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Diet regulates class II MHC expression in mouse intestinal epithelium through the regulatory molecule, CIITA
AGAA77
April 2000
579
581
DIET REGULATES CLASS II MHC EXPRESSION IN MOUSE INTESTINAL EPITHELIUM THROUGH THE REGULATORY MOLECULE, CIITA. Demetra S. Stamm, Suzan Dziennis, Stephen A. Bustin, Ian R. Sanderson, Harvard Clin Nutrition Research Ctr, Boston, MA; St Bartholomew's and The Royal London Sch of Medicine, London, United Kingdom. Class II MHC (Ia) and invariant (Ii) chain expression in the intestinal epithelium is induced by weaning onto normal complex diets. Weaning onto an elemental diet, composed of nutrients that are fully absorbed, results in no expression of la and Ii chain. We hypothesized that diet induced invariant chain expression through the class II transactivator protein CIITA. Methods: Wild-type mice and IFN-oy receptor knock out mice were weaned onto either normal complex (chow) diet or onto an elemental diet. CIITA mRNA was examined by RNase protection assay and subtypes of CIITA (CIIT A I, III, and IV) by real time PCR. Results: CIITA mRNA was induced in the intestinal epithelium of wild type mice by weaning onto a complex diet, mirroring the expression of invariant chain. Weaning onto an elemental diet did not increase CIITA mRNA. Complex diet did not induce CIITA I, but induced CIITA III and CIITA IV. Because stimulation of the intestinal epithelial cell line (lEC-6) with IFN-oy induced CIITA, we wished to exclude the possibility that the induction of invariant chain was through IFN-oy. We therefore studied the effect of diet in mice deficient in the IFN·oyreceptor. Like wild type mice, they expressed invariant chain in the intestinal epithelium, when weaned onto a complex diet. Furthermore, complex diet induced the accumulation of CIITA III mRNA, but not CIITA IV. Conclusion: The data show that dietary constituents induce the expression of a regulatory gene, CIITA, in the intestinal epithelium, whose function is to control Class II MHC and invariant chain expression. These mechanisms act through CIITA and are independent of IFN-oy.
DEPRIVATION OF NON·STARCH POLYSACCHARIDES IN THE RAT DIET CAUSES MUCOSAL DAMAGE. Vicki Strugala, Adrian Allen, Michael E. Havler, Peter W. Dettmar, Jeffrey P. Pearson, Univ of Newcastle Upon Tyne, Newcastle, United Kingdom; Reckitt & Colman Products Ltd, Hull, United Kingdom. Introduction & Aim Non-starch polysaccharide (NSP, dietary fibre) has many beneficial effects on the colon including decreasing transit time, increasing stool weight, stool water content and reducing intra-adbominal pressure. It is proposed to protect against a number of diseases such as constipation, colonic cancer, coronary heart disease and diverticultis. The aim of this study was to determine the effect of NSP, soluble and insoluble, on the colonic mucosa of rats in vivo. Methods Rats were fed either a) fibre deficient, b) ispaghula (psyllium) enriched (13.7% NSP, 11.9% soluble NSP), c) control diet (15.4% mixed NSP, 1.8% soluble NSP) ad libitum for 4 weeks. The rats were fasted overnight and anaesthetised. The colon was exteriorised, opened laterally and placed in a mucosal chamber bathed in saline. The mucosa was observed using intravital microscopy. Mucosal bleeding was assessed using a 4-point visual scale (O=no visual evidence of bleeding, I =mild, small bleed that clots quickly, 2=moderate, 1-2 areas of bleeding that clot slowly, 3= severe, 3+ areas of bleeding that clot slowly). Fibrinogen (and fibrin) content was measured in the bathing solution using an ELISA and immunohistochemically in sections of frozen colonic mucosa. Fibrinogen was measured using a polyclonal rabbit antihuman fibrinogen antibody (DAKO) and the levels present determined from a rat fibrinogen standard curve. Haemoglobin content of the bathing solution was assessed by measuring absorbance at 575nm. Results Mean (SE) mucosal bleeding score for the fibre deficient rats was 2.1 (0.3). This was significantly higher (p
