Imaging of chronic inflammatory bowel disease with 18F-FDG PET in children and adolescents.

Standard for diagnosis of inflammatory bowel disease (IBD) is the endoscopy of the stomach and the intestine. Aim of this study was to determine the value of 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) in pediatric patients with mild to moderate IBD.We included 23 children and adolescents between 8 and 17 years (median 15 years, 13 boys, 10 girls) in this retrospective study in a routine clinical setting. Diagnoses were Crohn's disease in 19 and ulcerative colitis in 4 cases.3 children had a conventional FDG-PET, 20 patients a combined FDG-PET-computed tomography exam. All children had upper and lower intestinal endoscopy with biopsy and a Hydro-MRI exam to assess the jejunum and proximal ileum. The gastrointestinal tract was divided in 7 segments: Stomach plus duodenum, jejunum and proximal ileum, terminal ileum, cecum plus ascending colon, transverse colon, descending colon, and rectosigmoid.Superficial gastric lesions were missed, gastric ulcerations were detected. For the stomach, the sensitivity was 0.25, the specificity was 1.00, the positive predictive value was 1.00, for the lower intestine (terminal ileum and colon) the values were 0.74, 0.88, and 0.96; for the terminal ileum 0.89, 0.75 and 0.94, respectively.The sensitivity and specificity for of ileal and colonic lesions is high. FDG-PET has to be discussed as a tool for the determination of extent and degree of inflammation, especially in those parts of the small bowel that are not accessible to endoscopy. This has to be weighed against the additional radiation exposure administrated.

Coeliac disease diagnosis: ESPGHAN 1990 criteria or need for a change? Results of a questionnaire.

BACKGROUND AND OBJECTIVES: A revision of criteria for diagnosing coeliac disease (CD) is being conducted by The European Society for Pediatric Gastroenterology, Hepatology, and Nutrition (ESPGHAN). In parallel, we have performed a survey aimed to evaluate present practices for CD among paediatric gastroenterologists and to learn their views on the need for modification of present criteria for CD diagnosis. PATIENTS AND METHODS: Questionnaires were distributed to experienced paediatric gastroenterologists (ESPGHAN members) via the Internet. RESULTS: Overall, 95 valid questionnaires were available for analysis, pertaining to 28 different countries, with the majority of responders treating patients with CD for >15 years. Only about 12% of the responders comply with present criteria, noncompliance being related mainly to the challenge policy. Approximately 90% request a revision and modification of the present criteria. Forty-four percent want to omit the small bowel biopsy in symptomatic children with positive anti-tissue transglutaminase immunoglobulin (Ig) A or endomysial IgA antibodies, especially if they are DQ2/DQ8 positive. For silent cases detected by screening with convincingly positive anti-tissue transglutaminase IgA or EMA IgA, about 30% consider that no small bowel biopsy should be required in selected cases. Adding human leukocyte antigen typing in the diagnostic workup was asked for by 42% of the responders. As for gluten challenge, a new policy is advocated restricting its obligation to cases whenever the diagnosis is doubtful or unclear. CONCLUSIONS: Based on these opinions, revision of the ESPGHAN criteria for diagnosing CD is urgently needed.

[Prophylactic prenatal therapy with intravenous immunoglobulins for patients at risk for neonatal haemochromatosis]. / Prophylaktische pränatale Therapie mit intravenösen Immunglobulinen bei Risiko für eine Neonatale Hämochromatose.

Neonatal haemochromatosis (NH) is a connatal hepatopathy that is lethal in 32% and necessitates liver transplantation in 63% of the survivors. The classical diagnostic criteria of extrahepatic siderosis do not apply in all patients who are suspected to have NH. The hypothesis of NH as an alloimmune disease is supported by the quantitative immunohistochemical proof of C5b-9 complement complexes on the hepatocytes of liver biopsy material. This has opened a new perspective in the therapy and prophylaxis for this severe disease. Prophylactic therapy with intravenous immunoglobulins (IVIG) for mothers at risk can prevent a relevant NH in most cases.

Partial splenic embolization as an alternative to splenectomy in hypersplenism--single center experience in 16 years.

