Sa2056 Comparison of the Effect of Prucalopride, Sodium Picosulfate and Placebo on Rectal Motor and...
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in the proximal colon, but was slightly elevated in the distal colon. In the presence of 5 μM propofol, the amplitude was significantly reduced in both distal and proximal colon (p<0.01, n=4). In both the distal and proximal colon, there was change in the frequency of the CMMCs in the presence of propofol. When both ondansetron and L-NNA were added to preparations in the presence of propofol, no differences were observed. In the presence of 1 μM propofol, fecal pellet motility was significantly reduced compared to control responses (p<0.001, n=5). Conclusions: Propofol significantly increased 5-HT levels in distal colonic but not the proximal mucosa and thus has site specific activity. This may in turn lead to the desensitization of 5-HT3Rs on IPANs. Such effects of propofol can explain the reduced amplitude of CMMCs in the proximal and distal colon and reduced fecal pellet motility. Such findings may provide more insight into intestinal side effects observed in patients utilising propofol for maintenance of anaesthesia and sedation and will provide guidance on approaches to therapeutically managed patients utilising this form of anaesthetic agent. Sa2054 Germ Free Mice Have Normal Epithelial Responses to Serotonin and Acetylcholine in the Proximal Colon John F. Rainey, David R. Linden, Madhusudan Grover, Charles E. Salmonson, Cheryl E. Bernard, Joseph H. Szurszewski, Gianrico Farrugia, Purna C. Kashyap Background: Gut microbiota play an important role in modulating gastrointestinal physiology including changes in intestinal transit and epithelial barrier function. Both serotonin (5HT) and acetylcholine (Ach) play important roles in regulation of gut function yet it is not known if absence of gut microbiota alters the host colonic responses to 5HT and acetylcholine. The aim of this to study was to determine host colonic epithelial function in the absence of gut microbiota. Methods: We used germ free (GF) and humanized mice (ex-GF colonized with human bacteria; HM). Two segments of proximal colon, stripped of external muscle layers from each animal were mounted in 0.3 cm2 area, 4 ml Ussing chambers. Colonic mucosal transepithelial resistance (TER) was measured at the beginning of each experiment and FITC Dextran (4 k Da) flux was determined from the linear fit of samples obtained from the serosal side every 5 min for 30 min after adding 1mg/ml FITC-Dextran to the mucosal side. Change in short circuit current (Δ Isc) was determined in response to increasing concentra- tions of ACh (0.01-300 μM) on the serosal side, 5HT (0.003-300μM) on the mucosal or serosal side and electrical field stimulation (EFS) using foil electrodes on the submucosal side (30-150V, 100 0.5ms pulses at 1-2Hz). Results: There were no differences in TER (50.2±1.7 vs 69.2±10.5V/cm2; n=4-5; p>0.05) or FITC dextran flux (2.4±0.5 vs 3.4±2 ng/ ml/min/cm2; n=4; p>0.05) between GF and HM mice, respectively. Cumulative concentration response curves induced by serosal ACh were not different between GF and HM as evident by similar Emax (261±38 vs 299±27μA/cm2; n=5; p>0.05) and EC50 (20.6±3 vs 22.7±9μM) values. Cumulative concentration response curves induced by serosal 5-HT were also not different between GF and HM (Emax: 210±14 vs 159±35μA/cm2; n=4; p>0.05) and EC50 (2.3±1.9 vs 2.6±1μM). While Δ Isc did not reach maximum response following mucosal application of 300μM 5HT, responses were similar between GF and HM(90±15 vs 85±2027μA/cm2; n=5; p>0.05) . There was no significant difference in Δ Isc between GF and HM in response to EFS at 1 Hz (138±42 vs 202±71μA/cm2; n=4; p>0.05) or 2 Hz (180±28 vs 252±70μA/cm2; n=4; p>0.05). Conclusion: GF mice have normal colonic epithelial cell function as well as normal responses to Ach and 5HT in the absence of gut microbiota making GF mice an excellent model system to study effects of