cholecystokinin (CCK) in eight male and six female prairie dogs fed a nonlithogenic diet. Ten days later, under alpha-chloralose anesthesia, sphincter of Oddi phasic wave activity was monitored for 10-min intervals before (control), during 20 min of CCK infusion, and for 20 min after infusion. Gallbladder emptying rate and ejection fraction and baseline sphincter of Oddi frequency, amplitude, and motility index (= frequency x amplitude) did not differ significantly between the sexes. Sphincter of Oddi phasic wave frequency was increased during CCK infusion in both males and females, but the change in amplitude was significantly greater in females, than males. We conclude that the increased incidence of biliary tract disease in women may be due to altered sphincter of Oddi hormonal response.
Influence of breathing pattern on the esophagogastric junction pressure and esophageal transit
Authors: Mittal R. K. , Shaffer H. A. , Parollisi S. , Baggett L.
Published: December 2017

The esophagogastric junction (EGJ) is guarded by two sphincters, a smooth muscle lower esophageal sphincter (LES) and a skeletal muscle crural diaphragm. The LES relaxes in response to a swallow but the crural diaphragm does not. Since contraction of the crural diaphragm is affected by the pattern of breathing, the latter may also influence the EGJ pressure and swallow-induced EGJ relaxation. Our aims were to study the effects of alterations of the breathing pattern on the EGJ pressure, swallow-induced EGJ relaxation, and esophageal transport of liquid bolus. Manometric, electromyographic, and videofluoroscopic studies were performed in 12 healthy subjects. The subjects were trained in two types of breathing patterns, hyperventilation and partial expiration, using the visual biofeedback from their own respiratory waveform. Hyperventilation increased the frequency of inspiratory pressure oscillations at the EGJ without affecting the end-expiratory EGJ pressure. Partial expiration resulted in an increase in the end-expiratory EGJ pressure. Swallow-induced relaxation at the EGJ was markedly reduced during partial expiration. An inspiration during a swallow (control breathing and hyperventilation) caused transient interruption of flow across the EGJ. Partial expiration resulted in cessation of the flow across the EGJ, failure of esophageal peristalsis to traverse the entire length of the esophagus, increased esophageal transit time, and incomplete esophageal clearance of a liquid bolus.(ABSTRACT TRUNCATED AT 250 WORDS)
Na(+)-K(+)-ATPase alpha 1- and beta 1-mRNA and protein levels in rat small intestine in experimental ileitis
Authors: Wild G. E. , Thomson A. B.
Published: December 2017

Na(+)-K(+)-adenosinetriphosphatase (ATPase) plays a key role in the absorption of electrolytes, water, and nutrients from the small intestine. The expression of Na(+)-K(+)-ATPase was examined in isolated enterocytes during the course of the ileal inflammatory response elicited by intraluminal administration of 2,4,6-trinitrobenzenesulfonic acid. The ileal inflammatory response was characterized by a marked cellular infiltrate, villous atrophy, and crypt hyperplasia along with fibrosis and smooth muscle hypertrophy. Peak levels of myeloperoxidase were observed at day 7, and ileal mucosal injury was paralleled by increases in ileal mucosal permeability. Ileal enterocytes were harvested from days 3 to 30 after the induction of ileitis. Decreases in Na(+)-K(+)-ATPase functional activity were observed from days 3 to 21 and were accompanied by corresponding decreases in Na(+)-K(+)-ATPase pump abundance, alpha 1- and beta 1-protein expression, and mRNA abundance, whereas Na(+)-K(+)-ATPase turnover, Michaelis-Menten constant values, and inhibition constant values for Na+ and ouabain, respectively, were unaltered. Alterations in transcriptional and posttranscriptional events may determine the changes in Na(+)-K(+)-ATPase activity in this particular model. Additionally observed increases in thymidine kinase and ornithine decarboxylase activities appear to signify alterations in the state of differentiation of the ileal epithelium and may determine the phenotypic expression of enterocyte transporters and permeability in the setting of inflammation.
Intermittence of blood flow in liver sinusoids, studied by high-resolution in vivo microscopy
Authors: MacPhee P. J. , Schmidt E. E. , Groom A. C.
Published: December 2017