580 DISTRIBUTION OF BILE ACID TRANSPORT CAPACITIES IN THE HUMAN ILEUM. Matthias Stelzner, Sivagurunathan Somasundaram, David J. Kearney, Univ of Washington/Seattle VAMC, Seattle, WA. The sodium-dependent bile acid transporter IBAT actively absorbs bile acids in the ileum. In rodents, uptake capacities, IBAT mRNA and IBAT protein concentrations follow distinct distribution curves. Such distributions have not been established in humans. We, therefore, mapped IBAT function, mRNA and IBAT protein in the ileum of 13 human volunteers. Methods: 10 - 30 biopsies were obtained from each subject during colonoscopy or surgical resection of healthy ileum 0 -120 cm proximal to the ileocecal valve. Active and passive bile acid uptake rates were measured using a miniaturized everted sleeve technique. IBAT mRNA concentrations were determined by competitive PCR, IBAT protein concentrations by semiquantitative immunohistology. Results.Bile acid uptake showed distinct variations. (Fig.; active= - , passive= •••). Maximal active uptake varied among II subjects from 0.6 - 3.5 pmol/min x mrrr'. 2 subjects showed no active uptake. IBAT mRNA and protein distributions corroborated uptake curves. Conclusion:IBAT function and gene expression vary throughout the human ileum. This data will be important in studying regulation of IBAT in humans.
2000
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c
'E
1500
x N
E 1000
.§
'0 E
c..
500 • • • • • • 6o 6o6o6o • • • • • •
o o
50 100 150 Distance from Ileocecal Valve [cm]
582 NICOTINE AND GASTRODUODENAL PERMEABILITY. Peter Suenaert, Veerle Bulteel, Elly Den Hond, Benny Geypens, Anja Lyupaerts, Fred Monsuur, Y vo Ghoos, Paul Rutgeerts, Univ Hospitals Gasthuisberg, Leuven, Belgium; Univ Hosp Gasthuisberg, Leuven, Belgium. Introduction and aim: Smoking exerts adverse effects on gastric mucosal physiology and is associated with peptic disease and aggravation of preexisting mucosallaesions. However, the effect of nicotine on the gastroduodenal permeability has never been studied. As sucrose is a reliable site-specific permeability probe of the gastroduodenal mucosa I, we investigated the influence of nicotine on the gastroduodenal permeability in healthy nonsmoker subjects. Method: A test solution containing 20g sucrose was ingested after an overnight fast, once with and once without application of transdermal nicotine patches. These nicotine patches were administered to 24 normal persons during eight days in a climbing dose from 5 to 15mg per 24 hours. Intake of NSAID's was prohibited one week before, alcohol 48 hours before the test. The urinespecimens were collected during 6 hours. The urine sugar concentration was measured using GCFID. The gastric emp7,ing time ti/2 was measured by the Na- 13C-octanoate breath test (lOOmg I C-octanoate mixed in a liquid meal). Results: We found a highly significant increase in sucrose permeability during nicotine administration compared with the pre-nicotine values in the same normal individuals, with a short interval of three weeks between the two tests. The gastric emptying time t'/2 did not differ significantly between these two conditions. Discussion: It is the first time that the effect of nicotine on the gastroduodenal mucosa is assessed with the site-specific marker sucrose. In this study nicotine increases the gastroduodenal permeability to sucrose in healthy individuals independently from gastric emptying or other permeation factors as mucosal probe degradation, systemic bloodflow or renal clearance since the test was executed in the same subjects. Conclusion: Nicotine seems to "widen" the upper intestinal barrier. The underlying mechanism warrants further investigation. 'Gastroenterology 1993; 104: 1619-26.
Sucrose permeability mean (%)±SD Gastric emplYlngl1/2 mean (min)±SD prenicotine postnicotine p
0006±0010 0.024±0.025 0.0038' (Wilcoxon)
37±141 37±13.7 0992 (paired T-test)
Mean urinary excretion fractions ofsucrose (%) and gastric emptying time t1/2 (minutes) in24 healthy volunteers." shows asignificant correlation (p<0.05).
