Previous data in our lab demonstrated an inability to induce oral tolerance when using a feeding needle gavage for 14 days. Given that the upper gastrointestinal (GI) tract is the site of antigen introduction, and the interplay between immune cells of the mucosal tissues, we questioned if inflammation in this tissue, induced by physical trauma, would affect oral tolerance induction. We performed studies on Balb/c mice using a needle gavage or syringe feeding method followed by doses of the immunogenic protein ovalbumin (OVA) to induce tolerance. Immunohistochemistry was used to assess inflammation in esophageal tissues and to correlate with an ability or inability to induce tolerance. Non-cellular alterations within the tissue were also assessed using a pathology grading score. Although fluctuations in cell populations were observed in both the syringe and gavage treated mice, the needle gavage caused significant noncellular damage to esophageal mucosal tissue, which is the most likely cause of failed tolerance induction to the OVA. / Department of Biology

Human milk contains ~ 25 µg/mL of soluble cluster of differentiation 14 (sCD14) protein, a pattern recognition receptor (PRR) that triggers the innate immune system to respond to bacterial lipopolysaccharide (LPS). To date, the role of CD14 in the digestive tract of breast fed infants has not been well characterized and is the subject of this thesis. To investigate the biodistribution of proteins such as CD14 in vivo, a novel method for 14C radiolabeling of proteins to high specific radioactivity was developed using in vacuo methylation. Bovine serum albumin (BSA) and casein were used as test proteins to determine the following: 1) The efficacy of the in vacuo radiolabeling procedure; 2) The extent of incorporation of the 14C-label into the organs of oro-gastric gavaged 10 day old Sprague Dawley rats. [14C]BSA, [14C]casein and [14C]CD14 were prepared with specific radioactivities of 10 400, 10 800 and 163 000 dpm/µg, respectively. After feeding 6.25 µg of 14C-labeled proteins, quantifiable levels of 14C were found in the stomach, jejunum, duodenum, ileum, large intestine, intestinal luminal flushes, blood, liver, spleen and kidneys of rats. The accumulation of radiolabel in the organs of [14C]CD14 fed rats was temporally and spatially distinct from [14C]BSA and [14C]casein. Most notably, the label persisted in the stomach 480 min post-gavage. To design a neonate animal model for biodistribution, the segmental and total gastrointestinal transit times (GItt) were measured in two litters of 10 and 15 day old Sprague Dawley rat pups using barium sulfate. Ten day old rat pups that remained with and without the dam had a total gastrointestinal transit time of 13.8 ± 0.9 hr and 9.3 ± 0.7 hr, respectively. This decrease (p<0.05) in total gastrointestinal transit time in the absence of the dam was age dependent, as it was not observed (p>0.05) in the 15 day old rat pup litter. The immunological impact of an exogenous sCD14 source was examined in human peripheral blood mononuclear cells (PBMC). Pre-treatment of CD14+ monocytes with sCD14 had a protective effect, one of reducing the production of proinflammatory cytokines (TNF-α, IL-6, IL-8, IL-1β) when challenged with LPS. 14C was absorbed by neonate rats upon ingestion of [14C]CD14 and exposure to relatively high concentrations of rCD14 led to a reduction in inflammation. This may be beneficial to initial gut colonization in breast-fed newborns. / Alexander Graham Bell NSERC CGS M scholarship. Japan Society for the Promotion of Sciences, Summer in Japan Fellowship. Funded by the Canadian Institutes of Health Research, Institute of Nutrition Metabolism and Diabetes Grant #82816 “Fate and function of breast milk and recombinant human CD14 at mammary and newborn gastrointestinal mucosal epithelia”.

Parkinson’s disease is a neurodegenerative disease resulting primarily from loss of dopaminergic innervation in the striatum subsequent to cell loss in the substantia nigra pars compacta. The abnormal accumulation of the normal pre-synaptic protein α-synuclein (αsyn) forms intraneuronal inclusions known as Lewy neurites and Lewy bodies. The origins of central Lewy pathology have been suggested to lie in the enteric nervous system, ascending through the vagus nerve to the dorsal motor nucleus of the vagus. To ascertain gastrointestinal regions most likely to be the source of central Lewy pathology, αsyn expression was evaluated in the neural elements of gastrointestinal regions receiving the densest vagal innervation. The vermiform appendix was found to have the densest αsyn-immunoreactive innervation in all layers of the gut wall. In addition, macrophages in the appendiceal mucosa were laden with αsyn within lysosomes, consistent with attempts to prevent the spread of disease or to correct synaptic dysfunction.

