Cell-based BSEP trans-inhibition: A novel, non-invasive test for diagnosis of antibody-induced BSEP deficiency.

Background & Aims: Antibody-induced bile salt export pump deficiency (AIBD) is an acquired form of intrahepatic cholestasis, which may develop following orthotopic liver transplantation (OLT) for progressive familial intrahepatic cholestasis type 2 (PFIC-2). Approximately 8-33% of patients with PFIC-2 who underwent a transplant develop bile salt export pump (BSEP) antibodies, which trans-inhibit this bile salt transporter from the extracellular, biliary side. AIBD is diagnosed by demonstration of BSEP-reactive and BSEP-inhibitory antibodies in patient serum. We developed a cell-based test directly measuring BSEP trans-inhibition by antibodies in serum samples to confirm AIBD diagnosis. Methods: Sera from healthy controls and cholestatic non-AIBD or AIBD cases were tested (1) for anticanalicular reactivity by immunofluorescence staining of human liver cryosections, (2) for anti-BSEP reactivity by immunofluorescence staining of human embryonic kidney 293 (HEK293) cells expressing BSEP-enhanced yellow fluorescent protein (EYFP) and immunodetection of BSEP-EYFP on Western blot, and (3) for BSEP trans-inhibition using HEK293 cells stably expressing Na+/taurocholate cotransporting polypeptide (NTCP)-mCherry and BSEP-EYFP. The trans-inhibition test uses [3H]-taurocholate as substrate and is divided into an uptake phase dominated by NTCP followed by BSEP-mediated export. For functional analysis, sera were bile salt depleted. Results: We found BSEP trans-inhibition by seven sera containing anti-BSEP antibodies, but not by five cholestatic or nine control sera, all lacking BSEP reactivity. Prospective screening of a patient with PFIC-2 post OLT showed seroconversion to AIBD, and the novel test method allowed monitoring of treatment response. Notably, we identified a patient with PFIC-2 post OLT with anti-BSEP antibodies yet without BSEP trans-inhibition activity, in line with asymptomatic presentation at serum sampling. Conclusions: Our cell-based assay is the first direct functional test for AIBD and allows confirmation of diagnosis as well as monitoring under therapy. We propose an updated workflow for AIBD diagnosis including this functional assay. Impact and Implications: Antibody-induced BSEP deficiency (AIBD) is a potentially serious complication that may affect patients with PFIC-2 after liver transplantation. To improve its early diagnosis and thus immediate treatment, we developed a novel functional assay to confirm AIBD diagnosis using a patient's serum and propose an updated diagnostic algorithm for AIBD.

[Fecal Microbiota Transfer (FMT) in Children and Adolescents - Review and statement by the GPGE microbiome working group]. / Fäkaler Mikrobiota Transfer (FMT) bei Kindern und Jugendlichen ­ Review und Stellungnahme der GPGE AG Mikrobiom.

The human microbiome and especially the gastrointestinal microbiota are associated with health and disease. Disturbance in the composition or function of fecal microbiota (dysbiosis) plays a role in the development of pediatric gastrointestinal diseases. Fecal microbiota transfer (FMT) is a special intervention, where microbiota are transferred from a healthy donor.In this review we describe the current state of knowledge for FMT in pediatric patients. There is satisfactory evidence concerning FMT in patients with recurrent C. difficile infection. For inflammatory bowel disease, few studies show a potential benefit.Adverse events occurred frequently in clinical studies, but were mostly mild and transient. There are hardly any data on long-term side effects of FMT, which are particularly significant for pediatrics. In practice, there is uncertainty as to which application route, dosage or frequency should be used. Legally, donor stool is considered a drug in German-speaking countries, for which no marketing authorization exists.In conclusion, knowledge about physiology, efficacy and side effects of FMT is insufficient and legal concerns complicate its implementation. More studies on this topic are needed urgently.

