bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of short-term endotoxemia and dopamine on mucosal oxygenation in porcine jejunum
Authors: Hasibeder W. , Germann R. , Wolf H. J. , Haisjackl M. , Hausdorfer H. , Riedmann B. , Bonatii J. , Gruber E. , Schwarz B. , Waldenberger P. , Friesenecker B. , Furtner B.
Published: December 2017

Effects of Escherichia coli lipopolysaccharide (2 micrograms.kg-1.20 min-1; LPS), given systemically (S) or via superior mesenteric artery (M), and consecutive dopamine infusion (16 micrograms.kg-1.20 min-1) on jejunal mucosal tissue O2 tension (PO2muc) and serosal tissue O2 tension (PO2ser; Clark-type surface electrodes) and jejunal mucosal microvascular hemoglobin O2 saturation (HbO2muc; tissue reflectance spectrophotometry) were investigated in a hemodynamically stable pig model. Twenty-one pigs were anesthetized, paralyzed, and mechanically ventilated. After laparotomy, a mesenteric venous catheter was inserted and a jejunal antimesenteric enterotomy performed. LPS-infused animals developed similar degrees of pulmonary hypertension. No differences in cardiac output and mean arterial blood pressure between groups were found. PO2muc and HbO2muc were significantly lower in M animals compared with control (C) [210 min; PO2muc: 7.12 +/- 1.81 (M), 19.01 +/- 3.12 mmHg (C); HbO2muc: 28.78 +/- 3.36 (M), 49.09 +/- 3.84% (C)], whereas S animals ranged in between (PO2muc: 13.36 +/- 2.2 mmHg; HbO2muc: 40.68 +/- 4.43%). Of measured PO2muc values, 12.6 (C), 20.6 (S), and 46.3% (M) ranged from 0 to 5 mmHg. PO2ser was lower in LPS animals compared with control [59.43 +/- 5.4 (C), 45.00 +/- 6.12 (S), 47.33 +/- 4.34 (M) mmHg]. Dopamine increased PO2muc and HbO2muc to similar absolute values and significantly decreased frequency of PO2muc (0-5 mmHg) in M animals. We conclude that LPS impairs mucosal tissue oxygenation independently of systemic hemodynamics. Mucosal microvascular dysfunction depends on regional LPS concentrations. Under conditions of compromised tissue oxygenation, dopamine significantly improves PO2muc and HbO2muc.
Role of prostaglandins and nitric oxide in gastrointestinal hyperemia of diabetic rats
Authors: Goldin E. , Casadevall M. , Mourelle M. , Cirera I. , Elizalde J. I. , Panes J. , Casamitjana R. , Guth P. , Pique J. M. , Teres J.
Published: December 2017

The aim of the study was to characterize the gastric and mesenteric vascular changes induced by diabetes and the implication of endothelial [nitric oxide (NO) and prostaglandins] and humoral (glucagon) factors in such changes. Diabetes was induced in rats by a single streptozotocin injection. Four weeks later, gastric mucosa, left gastric artery, and superior mesenteric artery blood flows were measured using hydrogen gas clearance and perivascular ultrasonic flowmeter techniques, respectively, in anesthetized and fasted diabetic and control rats. Blood pressure, hematocrit, blood volume, and blood viscosity were also measured. Left gastric (41 +/- 6 vs. 25 +/- 4 ml.min-1.100 g-1) and superior mesenteric artery blood flows (83 +/- 8 vs. 65 +/- 4 ml.min-1.100 g-1) were significantly higher in diabetic than in control rats. The increased blood flow in the left gastric artery was distributed to a hypertrophic mucosa in diabetic rats; therefore, the blood flow per 100 g tissue in the gastric mucosa was not significantly different in diabetic compared with control rats. Pretreatment with indomethacin reduced both increase gastric and mesenteric flows of the diabetic rats to the same levels as in control rats. NG-nitro-L-arginine methyl ester decreased gastric blood flow in a dose-dependent manner and to a similar extent in diabetic and control rats. In contrast, an increased sensitivity to the higher doses of the NO inhibitor was observed in the mesenteric vascular bed of diabetic rats. Glucagon reduction achieved by somatostatin infusion did not influence either gastric or mesenteric blood flow in diabetic rats. In summary, the present study revealed an increase in gastric and mesenteric arterial blood flows in streptozotocin-induced diabetic rats. The gastrointestinal hyperemia seems to be due, at least in part, to the increased demand of a hypertrophic mucosa and is mediated primarily by endogenous prostaglandins. Increased vascular sensitivity to NO may also contribute to the mesenteric vasodilation.
Effects of extracellular ATP on hepatic fatty acid metabolism
Authors: Guzman M. , Velasco G. , Castro J.
Published: December 2017

Incubation of rat hepatocytes with extracellular ATP inhibited acetyl-CoA carboxylase (ACC) activity and fatty acid synthesis de novo, with a concomitant decrease of intracellular malonyl-CoA concentration. However, both carnitine O-palmitoyltransferase I (CPT-I) activity and ketogenesis from palmitate were inhibited in parallel by extracellular ATP. The inhibitory effect of extracellular ATP on ACC and CPT-I activities was not evident in Ca2+ -depleted hepatocytes. Incubation of hepatocytes with thapsigargin, 2,5-di-(t-butyl)-1,4-benzohydroquinone (BHQ), or A-23187, compounds that increase cytosolic free Ca2+ concentration ([Ca2+]i), depressed ACC activity, whereas CPT-I activity was unaffected. The phorbol ester 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (PMA) increased ACC activity, whereas it decreased CPT-I activity in a nonaddictive manner with respect to extracellular ATP. The inhibitory effect of extracellular ATP on ACC activity was also evident in the presence of bisindolyl-maleimide, a specific inhibitor of protein kinase C (PKC), whereas this compound abolished the extracellular ATP-mediated inhibition of CPT-I. In addition, the PMA-induced inhibition of CPT-I was not potentiated by thapsigargin, BHQ, or A-23187. Results thus show 1) that the intracellular concentration of malonyl-CoA is not the factor responsible for the inhibition of hepatic long-chain fatty acid oxidation by extracellular ATP, and 2) that the inhibition of ACC by extracellular ATP may be mediated by an elevation of [Ca2+]i, whereas CPT-I may be inhibited by extracellular ATP through a PKC-dependent mechanism.
Defect in pancreatic exocrine and endocrine response to CCK in genetically diabetic OLETF rats
Authors: Tachibana I. , Akiyama T. , Kanagawa K. , Shiohara H. , Furumi K. , Watanabe N. , Otsuki M.
Published: December 2017