Entrotoxigenic Escherichia coli (ETEC) K88 is a causative agent of post-weaning diarrhea (PWD) in early-weaned pigs. This study investigated the efficacy of two alternative diets, charcoal (0.1, 0.5, 1, and 2%) and a low crude protein (CP) diet (17%) supplemented with probiotic E. coli strains (UM2 and UM7), against PWD infection in ETEC K88 challenged piglets. The present study found that charcoal had no effect on the challenged piglets’ performance, ileal and colonic microbiota or their fermentation end products. There was, however, a correlation between charcoal dosage and fecal consistency score. Charcoal reduced the ileal mucosal attached ETEC K88. Feeding a low-CP diet resulted in a lower ileal ammonia concentration. The low-CP diet reduced the E. coli populations in the ileal digesta as well as lowered mRNA expression of the IL-1ß. We concluded that the use of both 1-2% charcoal diet and a low-CP diet supplemented with probiotic E. coli strains were effective in reducing the incidence and severity of PWD infection.

Lange Zeit galt der Dünndarm als „Blackbox“ des Gastrointestinaltraktes. Seit Einführung der Videokapselendoskopie im Jahr 2001 eröffnete sich eine Methode, den Dünndarm zu visualisieren. An einem Kollektiv von 203 Patienten habe ich Indikationen, Ergebnisse und klinischen Nutzen von Dünndarmkapselendoskopien in einem Zeitraum von 4 Jahren untersucht. Der Dünndarm ist in der Gastroduodeno- und Koloskopie nicht komplett zugänglich, so dass bei entsprechender Indikation die nicht-invasive Videokapselendoskopie vorgenommen werden kann. Sie ist in der Lage 2-4 Bilder pro Sekunde in einem Zeitraum von 8-9 Stunden aufzunehmen, die als Film von ca. 50.000 Bildern zusammengestellt und interpretiert werden kann. Die Daten zur diagnostischen Ausbeute dieser Untersuchung variieren und sind abhängig von der entsprechenden Indikation. Zur Überprüfung des klinischen Nutzens habe ich daher in meiner Arbeit speziell die Passagezeiten und die erhobenen Befunde, wie Erosionen, Ulzerationen, Angiodysplasien, Petechien, Venektasien, Lymphangiektasien, Erytheme, Ödeme, Zottenreliefveränderungen, extrinsische Engen und Erhabenheiten im Hinblick für ihre diagnostische Bedeutung ausgewertet. Berücksichtigt wurden die Auswertbarkeit, Komplikationsrate sowie Vor- und Nachuntersuchungen. Das Aufklärungsgespräch erfolgte mindestens einen Tag vor der Videokapselendoskopie. Die Abführmaßnahmen entsprachen einer Koloskopievorbereitung. Das Studienkollektiv (203 Patienten) bestand aus 58% männlichen und 42% weiblichen Patienten. Der Altersdurchschnitt betrug 58 Jahre, die Altersspanne reichte von 8-90 Jahren. Über 93% nahmen die Videokapsel selbstständig ein, eine Applikation erfolgte bei 7% der Patienten in den Bulbus duodeni. Folgende Indikationen führten bei unserer Patientenklientel zu der Videokapselendoskopie: unklare gastrointestinale Blutung (45,3%), unklare abdominelle Schmerzen (24,1%), unklare Anämie (11,3%), Verdacht auf/ oder Komplikation bei Morbus Crohn (6,5%), unklare Diarrhoe (6,4%), Polyp- und Tumorsuche (5,4%), rezidivierendes unklares Erbrechen und Eiweißverlustsyndrom (jeweils 0,5%). Eine komplette Dünndarmpassage konnte innerhalb der Aufzeichnungszeit von 8-9 Stunden bei 84% der Patienten erreicht werden. Der Mittelwert der Magenpassagezeit lag bei 21 Minuten und der Dünndarmpassagezeit bei 6 Stunden. Die Komplikation Kapselretention trat bei 2% auf. Pathologische Befunde im Dünndarm wurden bei 85% detektiert. Die höchste diagnostische Ausbeute ergab sich bei der Abklärung der unklaren gastrointestinalen Blutung (80%) und bei der unklaren Anämie (78%), als häufigste Ursache wurden Schleimhautläsionen (43%) gefunden. Unklare abdominelle Schmerzen wiesen eine niedrigere diagnostische Ausbeute (41%) auf. Therapeutische Maßnahmen resultierten bei 73% der untersuchten Patienten aus den Kapselergebnissen. Eine medikamentöse Therapie wurde bei 66% eingeleitet oder verändert, Endoskopien wurden bei 4% und eine operative Therapie bei 4,4% durchgeführt. Damit ist die Dünndarmkapselendoskopie bei klarer Fragestellung und guter Darmvorbereitung eine sichere und sinnvolle Untersuchungsmethode, insbesondere zur Klärung unklarer gastrointestinaler Blutungen. Spezifische Dünndarmerkrankungen, wie der M. Crohn oder Tumore können relativ sicher ausgeschlossen werden.