individual microbes/microbial products on epithelial cell function. Sa2055 Alpha/Beta Hydrolase Domain 6 (ABHD6), A Novel Endocannabinoid Hydrolytic Enzyme, Regulates Contractility in the Mouse Intestine Mohammad Bashashati, Winnie Ho, Ken Mackie, Morley D. Hollenberg, Keith A. Sharkey Background: Endocannabinoids (ECs) regulate gastrointestinal (GI) motility by controlling acetylcholine release. The most abundant EC in the intestine is 2-arachidonoylglycerol (2-AG). The functional activity of 2-AG is governed in part by the degradative enzyme monoacylglycerol lipase. Recently, another degradative pathway was discovered in the brain that regulates 2-AG levels and involves the enzyme ABHD6. The distribution and function of ABHD6 in the GI tract has not been described. We tested whether: (a) ABHD6 is present in the GI tract and; (b) inhibiting ABHD6 attenuates GI contractility in the mouse. Methods: C57BL/6N and CB1-deficient mice (CB1−/−) were used. ABHD6 mRNA expression was studied in the frontal cortex and throughout the length of the gut using real-time PCR. The distribution of ABHD6 was examined in whole mount preparations of the myenteric plexus or sections throughout the GI tract by immunohistochemistry. The effects of WWL70, an inhibitor of ABHD6, were examined on contractility of ileal and colonic preparations evoked by electrical field stimulation (EFS), veratridine or a specific protease-activated receptor (PAR)-2 activating peptide (2-furoyl-LIGRLO-NH2; 2-fLIGRL). Contractility is presented as mean ± SEM of mg tension per mg tissue. Results: mRNA of ABHD6 is expressed throughout the GI tract with a significantly higher expression in the ileum compared to the colon or frontal cortex. ABHD6 immunoreactivity is expressed on intestinal smooth muscle, epithelium and in neurons of the myenteric plexus throughout the GI tract. Cumulative concentrations of WWL70 (1-10μM) did not change the amplitude of EFS (4-16Hz)-evoked contractility in either region of the gut. However, WWL70 (10μM) attenuated veratridine (1μM) induced contraction of the ileum (5.7±1.7 vs. 1.7±0.6, p<0.05). 2-fLIGRL (5μM) also contracted both ileal and colonic preparations (2±0.2 and 3.8±2.2, respectively). In the ileum, the contractility induced by 2-fLIGRL was tetrodotoxin (TTX) sensitive and was attenuated by pre-incubation with WWL70 (10μM) (1.1±0.3 after WWL70; p <0.05). The effects of 2- fLIGRL on colonic preparations were not TTX or WWL70 sensitive. The effects of WWL70 on veratridine or 2-fLIGRL induced contractility were abolished in the ileum of CB1−/− mice. Conclusions: ABHD6 is present throughout the GI tract. Inhibiting ABHD6 attenuates ileal contractility in the mouse. ABHD6 is a novel enzymatic target for modulating EC activity and motility in the GI tract. S-365 AGA Abstracts Sa2056 Comparison of the Effect of Prucalopride, Sodium Picosulfate and Placebo on Rectal Motor and Sensory Response to Distension Assessed by Electronic Barostat in Healthy Subjects Maura Corsetti, Jan F. Tack Prucalopride and sodium picosulfate have been demonstrated to be more effective than placebo in increasing bowel movements and reducing stool consistency, but also improving the overall satisfaction with abdominal bloating and discomfort of patients with chronic constipation (Ford AC et al 2011). Whether the effect on bloating and discomfort is related to the effect of the two drugs on motility or to a direct effect on sensory pathways is unknown. Aim of the study: to compare the effect of prucalopride, sodium picosulfate and placebo on the rectal sensory and motor response to distension assessed by the electronic barostat. Methods: After an overnight fast a polyvinyl tube with 700 ml plastic balloon was introduced in the rectum of 7 volunteers (4 females, 30±4 years) and connected to an electronic barostat (G&J Electronics Inc, Toronto, Ontario, Canada). A volume-controlled (20 ml/2 min) and a pressure-controlled (4 mm Hg/2 min) stepwise distensions were performed and the