Duodenal mast cells and eosinophils in children with celiac disease: occurrence and distribution pattern.

OBJECTIVE: The aim of this study was to characterize duodenal mast cell (MC) and eosinophil (EO) numbers, their distribution within the lamina propria and possible impact on disease severity of paediatric celiac patients compared to children without celiac disease (CD). METHODS: We analysed duodenal samples of 215 children (109 CD, 106 controls) who underwent esophagogastroduodenoscopy from 2010 to 2018. After immunohistochemical staining, average MC and EO counts were histologically examined in ten high-power-fields. Additionally, cell-distribution within the lamina propria was analysed. Possible influence of relevant clinical parameters was evaluated. STATISTICS: Student's-t-test, Mann-Whitney U-test, Chi-square-test, ANOVA, significance-level <.05. Trial registration-number: DRKS00024669. RESULTS: MC-density was higher in CD-patients compared to the control-group (23.7 (±12.1)/HPF versus 19.7 (±9.1)/HPF; p = .008), varying in number interindividually. Eosinophils were also increased in the duodenum of celiac patients (23.3 (±9.3)/HPF versus 12.2 (±6.3)/HPF; p= <.001). MCs were distributed more often homogenously in all parts of CD lamina propria (44 biopsies (40.4%), residing more distant from the intestinal lumen in controls (0 biopsies with homogenous distribution-pattern (0%); p= <.001). Regarding EOs no polarity was observable. Atopic diseases did not occur significantly more often in patients with elevated EO-counts. CONCLUSION: MC- and EO-numbers were increased in the duodenum of CD-patients and MCs showed a different distribution-pattern in the lamina propria of celiac patients. These findings support the concept that both cell-types contribute to disease-pathogenesis. However, functional studies highlighting both cell-types' and their mediators' role regarding mucosal alterations during the course of the inflammatory process in celiac patients are needed. TRIAL REGISTRATION NUMBER AND URL: DRKS00024669; https://www.drks.de/drks_web/.

Local and Systemic Alterations of the L-Arginine/Nitric Oxide Pathway in Sputum, Blood, and Urine of Pediatric Cystic Fibrosis Patients and Effects of Antibiotic Treatment.

Alterations in the L-arginine (Arg)/nitric oxide (NO) pathway have been reported in cystic fibrosis (CF; OMIM 219700) as the result of various factors including systemic and local inflammatory activity in the airways. The aim of the present study was to evaluate the Arg/NO metabolism in pediatric CF patients with special emphasis on lung impairment and antibiotic treatment. Seventy CF patients and 78 healthy controls were included in the study. CF patients (43% male, median age 11.8 years) showed moderately impaired lung functions (FEV1 90.5 ± 19.1% (mean ± SD); 21 (30%) had a chronic Pseudomonas aeruginosa (PSA) infection, and 24 (33%) had an acute exacerbation). Plasma, urinary, and sputum concentrations of the main Arg/NO metabolites, nitrate, nitrite, Arg, homoarginine (hArg), and asymmetric dimethylarginine (ADMA) were determined in pediatric CF patients and in healthy age-matched controls. Clinical parameters in CF patients included lung function and infection with PSA. Additionally, the Arg/NO pathway in sputum samples of five CF patients was analyzed before and after routine antibiotic therapy. CF patients with low fractionally exhaled NO (FENO) showed lower plasma Arg and nitrate concentrations. During acute exacerbation, sputum Arg and hArg levels were high and dropped after antibiotic treatment: Arg: pre-antibiotics: 4.14 nmol/25 mg sputum vs. post-antibiotics: 2.33 nmol/25 mg sputum, p = 0.008; hArg: pre-antibiotics: 0.042 nmol/25 mg sputum vs. post-antibiotics: 0.029 nmol/25 mg sputum, p = 0.035. The activated Arg/NO metabolism in stable CF patients may be a result of chronic inflammation. PSA infection did not play a major role regarding these differences. Exacerbation increased and antibiotic therapy decreased sputum Arg concentrations.