To test the hypothesis that maternal dietary glucose restriction throughout pregnancy and lactation would perturb glucose transporter (GLUT) protein levels in offspring, isoenergetic diets containing graded levels of glucose (0, 12, 24 and 60%) were fed to pregnant rats and their offspring from gestation day (gd) 0 through postnatal day (pd) 49. Diets were defined as deficient (0%), restricted (12, 24%) or adequate (60%) in glucose. Plasma, small intestine, liver and kidney tissues were collected perinatally (gd20, birth, 12--24hrs postnatal), during lactation (pd7, 15, 21), post-weaning (pd28, 35, 49) and in adult controls. The proximal and distal regions of the small intestine responded differently to the dietary glucose restriction. Proximal small intestine GLUT2 protein levels did not change throughout postnatal development and remained unaltered with dietary glucose restriction, while distal small intestine GLUT2 protein expression changed throughout postnatal development and with dietary glucose restriction. These findings, together with information from the literature, indicate a dissociation between small intestine GLUT2 mRNA expression, GLUT2 protein levels and small intestine glycogen reserves.

Gastrointestinal (GI) health is an important contributor to one’s overall well-being. In the past decade, the focus of this aspect of health has been on the organisms that inhabit the intestines: gut microbes. A concept central to understanding bacterial behavior for health or disease promotion is biological film (biofilm) formation. The predominant form of growth for bacteria is biofilm formation, an as yet poorly described phenomenon for gut microbes. In order to better understand bacterial behavior in response to nutrients that pass through the GI system, a high throughput system to assess biofilm formation was developed. Gastrointestinal-associated microbes, Escherichia coli and Enterococcus faecalis, were assayed for biofilm formation in 96-well plates after 24 hours of incubation. Nutrients, inter-and intrakingdom signaling molecules such as monosaccharides, calcium, insulin, endocannabinoids, and AI-1, AI-2 like signaling compounds, respectively, were added to cultures in order to test their effects on biofilm formation. Biofilm was quantified spectrophotometrically by the measurement of optical density of each well measured at 580nm following crystal violet staining of adherent biofilm. Values were expressed as means ± standard error of the mean (SEM); differences between means were assessed using t-testing and ANOVA using GraphPad Prism, with mean differences considered significant at P < 0.05. Results showed that biofilm formation by E. coli was enhanced by glucose, galactose, lactose, AI-1 like signaling compound and insulin at 50 and 100µU/ml, while AI-2 like compound and calcium inhibited biofilm formation. Biofilm formation by E. faecalis was also enhanced by AI-1 like compound and insulin at 50µU/ml in RPMI medium and inhibited in cultures grown in BHI medium or with added calcium. We conclude that gastrointestinal-associated microbes are influenced by nutrients as well as various factors, including the culture medium, signaling compounds, as well as host-signaling compounds such as insulin and calcium. This study provides a platform required for future studies involving nutrient effect on biofilm formation.