intra-balloon pressure then settled at minimal distending pressure (MDP) plus 2 mm Hg for 30 min before and 120 min after the oral administration of placebo, prucalopride (2 mg) and sodium picosulfate (10 mg) in a randomised, placebo-controlled, crossover fashion. Stepwise distensions were then repeated. Rectal tone, rectal distensibility (slope of the volume-pressure and pressure-volume relationship) and volume and corre- sponding pressure discomfort threshold during volume-controlled as well as pressure and corresponding volume discomfort threshold were compared by means of the Wilcoxon test. To take into account the rectal adaptive response to repeated distension, data are expressed by mean±SEM of the delta (Δ) between values after and before the administration of the drugs. Results: The MDP did not differ between arms, and rectal tone did not differ between treatments. During volume-controlled distension, rectal distensibility as well as volume and pressure thresholds for discomfort did not differ between the three treatments. During pressure-controlled distension, the treatment-induced Δ rectal distensibility was significantly lower after prucalopride as compared to both placebo (P= 0.02) and picosulfate (P= 0.04) (Figure). After prucalopride, discomfort occurred at lower Δ distending volumes compared to picosulfate (-136±61 ml vs. 59±12 ml, P= 0.008) and to placebo (27±10 ml, P= 0.06), and at significantly lower Δ pressures as compared to picosulfate (-10±5 mm Hg vs 2.5±2 mm Hg, P= 0.02) but not to placebo (-2.6±2 mm Hg, P= 0.3). Rectal distensibility and volume and pressure thresholds did not differ between picosulfate and placebo. Conclusions: In man, prucalopride decreases rectal distensibility, potentially decreasing the adaptation to rectal filling and sensitizing the rectum to pressure-controlled distension. Sa2057 Lactoferrin Enhances Intestinal Barrier Function by Counteracting Oxidative Stress and Inflammation While Upregulating Tight Junction Protein Expression and Lowering Permeability in CaCO-2/15 Cells Sylvain Mayeur, Ernest G. Seidman, Carole Garofalo, Schohraya Spahis, Valerie Marcil, Yves Pouliot, Jean-Francois Beaulieu, Emile Levy BACKGROUND: Lactoferrin (Lf), an iron-binding glycoprotein in human milk, is involved in multiple biological functions, including cellular proliferation and differentiation, host defense against microbial infection, anti-inflammatory activity and antioxidant defense in various systems. Several investigators have also reported its relative resistance to proteolysis in the intestinal lumen, but little is known on its effects and mechanisms of action towards oxidative stress (OxS) and inflammation in the gastrointestinal tract. OBJECTIVES: To determine the modulation of OxS and inflammation by Lf, as well as Lf transport and mechanisms of actions. METHODS: Differentiated Caco-2/15 cells were used to study the role of Lf beneficial effects against OxS and inflammation induced by iron-ascorbate oxygen radical-generating system (Fe/Asc, 200 μM/2 mM) and lipopolysaccharide (LPS, 150 μg/ ml), respectively. RESULTS: The addition of Fe/Asc to Caco-2/15 cells induced OxS as demonstrated by the rise in malondialdehyde and depletion of polyunsaturated fatty acids. LPS-treatment induced inflammation and abolished intracellular Lf protein expression. How- ever, preincubation with Lf prevented Fe/Asc-mediated lipid peroxidation and counteracted LPS-induced inflammation as evidenced by the down-regulation of inflammatory cytokines and NF-κB, as well as cyclooxygenase (COX)-2 and PGE2. In addition, Lf was able to restore intestinal barrier integrity affected by LPS, as noted by the raised expression of the tight junction occludin and by the lower permeability assessed by fluorescein isothiocyanate- dextran in polarized Caco-2/15 cells. CONCLUSIONS: Lf reduces OxS and inflammation, AGA Abstracts
Transcript of Sa2056 Comparison of the Effect of Prucalopride, Sodium Picosulfate and Placebo on Rectal Motor and...