Kupffer cell migration and leukocyte-vessel wall interactions cause temporary slowing and/or stoppage of blood flow through individual liver sinusoids. Such temporal heterogeneity of flow was quantified in anesthetized mice and rats. Video recordings of red blood cell flow in 44 networks containing 8-16 sinusoids each were analyzed for 5- to 10-min periods. Flow was graded "fast," "slow," "stopped," or "reversed" based on red blood cell velocity. The mean numbers of flow changes (between grades) per minute in zone 1 vs. zone 3 were 1.39 vs. 0.78 (mouse) and 1.25 vs. 0.09 (rat). The mean percentage of time for each flow grade differed significantly between zones 1 and 3 and between species. For example, fast flow was present in zone 1 sinusoids for 51% of the time in mice and for 74% in rats; in zone 3 the corresponding numbers were 76 and 95%. Flow stasis was present in zone 1 sinusoids for 19% of the time in mice and for 7% in rats; in zone 3 the corresponding numbers were 2 and 0%. Thus considerable intermittence of perfusion exists, and the flow conditions create very different microenvironments for hepatocytes in zone 1 vs. zone 3.
Gastric antisecretory and anti-ulcer actions of IL-1 in rat involve different IL-1 receptor types
Authors: Mugridge K. G. , Perretti M. , Ghiara P. , Galeotti C. L. , Melli M. , Parente L.
Published: December 2017

Limited knowledge exists concerning the interleukin-1 (IL-1) receptor type (IL-1RT) mediating the potent antisecretory and gastro-protective actions of IL-1. In the present study, the gastric actions of IL-1 beta and two related mutant proteins, yIL-1 beta delta 4, an analogue that preferentially binds to IL-1-RTII, and mutant yIL-1 beta N7/Q, an analogue that has equal affinity as IL-1 beta for IL-1RTI and IL-1RTII, have been compared. Modulation of IL-1 gastric actions were also investigated using monoclonal antibody (MAb) preparations raised against IL-1RTI or IL-1RTII. In the pylorus-ligated rat, yIL-1 beta delta 4, yIL-1 beta N7/Q, and IL-1 beta (all at 1 microgram/kg ip) reduced gastric acid secretion (50, 79, and 78%, respectively), indicating the importance of IL-1RTII binding for antisecretory activity. This was further substantiated in experiments using the MAb preparations, which showed that IL-1 beta (1 microgram/kg ip) antisecretory activity was reversed by MAb IL-1RTII (10-50 micrograms/kg sc) but not by MAb IL-1RTI (50 micrograms/kg sc). In contrast, at dosages 10-fold higher (10 micrograms/kg ip) than that used in the study to inhibit acid secretion, IL-1 beta and yIL-1N7/Q equally reduced (approximately 80%) indomethacin-induced gastric damage, but yIL-1 beta delta 4 was ineffective. The results using yIL-1 beta delta 4 indicated that impairment of IL-1RTI binding capacity appeared to be paralleled by a decreased gastroprotective effect.(ABSTRACT TRUNCATED AT 250 WORDS)
Octreotide reduces perception of rectal electrical stimulation by spinal afferent pathway inhibition
Authors: Chey W. D. , Beydoun A. , Roberts D. J. , Hasler W. L. , Owyang C.
Published: December 2017

Octreotide reduces perception of rectal distension in normal volunteers and irritable bowel patients. To localize octreotide's site of action, perceptual and evoked potential responses to rectal electrical stimulation were tested in seven normal volunteers after double-blind octreotide (100 micrograms 2) or placebo. After octreotide, the currents needed to elicit threshold perception of square-wave impulses delivered to the rectum were 29% higher than after placebo. When electrical stimulation was delivered at constant currents 50% above threshold, rectal perception scores were significantly reduced after octreotide compared with placebo. Rectal electrical stimulation led to characteristic and reproducible cerebral evoked potentials. Octreotide had no effect on latencies, but reduced peak-to-peak amplitudes by 35% compared with placebo. Rectal electrical stimulation also led to characteristic and reproducible spinal evoked potentials. Octreotide had no effect on spinal latencies, but reduced peak-to-peak amplitudes by 51%. In conclusion, octreotide reduces perception of rectal electrical stimulation, which is associated with inhibition of cerebral and spinal evoked potential amplitude, indicating effects on spinal afferent pathways.
Intra-islet regulation of hormone secretion by glucagon-like peptide-1-(7--36) amide
Authors: Heller R. S. , Aponte G. W.
Published: December 2017

Glucagon-like peptide (GLP)-1-(7--36) amide, a peptide product of the posttranslational processing of pancreatic and intestinal proglucagon, has been shown to regulate insulin secretion. Monoclonal antibodies to glucagon and GLP-1-(7--36) amide were generated to localize GLP-1-(7--36) amide in the pancreatic islets by immunocytochemistry and radioimmunoassay. GLP-1-(7--36) amide immunoreactivity was found in some, but not all, glucagon-containing alpha-cells. Displaceable receptor binding for GLP-1-(7--36) amide and nonamidated GLP-1-(7--37) on hormone secretion were investigated using isolated pancreatic islet preparations. GLP-1-(7--37) and -(7--36) amide significantly increased insulin and somatostatin release in the concentration range of 0.01-100 nM in 11.0 mM glucose. GLP-1-(7--37) and -(7--36) amide had no effect on glucagon secretion in the presence of 11.0 mM glucose. GLP-1-(7--36) amide was released from isolated islets in response to 2.25, 5.5, and 11.0 mM glucose. These results suggest that pancreatic GLP-1 may be important in the regulation of intra-islet hormone secretion.
PYY and GLP-1 contribute to feedback inhibition from the canine ileum and colon
Authors: Wen J. , Phillips S. F. , Sarr M. G. , Kost L. J. , Holst J. J.
Published: December 2017