[Crohn's Disease Exclusion Diet - an alternative to exlusive enteral nutritional therapy in children and adolescents with Crohn's disease? Statement of the GPGE working groups CEDATA and Nutrition/Nutrition Medicine]. / Crohnʼs Disease Exclusion Diet (CDED) als Alternative zur exklusiven Ernährungstherapie bei Kindern und Jugendlichen mit Morbus Crohn? Stellungnahme der Arbeitsgemeinschaften CEDATA und Ernährung/Diätetik/Ernährungsmedizin der Gesellschaft für pädiatrische Gastroenterologie und Ernährung e. V.

Epidemiological an clinical observations as well as results from animal studies indicate that nutrition can play a role in the development of inflammatory bowel disease (IBD). Exclusive enteral nutrition therapy represents an example for modulating inflammatory responses solely through diet modification. Therefore, caretakers, patients, families, doctors and nutritionists seek for more dietary options to control IBD. These options include partial enteral nutrition therapy as for example the socalled Crohn's disease exclusion diet. The following statement summarizes existing data and provides recommendations for the current management of enteral nutrition therapy in pediatric Crohn's disease.

Activated L-Arginine/Nitric Oxide Pathway in Pediatric Cystic Fibrosis and Its Association with Pancreatic Insufficiency, Liver Involvement and Nourishment: An Overview and New Results.

Cystic fibrosis (CF; OMIM 219700) is a rare genetic disorder caused by a chloride channel defect, resulting in lung disease, pancreas insufficiency and liver impairment. Altered L-arginine (Arg)/nitric oxide (NO) metabolism has been observed in CF patients' lungs and in connection with malnutrition. The aim of the present study was to investigate markers of the Arg/NO pathway in the plasma and urine of CF patients and to identify possible risk factors, especially associated with malnutrition. We measured the major NO metabolites nitrite and nitrate, Arg, a semi-essential amino acid and NO precursor, the NO synthesis inhibitor asymmetric dimethylarginine (ADMA) and its major urinary metabolite dimethylamine (DMA) in plasma and urine samples of 70 pediatric CF patients and 78 age-matched healthy controls. Biomarkers were determined by gas chromatography-mass spectrometry and high-performance liquid chromatography. We observed higher plasma Arg (90.3 vs. 75.6 µM, p < 0.0001), ADMA (0.62 vs. 0.57 µM, p = 0.03), Arg/ADMA ratio (148 vs. 135, p = 0.01), nitrite (2.07 vs. 1.95 µM, p = 0.03) and nitrate (43.3 vs. 33.1 µM, p < 0.001) concentrations, as well as higher urinary DMA (57.9 vs. 40.7 µM/mM creatinine, p < 0.001) and nitrate (159 vs. 115 µM/mM creatinine, p = 0.001) excretion rates in the CF patients compared to healthy controls. CF patients with pancreatic sufficiency showed plasma concentrations of the biomarkers comparable to those of healthy controls. Malnourished CF patients had lower Arg/ADMA ratios (p = 0.02), indicating a higher NO synthesis capacity in sufficiently nourished CF patients. We conclude that NO production, protein-arginine dimethylation, and ADMA metabolism is increased in pediatric CF patients. Pancreas and liver function influence Arg/NO metabolism. Good nutritional status is associated with higher NO synthesis capacity and lower protein-arginine dimethylation.

Dataset of clinical, immunohistopathological and laboratory features of patients with MHC II deficiency suffering from enteropathy.