in the proximal colon, but was slightly elevated in the distal colon. In the presence of 5μM propofol, the amplitude was significantly reduced in both distal and proximal colon(p<0.01, n=4). In both the distal and proximal colon, there was change in the frequencyof the CMMCs in the presence of propofol. When both ondansetron and L-NNA were addedto preparations in the presence of propofol, no differences were observed. In the presenceof 1 μM propofol, fecal pellet motility was significantly reduced compared to control responses(p<0.001, n=5). Conclusions: Propofol significantly increased 5-HT levels in distal colonicbut not the proximal mucosa and thus has site specific activity. This may in turn lead tothe desensitization of 5-HT3Rs on IPANs. Such effects of propofol can explain the reducedamplitude of CMMCs in the proximal and distal colon and reduced fecal pellet motility.Such findings may provide more insight into intestinal side effects observed in patientsutilising propofol for maintenance of anaesthesia and sedation and will provide guidanceon approaches to therapeutically managed patients utilising this form of anaesthetic agent.
Sa2054
Germ Free Mice Have Normal Epithelial Responses to Serotonin andAcetylcholine in the Proximal ColonJohn F. Rainey, David R. Linden, Madhusudan Grover, Charles E. Salmonson, Cheryl E.Bernard, Joseph H. Szurszewski, Gianrico Farrugia, Purna C. Kashyap
Background: Gut microbiota play an important role in modulating gastrointestinal physiologyincluding changes in intestinal transit and epithelial barrier function. Both serotonin (5HT)and acetylcholine (Ach) play important roles in regulation of gut function yet it is not knownif absence of gut microbiota alters the host colonic responses to 5HT and acetylcholine. Theaim of this to study was to determine host colonic epithelial function in the absence of gutmicrobiota. Methods: We used germ free (GF) and humanized mice (ex-GF colonized withhuman bacteria; HM). Two segments of proximal colon, stripped of external muscle layersfrom each animal were mounted in 0.3 cm2 area, 4 ml Ussing chambers. Colonic mucosaltransepithelial resistance (TER) was measured at the beginning of each experiment and FITCDextran (4 k Da) flux was determined from the linear fit of samples obtained from theserosal side every 5 min for 30 min after adding 1mg/ml FITC-Dextran to the mucosal side.Change in short circuit current (Δ Isc) was determined in response to increasing concentra-tions of ACh (0.01-300 μM) on the serosal side, 5HT (0.003-300μM) on the mucosal orserosal side and electrical field stimulation (EFS) using foil electrodes on the submucosalside (30-150V, 100 0.5ms pulses at 1-2Hz). Results: There were no differences in TER(50.2±1.7 vs 69.2±10.5V/cm2; n=4-5; p>0.05) or FITC dextran flux (2.4±0.5 vs 3.4±2 ng/ml/min/cm2; n=4; p>0.05) between GF and HM mice, respectively. Cumulative concentrationresponse curves induced by serosal ACh were not different between GF and HM as evidentby similar Emax (261±38 vs 299±27μA/cm2; n=5; p>0.05) and EC50 (20.6±3 vs 22.7±9μM)values. Cumulative concentration response curves induced by serosal 5-HT were also notdifferent between GF and HM (Emax: 210±14 vs 159±35μA/cm2; n=4; p>0.05) and EC50(2.3±1.9 vs 2.6±1μM). While Δ Isc did not reach maximum response following mucosalapplication of 300μM 5HT, responses were similar between GF and HM(90±15 vs85±2027μA/cm2; n=5; p>0.05) . There was no significant difference in Δ Isc between GFand HM in response to EFS at 1 Hz (138±42 vs 202±71μA/cm2; n=4; p>0.05) or 2 Hz (180±28vs 252±70μA/cm2; n=4; p>0.05). Conclusion: GF mice have normal colonic epithelial cellfunction as well as normal responses to Ach and 5HT in the absence of gut microbiotamaking GF mice an excellent model system to study effects of individual microbes/microbialproducts on epithelial cell function.
