- Email: [email protected]
Methotrexate as rescue therapy in pediatric Crohn's disease
was previously shown to be involved in the secretory effects of bile acids on the colonocytes, the potential involvement of histamine receptors was also examined in current studies. Both cimetidine (1 raM) and diphenhydramine (0.1raM), H2 and H1 histamine receptor antagonists respectively, had no effect on inhibition of 36C1 uptake by TDC. In conclusion, our studies, provide direct evidence for inhibition of human intestinal luminal CI'/OH (HCO~) exchange activity by bile acids. The results of our studies demonstrate that bile acid effects on the apical CI/OH exchange in Caco2 cells are not mediated via histamine receptors and involve a cAMP-independent but Ca + +, PKC and PI3 kinase-dependent pathways.
MeCP2 genotype and GI phenotype correlative clinical study in children with Rett syndrome. Methods: Thirty-six consecutive females with a mean (SD) age of 7.4 (3.4) years were ascertained and examined at The Rett Syndrome clinic at The Kennedy Krieger Institute. Genomic DNA was isolated from peripheral lymphocytes and mutation analysis was performed by using a combination of DHPLC (detection of heteroduplexes by ion-pair reverse phase HPLC) and direct sequencing (Genomics 1998;52:44-49). Mutation analysis of the MeCP2 gene was correlated with failure to thrive (z-score<-2SD), pathological GERD, constipation and oropharyngeal dysphagia. Results: Twenty-six patients with Rett syndrome showed a wide distribution of mutations throughout the exon 3 and 4 of the MeCP2 gene (Table 1). Among these patients, all but one manifested clinical symptoms associated with gastrointestinal dysfunction requiring medical intervention. Oropharyngeal dysphagia (4) associated well (p<0.05) with mutations localized to the proximal portion of the MeCP2 gene suggesting that more proximal mutations may predispose to neuroenteric dysfunction. A similar association has also been noted in the presence of more proximal MeCP2 mutations and the severity of central and autonomic neuronal function, including frequency and manageability of seizures, respiratory difficulties and the ability to walk. Conclusions: MeCP2 gene mutations commonly associate with the gastrointestinal manifestation of Rett syndrome. Proximal mutations within the MeCP2 gene may also correlate with more severe forms of swallowing dysfunction. Future challenges include the application of video fluoroscopy and esophageal manometry in the diagnosis and management of patients with Rett syndrome with symptoms of oropharyngeal dysphagia.
301 Mechanisms of Liquid and Gas Gastroesophageal Reflux in Healthy Preterm lafants. A Combined Manometric and Impedance Study Taher Omari, Nathalie Rommel, Esther Staunton, Louise Goodchild, Ross Lontis, Ross I-hslam, John Dent, Geoffrey Davidson Background: In preterm infants, transient lower esophageal sphincter relaxation (TLESR) is the predominant mechanism of gastroesophageal reflux (GER) detected manometrically, by the presence of esophageal common cavity episodes (CC), and/or by pH probe. TLESRs triggering CC, but not acid GER, are common during the early post-prandial period, however, the precise nature (liquid/gas) of these reflux episodes has not been previously determined. The aim of this study was to use a novel combined manometry and impedance assembly to charactense the mechanisms of liquid and gas GER in preterm infants. Methods: 5 preterm infants (4M: IF) ranging in weight from 2480-3180g were studied at a post menstrual age of 34 to 37 weeks. Combined manometry and multi-channel intralumina[ impedance (MII) was performed using a novel lower esophageal sphincter (LES) sleeve assembly (OD 2.5mm) incorporating a 9cm array of 6 impedance rings spaced at 1.5cm intervals for MII measurement. After placement of the assembly infants were fed with 45-80ml of expressed breast milk via an assembly infusion port and then esophageal manometry and MII were recorded for 4 hours post prandially. Episodes of liquid and gas GER were identified using standard MII cnteria and the esophageal motor mechanism(s) of GER triggering characterised. Results: 71 GER episodes were recorded consisting of 57(80%) liquid, 8(11%) gas and 6(8%) combined liqind/gas. The mean(SE) bolus clearance time of liquid GER was 13.8(1.1) sec. The proximal extent of GER was >9cm for 43(65%) of liquid or mixed GER episodes. TLESRwas the predominant mechanism of reflux, triggering 70% of all GER episodes. Other GERmechanisms identified included swallow related LES relaxation (10%), vomiting (6%) and straining in association with low LES pressure (3%). Of 70 TLESRs recorded, 20(29%) wereuneventful, 44(63%) triggered liquid GER, 3(4%) triggered gas GER and 3(4%) triggered combined liquid/gas GER. TLESR triggered exclusively liquid GER in the first postrandial hour. Conclusions: In preterm infants, the majority of GER is liquid in nature and usually extends into the proximal oesophagus. TLESR is the predominant mechanism of all GER in these infants.