Major histocompatibility complex class II (MHC II) is essential for adaptive immune response. We recently reported on disturbed adaptive mucosal immunity due to MHC II deficiency and prolonged enteropathy. Here, we share medical history, flow cytometric analysis of blood lymphocytes, immunohistopathology, and fecal analysis of seven genetically confirmed patients with MHC II deficiency suffering from enteropathy. Data on flow cytometric analysis of HLA-DR expression on monocytes and B cells before hematopoietic stem cell transplantation (HSCT) and after in-vitro stimulation is shown. The course of immune reconstitution after HSCT of MHC II deficient patients in comparison to severe combined immunodeficiency (SCID) patients is described. In addition, immunohistopathology illustrating CD4 and CD8 T cell infiltration, absence of B lymphocytes and plasma cells, and disturbed immunoglobulin expression in the gut as well as absent HLA-DR expression in the liver is shown. Furthermore, data from fecal analysis such as stool fat, nitrogen, and water fraction as well as faecal markers such as alpha-1-antitrypsin, pancreas specific elastase 1, eosinophilic protein X (EPX), and beta defensin 2 are presented. Altogether this data demonstrates the complex phenotype of MHC II deficiency. The data can be valuable for researchers interested in mucosal immunity. For further interpretation of the data presented in this article, please see the research article "Persisting enteropathy and disturbed adaptive mucosal immunity due to MHC class II deficiency" (Posovszky et al., 2019).

Persisting enteropathy and disturbed adaptive mucosal immunity due to MHC class II deficiency.

Intestinal epithelial cells (IECs) form a fundamental mucosal barrier and actively participate in tolerance and immunity against intestinal contents. Major histocompatibility complex class II (MHC II) and invariant chain (Ii) molecules are essential for adaptive immune response. MHC II deficiency often presents with gastrointestinal disorders. Intestinal biopsy samples revealed an absence of HLA-DR, Ii, and local immunoglobulins in both hematopoietic immune cells and IECs accompanied by a lack of faecal sIgA. After successful hematopoietic stem cell transplantation (HSCT) absent HLA-DR and Ii expression persisted in IECs and faecal stool analysis indicated inflammation and high microbial activity. We describe multifaceted disturbance of adaptive mucosal immunity in MHC II deficient patients suffering from enteropathy. HLA-DR and Ii expression on enterocytes is not restored by HSCT. This may account for increased susceptibility to enteric infections and intestinal inflammation leading to prolonged enteropathy reported in MHC II deficient patients.

The L-arginine/NO pathway, homoarginine, and nitrite-dependent renal carbonic anhydrase activity in young people with type 1 diabetes mellitus.