To explore mechanisms whereby unabsorbed nutrients in the ileum inhibit the upper gut ("ileal brake"), we perfused the canine ileum or colon and monitored phase 3 in the duodenum. Fasting motility was recorded when the ileum or colon was perfused with 154 mM NaCl, a mixed isotonic nutrient solution (Ensure), or individual nutrients (maltose, casein hydrolysates, or sodium oleate). Blood samples were collected before and during the perfusions. The ileum was also perfused with 154 mM NaCl while peptide YY (PYY) was infused by vein. In both sets of experiments, plasma levels of PYY, neurotensin, and glucagon-like peptide-1 (GLP-1) were measured. Ileal or colonic perfusion of Ensure delayed phase 3 [migrating motor complexes (MMC)] in the duodenum, inhibited ileal motility, and increased plasma levels of PYY and GLP-1. Ileal casein and oleate and colonic casein also delayed the duodenal MMC. The MMC cycle length and plasma levels of PYY were closely correlated. Intravenous PYY prolonged the MMC cycle; an intravenous dose of 100 pmol.kg-1.h-1 of PYY mimicked the effects of ileal Ensure. These results support the hypothesis that PYY, and possibly GLP-1, participate in the ileal brake. This negative feedback loop also affects the distal small bowel. The proximal colon also triggers the feedback inhibition of gut motility (colonic brake).
Resection-induced colonic adaptation is augmented by IGF-I and associated with upregulation of colonic IGF-I mRNA
Authors: Mantell M. P. , Ziegler T. R. , Adamson W. T. , Roth J. A. , Zhang W. , Frankel W. , Bain A. , Chow J. C. , Smith R. J. , Rombeau J. L.
Published: December 2017

The effects of exogenous insulin-like growth factor-I (IGF-I) on colonic adaptation were examined in male Sprague-Dawley rats (n = 60, 225-275 mg) after either a 60% small bowel and cecal resection (RX) or mid-small bowel transection with reanastomosis (TX). Animals received a 7-day treatment with either IGF-I (2.4 mg.kg-1.day-1) or vehicle (V; 0.1 mol/l acetic acid). Body weight decreased significantly after resection (-25.6 +/- 4.0 g; P < 0.05 vs. TX/V). IGF-I treatment significantly reduced weight loss after resection (-12.4 +/- 3.8 g; P < 0.01 vs. RX/V) and induced significant weight gain after transection (15.6 +/- 4.0 g; P < 0.05 vs. TX/V). Plasma IGF-I decreased with resection (526 +/- 41 TX/V vs. 344 +/- 17 ng/ml RX/V; P < 0.01). IGF-I treatment significantly increased plasma IGF-I levels (805 +/- 100 ng/ml TX/IGF, 677 +/- 56 ng/ml RX/IGF). After resection, IGF-I treatment significantly increased colonic mucosal weight, DNA, protein content, and crypt depth when compared with resection alone (P < 0.05). Colonic water absorption, measured by an in vivo [3H]polyethylene glycol assay, was significantly increased by IGF-I treatment in resected animals (399 +/- 23 RX/IGF vs. 306 +/- 32 microliter.cm-1.h-1 RX/V; P < 0.05). Resection resulted in increased steady-state colonic IGF-I mRNA (182% of TX/V; P < 0.01) without significantly affecting IGF-I receptor mRNA expression. Regulation of IGF binding protein (BP)-3 and -4 was discoordinate, with IGFBP-3 mRNA tending to decrease with resection (67% of TX/V; P is not significant) and IGFBP-4 increasing significantly (191% of TX/V; P < 0.05). An important role for IGF-I in colonic adaptation after massive intestinal resection is indicated by 1) significantly enhanced colonic mucosal growth and water absorption with IGF-I treatment and 2) postresection upregulation of colonic IGF-I mRNA and alteration of IGFBP-3 and IGFBP-4 mRNA expression.
Enterocytic gene expression in intestinal adaptation: evidence for a specific cellular response
Authors: Rubin D. C. , Swietlicki E. A. , Wang J. L. , Dodson B. D. , Levin M. S.
Published: December 2017