Clinical as well as experimental studies in insulinopenic diabetes mellitus have demonstrated abnormal pancreatic exocrine responses to cholecystokinin (CCK). In the present study, we examined pancreatic exocrine and endocrine function in the recently developed genetically diabetic Otsuka Long-Evans Tokushima fatty (OLETF) rats and compared them with those in the control Long-Evans Tokushima Otsuka (LETO) rats of the same age. Stepwise increasing doses of CCK octapeptide (CCK-8; 0.027-7.0 nmol.kg-1.h-1) evoked a characteristic biphasic dose-response curve for pancreatic juice and protein output in the LETO rats, whereas the OLETF rats were totally insensitive to CCK-8 stimulation. However, the responsiveness and the sensitivity to both carbamylcholine and secretin were similar in the two groups. Intraduodenal infusion of casein (500 mg/h) failed to stimulate pancreatic exocrine secretion in the OLETF rats despite a greater CCK response than in the LETO rats (peak response: 8.43 +/- 0.97 vs 5.12 +/- 0.30 pmol/l in LETO, P < 0.01). Intravenous infusion of CCK-8 (4.4 nmol.kg-1.20 min-1) caused a significant increase in serum insulin concentrations and a concomitant decrease in glucose levels in the LETO rats but not in the OLETF rats. On the other hand, an intravenous bolus injection of 1.1 mmol/kg glucose caused a greater insulin release in the OLETF rats than in the LETO rats. In contrast, gastric acid secretion in the OLETF rats was significantly high in basal and in response to intravenous infusion of CCK-8 compared with that in the LETO rats. Four subcutaneous injections of 20 micrograms/kg caerulein at hourly intervals over 3 h induced acute pancreatitis in the LETO rats but did not elicit any significant increase in serum amylase or lipase activities and pancreatic wet weight or histological evidence of acute pancreatitis in the OLETF rats. These results indicate that the exocrine and endocrine pancreas of the recently developed genetically diabetic OLETF rats are totally and specifically insensitive to exogenous and endogenous CCK stimulation, whereas parietal cells in these rats are sensitive to CCK stimulation.
Taurocholate induces directional excretion of bilirubin into bile in perfused rat liver
Authors: Yamaguchi T. , Wakabayashi Y. , Tanaka M. , Sano T. , Ishikawa H. , Nakajima H. , Suematsu M. , Ishimura Y.
Published: December 2017

The role of taurocholate, one of the major conjugated bile acids present in portal blood, in excretion of bilirubin from liver parenchyma to biliary and vascular compartments was studied in isolated perfused rat liver. Contents of bilirubin and carbon monoxide (CO) in the bile or venous effluents were determined by enzyme-linked immunosorbent microassay with the use of antibilirubin monoclonal antibody 24G7 and myoglobin-assisted spectrophotometry, respectively. In the presence of taurocholate, bilirubin excreted into the biliary compartment constituted greater than 90% of the total bilirubin excreted from the liver (0.26 nmol.min-1.g liver-1), corresponding to 60% of the outflow of CO into the venous effluents. In its absence, however, the total amount of bilirubin excreted into extrahepatic compartments was reduced to 27% of CO flux, and more than 90% of the excreted bilirubin was in the venous effluent. Thus a choleretic bile acid such as taurocholate is necessary for directional transport of bilirubin into bile in the perfused liver preparation.
Linkage of [Ca2+]i in single isolated D cells to somatostatin secretion induced by cholecystokinin
Authors: DelValle J. , Wakasugi J. , Takeda H. , Yamada T.
Published: December 2017

The Ca2+/inositol phospholipid signaling cascade has been implicated in the mechanism by which cholecystokinin (CCK) stimulates gastric somatostatin release, but a direct linkage between intracellular events in gastric D cells and somatostatin secretion has not been established. To address this problem we developed a method for correlating somatostatin release with the measurement of intracellular Ca2+ concentration ([Ca2+]i) in isolated D cells. Resting [Ca2+]i in single D cells was 100 +/- 5.7 nM (means +/- SE, n = 41), and CCK induced a rise in [Ca2+]i in a dose-dependent fashion, producing a maximal stimulatory effect (243 +/- 15% of control, n = 12) at a peptide concentration of 2 x 10(-8) M. The CCK-mediated increase in [Ca2+]i was biphasic, with a rapid, initial transient elevation followed by a sustained plateau. The rise in [Ca2+]i was accompanied by a concomitant increase in release of somatostatin-like immunoreactivity (SLI). Removal of extracellular Ca2+ had no effect on the initial transient elevation in [Ca2+]i induced by CCK but abolished both the sustained plateau in [Ca2+]i and the release of SLI. The selective CCK antagonist L-364, 718 (10(-7) M) inhibited the effects of CCK on both [Ca2+]i and SLI release. The nonspecific Ca2+ channel blocker NiCl2 (10(-3) M) and the L-type Ca2+ channel blocker nifedipine inhibited the sustained rise in [Ca2+]i and the release of SLI but left the initial transient increase in [Ca2+]i unaltered. These results indicate that CCK-stimulated release of SLI from D cells in the gastric fundus is linked to influx of extracellular Ca2+ via L-type Ca2+ channels.
Okadaic acid disrupts Golgi structure and impairs enzyme synthesis and secretion in the rat pancreas
Authors: Waschulewski I. H. , Kruse M. L. , Agricola B. , Kern H. F. , Schmidt W. E.
Published: December 2017