Sa2055
Alpha/Beta Hydrolase Domain 6 (ABHD6), A Novel EndocannabinoidHydrolytic Enzyme, Regulates Contractility in the Mouse IntestineMohammad Bashashati, Winnie Ho, Ken Mackie, Morley D. Hollenberg, Keith A. Sharkey
Background: Endocannabinoids (ECs) regulate gastrointestinal (GI) motility by controllingacetylcholine release. The most abundant EC in the intestine is 2-arachidonoylglycerol(2-AG). The functional activity of 2-AG is governed in part by the degradative enzymemonoacylglycerol lipase. Recently, another degradative pathway was discovered in the brainthat regulates 2-AG levels and involves the enzyme ABHD6. The distribution and functionof ABHD6 in the GI tract has not been described. We tested whether: (a) ABHD6 is presentin the GI tract and; (b) inhibiting ABHD6 attenuates GI contractility in the mouse. Methods:C57BL/6N and CB1-deficient mice (CB1−/−) were used. ABHD6 mRNA expression wasstudied in the frontal cortex and throughout the length of the gut using real-time PCR. Thedistribution of ABHD6 was examined in whole mount preparations of the myenteric plexusor sections throughout the GI tract by immunohistochemistry. The effects of WWL70, aninhibitor of ABHD6, were examined on contractility of ileal and colonic preparations evokedby electrical field stimulation (EFS), veratridine or a specific protease-activated receptor(PAR)-2 activating peptide (2-furoyl-LIGRLO-NH2; 2-fLIGRL). Contractility is presented asmean ± SEM of mg tension per mg tissue. Results: mRNA of ABHD6 is expressed throughoutthe GI tract with a significantly higher expression in the ileum compared to the colon orfrontal cortex. ABHD6 immunoreactivity is expressed on intestinal smooth muscle, epitheliumand in neurons of the myenteric plexus throughout the GI tract. Cumulative concentrationsof WWL70 (1-10μM) did not change the amplitude of EFS (4-16Hz)-evoked contractilityin either region of the gut. However, WWL70 (10μM) attenuated veratridine (1μM) inducedcontraction of the ileum (5.7±1.7 vs. 1.7±0.6, p<0.05). 2-fLIGRL (5μM) also contractedboth ileal and colonic preparations (2±0.2 and 3.8±2.2, respectively). In the ileum, thecontractility induced by 2-fLIGRL was tetrodotoxin (TTX) sensitive and was attenuated bypre-incubation with WWL70 (10μM) (1.1±0.3 after WWL70; p <0.05). The effects of 2-fLIGRL on colonic preparations were not TTX or WWL70 sensitive. The effects of WWL70on veratridine or 2-fLIGRL induced contractility were abolished in the ileum of CB1−/−mice. Conclusions: ABHD6 is present throughout the GI tract. Inhibiting ABHD6 attenuatesileal contractility in the mouse. ABHD6 is a novel enzymatic target for modulating EC activityand motility in the GI tract.