After massive small bowel resection, the remnant gut epithelium undergoes an adaptive response marked by an increase in villus height, crypt depth, and crypt cell production rate. Although morphological features of gut adaptation have been well characterized, the differentiation status and response of epithelial cells populating the adaptive villus is unclear. To address these issues, cell-specific and spatial patterns of expression of a set of enterocytic genes were characterized in rats after 70% small bowel resection. The liver and intestinal (I) fatty acid binding protein (FABP) and apolipoprotein A-I (apo A-I) and apo A-IV genes were studied because they exhibit unique regional and cell-specific patterns of expression in the developing and adult gut. At 48 h after surgery, apo A-IV and I-FABP mRNA levels were increased up to 3.5-fold in adaptive remnant ileum compared with sham-operated or sham-resected control ileum. In situ hybridization and immunohistochemical analyses revealed a marked increase in enterocytic apo A-IV mRNA and protein expression in the adaptive ileum, from villus base to tip but not in crypts. By 1 wk after resection, apo A-IV, but not I-FABP, mRNA levels remained elevated in remnant ileum, although duodenal I-FABP mRNA levels were still increased. In contrast, apo A-I mRNA levels were not significantly induced. These results indicate that the enterocyte can respond acutely to loss of small bowel surface area by increasing expression of several genes. This compensatory enterocytic response is spatially (from duodenum to ileum) and temporally regulated. These results suggest initiation of the adaptive response occurs by way of a complex set of molecular pathways involving villus and crypt cells.
Ion transport systems in human labial acinar cells
Authors: Paulais M. , Valdez I. H. , Fox P. C. , Evans R. L. , Turner R. J.
Published: December 2017

Human labial acini were assayed for the presence of ion transport systems associated with salivary fluid secretion using microfluorometric methods. Na(+)-K(+)-Cl- contransport and Na+/H+ exchange activities (determined by their bumetanide and amiloride sensitivities, respectively) were found at levels approximately 50% of those seen in similarly assayed rat parotid acini, but little, if any, C1-/HCO3-exchange activity was observed. Also, when human labial acini were stimulated with the muscarinic agonist carbachol, little evidence of the intracellular acidification associated with HCO3- secretion by other salivary glands was found. Na+/H+ exchange activity in human labial acini was downregulated (approximately 40%) by beta-adrenergic stimulation and upregulated (approximately threefold) by muscarinic stimulation. In contrast, beta-adrenergic stimulation produced only a marginally significant increase in Na(+)-K(+)-Cl- cotransport activity, and muscarinic stimulation was without effect. We include that basolateral Na(+)-K(+)-Cl- cotransport appears to be the dominant mechanism driving Cl- secretion and thereby fluid secretion in this tissue.
Identification and expression of prohormone-converting enzymes in the rat stomach
Authors: Macro J. A. , Dimaline R. , Dockray G. J.
Published: December 2017

The conversion of regulatory peptide precursors to their active forms usually involves limited proteolysis that may be mediated by subtilisin-like prohormone convertases (PC). We have examined the representation of this enzyme family in rat gastric mucosa. With the use of polymerase chain reaction, employing primers to conserved sequences, we identified from rat antrum clones corresponding to PC1/3, PC2, PC5, and furin. Northern blots indicated that the mRNAs for PC1/3 and PC2 were substantially more abundant in mucosa compared with muscle, and that there were differences in expression in antrum and corpus. In the antrum a PC1/3 probe identified hands of 3 and 4.5 kb that were of equal intensity and were both increased in fasted rats; in corpus, the latter mRNA species predominated and did not change with fasting. In rats treated with omeprazole, there was a preferential increase in the antral 3-kb band. In both antrum and corpus, a PC2 probe hybridized with a band of 2.8 kb that increased in omeprazole-treated rats. The data suggest that 1) PC1/3 and PC2 are expressed in antral mucosa and so are candidates for gastric regulatory peptide processing, 2) there is selective processing of the mRNAs encoding prohormone convertases in different gastric cell populations, and 3) the expression of these enzymes is physiologically regulated.
A cascade of degradative hydrolase activity contributes to hepatocyte necrosis during anoxia
Authors: Arora A. S. , de Groen P. , Emori Y. , Gores G. J.
Published: December 2017

Calpain proteases contribute to hepatocyte necrosis during anoxia. Our aim was to ascertain the mechanism causing calpain activation during anoxia. In rat hepatocytes, a twofold increase in calpain activity occurred despite the lack of an increase in cytosolic Ca2+ concentration ([Ca2+]i). The increase in calpain activity was not associated with an increase in calpain mRNA or a decrease in calpastatin mRNA expression. Because phospholipid degradation products generated by phospholipases can activate calpains at physiological [Ca2+]i, we determined the effect of phospholipase inhibitors and activators on calpain activity. Pretreatment of hepatocytes with fluphenazine, a phospholipase inhibitor, decreased calpain activation and improved cell survival. Melittin, a phospholipase A2 activator, increased calpain activity and potentiated cell killing. Finally, phospholipid degradation preceded the increase in calpain activity. Thus the enhanced calpain activity occurring in hepatocytes during anoxia 1) is regulated at the posttranslational level and 2) appears to be dependent on phospholipase activity. These data suggest a novel cascade for degradative hydrolase activity during hepatocyte necrosis by anoxia with phospholipase-mediated activation of calpains.
Pharyngeal swallowing elicited by fluid infusion: role of volition and vallecular containment
Authors: Pouderoux P. , Logemann J. A. , Kahrilas P. J.
Published: December 2017