Okadaic acid, a serine/threonine phosphatase inhibitor, has been shown to inhibit rat pancreatic enzyme secretion by interference with late processes in stimulus-secretion coupling. To further characterize its action, we studied the effect of okadaic acid on secretion of newly synthesized proteins, protein synthesis, and cellular ultrastructure in pancreatic lobules derived from rats stimulated in vivo by feeding the synthetic proteinase inhibitor FOY-305. Okadaic acid completely blocked protein secretion at concentrations that inhibit the Ca2+/calmodulin-dependent protein phosphatase 2b, calcineurin. Protein synthesis was abolished at 10(-6) mol/l and reduced by 60% at 5 x 10(-7) mol/l okadaic acid. Pancreatic lobules exposed to 5 x 10(-7) mol/l okadaic acid for 20 min fully restored their secretory capacity on removal of the drug; whereas, after a preincubation with okadaic acid for > 40 min, protein secretion remained impaired during the recovery period. Electron microscopic examination of pancreatic acinar cells treated with 5 x 10(-7) mol/l okadaic acid revealed a dilated Golgi complex after 15 and 30 min and a subsequent fragmentation of Golgi cisternae into clouds of small uniform vesicles after 60 min. Reassembly of Golgi stacks occurred after a 60-min recovery without okadaic acid. These data indicate that serine/threonine phosphatases play an important role not only in the regulation of pancreatic enzyme synthesis and exocytosis but also are crucial for the maintenance of normal Golgi architecture and function in the exocrine rat pancreas. These effects are probably not exclusively mediated via type 2b calcineurin-like protein phosphatases.
Inhibition of rat sinusoidal GSH transporter by thioethers: specificity, sidedness, and kinetics
Authors: Fernandez-Checa J. C. , Garcia-Ruiz C. , Colell A. , Yi J. R. , Kaplowitz N.
Published: December 2017

In isolated hepatocytes, cystathionine, methionine, and thioether analogues of methionine, cysteine, and homocysteine, including S-adenosyl derivatives, inhibited reduced glutathione (GSH) efflux. The potency of inhibition by thioethers with different S-alkyl moieties was methyl < ethyl < butyryl < aminoethyl < alpha-aminopropionyl. Inhibition of GSH efflux by cystathionine from hepatocytes that were allowed to resynthesize GSH resulted in greater repletion (30-40%) of GSH levels compared with absence of cystathionine. To address unequivocally the sidedness of inhibition, i.e., cis vs. trans, we examined the effect of cystathionine on the activity of GSH transport in Xenopus oocytes expressing the cRNA of cloned rat liver sinusoidal (RsGshT) and canalicular (RcGshT) GSH transporters. Cystathionine trans inhibited efflux of GSH and cis inhibited uptake of GSH by oocytes expressing RsGshT. Conversely, when oocytes expressing RsGshT were loaded with cystathionine, no inhibition of uptake or efflux was observed. The same structural requirement of a thioether bond to exert an inhibitory effect on GSH transport was observed in oocytes expressing RsGshT. Oocytes expressing RsGshT do not transport methionine, whereas oocytes expressing total rat liver mRNA express methionine transport. Inhibition of both GSH efflux from and uptake by oocytes expressing RsGshT exhibited a competitive type of kinetics: cystathionine increased the Michaelis constant for GSH transport (4.5 +/- 0.9 vs. 10 +/- 2.5 mM and 7.5 +/- 0.6 vs. 12.9 +/- 1.5 mM for uptake and efflux, respectively) without affecting the maximal velocity for transport. Thus thioethers such as methionine and cystathionine inhibit the transport of GSH by interacting in a competitive and specific fashion with the sinusoidal GSH transporter without themselves being transported by this carrier.
Regulation of sucrase-isomaltase and hexose transporters in Caco-2 cells: a role for cytochrome P-4501A1?
Authors: Carriere V. , Barbat A. , Rousset M. , Brot-Laroche E. , Dussaulx E. , Cambier D. , De Waziers I. D. , Beaune P. , Zweibaum A.
Published: December 2017

Involvement of cytochrome P-4501A1 (CYP1A1) in the regulation of sucrase-isomaltase and hexose transporters was analyzed in low (TC7)- and high (PF11)-glucose-consuming Caco-2 clones. CYP1A1 mRNA is elevated in exponentially growing cells concomitantly with high rates of glucose consumption and high levels of GLUT-1 and GLUT-3 mRNA. After confluency, CYP1A1 is not detectable in TC7 cells; this is associated with a decreased glucose consumption, a downregulation of GLUT-1 and GLUT-3, and an upregulation of sucrase-isomaltase, SGLT-1, GLUT-2, and GLUT-5. In PF11 cells CYP1A1 mRNA remains elevated, along with high glucose consumption, high levels of GLUT-1 and GLUT-3, and minimal expression of sucrase-isomaltase, SGLT-1, GLUT-2, and GLUT-5. Exposure of TC7 cells to inducers of CYP1A1 results in high levels of CYP1A1 mRNA, a 10-fold increase of glucose consumption after confluency, an upregulation of GLUT-1 and GLUT-3, and a downregulation of sucrase-isomaltase, GLUT-2, and, to a lesser extent, SGLT-1 and GLUT-5. These results suggest that activation of CYP1A1, whether spontaneous or drug induced, is involved in the variations of glucose utilization and in the associated modifications of expression of sucrase-isomaltase and hexose transporters.
Expression of PRL-1 nuclear PTPase is associated with proliferation in liver but with differentiation in intestine
Authors: Diamond R. H. , Peters C. , Jung S. P. , Greenbaum L. E. , Haber B. A. , Silberg D. G. , Traber P. G. , Taub R.
Published: December 2017