PROBLEM: In young patients with hypersplenism splenectomy implies a lifelong increased risk for post-splenectomy infection. Especially in children, whose immune system is not yet completely matured, the risk for some bacterial infection may increase after splenectomy because the spleen helps to defend against encapsulated bacteria like pneumococci, meningococci and haemophilus influenzae. We present partial splenic embolization as an alternative to surgical splenectomy. METHOD: Partial splenic embolization was performed in 17 patients from 1-31 years with hypersplenism of various etiologies and was achieved by selective catheterization of splenic arteries and injection of 150-355 µm polyvinyl alcohol particles (Ivalon (®)). After the intervention the patients received an intensified analgesic regimen and antibiotics to avoid concurrent infectious complications. RESULTS: Partial splenic embolization represented between 30-60% of the splenic volume and was followed in general by an immediate increase of all blood cells and symptoms of hypersplenism were reduced. In 2 patients the procedure was repeated because the result of the first embolization was insufficient in one patient and became necessary in another in the long run. Post-procedural side effects included fever, abdominal pain, ascites and pleural effusions. There were no acute infections in any patient. CONCLUSION: Our monoinstitutional experiences over 16 years offer, partial splenic embolization in patients with hypersplenism from miscellaneous reasons as a low-risk alternative to surgical splenectomy. The procedure can be repeated as necessary, but it is always a temporary palliation depending on the underlying disease which often leads to liver transplantation. Using intensive analgesia and antibiotics side effects were tolerable, and patients could be discharged after a few days.

High diagnostic value of 18F-FDG-PET in pediatric patients with chronic inflammatory bowel disease.

Diagnosis of chronic inflammatory bowel disease (IBD) in children requires noninvasive, atraumatic diagnostic tools that depict localization and acuity of inflammation and yield only a low radiation dose. This retrospective analysis evaluates the diagnostic potential of FDG-PET. Twenty-six consecutive FDG-PET scans of 23 patients (age: 2-16, years, 14 M, 9 F) with suspected IBD were analyzed in this retrospective study. Results were compared to endoscopic, histologic, and abdominal ultrasound (US) finding. In these examinations, presence of inflammation was evaluated in each patient in 8 bowel segments (score 1-4). Standardized uptake values (SUVs) for FDG-PET were measured for all segments. Sensitivity, specificity, and accuracy were calculated using histology as the standard of reference on a segment-based analysis (pathologic if inflammation score > or = 3 or SUV(max)/SUV(liver)>1.2). With histology as the standard of reference, FDG-PET showed a sensitivity/specificity/accuracy of 98%/68%/8%/3 as compared to endoscopy (90%/75%/82%) and US (56%/92%/75%). For the small bowel, FDG-PET was even more reliable (100%/86%/90%). Because of its high sensitivity and accuracy,FDG-PET is an excellent, noninvasive diagnostic tool for IBD. Depicting inflammation in the whole bowel, while being not traumatic, it is attractive for use especially in children. FDG-PET is especially reliable for the small bowel and can inform application of topical therapy.

Autophagocytosis of the apical membrane in microvillus inclusion disease.

BACKGROUND: Microvillus inclusion disease (MID) is a disorder with the clinical signs of intractable diarrhoea in the newborn and infancy. The typical pathological features of the disease are well known whereas the pathophysiology is still unclear. AIM: This study was performed to define possible alterations of the cytoskeleton and exocytic as well as endocytic pathways within enterocytes in MID. PATIENTS: Four patients with MID were studied. Three had a congenital onset of diarrhoea and one patient had a late onset form. METHODS: Thin frozen sections of small bowel biopsies of patients were labelled by antibodies against the cytoskeleton and the brush border enzyme sucrase-isomaltase. The binding sites of the primary antibodies were visualised by immunogold particles in the electron microscope. Biopsies were labelled in organ culture to analyse the biosynthetic and endocytic pathways within the enterocytes. RESULTS: Labelling with antibodies against actin and villin did not differ significantly in control and patient biopsies. Biosynthetic labelling revealed normal intracellular processing and transport of the brush border enzyme sucrase-isomaltase. Secretory granules in crypt epithelial cells were positive for sucrase-isomaltase, differing in its labelling density between patients. Patient biopsies showed microvillus inclusion bodies which endocytosed cationised ferritin within five minutes after uptake as well as ovalbumin after incubation for 10 minutes. These microvillus inclusion bodies correspond to early endosomes because they lack lysosome associated membrane proteins. Late endosomes and lysosomes containing sucrase-isomaltase did not reveal microvillus-like structures. CONCLUSION: Microvillus inclusion bodies in MID originate from autophagocytosis of the apical membrane of enterocytes with engulfing of microvilli.

Differences in the human and mouse amino-terminal leader peptides of ornithine transcarbamylase affect mitochondrial import and efficacy of adenoviral vectors.