Nonalimentary swallows minimize aspiration by clearing accumulated fluid from the pharynx. This study aimed to define 1) the pharyngeal sensory field to elicit swallowing and 2) the effect of infusion rate, volition, taste, and temperature on pharyngeal swallows. Test solutions were directed into the valleculae at 6.5, 11.5, and 32 ml/min through a catheter in eight healthy volunteers. Deglutition was signaled with electromyography and electroglottography. Spatial distribution of infusate before swallowing was studied using videofluoroscopy coupled with a video timer. Volitional control was assessed with rapid or restrained swallows. Pharyngeal swallow latency decreased as the instillation rate increased, was potently modified with volition, and was unchanged by infusate taste or temperature. Water infusion into the valleculae did not trigger pharyngeal swallowing until liquids overflowed and reached the aryepiglottic folds or pyriform sinuses. The variation in swallow latency among flow rates was mainly due to the duration of liquid containment within the valleculae. This suggests that the valleculae act to contain pharyngeal secretions and residue and prevent aspiration by diverting their contents around the larynx before swallowing.
Extracellular ATP mediates Ca2+ signaling in cultured myenteric neurons via a PLC-dependent mechanism
Authors: Kimball B. C. , Yule D. I. , Mulholland M. W.
Published: December 2017

In the myenteric plexus, ATP is released as a neurotransmitter by "purinergic" nerves, relaxing visceral smooth muscle. We report a signal transduction mechanism for ATP in cultured myenteric neurons involving receptor-mediated release of intracellular Ca2+ stores. Primary cultures of myenteric neurons from guinea pigs taenia coli were loaded with the Ca2+ indicator fura 2-acetoxymethyl ester (AM) and examined using digital imaging microscopy. Superfusion of single neurons with ATP (0.01-1,000 microM) resulted in concentration-dependent increases in intracellular Ca2+ concentration ([Ca2+]i) that were independent of extracellular Ca2+. Decrements in peak [Ca2+]i were seen with repetitive ATP exposure. Responsiveness of myenteric neurons to purinergic agonists (100 microM) was consistent with action at a neuronal P 2y purinoceptor: 2-chloro-ATP = ATP = 2-methyl-thio-ATP (MeSATP) > ADP > alpha, beta-MeATP = beta,gamma-MeATP > AMP > adenosine. ATP-evoked Ca2+ transients were inhibited dose dependently by suramin, a nonspecific P2 antagonist, and reactive blue 2, a specific P 2y antagonist. ATP and cyclopiazonic acid (30 microM) appear to release an identical intracellular Ca2+ store. Preincubation with the aminosteroid U-73122 (10 microM) inhibited ATP-evoked Ca2+ transients by 71 +/- 7%, whereas phorbol ester pretreatment (phorbol 12-myristate 13-acetate, 100 nM, 5 min) caused a 76 +/- 4% inhibition. Peak [Ca2+]i evoked by ATP was not affected by preincubation with pertussis toxin (100 ng/ml, 24 h) or nifedipine (10 microM). These data suggest a signal transduction mechanism for ATP in cultured myenteric neurons involving purinoceptor-mediated activation of phospholipase C (PLC), with release of D-myo-inositol 1,4,5-trisphosphate-sensitive intracellular Ca2+ stores.
Infection of T84 cells with enteropathogenic Escherichia coli alters barrier and transport functions
Authors: Philpott D. J. , McKay D. M. , Sherman P. M. , Perdue M. H.
Published: December 2017

The effect of enteropathogenic Escherichia coli (EPEC) infection on electrophysiology of T84 cell monolayers was examined. After 18 h of infection with EPEC (E2348), transepithelial electrical resistance was decreased (30 +/- 5% of uninfected values) compared with monolayers infected with a nonpathogenic E. coli strain (104 +/- 13%). Resistance of monolayers infected with EPEC mutant strain CVD206, deficient in attaching and effacing lesion formation, was partially reduced (66 +/- 10%). In addition, permeability of EPEC-infected T84 monolayers increased compared with uninfected cells. Associated with these changes was an altered distribution of the tight junction protein, ZO-1. Taken together, these findings suggest that the barrier defect induced by EPEC was at the level of the tight junction. Adenosine 3'5'-cyclic monophosphate-stimulated chloride secretion was also diminished in EPEC-infected cells, whereas Ca2+ -dependent chloride secretion was not different from uninfected cells. These findings indicate that EPEC infection alters intestinal epithelial barrier and transport functions. Furthermore, these results provide a possible mechanism for EPEC-induced diarrheal disease.
Endotoxin stimulates gene expression of ROS-eliminating pathways in rat hepatic endothelial and Kupffer cells
Authors: Spolarics Z.
Published: December 2017