Mechanisms controlling the tyrosine phosphorylation of cellular proteins are important in the regulation of cellular processes including growth and differentiation. It has become clear that a number of protein tyrosine phosphatases (PTPases) that dephosphorylate tyrosyl residues may play a role in the growth response, both in growth-promoting and growth-inhibiting capacities. We identified PRL-1, a unique nuclear PTPase that is an immediate-early gene in liver regeneration and is positively associated with growth, including fetal and neoplastic hepatic growth and anchorage-independent growth after overexpression in fibroblasts. In this study, we show that PRL-1 nuclear protein levels in regenerating liver parallel those of its mRNA, although the peak occurs later, just before the onset of DNA synthesis. We further show that PRL-1 is significantly expressed in intestinal epithelia and that, in contrast to the expression pattern of PRL-1 in liver, its expression is associated with cellular differentiation in intestine. Specifically, PRL-1 is expressed in villus but not crypt enterocytes and in confluent differentiated but not undifferentiated proliferating Caco-2 colon carcinoma cells. The expression of PRL-1 in intestine shows inverse correlation with proliferating cell nuclear antigen expression, a marker for S-phase cells. These results suggest that PRL-1 may play different roles in these two digestive tissues. Such a dichotomy of roles has previously been described for some protein tyrosine kinases and might be due to the availability of alternate substrates in different tissues.
Pharmacokinetics and organ metabolism of carboxyamidated and glycine-extended gastrins in pigs
Authors: Hansen C. P. , Stadil F. , Yucun L. , Rehfeld J. F.
Published: December 2017

The elimination of carboxyamidated gastrin-17 and its glycine-extended precursor was studied in anesthetized pigs during constant-rate infusion. Extraction of amidated gastrin-17 was recorded in the hindlimb (42%), kidney (40%), head (32%, P < 0.001), and the gut (13%, P < 0.01). Elimination was not recorded in the liver, lungs, or heart. Extraction of glycine-extended gastrin-17 was measured in the kidney (36%), hindlimb (31%, P < 0.001), head (26%), and the gut (16%, P < 0.01), but not in the liver or the lungs. Glycine-extended gastrin-17 was not processed to amidated gastrin during infusion. The half-life, metabolic clearance rate, and apparent volume of distribution for amidated gastrin-17 were 3.5 +/- 0.4 min, 15.5 +/- 1.1 ml.kg-1.min-1, and 76.5 +/- 9.9 ml/kg, respectively, and for glycine-extended gastrin-17 were 4.3 +/- 0.6 min, 17.4 +/- 0.9 ml.kg-1.min-1, and 104.7 +/- 11.9 ml/kg, respectively. We conclude that extraction of amidated and glycine-extended gastrin-17 varies in the vascular beds, with elimination mainly confined to nonorgan tissues and the kidneys.
Adaptations of intestinal nutrient transport to chronic caloric restriction in mice
Authors: Casirola D. M. , Rifkin B. , Tsai W. , Ferraris R. P.
Published: December 2017

Lifelong caloric restriction increases median and maximum life span and retards the aging process in many organ systems of rodents. Because the small intestine absorbs a reduced amount of nutrients each day, does lifelong caloric restriction induce adaptations in intestinal nutrient transport? We initially compared intestinal transport of sugars and amino acids between 24-mo-old mice allowed free access to food [ad libitum (AL)] and those provided a calorically restricted [40% less than ad libitum (CR)] diet since 3 mo of age. We found that CR mice had significantly greater transport rates for D-glucose, D-fructose, and several amino acids and had significantly lower villus heights. Total intestinal absorptive capacities for D-glucose, D-fructose, and L-proline were each 40-50% greater in CR mice; absorptive capacity normalized to metabolic mass (body weight 0.75) was approximately 80% greater in CR mice. Comparison of uptakes in aged AL and CR mice with previously published results in young AL mice suggests that caloric restriction delays age-related decreases in nutrient transport. In contrast to published studies in hibernation and starvation, chronic caloric restriction enhances not only uptake per milligram but also uptake per centimeter. We then switched 24-mo-old AL mice to a calorie-restricted diet for 1 mo and found that short-term caloric restriction has no effect on intestinal nutrient transport, intestinal mass, and total absorptive capacity. Thus chronic but not short-term caloric restriction increases intestinal nutrient transport rates in aged mice, and the main mechanism underlying these increases is enhanced transport rates per unit intestinal tissue weight.
Dynamics of arterial and portal venous flow interactions in perfused rat liver: an intravital microscopic study
Authors: Sherman I. A. , Dlugosz J. A. , Barker F. , Sadeghi F. M. , Pang K. S.
Published: December 2017

Intravital epifluorescent microscopy was used to quantitate microvascular parameters in the single-pass, dually perfused rat liver preparation. Livers perfused via the hepatic artery (HA) and portal vein (PV) at physiological pressures and perfusion rates responded to vasoactive agents and exhibited the HA buffer response. The distribution of arterial blood was found to be highly heterogeneous, whereas PV flow was distributed uniformly. The intrasinusoidal velocity of fluorescein isothiocyanate (FITC)-labeled red blood cells (RBCs) arriving from the HA was higher than that for RBCs arriving from the PV, indicating a shorter transit time for the arterially delivered FITC-RBCs. Experiments on livers perfused simultaneously via the HA and retrogradely via the hepatic vein revealed the presence of arteriovenous shunts, with some of the arterially delivered FITC-RBCs reaching the terminal hepatic venules via direct channels without traversing the sinusoidal bed. In livers perfused portally only, changes in PV flow rate (from 8 to 20 ml/min) produced small changes in perfusion pressure but large changes in vascular diameters, while portal pressure and transit time of portal blood remained relatively constant. In experiments designed to identify the location of hepatic vascular resistance, it was observed that hepatic venular diameters measured in the preparation under identical pressure and flow conditions were greater during retrograde than during prograde perfusion, suggesting that the site of hepatic vascular resistance is presinusoidal or sinusoidal.
Hexose transporter expression in rat small intestine: effect of diet on diurnal variations
Authors: Corpe C. P. , Burant C. F.
Published: December 2017