MeCP2
N
Age Range G E R D
Constipation
(~) R I ~ (Q/W) T158M RtflSX R255X R270X P,294X R306C 79Md(1)/1161d~(6) 801dd(C) No mutation=
2 3 5 4 5 2 2 2 1 10
4-8 4-7 2-16 3,5-11 3,5-10 4-13 4-5 4 3.5 3-15
Failure to
Dysphagla
~,~,e 0 0 2 1 3 0 0 1 0 I
1 2 3 2 3 2 1 1 1 4
1 3 4 3 3 2 1 1 0 5
2 0 2 0 0 0 0 0 0 0
304 Regional Variations in SP, VIP and NOS in Colon Circular muscle from Children With Slow Transit Constipation Bridget R. Southwell, Pare Farmer, John H. Hutson Coordinated firing of inhibitory and excitatory motor nerves controls contraction and relaxation of muscle producing movement of stools through the colon. Reductions in substance P (SP) and increases in vasoactive intestinal peptide (VIP) and have been observed in nerve fibres in colonic circular muscle (CCM) in children and adults with slow transit constipation (STC), suggesting a relationship between neurotransmitter levels and slowed colonic motility t. Aim: To determine regional variations in excitatory and inhibitory transmitters in nerve fibres in colon circular muscle CCM) from children with STC The density of nerve fibres containing SP, VIP and nitric oxide synthase (NOS) was quantified. Methods: STC was diagnosed using scintigraphy (radionncleide marker and multiple images) as radioactivity retained in the right and transverse colon at 48 hr. Seromuscular biopsies of CCM were collected laparascopically from right transverse (RT), left transverse (LT) and sigmoid colon and the location and density of neurotransmitters was determined by fluorescence immunohtstochemistry and confocal microscopy, n = 19. Results: There was regional variation in the density of transmitters in CCM. Density (/0.025 mm 2) of SP-1R fibres was 9 -+ 1 (mean _+SEM) in RT, < 12 _+2 in LT, < sigmoid colon (17 +- 2). There was bimodal distribution of density, with 84, 42 & 36% of patients with low SP in RT, LT and sigmoid colon resp. 21% had low SP in all 3 regions. VIP density was > in LT (33+3) than sigmoid (27+_3) or RT (21+_3) colon. NOS density was > in LT (37+_6) than sigmoid (32+_4) or RT (26+_3) colon. Ratios of NOS:SP and VIP:SP were highest in LT and lowest in sigmoid colon. Conc[: There is regional variation in the density of SP, VIP and NOS in CCM from children with STC. SP (excitatory transmitter) was lowest in RT, while inhibitory transmitters VIP and NOS were highest in LT colon. The ratio of inhibitory:excitatory transmitter density was highest in the LT colon. This could result in greater relaxation, reduced contraction and reduced motility in the transverse colon. 1. Hutson, J.M., McNamara, J., Gibb, S. & Shin, YM. Slow transit constipation in children. J Paediatr Child Health 37, 426-30 (2001).
302 Baclofen Reduces Emesis and Gastroesophageal Reflux in Neurologically Impaired Children with Gastroesophageal Reflux Disaese Masanobu Kawai, Shinobu Ida, Norikazu Yoshimura, Hisayoshi Kawahara Neurologically impaired (NI) children frequently have various symptoms caused by gastroesophageal reflux disease (GERD). Therapeutic options for GERD have been limited in these patients. The GABAB agonist baclofen has recently been shown to reduce postprandial acid reflux in adult patients with GERD by reducing the incidence of transient lower esophageal sphincter relaxations. The present study was undertaken to investigate the effects of badofen on clinical symptoms and acid refiuxes in pediatric N[ patients. Eight NI children with cerebral palsy, aged from 2 months to 16 year of age(median age: 5 years), were studied. All patients showed frequent emesis and were diagnosed with GERD by 24-hr esophageal pH data (the percentage time of esophageal pH<4.0 over 5.0). Baclofen (0.7mg/kg/day) was administered orally or via naso-gasmc tube in three divided doses 30 minutes before meals for 7 days. The frequency of emesis was graded as follows: grade I > twice a week; grade I1 > once a day; grade III> twice a day; grade IV every time after meal. The clinical symptom was scored on a scale of 1 to 4 according to the grading of emesis (grade I was score 1). The esophageal pH data were analyzed for the number of acid refluxes and the percentage time of esophageal pH < 4 0 . The symptom scores and the esophageal pH data were compared between the values before the administration of baclofen and those on the last day of this trial. Data are expressed as medians and interquartile ranges. Statistical analystswas performed with Wilcoxon's signed rank test. The symptom scores decreased significantly from 4 (3-4) to 1 (1-3) (p=0.02). The number of acid refluxes decreased signihcantly from 193 (149-295) to 124 (67-211) in 24 hours (p=0.01) and from 94(66182) to 62(42-109) in the postprandial 2 hours (p<0.05). The percentage time of esophageal pH<4.0 did not show any sigmficant difference in 24 hours (17(13-24) vs 11(4-24),p = 0.21) and in the postprandial 2 hours (26(16-35) vs 14(8-31),p =0.40). No adverse effects were noted except mild hypotonia in one subject during this trial. In conclusion, a short-term baclofen administration is effective to reduce the frequency of emesis without significant adverse effects and the number of acid refluxes in NI children with GERD.