Reactive oxygen species (ROS) are mediators of cellular injury and play a putative role in the onset of hepatic damage during endotoxemia or sepsis. It has been suggested that induction of glucose-6-phosphate (G-6-P) dehydrogenase, the key enzyme of the hexose monophosphate shunt (HMS), may support ROS-producing or ROS-eliminating pathways in hepatic endothelial and Kupffer cells during endotoxemia. The aim of the study was to assess in vivo lipopolysaccharide (LPS)-induced alterations in rat gene expression of selected enzymes that are in functional relationship with the HMS. mRNA levels and activities of glucose transporter GLUT-1, Mn- and CuZn-dependent superoxide dismutases (Mn-SOD and CuZn-SOD), and Se-dependent glutathione peroxidase (Se-GPX) were determined. Cellular extracts were analyzed 7 or 22 h after injection of LPS (Escherichia coli, 2 mg/kg ip) or injection of saline. Exposure to LPS for 7 or 22 h caused a 10- to 25-fold increase in GLUT-1 mRNA levels in endothelial and Kupffer cells. In parenchymal cells, GLUT-1 mRNA expression was low, and LPS caused no marked changes. Cellular levels of Mn-SOD mRNA were 20-40 times greater in all hepatic cells from LPS-treated animals than in cells from control rats. LPS at 22 h increased Mn-SOD activity by 45% in endothelial cells but caused no significant changes in Kupffer or parenchymal cells. Message levels and enzyme activities of CuZn-SOD and Se-GPX were significantly elevated 22 h after LPS injection in endothelial cells only. Thus LPS results in marked upregulation of functionally related genes in hepatic cells. In endothelial cells, the simultaneous upregulation of GLUT-1, G-6-P dehydrogenase, Mn-SOD, CuZn-SOD, and Se-GPX may represent an important mechanism for accelerated elimination of ROS released from activated sinusoidal phagocytes. In Kupffer cells, upregulated GLUT-1 and G-6-P dehydrogenase, together with constitutively present SOD and lack of upregulated Se-GPX, suggest an elevated capacity to produce O2- and H2O2 that is consistent with primed bacterial killing.
Hepatic oxidant injury and glutathione depletion during total parenteral nutrition in weanling rats
Authors: Sokol R. J. , Taylor S. F. , Devereaux M. W. , Khandwala R. , Sondheimer N. J. , Shikes R. H. , Mierau G.
Published: December 2017

Hepatobiliary dysfunction occurs commonly in infants on prolonged parenteral nutrition alimentation; however, the underlying mechanisms causing liver injury are poorly understood. We postulated that oxidant stress played a significant role in parenteral nutrition-induced liver abnormalities and tested this hypothesis in a rat model. Weanling male rats received 8 days of total parenteral nutrition (TPN) through a central venous catheter (TPN group), pair feeding of rat chow and placement of a central venous catheter (sham group), or ad libitum feedings of rat chow (control group). After 8 days of TPN, serum alanine aminotransferase and cholylglycine levels were elevated, hepatocellular steatosis was present, hepatic mitochondria had dilated intracristal spaces, and lipid peroxidation of mitochondria was increased compared with sham and control groups. Hepatic glutathione levels decreased to 16% of control values after 5 days of TPN; this was followed by mitochondrial lipid peroxidation and elevated serum cholylglycine levels after 8 days of TPN. Sham and control rats showed no evidence of mitochondrial lipid peroxidation or liver injury after 8 days. Removal of metabisulfate from TPN solutions and addition of cysteine HCl or choline had no major effect on these findings. Bacterial translocation was not increased in TPN rats. These data suggest that glutathione depletion and oxidant stress are important factors in the pathogenesis of TPN-induced liver abnormalities in the weanling rats.
Inhibition of nitric oxide synthase in the brain suppresses colonic motor activity
Authors: Ohta D. , Sarna S. K. , Condon R. E. , Lang I. M.
Published: December 2017