In rodents, a number of intestinal digestive and absorptive processes demonstrate a diurnal pattern of activity. To investigate if the jejunal hexose transporters are regulated in such a diurnal fashion, the levels for the glucose and fructose transporter mRNA and proteins were determined at 6-h intervals over a 24-h control fed period. SGLT-1, GLUT-2, and GLUT-5 mRNA levels increased between two- and eightfold before the onset of peak feeding. GLUT-5 protein levels also varied in a diurnal fashion but were out of phase with the observed changes in GLUT-5 mRNA levels. In contrast, GLUT-2 protein levels remained relatively constant during the control fed 24-h period. The effect of dietary manipulations on the observed diurnal variation was also investigated. After only 3 h of feeding a 60% fructose-enriched diet, the levels of GLUT-5 mRNA and protein were significantly elevated. GLUT-5 mRNA and protein levels remained elevated relative to the level of control diet-fed animals over the ensuing 24 h and during the 7th day of fructose feeding. Exposure to elevated levels of fructose had no significant effect on the diurnal pattern of GLUT-2 and SGLT-1 mRNA. In contrast, GLUT-2 protein was rapidly downregulated during the length of the fructose feeding study. In conclusion, the data demonstrate a normal daily variation in the level of hexose transporter expression that can be rapidly modulated by diet.
Positional specificity of defensin gene expression reveals Paneth cell heterogeneity in mouse small intestine
Authors: Darmoul D. , Ouellette A. J.
Published: December 2017

Cryptdins are antimicrobial peptides of the defensin family that are expressed specifically by Paneth cells in small intestinal crypts (M.E. Selsted, S.I. Miller, A.H. Henschen, and A.J. Ouellette. J. Cell Biol. 118: 929-936, 1992), and at least 17 cryptdin isoforms have been reported in mouse small intestine (A.J. Ouellette, M.M. Hsieh, M.T. Nosek, D.F. Cano-Gauci, K.M. Huttner, R.N. Buick, and M.E. Selsted. Infect. Immun. 62: 5040-5047, 1994). Analysis of cryptdin gene expression in adult mouse small bowel revealed that the cryptdin-4 isoform is differentially expressed along the proximal-to-distal intestinal axis. By peptide-specific reverse transcriptase-polymerase chain reaction-based assays, cryptdin-4 mRNA was found to be absent from the proximal small bowel, increasing to maximal levels in the ileum. In contrast, intestinal content of cryptdin-1 and -5 mRNAs was equivalent in duodenum, jejunum, and ileum, and Northern blot hybridization experiments were consistent with both sets of data. Similarly, individual crypts isolated from duodenum contain cryptdin-1 mRNA but not cryptdin-4 mRNA. Taken together, the results show that Paneth cells are heterogeneous, depending on their position along the longitudinal axis of the small bowel. The positional specificity of defensin gene expression suggests that cryptdins may be useful markers for investigating the establishment and maintenance of this epithelial lineage in the mouse small intestine.
Ca(2+)-activated C1- channels in a human biliary cell line: regulation by Ca2+/calmodulin-dependent protein kinase
Authors: Schlenker T. , Fitz J. G.
Published: December 2017

Biliary epithelial cells contribute to bile formation through absorption and secretion of fluid and electrolytes. Recent studies indicate that membrane Cl- permeability is regulated in part by increases in intracellular Ca2+ concentration. The purpose of these studies was to evaluate the effects of intracellular Ca2+ on channel activity, using the human Mz-ChA-1 cholangiocarcinoma cell line as a model, and to assess the possible roles of Ca(2+)-dependent kinases in channel regulation. Exposure to ionomycin (1 microM) activated ion channels in the cell-attached configuration in 63 of 74 attempts, increasing open probability (NPo) from 0 to 0.26 +/- 0.15 (n = 17). Multiple channels were present in each patch, and the effects of ionomycin were reversed by subsequent addition of ethylene glycol-bis (beta-aminoethyl ether)-N,N,N',N'- tetraacetic acid (2 mM) to the bath. With Cl(-)-containing solutions, channels had a slope conductance of 14 +/- 4 pS (n = 11), and the mean open time was estimated to be 5.3 +/- 1.8 ms. These channels were anion selective, and currents were carried by efflux of Cl- at the resting potential. Exposure to the Ca2+/calmodulin-dependent protein kinase II (CaMKII) antagonist calmidazolium (100 microM) decreased NPo in ionomycin-stimulated cells to 0.02 +/- 0.06 (n = 19). The protein kinase C antagonist chelerythrine (50 microM) was without effect. In parallel studies in subconfluent cell monolayers, CaMKII antagonists were also potent inhibitors of ionomycin-stimulated 125I efflux. These findings indicate that Ca(2+)-dependent increases in membrane Cl- permeability are related in part to opening of 14.pS anion channels through a mechanism that depends on both Ca2+ and CaMKII. These channels represent a potential target for pharmacological modulation of biliary cell transport and function.
Analysis of cloned cDNAs differentially expressed in adapting remnant small intestine after partial resection
Authors: Dodson B. D. , Wang J. L. , Swietlicki E. A. , Rubin D. C. , Levin M. S.
Published: December 2017