April 2000
579
581
DIET REGULATES CLASS II MHC EXPRESSION IN MOUSE INTESTINAL EPITHELIUM THROUGH THE REGULATORY MOLECULE, CIITA. Demetra S. Stamm, Suzan Dziennis, Stephen A. Bustin, Ian R. Sanderson, Harvard Clin Nutrition Research Ctr, Boston, MA; St Bartholomew's and The Royal London Sch of Medicine, London, United Kingdom. Class II MHC (Ia) and invariant (Ii) chain expression in the intestinal epithelium is induced by weaning onto normal complex diets. Weaning onto an elemental diet, composed of nutrients that are fully absorbed, results in no expression of la and Ii chain. We hypothesized that diet induced invariant chain expression through the class II transactivator protein CIITA. Methods: Wild-type mice and IFN-oy receptor knock out mice were weaned onto either normal complex (chow) diet or onto an elemental diet. CIITA mRNA was examined by RNase protection assay and subtypes of CIITA (CIIT A I, III, and IV) by real time PCR. Results: CIITA mRNA was induced in the intestinal epithelium of wild type mice by weaning onto a complex diet, mirroring the expression of invariant chain. Weaning onto an elemental diet did not increase CIITA mRNA. Complex diet did not induce CIITA I, but induced CIITA III and CIITA IV. Because stimulation of the intestinal epithelial cell line (lEC-6) with IFN-oy induced CIITA, we wished to exclude the possibility that the induction of invariant chain was through IFN-oy. We therefore studied the effect of diet in mice deficient in the IFN·oyreceptor. Like wild type mice, they expressed invariant chain in the intestinal epithelium, when weaned onto a complex diet. Furthermore, complex diet induced the accumulation of CIITA III mRNA, but not CIITA IV. Conclusion: The data show that dietary constituents induce the expression of a regulatory gene, CIITA, in the intestinal epithelium, whose function is to control Class II MHC and invariant chain expression. These mechanisms act through CIITA and are independent of IFN-oy.
DEPRIVATION OF NON·STARCH POLYSACCHARIDES IN THE RAT DIET CAUSES MUCOSAL DAMAGE. Vicki Strugala, Adrian Allen, Michael E. Havler, Peter W. Dettmar, Jeffrey P. Pearson, Univ of Newcastle Upon Tyne, Newcastle, United Kingdom; Reckitt & Colman Products Ltd, Hull, United Kingdom. Introduction & Aim Non-starch polysaccharide (NSP, dietary fibre) has many beneficial effects on the colon including decreasing transit time, increasing stool weight, stool water content and reducing intra-adbominal pressure. It is proposed to protect against a number of diseases such as constipation, colonic cancer, coronary heart disease and diverticultis. The aim of this study was to determine the effect of NSP, soluble and insoluble, on the colonic mucosa of rats in vivo. Methods Rats were fed either a) fibre deficient, b) ispaghula (psyllium) enriched (13.7% NSP, 11.9% soluble NSP), c) control diet (15.4% mixed NSP, 1.8% soluble NSP) ad libitum for 4 weeks. The rats were fasted overnight and anaesthetised. The colon was exteriorised, opened laterally and placed in a mucosal chamber bathed in saline. The mucosa was observed using intravital microscopy. Mucosal bleeding was assessed using a 4-point visual scale (O=no visual evidence of bleeding, I =mild, small bleed that clots quickly, 2=moderate, 1-2 areas of bleeding that clot slowly, 3= severe, 3+ areas of bleeding that clot slowly). Fibrinogen (and fibrin) content was measured in the bathing solution using an ELISA and immunohistochemically in sections of frozen colonic mucosa. Fibrinogen was measured using a polyclonal rabbit antihuman fibrinogen antibody (DAKO) and the levels present determined from a rat fibrinogen standard curve. Haemoglobin content of the bathing solution was assessed by measuring absorbance at 575nm. Results Mean (SE) mucosal bleeding score for the fibre deficient rats was 2.1 (0.3). This was significantly higher (p