The effects of inhibition of nitric oxide (NO) synthase in the brain on colonic motor activity were investigated in conscious dogs. Intracerebroventricular (ICV) infusion of N omega-nitro-L-arginine methyl ester (L-NAME) and N omega-nitro-L-arginine (L-NNA) significantly suppressed colonic motor activity. The inhibitory effects occurred 1 h after the end of the infusion and lasted for at least 4 h. This suppression was blocked by a concurrent infusion of L-arginine but not D-arginine. The suppression of colonic motor activity was not blocked by bilateral truncal vagotomy or intravenous administration of phentolamine, an alpha-receptor antagonist, and propranolol, a beta-receptor antagonist. ICV infusion of L-NAME had no significant effect on the occurrence of giant migrating contractions in the colon. By contrast, intravenous administration of L-NAME at higher doses significantly enhanced colonic motor activity. We conclude that the inhibition of NO synthesis in the brain suppresses, whereas the inhibition of NO synthesis in peripheral neurons stimulates, colonic motor activity. The central effect may be mediated by a decrease in cerebral blood flow due to the inhibition of NO synthesis in cerebral arteries.
Role of prostaglandins in intestinal epithelial restitution stimulated by growth factors
Authors: Zushi S. , Shinomura Y. , Kiyohara T. , Minami T. , Sugimachi M. , Higashimoto Y. , Kanayama S. , Matsuzawa Y.
Published: December 2017

Mucosal integrity is reestablished after superficial injuries by a rapid resealing process, termed epithelial restitution, that is regulated by several growth factors and cytokines. Growth factors are also known to stimulate the synthesis of endogenous prostaglandins that mediate important functions in intestinal epithelial cells. Therefore, we examined the effect of endogenous eicosanoid production modulators, piroxicam, dexamethasone, and nordihydroguaiaretic acid (NDGA) on intestinal epithelial restitution using two cultured cell wound-resealing models, IEC-6 and Caco-2 cells. Epidermal growth factor, transforming growth factor-beta, hepatocyte growth factor, and fetal calf serum (FCS) accelerated intestinal epithelial restitution, and piroxicam significantly suppressed these stimulatory effects. Dexamethasone mimicked the action of piroxicam. No additive effect of piroxicam and dexamethasone was observed. NDGA did not affect epithelial restitution. Piroxicam abolished the increase in 6-ketoprostaglandin F1 alpha (PGF1 alpha) release induced by FCS. Furthermore, addition of a stable PGI2 analogue, OP-41483 [5(E)-6,9-deoxa-6,9-methylene-15-cyclopentyl-16,17,18,19,20-pen tanor-PGI2], reversed the slowing of epithelial restitution induced by piroxicam. These results suggest that endogenous prostaglandins play an important role in regulating intestinal epithelial restitution.
Role of ornithine decarboxylase in enterocyte mitochondrial function and integrity
Authors: Madsen K. L. , Brockway P. D. , Johnson L. R. , Hardin J. A. , Gall D. G.
Published: December 2017

We examined the role of ornithine decarboxylase (ODC) and polyamine biosynthesis in regulating mitochondrial function and integrity along the crypt-villus axis in male Sprague-Dawley rats. Isolated villus tip enterocytes from control rats demonstrated a greater cellular capacity for glucose oxidation than crypt enterocytes. Mitochondrial enzyme activities were similar along the crypt-villus axis. The role of ODC was assessed by treating experimental rats with the irreversible ODC inhibitor alpha-difluoromethylornithine (DFMO) for 24 h. Animals receiving DFMO demonstrated a decreased CO2 production from [2-(14)C]pyruvate along the entire crypt-villus axis coupled with an increase in lactate production in the upper cell populations. CO2 production from [14C]glucose and total ATP levels were not affected by DFMO treatment. Ultrastructural examination revealed localized mitochondrial swelling and bursting only in enterocytes corresponding to the population of cells newly emerged from the crypt during DFMO treatment. In DFMO-treated animals, 2 microM spermine completely prevented the structural mitochondrial injury and restored the metabolic crypt-villus gradient. These results suggest that as enterocytes migrate from the crypt up the villus, mitochondrial function increases to handle the increased metabolic demands placed on the cell by nutrient absorption. ODC activity and polyamines are necessary for this increased mitochondrial function and have a role in the maintenance of mitochondrial integrity in maturing enterocytes migrating from the crypt onto the villus.
Contribution of surface epithelial cells to total conductance of Necturus gastric fundus mucosa
Authors: Kottra G. , Iacovelli C. , Caroppo R. , Curci S. , Bakos P. , Fromter E.
Published: December 2017

Microelectrode techniques were used to quantify the contribution of surface epithelial cells (SEC) to transepithelial conductance (gt) of Necturus gastric fundus mucosa. Transepithelial voltage (Vt) and resistance (Rt) as well as the basolateral cell membrane potential (Vb) and voltage divider ratio of SEC were measured. Freshly mounted preparations did not respond to luminal amiloride (10 microM), but within 2-3 h a significant response developed (delta Vt = 3.8 +/- 1.2 mV, delta Rt = 63 +/- 23 omega cm2, and delta Vb = -6.9 +/- 1.3 mV), indicating activation of an apical Na+ conductance in SEC. Using circuit analysis equations, we calculate that SEC contribute 10.4% to gt under control conditions and 13.0% after Na+ conductance activation. Histamine (0.1 mM), which stimulates the oxyntopeptic cells (OC), increased Vt and decreased Rt but did not significantly alter the membrane resistances of SEC. As a result, the contribution of SEC to gt fell to 7.4 or 9.3%, respectively. The data confirm that SEC are poorly permeable and that the major conductance path across gastric mucosa leads through OC in the glands. The reason for the protracted in vitro activation of the apical Na+ conductance in SEC is not known.
Kupffer cell depletion by gadolinium chloride enhances liver regeneration after partial hepatectomy in rats
Authors: Rai R. M. , Yang S. Q. , McClain C. , Karp C. L. , Klein A. S. , Diehl A. M.
Published: December 2017