High circulating levels of asymmetric dimethylarginine (ADMA) and low circulating levels of homoarginine (hArg) are known cardiovascular risk factors in adults. While in adults with type 1 diabetes mellitus (T1DM) circulating ADMA is significantly elevated, in children and adolescents the reported ADMA data are contradictory. In 102 children with T1DM and 95 healthy controls (HC) serving as controls, we investigated the L-arginine (Arg)/nitric oxide (NO) pathway. Children with T1DM were divided into two groups, i.e., in children with newly diagnosed diabetes mellitus [T1DM-ND; n = 10; age, 8.8 (4.4-11.2) years; HbA1c, 13 (8.9-13.9) %] and in those with long-term treatment [T1DM-T; n = 92; age, 12.5 (10.5-15.4) years; HbA1c, 8.0 (7.2-8.6) %]. The age of the HC was 11.3 (8-13.3) years. Amino acids and NO metabolites of the Arg/NO pathway, creatinine and the oxidative stress biomarker malondialdehyde (MDA) were measured by GC-MS or GC-MS/MS. Plasma hArg, ADMA and the hArg/ADMA molar ratio did not differ between the T1DM and HC groups. There was a significant difference between T1DM-T and HC with regard to plasma nitrite [0.53 (0.48-0.61) vs 2.05 (0.86-2.36) µM, P < 0.0001] as well as to urinary nitrite [0.09 (0.06-0.17) vs 0.22 (0.13-0.37) µmol/mmol creatinine, P < 0.0001]. Plasma, but not urinary nitrite, differed between T1DM-ND and HC [0.55 (0.50-0.66) vs 2.05 (0.86-2.36) µM, P < 0.0001]. Plasma MDA did not differ between the groups. The urinary nitrate-to-nitrite molar ratio (UNOXR), a measure of nitrite-dependent renal carbonic anhydrase (CA) activity, was higher in T1DM-T [1173 (738-1481), P < 0.0001] and T1DM-ND [1341 (1117-1615), P = 0.0007] compared to HC [540 (324-962)], but did not differ between T1DM-T and T1DM-ND (P = 0.272). The lower nitrite excretion in the children with T1DM may indicate enhanced renal CA-dependent nitrite reabsorption compared with healthy children. Yet, lower plasma nitrite concentration in the T1DM patients may have also contributed to the higher UNOXR. Patients' age correlated positively with plasma hArg and hArg/ADMA and urinary DMA/ADMA. Plasma ADMA and urinary ADMA, DMA, nitrite and nitrate correlated negatively with age of the T1DM-T children. Significant correlations were found between plasma hArg and plasma Arg (r = 0.468, P < 0.0001), and urinary DMA (r = -0.426, P = 0.0001), ADMA (r = -0.266, P = 0.021) and nitrate (r = -0.234, P = 0.043). Plasma hArg correlated positively with age at diagnosis (r = +0.337, P = 0.002). ADMA, but not hArg, correlated with HbA1c in T1DM-T (r = -0.418, P < 0.0001) and T1DM-ND (r = +0.879, P = 0.0016). The greatest differences between T1DM-T and T1DM-ND were observed for urinary ADMA, DMA/ADMA ratio, nitrite and nitrate. The Arg/NO pathway is altered in T1DM in childhood and adolescence, yet the role and the importance of hArg and ADMA in T1DM remain to be elucidated. In young T1DM patients, oxidative stress (lipid peroxidation) is not elevated.

The role of baseline impedance as a marker of mucosal integrity in children with gastro esophageal reflux disease.

OBJECTIVES: Diagnosis of gastro esophageal reflux disease (GERD) in children is challenging. 24-h-pH-multichannel-intraluminal-impedance measurement (pH-MII) is the best diagnostic tool to display gastro esophageal reflux whereas esophageal endoscopy indicates mucosal lesions. The aim of this study was to compare esophageal endoscopy results with reflux parameters such as acid exposure time (reflux index RI), bolus exposure time (bolus index BI), baseline impedance level (BIL) detected by pH-MII in children with suspected GERD. METHODS: Analysis of data from 285 children (38 infants) referred to our hospital with suspected GERD. Division into three 'reflux esophagitis' (RE)-stages depending on the severity of endoscopic and histological findings and comparison with reflux parameters in these stages. Further categorization into four groups based on the pH-MII-results. RESULTS: Children with high-grade esophagitis had a significantly lower BIL; otherwise there was no significant association between elevated reflux parameters and esophagitis. Pathological pH-MII results (RI and BI) were associated with lower BIL in the distal impedance channel. The BIL was significantly lower in infants compared to children >1 year regardless of the RI or BI. The main difference between these groups regarding reflux parameters was a longer BI and a higher number of retrograde bolus movements. CONCLUSION: Pathologic pH-MII results are not predictive for an erosive esophagitis and vice versa. Therefore, these two procedures cannot replace each other. A lower BIL is associated with esophagitis ≥ LA-grade B and may be caused by longer acid but also by longer bolus exposure and thus may be another useful parameter in GERD monitoring.

Inter- and intraobserver agreement in 24-hour combined multiple intraluminal impedance and pH measurement in children.