3O5 Methotrexate as Rescue Therapy in Pediatric Crohn's Disease Joel R. Rosh, Nader N. Yonssef, Stephanie Schuckalo, Jaya Punati, Barbara Fehling, Richard L. Mones AIMS: There is no clearly preferred therapy for children with Crohn's disease who develop intolerance or lack of clinical response to 6-mercaptopurine (6MP). This study looks at the efficacy of immune-modulation with methotrexate (MTX) as maintenance therapy in a cohort of such patients. Primary outcome measure was a significant reduction in the Pediatric Crohn's Disease Activity Index (PCDAI) score. Secondary measures were need for surgical resection and continued need for infliximab administration. SUBJECTS & METHODS: Of the 240 children with inflammatory bowel disease actively followed at our center, 12 with biopsy-proven Crohn's disease (6 male, aged 13.7 +/- 5.7 years) were withdrawn from 6MP therapy and started on weekly subcutaneous MTX. 6MP was withdrawn due to lack of clinical response or drug intolerance (6 patients). Intolerance was defined as nausea, vomiting and increased abdominal pain (5) and fever with severe arthralgia (1) with drug challenge and withdrawal. Lack of clinical response was defined by elevated PCDA1 scores and/or need for surgical intervention. At the time of MTX introduction, all patients were requiring multiple infusions of infliximab (5.6 +/- ].7 infusions / patient). Time from diagnosis of Crohn's disease until introduction of MTX was 4.3 +/- 2.4 years. Prior to MTX,
303 X-linked Methyl CpG Binding Protein (MeCP2) Gene Mutations in Children with Rett Syndrome: Correlation with Clinical Severity of Gastrointestinal Symptoms Carmen Cuffari, Anil Darbari, Genila Bibat, Sakkubai Naidu Background: Rett syndrome is a neurodevelopmental disorder that is caused by a number of mutations in the MeCP2 gene located on chromosome Xq28. Mutations in the MeCP2 proteinresults in a loss of function leading to developmental regression. The GI manifestations in these children vary in clinical severity and may be genotype dependent. Aim: To report a
A-41
AGA Abstracts
308
6MP had been used 2.3 years +/- 2.9 years. The final dose of 6MP was 1.5 +/- 0.5 rag/ kg / day. RESULTS: Eleven of 12 patients (92%) responded to MTX. The duration of MTX therapy was 7.8 +/- 5.5 months at time of follow-up. The effective dose was 0.4 +/- 0.2 m g / kg / weekly. MTX maintained remission as demonstrated by all outcome measures including: decreased PCDA1, 30.5 v. 13.1 (p<0.05); decreased need for surgical intervention 50% v. 8.5% (p = 0.07); decreased need for mfliximab 100% v. 50% (p<0.02). MTX was discontinued in 2 patients due to eventual relapse (MTX given 6.7 +/- 6.1 months) and 1 patient due to refusal to continue injections. Response to MTX was unrelated to gender, disease distribution, or prior need for surgical resection. There were no side effects attributed to MTX. CONCLUSIONS: MIX is a safe and effective maintenance agent for 6MP intolerant or unresponsive pediatric Crohn's disease patients. This immune-modulating therapy may be used in low doses, once .weekly and appears to be well tolerated. In these patients MTX allowed for decrease in infliximab requirement as well as the need for surgical intervention. The long-term efficacy of MTX in pediatric Crohn's disease requires further study.
Different Effects of Nutrients on Intestinal Gas Dynamics and Tolerance Influenced by the Small Intestine Ana C. Hemando-Harder, Hermann Harder, Heinz J. Krammer, Manfred V. Singer Background: Depending on food composition, postprandial gas related abdominal symptoms are frequendy reported. Therefore we compared the effects of a mixed liquid meal and specific nutrient components on intestinal gas dynamics and tolerance evaluating influences of the proximal vs. distal small intestine. Methods: Forty healthy subjects were studied twice in randomised order on different days. Either a mixed liquid meal, lipids, proteins, glucose at lkcal/min or a control solution of saline (n = 8 for each group) were infused with 2 ml / rain into the proximal duodenum. Intestinal gas transit and tolerance were evaluated using a previously validated technique (Gastroenterology,1998; 115:542-50): After 30 mmntes of basal nutrient infusion, a gas mixture infusion (N2, 02 and CO2 in venous proportions) was started at 12 ml / min into the proximal duodenum or 120 cm distal the figamentum of Treitz in randomised order for 150 min; gas evacuation was measured via an intrarectal cannula by a barostat, perception was assessed with a 0-6 score scale and girth changes using a memc band. Results: Both fipids (733 +-- 26 ml / 271 -+ 78 ml) and proteins (271 --- 78 ml / 96 +- 51 ml) lead to significant abdominal gas retention, p < 0.05 compared to normal saline (8 + 44 ml / -63 +- 28 ml) as control (p < 0.05; proximal / distal gas infusion), intraduodenal lipids slightly increased abdominal perception of proximal and distal gas infusion (2.0 -+ 0.3 score and 2.3 - 0.6 score, p < 0.05 vs. control). Lipids also caused abdominal girth changes correlating with intestinal gas retention (r = 0.79; p < 0.001). In contrast, no or even negative gas retention was seen with both, the mixed meal (-7 - 58 ml / -92 + 44 ml) and glucose (35 - 126 ml / -157 -4- 57 ml) with either proximal or distal gas infusion, respectively. There were no sigmficant changes in abdominal girth and abdominal or rectal perception. Conclusion: The composition of a meal is a major factor governing intestinal gas dynamics and perception threshold. Abdominal and rectal symptoms depend only partly on the total volume of gas retained into the gut, initiated by the specific nutrient ingested. Different intestinal sites pronounce the effects of nutrients by regulatory mechanisms which need further evaluation.