S-365 AGA Abstracts
Sa2056
Comparison of the Effect of Prucalopride, Sodium Picosulfate and Placebo onRectal Motor and Sensory Response to Distension Assessed by ElectronicBarostat in Healthy SubjectsMaura Corsetti, Jan F. Tack
Prucalopride and sodium picosulfate have been demonstrated to be more effective thanplacebo in increasing bowel movements and reducing stool consistency, but also improvingthe overall satisfaction with abdominal bloating and discomfort of patients with chronicconstipation (Ford AC et al 2011). Whether the effect on bloating and discomfort is relatedto the effect of the two drugs on motility or to a direct effect on sensory pathways isunknown. Aim of the study: to compare the effect of prucalopride, sodium picosulfate andplacebo on the rectal sensory and motor response to distension assessed by the electronicbarostat. Methods: After an overnight fast a polyvinyl tube with 700 ml plastic balloon wasintroduced in the rectum of 7 volunteers (4 females, 30±4 years) and connected to anelectronic barostat (G&J Electronics Inc, Toronto, Ontario, Canada). A volume-controlled(20 ml/2 min) and a pressure-controlled (4 mm Hg/2 min) stepwise distensions wereperformed and the intra-balloon pressure then settled at minimal distending pressure (MDP)plus 2 mm Hg for 30 min before and 120 min after the oral administration of placebo,prucalopride (2 mg) and sodium picosulfate (10 mg) in a randomised, placebo-controlled,crossover fashion. Stepwise distensions were then repeated. Rectal tone, rectal distensibility(slope of the volume-pressure and pressure-volume relationship) and volume and corre-sponding pressure discomfort threshold during volume-controlled as well as pressure andcorresponding volume discomfort threshold were compared by means of the Wilcoxon test.To take into account the rectal adaptive response to repeated distension, data are expressedby mean±SEM of the delta (Δ) between values after and before the administration of thedrugs. Results: The MDP did not differ between arms, and rectal tone did not differ betweentreatments. During volume-controlled distension, rectal distensibility as well as volume andpressure thresholds for discomfort did not differ between the three treatments. Duringpressure-controlled distension, the treatment-induced Δ rectal distensibility was significantlylower after prucalopride as compared to both placebo (P= 0.02) and picosulfate (P= 0.04)(Figure). After prucalopride, discomfort occurred at lower Δ distending volumes comparedto picosulfate (-136±61 ml vs. 59±12 ml, P= 0.008) and to placebo (27±10 ml, P= 0.06),and at significantly lower Δ pressures as compared to picosulfate (-10±5 mm Hg vs 2.5±2mm Hg, P= 0.02) but not to placebo (-2.6±2 mm Hg, P= 0.3). Rectal distensibility andvolume and pressure thresholds did not differ between picosulfate and placebo. Conclusions:In man, prucalopride decreases rectal distensibility, potentially decreasing the adaptation torectal filling and sensitizing the rectum to pressure-controlled distension.
Sa2057
Lactoferrin Enhances Intestinal Barrier Function by Counteracting OxidativeStress and Inflammation While Upregulating Tight Junction ProteinExpression and Lowering Permeability in CaCO-2/15 CellsSylvain Mayeur, Ernest G. Seidman, Carole Garofalo, Schohraya Spahis, Valerie Marcil,Yves Pouliot, Jean-Francois Beaulieu, Emile Levy
BACKGROUND: Lactoferrin (Lf), an iron-binding glycoprotein in human milk, is involvedin multiple biological functions, including cellular proliferation and differentiation, hostdefense against microbial infection, anti-inflammatory activity and antioxidant defense invarious systems. Several investigators have also reported its relative resistance to proteolysisin the intestinal lumen, but little is known on its effects and mechanisms of action towardsoxidative stress (OxS) and inflammation in the gastrointestinal tract. OBJECTIVES: Todetermine the modulation of OxS and inflammation by Lf, as well as Lf transport andmechanisms of actions. METHODS: Differentiated Caco-2/15 cells were used to study therole of Lf beneficial effects against OxS and inflammation induced by iron-ascorbate oxygenradical-generating system (Fe/Asc, 200 μM/2 mM) and lipopolysaccharide (LPS, 150 μg/ml), respectively. RESULTS: The addition of Fe/Asc to Caco-2/15 cells induced OxS asdemonstrated by the rise in malondialdehyde and depletion of polyunsaturated fatty acids.LPS-treatment induced inflammation and abolished intracellular Lf protein expression. How-ever, preincubation with Lf prevented Fe/Asc-mediated lipid peroxidation and counteractedLPS-induced inflammation as evidenced by the down-regulation of inflammatory cytokinesand NF-κB, as well as cyclooxygenase (COX)-2 and PGE2. In addition, Lf was able to restoreintestinal barrier integrity affected by LPS, as noted by the raised expression of the tightjunction occludin and by the lower permeability assessed by fluorescein isothiocyanate-dextran in polarized Caco-2/15 cells. CONCLUSIONS: Lf reduces OxS and inflammation,
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