Mouse models of ornithine transcarbamylase (OTC) deficiency are being used to test the efficacy of viral vectors as possible vehicles for gene therapy. However, it has been demonstrated that virus containing the human OTC cDNA failed to express functional OTC enzyme in the recipient animals. Because functional OTC is assembled as a homotrimer in the mitochondria, there are at least two possible explanations for these results. Either endogenous mutant protein coassembles with the human OTC and has a "dominant-negative effect," or the human version of the protein is not appropriately imported or processed in the mouse mitochondria. To test the importance of processing, which in rodents is thought to depend on the leader peptide, adenoviral vectors containing chimeric OTC cDNAs were prepared. These vectors were evaluated in the OTC-deficient sparse fur mouse models. Although comparable levels of transgene expression were observed in all groups of mice, the only mice that had high levels of OTC activity and mitochondrial OTC immunoreactivity were those mice injected with the vectors containing the mouse leader peptide (mouse OTC and a mouse-human chimera of OTC). To address possible dominant-negative effects, adenoviruses containing mutant human or mouse OTC cDNAs were prepared and evaluated in cell lines or normal C3H mice, respectively. No inhibition of normal OTC activity was observed in either model system. Together, these studies provide no evidence of a dominant-negative effect and suggest that the human and rodent enzymes responsible for transporting of OTC and possibly other mitochondrial proteins have different specificity.

Congenital sucrase-isomaltase deficiency arising from cleavage and secretion of a mutant form of the enzyme.

Congenital sucrase-isomaltase deficiency (CSID) is an autosomal recessive human intestinal disorder that is clinically characterized by fermentative diarrhea, abdominal pain, and cramps upon ingestion of sugar. The symptoms are the consequence of absent or drastically reduced enzymatic activities of sucrase and isomaltase, the components of the intestinal integral membrane glycoprotein sucrase-isomaltase (SI). Several known phenotypes of CSID result from an altered posttranslational processing of SI. We describe here a novel CSID phenotype, in which pro-SI undergoes an unusual intracellular cleavage that eliminates its transmembrane domain. Biosynthesis of pro-SI in intestinal explants and in cells transfected with the SI cDNA of this phenotype demonstrated a cleavage occurring within the endoplasmic reticulum due to a point mutation that converts a leucine to proline at residue 340 of isomaltase. Cleaved pro-SI is transported to and processed in the Golgi apparatus and is ultimately secreted into the exterior milieu as an active enzyme. To our knowledge this is the first report of a disorder whose pathogenesis results not from protein malfolding or mistargeting, but from the conversion of an integral membrane glycoprotein into a secreted species that is lost from the cell surface.

[Arthrogryposis, renal tubular dysfunction, cholestasis (ARC) syndrome: case report and review of the literature]. / Arthrogryposis, renale tubuläre Dysfunktion, Cholestase (ARC)-Syndrom: Fallbeschreibung und Literaturübersicht.

The ARC-syndrome is a rare disease with the obligatory symptoms arthrogryposis, renal tubular dysfunction and cholestasis. Optional further symptoms like ichthyosis, diarrhea, central nervous system defects and recurrent infections have been reported. The ARC-syndrome was first reported by Lutz-Richner and Landolt in 1973. The pathophysiology is still unknown, an autosomal recessive inheritance is postulated. Patients rarely exceed an age of six month. We report a boy of consanguineous Turkish parents who suffered from congenital deformities of the lower extremities, a metabolic acidosis and failure to thrive. In the sequel he developed a renal Fanconi syndrome and cholestasis. Histology of liver and muscle biopsy specimen showed the typical findings of the disease with giant cell hepatitis and neurogenous muscle atrophy. His condition could be stabilized and he increased in weight by substituting fluid, electrolytes, buffer and parenteral nutrition. Total enteral nutrition of the 280 ml/kg/d he required failed even by nasogastric tube and percutaneous endoscopic gastrostomy. Additional fluid substitution by central venous catheter remained necessary. At the age of 7 month he died.

Modulation of antigen trafficking to MHC class II-positive late endosomes of enterocytes.

BACKGROUND & AIMS: Oral tolerance is recognized as a central immunoregulatory phenomenon. The mechanisms of its induction remain unclear, and the role of the intestinal epithelial cells that are able to present antigens to T lymphocytes is poorly understood. In this report, we analyze under in vivo conditions the intracellular targeting of mucosally administered ovalbumin (OVA) to major histocompatibility complex (MHC) class II antigen containing compartments of enterocytes and compare these pathways between BALB/c and SCID mice, the latter being unable to generate a transferable tolerogenic moiety after a feed of OVA. METHODS: OVA, lysosome-associated membrane proteins (LAMP-1), and MHC class II antigens were localized in jejunal biopsy specimens of BALB/c and SCID mice at 0, 5, 10, 20, 40, 60, and 120 minutes after a single feed with OVA by fluorescence and electron microscopy. RESULTS: Ten minutes after oral administration, OVA was transported to the proximity of MHC class II antigens within LAMP-1-positive vacuoles and to the basolateral membrane of enterocytes from BALB/c strain mice. However, in SCID mice, OVA reached the paracellular spaces during the same time period through LAMP-1-negative vacuoles of enterocytes, which lacked MHC class II antigens. CONCLUSIONS: Orally administered OVA is rapidly targeted to late endosomes containing LAMP-1 and MHC class II antigens in enterocytes of BALB/c mice but not in SCID mice bred on a BALB/c background. We suggest that this targeting process within the enterocytes is one of the requirements for the induction of oral tolerance.