306
The Mismatch Repair (MMR) Protein hMSH3 Is Lost From Polyp Epithelium of Patients with PTEN Germtine llamartomatons Syndromes Sherry C. Huang, Arun Swaminath, E. Jnlieta Smith, Ryan T. Doctolero, Anna Dredinger, John M Carethers BACKGROUND&AIMS: The mechanism for cancer formation in hamartomatous syndromes has not been established. In juvenile polyposis syndrome (JPS), there is up to 12-fold increased risk for co[orectal cancer. In Cowden disease (CD), there is increased risk for breast, endometrial, and thyroid cancers, but the risk is unclear for colon cancer. Both syndromes share polyposis and germline mutations in the PTEN phosphatase tumor suppressor gene in some cases. PTEN may be a target for mutation in cells with defective MMR due to a coding microsatelhte. Because of the increased risk for cancer in these syndromes, we hypothesized that defects in the DNA MMR system might contribute to their cancer potential. METHODS: We examined polyps from families with germline PTEN mutations. A JPS kindred had members with multiple juvenile polyps. A CD kindred had a member who developed facial trichilemmomas and had a thyroid resection. A 1 cm adenoma with was excised from the CD patient. Polyps were microdissected into epithelial and lamina propna domains. Microsatellite analyses were performed using five standardized markers. lmmunohistochemistry was performed on sections using specific antibodies to PTEN and the DNA MMR proteins hMSH2, hMLH1, hMSH6, and hMSH3. Additional polyps from patients without germline PTEN mutations were immunostamed for comparison. RESULTS: Both kindreds had germline PTEN mutations: the JPS family had a splice alteration at the intron-exon boundary of exon 4, and the CD patient had a R130X truncating mutation. Microsatellite analysis revealed polyps from both kindreds had microsatellite instability-high (MS[), indicative of a MMR gene defect. The MSI was limited to the epithelium and not other polyp components, lmmunostaming revealed complete loss of PTEN expression from the epithelium of the hamartomas and adenoma, indicative of somatic reactivation of the second allele. Polyps from patients without PTEN germline mutations or sporadic cases still expressed PTEN. The epithelial components from both types of polyps with germhne PTEN mutations showed loss of hMSH3, but not hMLH1, hMSH2, or hMSH6. CONCLUSIONS: Polyps from patients with hamartomatous syndromes due to germline PTEN mutations lose PTEN somatically from the epithelial components of the polyp. Additionally, the DNA MMR protein hMSH3 is absent from the epithelium. Loss of hMSH3 is a potential mechanism for cancer transformation in these polyps and may be responsible for inactivating the second ailde of PTEN
309 Characterization of Fasting and Postprandial Flow Patterns in the Small Intestine: Comparison of Muhichannel Intraluminal Impedance (MII) with Manometry Hala M. Imam, Claudia P. Sanmigue[, Yasser Bhat, Brett Larive, Edy Softer Background: Adequate flow of intestinal contents is critical for normal gut homeostasis. However, understanding of intestinal patterns of flow remains poor because of limitations of available techniques such as manometry or transit studies. MII is a new technique that detects flow in a viscous organ by measuring changes in intrahiminal impedance. Using combined videofluornscopy, MII and manonetry, we found that MII is more sensitive than manometry in detecting intestinal flow (Neurogastro 2002; 14:430). Aim: To determine patterns of intestinal flow and their relation to pressure events in fasting (F) and postprandial (PP) periods. Methods: 12 healthy subjects had simultaneous manometry and Mr[ of the duodenum by a probe incorporating 5 pressure sensors and 5 pairs of electrodes spaced 5 cm apart. Tracings inspected visually for presence and distance of propagation of sequences of impedance and pressure events. We analyzed 30 minutes in phase ll, following the first phase lI[, and 30 minutes after ingestion of 500ml of BOOST. Short sequences defined as present in I and 2 channels, long sequences defined as propagated over 4 and 5 channels. Percentages of Mll and pressure events having differing lengths were compared between periods using Cochran-Mantel-Haemzel mean scores tests. Other tests used mixed models ANOVA, p<0.05. Results: There were more events in the PP period compared to F. The distribution of MII sequence lengths differed significantly between the periods, with PP having more short sequences and F having longer ones (table). MII sequences, of various lengths, showed a significantly higher association with corresponding pressure sequences in the PP period as compared to F. 20% and 36% of long Mr[ sequences were not associated with contractions in F and PP periods respectively. Motility index values were significantly associated with increasing le'ngcha of MII propagation in F (P
307
Nutrient Modulation of Intestinal Gas Dynamics in Healthy Humans Depends on Caloric Content and Physical Properties of the Meal Sutep Gonlachanvit, Radoslav Coleski, William L. Hasler Patients with gas and bloating report increased symptoms after ingestion of meals with specific properties~ We have showed that liquid caloric meals stimulate transit of jejunallyperfused gas, but the effects of solid or non-caloric meals are unexplored. We hypothesized that meals of different caloric density and physical properties modulate gas transit to different degrees. 10 healthy subjects underwent jejunal perfusion of physiologic gas mixtures (88% Nz, 5.5% 02, 6.5% COL) at 12 ml/min x 3 hr with ingestion of nothing (control), water (240 ml), a 240 kcal liquid meal (240 ml, 41 gm CHO, 4 gm fat, and 10 gm protein), and a 240 kcal solid meal (toast and scrambled eggs) at the end of the second hour in separate studies Gas was collected from an mtrarectal catheter to bypass the anus and evacuation was quantified in rcal-time using a harostat. An initial lag phase was noted after starting gas perfusmn, then gas was evacuated from the rectum at i2.7+-0.7, 12.7_+ 1.2, 13.7_+0.8, and 137 _+1.3 ml/min (P=NS) for the control, water, liquid meal, and solid meal arms. Solid and liquid meals increased cumulative gas volumes evacuated during the initial 20 minutes after eating (451 _+128 and 410_+51 ml vs. 254_+54 ml for control; P