Although previous work suggests that tumor necrosis factor-alpha (TNF) promotes liver regeneration after partial hepatectomy (PH), the source of TNF is unknown. If Kupffer cells release TNF after PH, then Kupffer cell depletion by gadolinium chloride (GdCl) should inhibit liver regeneration. To test this hypothesis, cytokine expression and regenerative events were compared in GdCl-treated and control rats. Functional assays and Northern blot analysis of a Kupffer cell-specific mRNA confirmed that GdCl depleted Kupffer cells. Despite this, semiquantitative reverse transcription-polymerase chain reaction analysis of total hepatic RNA showed six- to eightfold higher levels of TNF transcripts in GdCl-treated rats. In this group, PH caused 12-to 16-fold greater induction of interleukin-6, a TNF-inducible cytokine, and two- to threefold greater induction of several cytokine-regulated genes (c-jun, C/EBP-beta, and C/EBP-delta). GdCl also amplified regeneration-associated increases in the DNA binding activity of AP-1, a growth regulatory transcription factor. Furthermore, hepatic incorporation of [3H]thymidine, expression of the S-phase antigen, proliferating cell nuclear antigen, and the hepatocyte mitotic index were each significantly greater in GdCl-treated rats. Thus, although GdCl causes Kupffer cell depletion, it does not decrease liver TNF and actually enhances liver regeneration after PH.
Tumor necrosis factor-alpha stimulates adrenal glucocorticoid secretion in cholestatic rats
Authors: Swain M. G. , Maric M.
Published: December 2017

Tumor necrosis factor-alpha (TNF-alpha ) is capable of activating the hypothalamic-pituitary-adrenal (HPA) axis. We have recently documented altered activation of this axis by endotoxin and interleukin-1 in cholestatic rats. Therefore, in this study, we examined TNF-alpha-induced activation of the HPA axis, in rats with cholestasis due to bile duct resection, with the use of sham-resected rats as controls. Administration of TNF-alpha to bile duct- and sham-resected rats in vivo resulted in a similar increase in plasma adrenocorticotropic hormone levels in both groups of animals but significantly higher corticosterone levels in cholestatic rats, suggesting a direct steroidogenic effect of TNF-alpha in cholestatic rats. This suggestion was confirmed in further experiments by the demonstration of TNF-alpha-induced corticosterone secretion in hypophysectomized cholestatic but not control rats. Furthermore, a direct stimulatory effect of TNF-alpha on adrenal corticosterone secretion in vitro was noted only for cholestatic rats, possibly via augmented adrenal prostaglandin E2 (PGE2) production. These results indicate that TNF-alpha has a direct stimulatory effect on adrenal corticosterone secretion in cholestatic rats, possibly due to augmented adrenal PGE2 release.
5-HT-induced jejunal motor activity: enteric locus of action and receptor subtypes
Authors: Graf S. , Sarna S. K.
Published: December 2017

The role of 5-hydroxytryptamine (5-HT), its enteric locus of action, and receptor subtypes involved in the regulation of jejunal contractions were investigated by close intra-arterial infusions in conscious dogs. Close intra-arterial infusions of 5-HT in short segments of the jejunum stimulated phasic contractions that were blocked completely by atropine, partially by tetrodotoxin, and not affected by hexamethonium. This response was also blocked significantly by 5-HT2A and 5-HT2C receptor antagonists but was not affected by 5-HT1A/5-HT1B, 5-HT3, and 5-HT4 receptor antagonists. Spontaneous phase III contractions were inhibited significantly by 5-HT2A and 5-HT2C receptor antagonists, not affected by 5-HT1A/5-HT1B and 5-HT3 receptor antagonists, and enhanced by 5-HT4 receptor antagonists. Repeated close intra-arterial infusions of 5-HT over several days stimulated giant migrating contractions. We conclude that in the conscious state, 5-HT acts on 5-HT2A and 5-HT2C receptors located on postsynaptic cholinergic neurons in the canine jejunum to stimulate phasic contractions and phase III activity. The 5-HT4 receptors in the canine small intestine may be localized on nonadrenergic, noncholinergic inhibitory neurons; these receptors suppress the amplitude and duration of phase III activity.