OBJECTIVES: Assessment of intra- and interobserver agreement in multiple intraluminal impedance (MII) measurement between investigators from different institutions. METHODS: Twenty-four 18- to 24-hour MII tracings were randomly chosen from 4 different institutions (6 per center). Software-aided automatic analysis was performed. Each result was validated by 2 independent investigators from the 4 different centers (4 investigator combinations). For intraobserver agreement, 6 measurements were analyzed twice by the same investigator. Agreement between investigators was calculated using the Cohen kappa coefficient. RESULTS: Interobserver agreement: 13 measurements showed a perfect agreement (kappa > 0.8); 9 had a substantial (kappa 0.61-0.8), 1 a moderate (kappa coefficient 0.41 to 0.6), and 1 a fair agreement (kappa coefficient 0.11-0.4). Median kappa value was 0.83. Intraobserver agreement: 5 tracings showed perfect and 1 showed a substantial agreement. The median kappa value was 0.88. CONCLUSIONS: Most measurements showed substantial to perfect intra- and interobserver agreement. Still, we found a few outliers presumably caused by poorer signal quality in some tracings rather than being observer dependent. An improvement of analysis results may be achieved by using a standard analysis protocol, a standardized method for judging tracing quality, better training options for method users, and more interaction between investigators from different institutions.

Detection of gastroesophageal reflux in children using combined multichannel intraluminal impedance and pH measurement: data from the German Pediatric Impedance Group.

OBJECTIVE: To validate the use of combined multichannel intraluminal impedance (MII)-pH (MII-pH) monitoring for detecting gastroesophageal reflux (GER) in children in daily clinical practice. STUDY DESIGN: The patients were divided into 3 symptom groups based on the main indication for the procedure. MII-pH monitoring was performed in 700 children presenting with symptoms suggestive of GER, including 329 children with pulmonary symptoms, 325 with gastrointestinal symptoms, and 46 with neurologic symptoms. The MII-pH results were compared with pH monitoring alone, and retrograde bolus movements, symptom index, and symptom association probability were compared. RESULTS: Overall, 270 measurements were abnormal: 101 (37%) showed abnormal MII-pH study, 49 (18%) showed only pathological pH measurements and 120 measurements (45%) had an abnormal MII recording only. Extraintestinal symptoms of GER were seen more often in younger children and were more often related to a normal pH study but an abnormal MII study. Infants had a significantly higher number of retrograde bolus movements than older children. Symptom index and symptom association probability showed moderate agreement (Cohen kappa, 0.54). CONCLUSIONS: From this large systematically standardized data collection of MII-pH measurements in children, we conclude that 45% of the patients with abnormal GER would not have been recognized by 24-hour pH measurement alone. Our findings confirm that MII-pH is superior to pH monitoring alone in detecting GER.

Pituitary adenylate cyclase activating polypeptide: an important vascular regulator in human skin in vivo.

Pituitary adenylate cyclase-activating peptide (PACAP) is an important neuropeptide and immunomodulator in various tissues. Although this peptide and its receptors (ie, VPAC1R, VPAC2R, and PAC1R) are expressed in human skin, their biological roles are unknown. Therefore, we tested whether PACAP regulates vascular responses in human skin in vivo. When injected intravenously, PACAP induced a significant, concentration-dependent vascular response (ie, flush, erythema, edema) and mediated a significant and concentration-dependent increase in intrarectal body temperature that peaked at 2.7°C. Topical application of PACAP induced marked concentration-dependent edema. Immunohistochemistry revealed a close association of PACAP-immunoreactive nerve fibers with mast cells and dermal blood vessels. VPAC1R was expressed by dermal endothelial cells, CD4+ and CD8+ T cells, mast cells, and keratinocytes, whereas VPAC2R was expressed only in keratinocytes. VPAC1R protein and mRNA were also detected in human dermal microvascular endothelial cells. The PACAP-induced change in cAMP production in these cells demonstrated VPAC1R to be functional. PACAP treatment of organ-cultured human skin strongly increased the number of CD31+ vessel cross-sections. Taken together, these results suggest that PACAP directly induces vascular responses that may be associated with neurogenic inflammation, indicating for the first time that PACAP may be a crucial vascular regulator in human skin in vivo. Antagonists to PACAP function may be beneficial for the treatment of inflammatory skin diseases with a neurogenic component.