AGA Abstracts
Table: MII and pressuresequencesin F and PP pedods ,r,,~ MH Pedods
F PP *P<0.05
Total
Short Mil
prmm
~qmma
~ 521 1187
511 1264
~) 64,1 70.1"
Short~
LongMII
sure~
q ~ 75.9 78.1
(%)
Long pre~
Nq-an,m sure~. (~) q u m ~ (~) 20.7" 15.5
15.8 10.2
310 Fat-Induced Ileal Brake Depends on an Opioid Pathway Acting on Kappa Receptors Henry C. Lin, Jin Hal Chen, Corynn Neeve| Background: Slowing of intestinal transit by fat as the ileal brake depends on a naloxone (nonspecific antagonist)-sensitive opioid pathway (Zhao et al. AJP 278: G866-70, 2000). Dynorphin (acting on kappa opioid receptors) immunoreactive nerves are found in the intestine (Costa & Fumess. Neuropeptides 5: 445, 1985). However, the exact role of kappa opinid receptors is unknown. Aim: To test the hypothesis that slowing of intestinal transit by fat may depend on an opioid pathway acting on kappa opioid receptors, we compared the fat-induced ileal brake with and without a kappa opioid receptor antagonist. Methods: 4 dogs were equipped with duodenal and midgnt fistulas. With occluding Foley catheters in the distal limb of the 2 fistulas, the small intestine was compartmentalized into the
A-42
MeCP2 genotype and GI phenotype correlative clinical study in children with Rett syndrome. Methods: Thirty-six consecutive females with a mean (SD) age of 7.4 (3.4) years were ascertained and examined at The Rett Syndrome clinic at The Kennedy Krieger Institute. Genomic DNA was isolated from peripheral lymphocytes and mutation analysis was performed by using a combination of DHPLC (detection of heteroduplexes by ion-pair reverse phase HPLC) and direct sequencing (Genomics 1998;52:44-49). Mutation analysis of the MeCP2 gene was correlated with failure to thrive (z-score<-2SD), pathological GERD, constipation and oropharyngeal dysphagia. Results: Twenty-six patients with Rett syndrome showed a wide distribution of mutations throughout the exon 3 and 4 of the MeCP2 gene (Table 1). Among these patients, all but one manifested clinical symptoms associated with gastrointestinal dysfunction requiring medical intervention. Oropharyngeal dysphagia (4) associated well (p<0.05) with mutations localized to the proximal portion of the MeCP2 gene suggesting that more proximal mutations may predispose to neuroenteric dysfunction. A similar association has also been noted in the presence of more proximal MeCP2 mutations and the severity of central and autonomic neuronal function, including frequency and manageability of seizures, respiratory difficulties and the ability to walk. Conclusions: MeCP2 gene mutations commonly associate with the gastrointestinal manifestation of Rett syndrome. Proximal mutations within the MeCP2 gene may also correlate with more severe forms of swallowing dysfunction. Future challenges include the application of video fluoroscopy and esophageal manometry in the diagnosis and management of patients with Rett syndrome with symptoms of oropharyngeal dysphagia.
301 Mechanisms of Liquid and Gas Gastroesophageal Reflux in Healthy Preterm lafants. A Combined Manometric and Impedance Study Taher Omari, Nathalie Rommel, Esther Staunton, Louise Goodchild, Ross Lontis, Ross I-hslam, John Dent, Geoffrey Davidson Background: In preterm infants, transient lower esophageal sphincter relaxation (TLESR) is the predominant mechanism of gastroesophageal reflux (GER) detected manometrically, by the presence of esophageal common cavity episodes (CC), and/or by pH probe. TLESRs triggering CC, but not acid GER, are common during the early post-prandial period, however, the precise nature (liquid/gas) of these reflux episodes has not been previously determined. The aim of this study was to use a novel combined manometry and impedance assembly to charactense the mechanisms of liquid and gas GER in preterm infants. Methods: 5 preterm infants (4M: IF) ranging in weight from 2480-3180g were studied at a post menstrual age of 34 to 37 weeks. Combined manometry and multi-channel intralumina[ impedance (MII) was performed using a novel lower esophageal sphincter (LES) sleeve assembly (OD 2.5mm) incorporating a 9cm array of 6 impedance rings spaced at 1.5cm intervals for MII measurement. After placement of the assembly infants were fed with 45-80ml of expressed breast milk via an assembly infusion port and then esophageal manometry and MII were recorded for 4 hours post prandially. Episodes of liquid and gas GER were identified using standard MII cnteria and the esophageal motor mechanism(s) of GER triggering characterised. Results: 71 GER episodes were recorded consisting of 57(80%) liquid, 8(11%) gas and 6(8%) combined liqind/gas. The mean(SE) bolus clearance time of liquid GER was 13.8(1.1) sec. The proximal extent of GER was >9cm for 43(65%) of liquid or mixed GER episodes. TLESRwas the predominant mechanism of reflux, triggering 70% of all GER episodes. Other GERmechanisms identified included swallow related LES relaxation (10%), vomiting (6%) and straining in association with low LES pressure (3%). Of 70 TLESRs recorded, 20(29%) wereuneventful, 44(63%) triggered liquid GER, 3(4%) triggered gas GER and 3(4%) triggered combined liquid/gas GER. TLESR triggered exclusively liquid GER in the first postrandial hour. Conclusions: In preterm infants, the majority of GER is liquid in nature and usually extends into the proximal oesophagus. TLESR is the predominant mechanism of all GER in these infants.