580 DISTRIBUTION OF BILE ACID TRANSPORT CAPACITIES IN THE HUMAN ILEUM. Matthias Stelzner, Sivagurunathan Somasundaram, David J. Kearney, Univ of Washington/Seattle VAMC, Seattle, WA. The sodium-dependent bile acid transporter IBAT actively absorbs bile acids in the ileum. In rodents, uptake capacities, IBAT mRNA and IBAT protein concentrations follow distinct distribution curves. Such distributions have not been established in humans. We, therefore, mapped IBAT function, mRNA and IBAT protein in the ileum of 13 human volunteers. Methods: 10 - 30 biopsies were obtained from each subject during colonoscopy or surgical resection of healthy ileum 0 -120 cm proximal to the ileocecal valve. Active and passive bile acid uptake rates were measured using a miniaturized everted sleeve technique. IBAT mRNA concentrations were determined by competitive PCR, IBAT protein concentrations by semiquantitative immunohistology. Results.Bile acid uptake showed distinct variations. (Fig.; active= - , passive= •••). Maximal active uptake varied among II subjects from 0.6 - 3.5 pmol/min x mrrr'. 2 subjects showed no active uptake. IBAT mRNA and protein distributions corroborated uptake curves. Conclusion:IBAT function and gene expression vary throughout the human ileum. This data will be important in studying regulation of IBAT in humans.
2000
• •
c
'E
1500
x N
E 1000
.§
'0 E
c..
500 • • • • • • 6o 6o6o6o • • • • • •
o o
50 100 150 Distance from Ileocecal Valve [cm]
582 NICOTINE AND GASTRODUODENAL PERMEABILITY. Peter Suenaert, Veerle Bulteel, Elly Den Hond, Benny Geypens, Anja Lyupaerts, Fred Monsuur, Y vo Ghoos, Paul Rutgeerts, Univ Hospitals Gasthuisberg, Leuven, Belgium; Univ Hosp Gasthuisberg, Leuven, Belgium. Introduction and aim: Smoking exerts adverse effects on gastric mucosal physiology and is associated with peptic disease and aggravation of preexisting mucosallaesions. However, the effect of nicotine on the gastroduodenal permeability has never been studied. As sucrose is a reliable site-specific permeability probe of the gastroduodenal mucosa I, we investigated the influence of nicotine on the gastroduodenal permeability in healthy nonsmoker subjects. Method: A test solution containing 20g sucrose was ingested after an overnight fast, once with and once without application of transdermal nicotine patches. These nicotine patches were administered to 24 normal persons during eight days in a climbing dose from 5 to 15mg per 24 hours. Intake of NSAID's was prohibited one week before, alcohol 48 hours before the test. The urinespecimens were collected during 6 hours. The urine sugar concentration was measured using GCFID. The gastric emp7,ing time ti/2 was measured by the Na- 13C-octanoate breath test (lOOmg I C-octanoate mixed in a liquid meal). Results: We found a highly significant increase in sucrose permeability during nicotine administration compared with the pre-nicotine values in the same normal individuals, with a short interval of three weeks between the two tests. The gastric emptying time t'/2 did not differ significantly between these two conditions. Discussion: It is the first time that the effect of nicotine on the gastroduodenal mucosa is assessed with the site-specific marker sucrose. In this study nicotine increases the gastroduodenal permeability to sucrose in healthy individuals independently from gastric emptying or other permeation factors as mucosal probe degradation, systemic bloodflow or renal clearance since the test was executed in the same subjects. Conclusion: Nicotine seems to "widen" the upper intestinal barrier. The underlying mechanism warrants further investigation. 'Gastroenterology 1993; 104: 1619-26.
Sucrose permeability mean (%)±SD Gastric emplYlngl1/2 mean (min)±SD prenicotine postnicotine p
0006±0010 0.024±0.025 0.0038' (Wilcoxon)
37±141 37±13.7 0992 (paired T-test)
Mean urinary excretion fractions ofsucrose (%) and gastric emptying time t1/2 (minutes) in24 healthy volunteers." shows asignificant correlation (p<0.05).