No description available.

No description available.

Efter operation och anestesi får patienter ofta en negativ påverkan på magsäck och tarmar. Illamående och kräkningar är ett stort problem och många har svårt att komma igång med intag av föda och normal tarmfunktion då magsäcken och tarmarna ”står stilla”. Flera faktorer bidrar- bl.a. smärtan, det kirurgiska traumat och de läkemedel vi ger i samband med anestesin. Av de senare är opioider, d.v.s morfin och morfinliknande läkemedel, starkt bidragande. I detta avhandlings- arbete har opioiders effekter på magsäckens motilitet studerats. Med ett absorptionstest (paracetamolmetoden) studerades hos frivilliga hur opioiden remifentanil påverkar magsäckstömning och om kroppspositionen har betydelse för tömningshastigheten ut i tarmen. Remifentanil fördröjde magsäcks-tömningen och under pågående opioid behandling hade kroppspositionen ingen större betydelse, vilket det däremot hade under kontrollförsöken. Med samma metod jämförde vi hos patienter två anestesimetoder och studerade magsäcks-tömning direkt efter en operation. Ingen skillnad kunde påvisas mellan en opioidbaserad och en opioidfri anestesi, men inom respektive grupp var det en stor variation i magsäckstömning mellan individerna. Med en barostat studerades tonus i övre delen av magsäcken. Hos hälften av de frivilliga orsakade remifentanil en ökning av tonus och hos den andra hälften en minskning av tonus. Vidare undersöktes hos en grupp patienter opioiden fentanyls påverkan på den elektriska aktiviteten i magsäcken. Med en elekroga-strograf (EGG) registrerades de långsamma elektriska vågor som koordinerar muskelrörelserna i magsäcken. Hos hälften av de undersökta påverkades aktiviteten av fentanyl med en sänkt vågfrekvens eller upphörande av vågor, medan aktiviteten var opåverkad hos den övriga hälften. För att finna en förklaring till variationen gjordes genetiska analyser av genen för opioidreceptorn hos de undersökta i barostat och EGG studierna. Variationer i genomet, s.k. polymorfism, var inte associerad till utfallen i studierna. Studierna har visat på att opioider har en uttalad effekt på magsäckens motilitet och att den varierar kraftigt mellan individer. Polymorfism i genen för opioid- receptorn förklarade inte skillnaden mellan individer. Direkt efter operation bidrar sannolikt andra faktorer än anestesimetod till det variabla utfallet i magsäckstömning. / After anesthesia and/or surgical procedures, gastrointestinal motility is commonly impaired. The causes are multifactorial, with surgical trauma, pain and perioperative drugs playing a major role. This thesis explores opioid effects on gastric motility in healthy volunteers and patients undergoing surgery. Gastric emptying was studied by an absorption test (paracetamol method), and in healthy volunteers a remifentanil infusion delayed gastric emptying. Body position altered emptying during the control situations, but not during the remifentanil infusion. Further, two anesthetic methods were compared and no differences were found in immediate postoperative gastric emptying between a remifentanil/propofol based intravenous anesthesia and an opioid free inhalational anesthesia, although the interindividual variability was high. Proximal gastric tone was studied using a gastric barostat. An infusion of remifentanil caused two patterns of reaction regarding gastric tone, with half of the subjects increasing and half decreasing in gastric tone. Gastric myoelectrical activity was evaluated with electrogastrography (EGG), and a bolus dose of fentanyl caused a decrease in frequency of the gastric slow waves or disrupted this activity. However, the activity was unaffected in half of the investigated subjects. Analysis of polymorphisms (A118G and G691C) in the µ-opioid receptor gene was performed to find an explanation for the great interindividual variations seen in the barostat and EGG studies, but no association could be found. These studies have shown that opioids have pronounced effects on gastric motility with variable individual responses that are difficult to predict. Polymorphisms in the µ-opioid receptor gene could not explain the variations. Postoperatively, other factors might contribute more than opioids to the impairment in gastric motility. / ISSN 1652-4063