MeCP2
N
Age Range G E R D
Constipation
(~) R I ~ (Q/W) T158M RtflSX R255X R270X P,294X R306C 79Md(1)/1161d~(6) 801dd(C) No mutation=
2 3 5 4 5 2 2 2 1 10
4-8 4-7 2-16 3,5-11 3,5-10 4-13 4-5 4 3.5 3-15
Failure to
Dysphagla
~,~,e 0 0 2 1 3 0 0 1 0 I
1 2 3 2 3 2 1 1 1 4
1 3 4 3 3 2 1 1 0 5
2 0 2 0 0 0 0 0 0 0
304 Regional Variations in SP, VIP and NOS in Colon Circular muscle from Children With Slow Transit Constipation Bridget R. Southwell, Pare Farmer, John H. Hutson Coordinated firing of inhibitory and excitatory motor nerves controls contraction and relaxation of muscle producing movement of stools through the colon. Reductions in substance P (SP) and increases in vasoactive intestinal peptide (VIP) and have been observed in nerve fibres in colonic circular muscle (CCM) in children and adults with slow transit constipation (STC), suggesting a relationship between neurotransmitter levels and slowed colonic motility t. Aim: To determine regional variations in excitatory and inhibitory transmitters in nerve fibres in colon circular muscle CCM) from children with STC The density of nerve fibres containing SP, VIP and nitric oxide synthase (NOS) was quantified. Methods: STC was diagnosed using scintigraphy (radionncleide marker and multiple images) as radioactivity retained in the right and transverse colon at 48 hr. Seromuscular biopsies of CCM were collected laparascopically from right transverse (RT), left transverse (LT) and sigmoid colon and the location and density of neurotransmitters was determined by fluorescence immunohtstochemistry and confocal microscopy, n = 19. Results: There was regional variation in the density of transmitters in CCM. Density (/0.025 mm 2) of SP-1R fibres was 9 -+ 1 (mean _+SEM) in RT, < 12 _+2 in LT, < sigmoid colon (17 +- 2). There was bimodal distribution of density, with 84, 42 & 36% of patients with low SP in RT, LT and sigmoid colon resp. 21% had low SP in all 3 regions. VIP density was > in LT (33+3) than sigmoid (27+_3) or RT (21+_3) colon. NOS density was > in LT (37+_6) than sigmoid (32+_4) or RT (26+_3) colon. Ratios of NOS:SP and VIP:SP were highest in LT and lowest in sigmoid colon. Conc[: There is regional variation in the density of SP, VIP and NOS in CCM from children with STC. SP (excitatory transmitter) was lowest in RT, while inhibitory transmitters VIP and NOS were highest in LT colon. The ratio of inhibitory:excitatory transmitter density was highest in the LT colon. This could result in greater relaxation, reduced contraction and reduced motility in the transverse colon. 1. Hutson, J.M., McNamara, J., Gibb, S. & Shin, YM. Slow transit constipation in children. J Paediatr Child Health 37, 426-30 (2001).
302 Baclofen Reduces Emesis and Gastroesophageal Reflux in Neurologically Impaired Children with Gastroesophageal Reflux Disaese Masanobu Kawai, Shinobu Ida, Norikazu Yoshimura, Hisayoshi Kawahara Neurologically impaired (NI) children frequently have various symptoms caused by gastroesophageal reflux disease (GERD). Therapeutic options for GERD have been limited in these patients. The GABAB agonist baclofen has recently been shown to reduce postprandial acid reflux in adult patients with GERD by reducing the incidence of transient lower esophageal sphincter relaxations. The present study was undertaken to investigate the effects of badofen on clinical symptoms and acid refiuxes in pediatric N[ patients. Eight NI children with cerebral palsy, aged from 2 months to 16 year of age(median age: 5 years), were studied. All patients showed frequent emesis and were diagnosed with GERD by 24-hr esophageal pH data (the percentage time of esophageal pH<4.0 over 5.0). Baclofen (0.7mg/kg/day) was administered orally or via naso-gasmc tube in three divided doses 30 minutes before meals for 7 days. The frequency of emesis was graded as follows: grade I > twice a week; grade I1 > once a day; grade III> twice a day; grade IV every time after meal. The clinical symptom was scored on a scale of 1 to 4 according to the grading of emesis (grade I was score 1). The esophageal pH data were analyzed for the number of acid refluxes and the percentage time of esophageal pH < 4 0 . The symptom scores and the esophageal pH data were compared between the values before the administration of baclofen and those on the last day of this trial. Data are expressed as medians and interquartile ranges. Statistical analystswas performed with Wilcoxon's signed rank test. The symptom scores decreased significantly from 4 (3-4) to 1 (1-3) (p=0.02). The number of acid refluxes decreased signihcantly from 193 (149-295) to 124 (67-211) in 24 hours (p=0.01) and from 94(66182) to 62(42-109) in the postprandial 2 hours (p<0.05). The percentage time of esophageal pH<4.0 did not show any sigmficant difference in 24 hours (17(13-24) vs 11(4-24),p = 0.21) and in the postprandial 2 hours (26(16-35) vs 14(8-31),p =0.40). No adverse effects were noted except mild hypotonia in one subject during this trial. In conclusion, a short-term baclofen administration is effective to reduce the frequency of emesis without significant adverse effects and the number of acid refluxes in NI children with GERD.