Defensin-mRNA expression in the upper gastrointestinal tract is modulated in children with celiac disease and Helicobacter pylori-positive gastritis.

OBJECTIVES: Defensins are expressed in epithelial cells as cationic peptides with antimicrobial properties. Because of their role as immunologically important effector molecules, their contribution in maintaining a stable microenvironment in the gastrointestinal tract has recently received much attention. The present study was designed to further characterize expression patterns of defensins in diseases of the upper gastrointestinal tract in children, particularly in Helicobacter pylori (Hp)-associated gastritis or celiac disease (CD). PATIENTS AND METHODS: Semiquantitative real-time reverse transcriptase-polymerase chain reaction (PCR) was carried out with gene-specific primers for human beta-defensin 1 to 6 (hBD1 to 6) and human alpha-defensin 5 and 6 (hD5 and 6) in mucosal biopsies of children diagnosed as having CD (n = 11; 4.2-16.2 years) or Hp gastritis (n = 18; 3.2-16.7 years). Levels of expression were compared with those of healthy individuals (n = 21; 2.8-14.6 years). Expression levels in Hp-infected specimens were furthermore compared with those with histologic inflammation not associated with Hp infection (n = 30; 3.6-15.7 years). RESULTS: Expression of hBD2 was upregulated in the antrum and corpus of patients with Hp gastritis, whereas inflammation without detection of Hp was not associated with any change in defensin gene expression. In patients with CD, expression of hBD2 was upregulated in the antrum, whereas hBD1 and 4 were downregulated in duodenal biopsies. CONCLUSIONS: Different pathological conditions of the upper gastrointestinal tract lead to specific modulations of defensin gene expression in children. Especially the pathophysiological role of hBD2 in Hp infection and hBD1 and 4 in CD warrant further attention.

Defensins are differentially expressed with respect to the anatomic region in the upper gastrointestinal tract of children.

Defensins are antimicrobial peptides expressed on various epithelial surfaces. Although they are believed to contribute to intestinal homeostasis, their expression pattern in children is not well characterized. As determined by real-time polymerase chain reaction, amount of human alpha-defensins (hD)-5 and -6 mRNA in duodenal biopsies were significantly higher than in biopsies taken from the gastric mucosa. On the contrary, expression of human beta-defensins (hBD)-1 and -2 mRNA showed a significantly higher expression in the stomach. Expression of hBD3 to 6 was inconsistently detected. These results suggest a distinct role for various defensins in host defense in the upper gastrointestinal tract of children.

Combined esophageal multichannel intraluminal impedance and pH monitoring after repair of esophageal atresia.

OBJECTIVE: Gastroesophageal reflux disease and impaired esophageal motility is a frequent problem after repair of esophageal atresia (EA). Combined multichannel intraluminal impedance (MII) and pH measurement identifies acidic and weakly acidic esophageal bolus movements. Our aim was to evaluate gastroesophageal reflux and its association with gastrointestinal or extraintestinal symptoms using combined MII-pH measurement. PATIENTS AND METHODS: In all, 24 patients (4 months-23 years; 17 male, 7 female) with repaired EA were included in this study. A single catheter with 6 impedance channels and 1 pH channel was used to perform the 24-hour MII-pH study. Symptoms were recorded during the study as events and with a standardized questionnaire. In a subgroup of patients, an impedance swallowing test evaluating liquid and viscous bolus movements was performed. RESULTS: We detected a total of 911 episodes of retrograde bolus movement, 379 acidic and 532 weakly acidic bolus movements. In all, 201 symptom events were recorded. Of these events, 42% were associated with retrograde bolus movements. More symptom events were associated with weakly acidic reflux (26%) than with acid reflux episodes (16%). There was a poor correlation between symptoms and MII-pH study findings. In comparison with patients without esophageal surgery, EA patients showed significantly fewer complete swallows of liquid (42% vs 98%) and viscous material (18% vs 87%). CONCLUSIONS: In patients with corrected EA, half of the reflux events could be detected only by MII. We also demonstrated that weakly acidic reflux can be responsible for the patients' symptoms. However, patients may have few or no symptoms despite poor esophageal function and extensive gastroesophageal reflux disease.