After partial resection, the remnant small intestine undergoes an adaptive response. Little is known about the molecular and cellular basis of intestinal adaptation. To identify genes transcriptionally regulated in response to loss of functional bowel surface area, we have isolated cDNAs differentially expressed in the adaptive ileum 48 h after 70% proximal small intestinal resection. A cDNA library constructed from the remnant ileum of rats subjected to resection was screened using subtractive hybridization techniques. Several groups of cDNA clones that were induced during intestinal adaptation were isolated. The first included liver fatty acid binding protein, apolipoprotein A-IV, cellular retinol binding protein II, and ileal lipid binding protein. These all encode proteins involved in the absorption, metabolism, and trafficking of nutrients. A second group included the catalytic subunit of protein phosphatase 1 delta, a 78-kDa glucose-regulated protein (grp 78; a glucose-regulated member of the 70-kDa heat-shock protein family), and several pancreatitis-associated proteins. A third group of induced genes contained novel cDNAs. To better characterize the adaptive response, the temporal, spatial, and cellular patterns of expression of several of these genes were analyzed with the use of immunohistochemical and in situ hybridization techniques. These studies indicate that during early adaptation, genes involved in nutrient trafficking, protein processing, and cell cycle regulation are transcriptionally regulated in the residual small intestine in distinct temporal and regional patterns consistent with a complex multifaceted response to intestinal resection.
Nongenomic effects of progesterone on human intestinal smooth muscle cells
Authors: Bielefeldt K. , Waite L. , Abboud F. M. , Conklin J. L.
Published: December 2017

Previous experiments demonstrated that progesterone affects intestinal smooth muscle cells through genomic and nongenomic pathways. We hypothesized that the nongenomic effect was mediated by changes in membrane excitability. We studied the effects of progesterone and other steroid hormones on a human intestinal smooth muscle cell line, using the whole cell patch-clamp technique. Ionic currents were elicited through steps from -70 mV to various test potentials. Progesterone dose-dependently reduced calcium currents. The decrease in inward current was partly due to a shift in the steady-state inactivation to more hyperpolarized potentials. This effect did not involve gene transcription, since it was not blocked by the progesterone antagonist ZK-98-299. The progesterone analogue 5-beta-dihydroprogesterone also decreased calcium currents, whereas its stereoisomer, 5-alpha- dihydroprogesterone, did not affect the properties of voltage-sensitive ion channels. Similarly, estradiol and dexamethasone did not alter inward currents. We conclude that progestins exert their nongenomic effects on intestinal smooth muscle cells by decreasing calcium currents. The change in the calcium signal may contribute to the reduction in muscle contraction observed after progesterone.
Autoregulatory capacity in the superior mesenteric artery is attenuated by nitric oxide
Authors: Macedo M. P. , Lautt W. W.
Published: December 2017

The hypothesis that nitric oxide antagonizes pressure-flow autoregulation in the superior mesenteric artery was tested with the use of a nitric oxide synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), and L-arginine. Autoregulation was assessed with the use of two indexes: 1) the autoregulatory index (ARI), expressed as the ratio of change in flow to change in pressure tested over the nonautoregulatory range, 40-70 mmHg, and over the autoregulatory range, 70-140 mmHg; and 2) the slope index, an index of linearity of the pressure-flow curve calculated by dividing the slope of the pressure-flow curve over the autoregulatory range by the slope of the curve over the nonautoregulatory range, expressed as percent. L-NAME significantly increased autoregulation from an ARI of 0.06 +/- 0.05 to 0.28 +/- 0.09 over the autoregulatory range (P < 0.003), and the nitric oxide synthase substrate L-arginine reversed ARI to 0.12 +/- 0.07 (mean +/- SE, n = 7). ARI in the nonautoregulatory range was not altered. The slope index revealed that L-NAME enhanced autoregulation (61 +/- 5.6 compared with control, 93.4 +/- 7.9; P < 0.003), which was reversed by the action of L-arginine (88.5 +/- 5.5, P < 0.007). The data were consistent with the hypothesis that increased flow leads to nitric oxide-induced dilation, which counteracts autoregulation.
Bidirectional transport of cholesterol between gallbladder epithelial cells and model bile
Authors: Hayashi A. , Lee S. P.
Published: December 2017

Lipids in hepatic bile may be modified by the gallbladder epithelium. The purpose of this study was to demonstrate a bidirectional exchange of cholesterol between biliary lipid carriers and gallbladder epithelial cells and to determine the factors regulating this cholesterol transfer. Gallbladder epithelial cells were cultured to confluent monolayers, their membranes were labeled with endogenously synthesized [14C]cholesterol, and the cells were incubated with model bile introduced into the apical membrane compartment. Similarly, model bile with different lipid composition containing [3H]cholesterol was incubated with the unlabeled monolayers. We found that cholesterol in the apical membrane bilayer of the epithelial cells exchanged readily with that in bile, but only in the presence of bile salts. The rate of exchange is dependent on the concentration and species of bile salts. The net gain of cholesterol (absorption) or net loss of cholesterol (cytotoxicity) exhibited by the epithelial cells was regulated by the thermodynamic stability of cholesterol and the detergent effect of mixed micelles in bile. It is also possible that the physicochemical composition of lipids in bile may modify the cellular function of the gallbladder epithelium.
Neural control of canine small intestinal motility during nonnutrient infusion
Authors: Behrns K. E. , Sarr M. G. , Hanson R. B. , Zinsmeister A. R.
Published: December 2017