3O5 Methotrexate as Rescue Therapy in Pediatric Crohn's Disease Joel R. Rosh, Nader N. Yonssef, Stephanie Schuckalo, Jaya Punati, Barbara Fehling, Richard L. Mones AIMS: There is no clearly preferred therapy for children with Crohn's disease who develop intolerance or lack of clinical response to 6-mercaptopurine (6MP). This study looks at the efficacy of immune-modulation with methotrexate (MTX) as maintenance therapy in a cohort of such patients. Primary outcome measure was a significant reduction in the Pediatric Crohn's Disease Activity Index (PCDAI) score. Secondary measures were need for surgical resection and continued need for infliximab administration. SUBJECTS & METHODS: Of the 240 children with inflammatory bowel disease actively followed at our center, 12 with biopsy-proven Crohn's disease (6 male, aged 13.7 +/- 5.7 years) were withdrawn from 6MP therapy and started on weekly subcutaneous MTX. 6MP was withdrawn due to lack of clinical response or drug intolerance (6 patients). Intolerance was defined as nausea, vomiting and increased abdominal pain (5) and fever with severe arthralgia (1) with drug challenge and withdrawal. Lack of clinical response was defined by elevated PCDA1 scores and/or need for surgical intervention. At the time of MTX introduction, all patients were requiring multiple infusions of infliximab (5.6 +/- ].7 infusions / patient). Time from diagnosis of Crohn's disease until introduction of MTX was 4.3 +/- 2.4 years. Prior to MTX,
303 X-linked Methyl CpG Binding Protein (MeCP2) Gene Mutations in Children with Rett Syndrome: Correlation with Clinical Severity of Gastrointestinal Symptoms Carmen Cuffari, Anil Darbari, Genila Bibat, Sakkubai Naidu Background: Rett syndrome is a neurodevelopmental disorder that is caused by a number of mutations in the MeCP2 gene located on chromosome Xq28. Mutations in the MeCP2 proteinresults in a loss of function leading to developmental regression. The GI manifestations in these children vary in clinical severity and may be genotype dependent. Aim: To report a
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6MP had been used 2.3 years +/- 2.9 years. The final dose of 6MP was 1.5 +/- 0.5 rag/ kg / day. RESULTS: Eleven of 12 patients (92%) responded to MTX. The duration of MTX therapy was 7.8 +/- 5.5 months at time of follow-up. The effective dose was 0.4 +/- 0.2 m g / kg / weekly. MTX maintained remission as demonstrated by all outcome measures including: decreased PCDA1, 30.5 v. 13.1 (p<0.05); decreased need for surgical intervention 50% v. 8.5% (p = 0.07); decreased need for mfliximab 100% v. 50% (p<0.02). MTX was discontinued in 2 patients due to eventual relapse (MTX given 6.7 +/- 6.1 months) and 1 patient due to refusal to continue injections. Response to MTX was unrelated to gender, disease distribution, or prior need for surgical resection. There were no side effects attributed to MTX. CONCLUSIONS: MIX is a safe and effective maintenance agent for 6MP intolerant or unresponsive pediatric Crohn's disease patients. This immune-modulating therapy may be used in low doses, once .weekly and appears to be well tolerated. In these patients MTX allowed for decrease in infliximab requirement as well as the need for surgical intervention. The long-term efficacy of MTX in pediatric Crohn's disease requires further study.
Different Effects of Nutrients on Intestinal Gas Dynamics and Tolerance Influenced by the Small Intestine Ana C. Hemando-Harder, Hermann Harder, Heinz J. Krammer, Manfred V. Singer Background: Depending on food composition, postprandial gas related abdominal symptoms are frequendy reported. Therefore we compared the effects of a mixed liquid meal and specific nutrient components on intestinal gas dynamics and tolerance evaluating influences of the proximal vs. distal small intestine. Methods: Forty healthy subjects were studied twice in randomised order on different days. Either a mixed liquid meal, lipids, proteins, glucose at lkcal/min or a control solution of saline (n = 8 for each group) were infused with 2 ml / rain into the proximal duodenum. Intestinal gas transit and tolerance were evaluated using a previously validated technique (Gastroenterology,1998; 115:542-50): After 30 mmntes of basal nutrient infusion, a gas mixture infusion (N2, 02 and CO2 in venous proportions) was started at 12 ml / min into the proximal duodenum or 120 cm distal the figamentum of Treitz in randomised order for 150 min; gas evacuation was measured via an intrarectal cannula by a barostat, perception was assessed with a 0-6 score scale and girth changes using a memc band. Results: Both fipids (733 +-- 26 ml / 271 -+ 78 ml) and proteins (271 --- 78 ml / 96 +- 51 ml) lead to significant abdominal gas retention, p < 0.05 compared to normal saline (8 + 44 ml / -63 +- 28 ml) as control (p < 0.05; proximal / distal gas infusion), intraduodenal lipids slightly increased abdominal perception of proximal and distal gas infusion (2.0 -+ 0.3 score and 2.3 - 0.6 score, p < 0.05 vs. control). Lipids also caused abdominal girth changes correlating with intestinal gas retention (r = 0.79; p < 0.001). In contrast, no or even negative gas retention was seen with both, the mixed meal (-7 - 58 ml / -92 + 44 ml) and glucose (35 - 126 ml / -157 -4- 57 ml) with either proximal or distal gas infusion, respectively. There were no sigmficant changes in abdominal girth and abdominal or rectal perception. Conclusion: The composition of a meal is a major factor governing intestinal gas dynamics and perception threshold. Abdominal and rectal symptoms depend only partly on the total volume of gas retained into the gut, initiated by the specific nutrient ingested. Different intestinal sites pronounce the effects of nutrients by regulatory mechanisms which need further evaluation.