Tailored resective pancreatic surgery for pediatric patients with chronic pancreatitis.

BACKGROUND: Surgical treatment for chronic pancreatitis (CP) in children comprises predominantly nonresective draining procedures. The purpose of this study was to identify indications, techniques, and results of organ-preserving resective pancreatic procedures for pediatric CP at our institution. PATIENTS AND METHODS: A retrospective chart review was performed of all children undergoing pancreatic surgery for CP over a period of 4 years. RESULTS: Overall, 6 pediatric patients (3 male, 3 female, ages 7-18 years) underwent a duodenum-preserving pancreatic head resection (3), a middle segmental pancreatic resection (2), or a distal pancreatectomy (1) for CP of different etiologies (idiopathic 2, posttraumatic 2, pancreas divisum 1, situs inversus 1). No mortality or major surgical complication occurred. Mean operative time was 294 min (207-412 min) and intraoperative blood loss was 541 mL (100-1300 mL). Postoperative hospital stay was 13 days (10-18 days). No endocrine or exocrine insufficiency occurred during follow up of 46 months (25-50 m), and pain control was improved in 5 of 6 patients. CONCLUSIONS: Tailored organ-preserving resective pancreatic surgery can be performed with low morbidity and mortality in pediatric patients with CP and not responding to conservative treatment.

Sensory neuropeptides and epithelial cell restitution: the relevance of SP- and CGRP-stimulated mast cells.

BACKGROUND: Calcitonin-gene-related peptide (CGRP) and substance P (SP) are neurotransmitters of extrinsic primary afferent neurons located within the dorsal root ganglia. In experimental models of colitis in rats and rabbits, a protective role of SP and CGRP on intestinal mucosa was presumed. The mucosal protection partly depends on a CGRP-mediated modulation of mucosal blood flow. Limited data are available regarding CGRP- or SP-mediated effects on epithelial cell restitution. Having shown earlier that SP-stimulated fibroblasts but not CGRP-stimulated fibroblasts induce epithelial cell migration in vitro, the aim of this study was to explore whether mast cells mediate effects of SP and CGRP on epithelial cell restitution in vitro. METHODS: Mast cells (C57) were exposed to SP [10(-12)-10(-6 M)] and CGRP [10(-12)-10(-7 M)]. After a 24-h incubation period, the cell supernatants (conditioned media, CDM) were taken from mast cell cultures and directly applied to rat intestinal epithelial cell lines-18 or Caco-2 monolayers, which had been wounded with a razor blade 24 h prior to the experiments. Epithelial cell migration was assessed by counting cells across the wound edge and epithelial cell proliferation was measured using 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl-tetrazolium bromide test. RESULTS: CGRP significantly induced epithelial cell migration and proliferation via mast cells when supernatants were directly applied to epithelial cells in vitro. The effects on epithelial cell migration were abolished after neutralizing anti-transforming growth factor-beta (TGF-beta) had been added to the cell cultures. SP had no effects on epithelial cells following stimulation of mast cells. CONCLUSION: CGRP modulates epithelial cell restitution in vitro mediated by mast cells. The CGRP- and mast-cell-induced epithelial cell migration is TGF-beta dependent. This observation underlines an important role for extrinsic primary afferent neurons in mucosal defence and repair and in keeping the mucosal homeostasis. This knowledge leads to a better understanding of the interaction of the enteric nervous system and wound healing and may, in the future, lead to new therapeutic approaches to inflammatory diseases of the intestine.