Our aim was to determine the effect of in situ neural isolation of the jejunoileum (extrinsic denervation and disruption of enteric neural continuity with the duodenum) on the spread of single pressure waves (SPW) and clustered contractions (CC) in response to increasing rates of isolated duodenal and jejunoileal nonnutrient infusions. Ten dogs were prepared with duodenal and jejunal infusion and manometry catheters and a diverting proximal jejunal cannula. Five of the dogs also underwent in situ neural isolation of the entire jejuno- ileum A noncaloric solution was infused at 0-15 ml/min into proximal duodenum or jejunum while manometric data were collected. Alterations in direction, distance, and velocity of spread of SPW and CC with increasing rates of intestinal infusion were analyzed by linear regression of responses to increasing infusion rates. Neural isolation of the jejunoileum did not markedly alter characteristics of duodenal or jejunal SPW or CC under conditions of no intestinal infusion. After neural isolation of jejunoileum, increasing rates of jejunal infusion decreased both the proportion and distance of antegrade spread of SPW in duodenum. These findings suggest that extrinsic innervation to the jejunoileum and enteric neural continuity with the duodenum do not regulate jejunal SPW or CC. Increasing rates of nonnutrient intestinal infusions do not alter local motor patterns in the innervated or neurally isolated jejunum, but after neural isolation of the jejunoileum, these infusions do alter characteristics of duodenal SPW by mechanisms independent of neural pathways.
Decreased ileal muscle contractility and increased NOS II expression induced by lipopolysaccharide
Authors: Weisbrodt N. W. , Pressley T. A. , Li Y. F. , Zembowicz M. J. , Higham S. C. , Zembowicz A. , Lodato R. F. , Moody F. G.
Published: December 2017

This study was designed to determine if an increase in nitric oxide synthase (NOS) activity induced by lipopolysaccharide (LPS) is associated with increases in NOS II protein and mRNA abundance and with altered ileal longitudinal muscle contractility. Strips of muscle taken from LPS-treated, but not control, animals exhibited reduced in vitro contractility when L-arginine was a component of the physiological salt solution. This reduction was reversed by N omega-nitro-L-arginine (L-NNA), a competitive inhibitor of NOS. Full-thickness segments of jejunum, ileum, and colon taken 5 h after LPS injection exhibited increased NOS activity, NOS II immunoreactivity, and NOS II mRNA abundance. Increased NOS II immunoreactivity and mRNA abundance also were detected in ileal muscle strips taken from LPS-treated animals. These data confirm the reported effects of LPS on intestinal NOS activity and indicate that it can be attributed, at least in part, to an increase in NOS II mRNA and protein abundance. Furthermore, the data suggest that an LPS-induced increase in NOS II may lead to a decrease in ileal muscle contractility.
Effect of luminal epidermal growth factor on enterocyte glucose and proline transport
Authors: Hardin J. A. , Wong J. K. , Cheeseman C. I. , Gall D. G.
Published: December 2017

The effect of luminal epidermal growth factor (EGF; 60 ng/ml) and tyrphostin-51 (TYR; 10 microM), a tyrosine kinase inhibitor, on rabbit jejunal brush-border and basolateral membrane transport was investigated. In separate experiments, the effect of EGF, EGF and TYR, or TYR alone was examined in in vivo loops. In addition, Na+ permeability in brush-border membrane vesicles (BBMV) and the effect of Ca2+ channel blockade on EGF-stimulated glucose uptake were examined. Luminal EGF significantly (P < 0.0001) increased the maximal rate of transport (Vmax) for glucose and proline uptake in BBMV. TYR and Ca2+ channel blockade completely abolished the EGF-induced increase in glucose transport and in the case of TYR resulted in a significant reduction in Vmax compared with controls (P < 0.0001). The Michaelis-Menten constant did not differ in any experimental group. EGF had no effect on brush-border Na+ permeability or basolateral membrane glucose transport. The findings indicate a role for EGF in the acute regulation of jejunal brush-border membrane nutrient uptake. Furthermore, tyrosine kinase activity appears to be involved both in mediating EGF-induced alterations in transport function and in the maintenance of basal brush-border membrane function.
Pituitary adenylate cyclase-activating polypeptide stimulates cholecystokinin secretion in STC-1 cells
Authors: Chang C. H. , Chey W. Y. , Braggins L. , Coy D. H. , Chang T. M.
Published: December 2017

Secretion of cholecystokinin (CCK) from the endocrine cells of small intestinal mucosa and the murine intestinal tumor cell line STC-1 is known to involve both adenosine 3',5'-cyclic monophosphate (cAMP)-and Ca(2+)-dependent signal transduction pathways. However, the endogenous stimulant(s) that acts through the cAMP-dependent cascade has not been identified. We determined the effect of pituitary adenylate cyclase-activating polypeptide (PACAP) on CCK secretion and cAMP production and its interaction with other CCK secretagogues in STC-1 cells. At concentrations > 10 nM, PACAP-27 stimulated the release of large intestinal CCK from STC-1 cells in a time- and dose-dependent manner. The stimulatory effect of PACAP-27 was enhanced by the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX). PACAP-27, PACAP-38, and vasoactive intestinal polypeptide (VIP), with or without IBMX, were equally effective and potent to elicit CCK release with similar half-maximal doses and maximal levels of stimulation. Both forms of PACAP and VIP stimulated a transient but not significant increase in the cellular cAMP level. In the presence of IBMX, all three peptides increased significantly the cellular cAMP level between 2 and 5 min, but PACAP produced a two times higher level than VIP. The stimulatory effect of PACAP-27 on CCK release was also potentiated by bombesin and KCl but without a synergistic production of cAMP. With or without IBMX, PACAP-27-stimulated CCK secretion was not affected by the Ca2+ channel blocker diltiazem (1 microM), the cell-permeable Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester (BAPTA-AM; 25 microM), or by downregulation of protein kinase C. The stimulatory effects of KCl and bombesin were either reduced or abolished by these treatments. The synergistic effect of bombesin with PACAP was abolished by diltiazem and BAPTA-AM but not by downregulation of protein kinase C, whereas KCl remained synergistic with PACAP after these treatments. Taken together, these results indicate that PACAP may be a neuromodulator of CCK secretion that acts through activation of adenylate cyclase and may function as a coregulator with other CCK secretagogues that are known to increase intracellular Ca2+ concentration.
Inhibition of acid secretion by electrical acupuncture is mediated via beta-endorphin and somatostatin
Authors: Jin H. O. , Zhou L. , Lee K. Y. , Chang T. M. , Chey W. Y.
Published: December 2017