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The Mismatch Repair (MMR) Protein hMSH3 Is Lost From Polyp Epithelium of Patients with PTEN Germtine llamartomatons Syndromes Sherry C. Huang, Arun Swaminath, E. Jnlieta Smith, Ryan T. Doctolero, Anna Dredinger, John M Carethers BACKGROUND&AIMS: The mechanism for cancer formation in hamartomatous syndromes has not been established. In juvenile polyposis syndrome (JPS), there is up to 12-fold increased risk for co[orectal cancer. In Cowden disease (CD), there is increased risk for breast, endometrial, and thyroid cancers, but the risk is unclear for colon cancer. Both syndromes share polyposis and germline mutations in the PTEN phosphatase tumor suppressor gene in some cases. PTEN may be a target for mutation in cells with defective MMR due to a coding microsatelhte. Because of the increased risk for cancer in these syndromes, we hypothesized that defects in the DNA MMR system might contribute to their cancer potential. METHODS: We examined polyps from families with germline PTEN mutations. A JPS kindred had members with multiple juvenile polyps. A CD kindred had a member who developed facial trichilemmomas and had a thyroid resection. A 1 cm adenoma with was excised from the CD patient. Polyps were microdissected into epithelial and lamina propna domains. Microsatellite analyses were performed using five standardized markers. lmmunohistochemistry was performed on sections using specific antibodies to PTEN and the DNA MMR proteins hMSH2, hMLH1, hMSH6, and hMSH3. Additional polyps from patients without germline PTEN mutations were immunostamed for comparison. RESULTS: Both kindreds had germline PTEN mutations: the JPS family had a splice alteration at the intron-exon boundary of exon 4, and the CD patient had a R130X truncating mutation. Microsatellite analysis revealed polyps from both kindreds had microsatellite instability-high (MS[), indicative of a MMR gene defect. The MSI was limited to the epithelium and not other polyp components, lmmunostaming revealed complete loss of PTEN expression from the epithelium of the hamartomas and adenoma, indicative of somatic reactivation of the second allele. Polyps from patients without PTEN germline mutations or sporadic cases still expressed PTEN. The epithelial components from both types of polyps with germhne PTEN mutations showed loss of hMSH3, but not hMLH1, hMSH2, or hMSH6. CONCLUSIONS: Polyps from patients with hamartomatous syndromes due to germline PTEN mutations lose PTEN somatically from the epithelial components of the polyp. Additionally, the DNA MMR protein hMSH3 is absent from the epithelium. Loss of hMSH3 is a potential mechanism for cancer transformation in these polyps and may be responsible for inactivating the second ailde of PTEN
309 Characterization of Fasting and Postprandial Flow Patterns in the Small Intestine: Comparison of Muhichannel Intraluminal Impedance (MII) with Manometry Hala M. Imam, Claudia P. Sanmigue[, Yasser Bhat, Brett Larive, Edy Softer Background: Adequate flow of intestinal contents is critical for normal gut homeostasis. However, understanding of intestinal patterns of flow remains poor because of limitations of available techniques such as manometry or transit studies. MII is a new technique that detects flow in a viscous organ by measuring changes in intrahiminal impedance. Using combined videofluornscopy, MII and manonetry, we found that MII is more sensitive than manometry in detecting intestinal flow (Neurogastro 2002; 14:430). Aim: To determine patterns of intestinal flow and their relation to pressure events in fasting (F) and postprandial (PP) periods. Methods: 12 healthy subjects had simultaneous manometry and Mr[ of the duodenum by a probe incorporating 5 pressure sensors and 5 pairs of electrodes spaced 5 cm apart. Tracings inspected visually for presence and distance of propagation of sequences of impedance and pressure events. We analyzed 30 minutes in phase ll, following the first phase lI[, and 30 minutes after ingestion of 500ml of BOOST. Short sequences defined as present in I and 2 channels, long sequences defined as propagated over 4 and 5 channels. Percentages of Mll and pressure events having differing lengths were compared between periods using Cochran-Mantel-Haemzel mean scores tests. Other tests used mixed models ANOVA, p<0.05. Results: There were more events in the PP period compared to F. The distribution of MII sequence lengths differed significantly between the periods, with PP having more short sequences and F having longer ones (table). MII sequences, of various lengths, showed a significantly higher association with corresponding pressure sequences in the PP period as compared to F. 20% and 36% of long Mr[ sequences were not associated with contractions in F and PP periods respectively. Motility index values were significantly associated with increasing le'ngcha of MII propagation in F (P
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Nutrient Modulation of Intestinal Gas Dynamics in Healthy Humans Depends on Caloric Content and Physical Properties of the Meal Sutep Gonlachanvit, Radoslav Coleski, William L. Hasler Patients with gas and bloating report increased symptoms after ingestion of meals with specific properties~ We have showed that liquid caloric meals stimulate transit of jejunallyperfused gas, but the effects of solid or non-caloric meals are unexplored. We hypothesized that meals of different caloric density and physical properties modulate gas transit to different degrees. 10 healthy subjects underwent jejunal perfusion of physiologic gas mixtures (88% Nz, 5.5% 02, 6.5% COL) at 12 ml/min x 3 hr with ingestion of nothing (control), water (240 ml), a 240 kcal liquid meal (240 ml, 41 gm CHO, 4 gm fat, and 10 gm protein), and a 240 kcal solid meal (toast and scrambled eggs) at the end of the second hour in separate studies Gas was collected from an mtrarectal catheter to bypass the anus and evacuation was quantified in rcal-time using a harostat. An initial lag phase was noted after starting gas perfusmn, then gas was evacuated from the rectum at i2.7+-0.7, 12.7_+ 1.2, 13.7_+0.8, and 137 _+1.3 ml/min (P=NS) for the control, water, liquid meal, and solid meal arms. Solid and liquid meals increased cumulative gas volumes evacuated during the initial 20 minutes after eating (451 _+128 and 410_+51 ml vs. 254_+54 ml for control; P
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Table: MII and pressuresequencesin F and PP pedods ,r,,~ MH Pedods
F PP *P<0.05
Total
Short Mil
prmm
~qmma
~ 521 1187
511 1264
~) 64,1 70.1"
Short~
LongMII
sure~
q ~ 75.9 78.1
(%)
Long pre~
Nq-an,m sure~. (~) q u m ~ (~) 20.7" 15.5
15.8 10.2
310 Fat-Induced Ileal Brake Depends on an Opioid Pathway Acting on Kappa Receptors Henry C. Lin, Jin Hal Chen, Corynn Neeve| Background: Slowing of intestinal transit by fat as the ileal brake depends on a naloxone (nonspecific antagonist)-sensitive opioid pathway (Zhao et al. AJP 278: G866-70, 2000). Dynorphin (acting on kappa opioid receptors) immunoreactive nerves are found in the intestine (Costa & Fumess. Neuropeptides 5: 445, 1985). However, the exact role of kappa opinid receptors is unknown. Aim: To test the hypothesis that slowing of intestinal transit by fat may depend on an opioid pathway acting on kappa opioid receptors, we compared the fat-induced ileal brake with and without a kappa opioid receptor antagonist. Methods: 4 dogs were equipped with duodenal and midgnt fistulas. With occluding Foley catheters in the distal limb of the 2 fistulas, the small intestine was compartmentalized into the
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