Electroacupuncture (EAP) was shown to inhibit basal gastric acid secretion in dogs and sham feeding-stimulated acid secretion in humans. However, its effect on a meal-stimulated acid secretion in dogs and the mechanisms involved remain unclear. In five dogs prepared with gastric cannulas, gastric acid secretion was determined by a dye-dilution technique for 60 min after intragastric administration of 200 ml of 4% mixed amino acid meal in six different experiments: study 1, no acupuncture; study 2, sham acupuncture (SAP); study 3, EAP; study 4, EAP plus naloxone; study 5, naloxone alone; and study 6, intravenous infusion of somatostatin (SS) and vasoactive intestinal peptide (VIP) at doses of 0.5 and 1.0 micrograms.kg-1.h-1, respectively. EAP was performed on three different points including Pishu, ZusanLi, and Neiguan. Biphasic electrical pulse (25-100 Hz, 12-16 mA) was applied continuously via needles for 75 min starting 15 min before meal. SAP on nonacupoints in hind- and forelegs was performed with the same electrical pulse. Plasma SS, VIP, beta-endorphin, and gastrin were determined by specific radioimmunoassays. EAP significantly inhibited acid secretion (75%; P < 0.01), which coincided with significant increases in plasma SS, VIP, and beta-endorphin and a significant decrease in plasma gastrin. Naloxone completely reversed EAP-induced inhibition of acid secretion and changes in plasma concentration of peptides. SAP also significantly suppressed acid output (30%; P < 0.05), with a modest but significant increase in plasma beta-endorphin. However, the inhibition by EAP on the acid output was significantly greater than that by SAP (P < 0.01). Furthermore, exogenous SS (0.5 microgram.kg-1.h-1) significantly inhibited acid output (78%), whereas VIP failed to inhibit gastric acid secretion. We conclude that, in dogs, EAP significantly inhibits meal-stimulated acid secretion. This acid inhibition is mediated by the release of beta-endorphin and somatostatin, and an endogenous opiate or opiates appear to play an important role in the release of SS, VIP, and beta-endorphin.
Hydrolysis of membrane-bound liver alkaline phosphatase by GPI-PLD requires bile salts
Authors: Deng J. T. , Hoylaerts M. F. , De Broe M. E. , van Hoof V. O.
Published: December 2017

Circulating liver plasma membrane fragments (LPMF) were purified from human serum by means of a monoclonal antileucine aminopeptidase antibody, AD-1. This was done by immunoaffinity chromatography or by incubating the sera with AD-1-coated nitrocellulose disks. Alkaline phosphatase (ALP, EC 3.1.3.1) is bound to these LPMF through a glycosylphosphatidylinositol (GPI) anchor and is referred to as membrane-bound liver ALP (Mem-LiALP). Low concentrations of Triton X-100 or high bile salt concentrations released GPI anchor-bearing LiALP (Anch-LiALP) from purified LPMF; once released, Anch-LiALP was slowly and progressively converted to hydrophilic dimeric LiALP [soluble LiALP (Sol-LiALP)], free from its GPI anchor. Low levels of GPI-specific phospholipase D (GPI-PLD) activity were measured in the pure LPMF. Apparently, this membrane-associated GPI-PLD was released by the action of detergents and contributed to the spontaneous conversion of Anch-LiALP to Sol-LiALP. In the absence of detergents, GPI-PLD had little effect on Mem-LiALP, both in purified form as well as in serum. In vitro, isolated Anch-LiALP was converted to Sol-LiALP by both GPI-specific phospholipase C and GPI-PLD. Sol-LiALP in serum, however, appeared to be the product of GPI-PLD activity only. Five- to tenfold higher concentrations of Triton X-100 were needed to release Anch-LiALP from LPMF in serum, compared with those required in a solution of purified LPMF. In serum, as well as in purified conditions, only a small range of detergent of bile salt concentrations permitted the conversion of Mem-LiALP to Sol-LiALP. A model is proposed for the release in the circulation of Mem-LiALP, Anch-LiALP, and Sol-LiALP, involving both LPMF-associated GPI-PLD and liver sinusoid bile salts.
Hyperglycemia affects proximal gastric motor and sensory function during small intestinal triglyceride infusion
Authors: Hebbard G. S. , Samsom M. , Sun W. M. , Dent J. , Horowitz M.
Published: December 2017

Hyperglycemia slows gastric emptying in normal individuals and patients with diabetes mellitus and may affect both somatic and visceral sensation. The effects of hyperglycemia on proximal gastric motility and sensation during intraduodenal infusion of a triglyceride emulsion were evaluated using a barostat in six normal subjects during euglycemia and hyperglycemia (approximately 15 mmol/l). Isobaric distension induced greater bag volumes during hyperglycemia compared with euglycemia at 3 (452 +/- 26 vs. 343 +/- 12 ml, P< 0.05) and 4 mmHg (600 +/- 55 vs. 497 +/- 50 ml, P < 0.05) above basal pressure. During isovolumetric distension, intrabag pressure was less during hyperglycemia at 500 (2.5 +/- 0.3 vs. 3.5 +/- 0.5 mmHg above basal pressure, P < 0.05) and 600 ml (3.0 +/- 0.4 vs. 4.5 +/- 0.5 mmHg above basal pressure, P < 0.05). Perception of nausea (P < 0.05) and fullness (P < 0.05) was increased during hyperglycemia compared with euglycemia. We conclude that hyperglycemia 1) reduces proximal gastric tone during intraduodenal triglyceride infusion, an effect that may contribute to delayed gastric emptying, and 2) increases the intensity of nausea and fullness during intraduodenal triglyceride infusion and proximal gastric distension, indicative of an effect on visceral sensation.