Intracellular transport of acid beta-glucosidase and lysosome-associated membrane proteins is affected in Gaucher's disease (G202R mutation).

Gaucher's disease (GD) is caused by an inherited deficiency of acid beta-glucosidase with storage of glucosylceramides in the lysosomes of macrophages. This study identifies a G202R mutation in the acid beta-glucosidase gene in an infant with severe neuronopathic (type 2) GD and only slightly reduced acid beta-glucosidase activity. Western blot analysis, pulse chase experiments, and the thin frozen section immunogold method were used to analyse the implications of this mutation on the pathogenesis, clinical heterogeneity and diagnostic evaluation of GD. The results show that acid beta-glucosidase persists in the patient's fibroblasts as a mannose-rich polypeptide in the endoplasmic reticulum and is not transported to the lysosomes. By contrast, high expression of the lysosome-associated membrane proteins LAMP-1 and LAMP-2, saposin C, and cathepsin D was observed in the patient's lysosomes. Immunogold labelling of the integral membrane proteins LAMP-1 and LAMP-2 increases significantly at the cell surface of Kupffer cells and fibroblasts as well as at the apical membrane of hepatocytes. In addition, LAMP-1 and LAMP-2 associate with the bilayer of stored glucosylceramide. It is concluded that defective intracellular transport of mutant acid beta-glucosidase from the endoplasmic reticulum to lysosomes leads to a more severe clinical phenotype than the residual enzyme activity may indicate. Furthermore, the detection of LAMP in the tubular bundles of undigested glucosylceramides, as well as their increased concentration at the surfaces of the affected cells, suggests that these proteins play a role in the storage or removal of substrate in GD. Intracellular targeting of acid beta-glucosidase and LAMP contributes to the broad phenotypic heterogeneity of GD.

Efficient mitochondrial import of newly synthesized ornithine transcarbamylase (OTC) and correction of secondary metabolic alterations in spf(ash) mice following gene therapy of OTC deficiency.

BACKGROUND: The mouse strain sparse fur with abnormal skin and hair (spf(ash)) is a model for the human ornithine transcarbamylase (OTC) deficiency, an X-linked inherited urea cycle disorder. The spf(ash) mouse carries a single base-pair mutation in the OTC gene that leads to the production of OTC enzyme at 10% of the normal level. MATERIALS AND METHODS: Recombinant adenoviruses carrying either mouse (Ad.mOTC) or human (Ad.hOTC) OTC cDNA were injected intravenously into the spf(ash) mice. Expression of OTC enzyme precursor and its translocation to mitochondria in the vector-transduced hepatocytes were analyzed on an ultrastructural level. Liver OTC activity and mitochondrial OTC concentration were significantly increased (300% of normal) in mice treated with Ad.mOTC and were moderately increased in mice receiving Ad.hOTC (34% of normal). The concentration and subcellular location of OTC and associated enzymes were studied by electron microscope immunolocalization and quantitative morphometry. RESULTS: Cytosolic OTC concentration remained unchanged in Ad.mOTC-injected mice but was significantly increased in mice receiving Ad.hOTC, suggesting a block of mitochondria translocation for the human OTC precursor. Mitochondrial ATPase subunit c [ATPase(c)] was significantly reduced and mitochondrial carbamy delta phosphate synthetase I (CPSI) was significantly elevated in spf(ash) mice relative to C3H. In Ad.mOTC-treated mice, the hepatic mitochondrial concentration of ATPase(c) was completely normalized and the CPSI concentration was partially corrected. CONCLUSIONS: Taken together, we conclude that newly synthesized mouse OTC enzyme was efficiently imported into mitochondria following vector-mediated gene delivery in spf(ash) mice, correcting secondary metabolic alterations.

Hereditary pancreatitis and mutation of the trypsinogen gene.

Hereditary pancreatitis is a rare form of chronic recurrent pancreatitis. A family, in which 11 members had chronic pancreatitis, five had diabetes, and two had pancreatic cancer, was studied, and hereditary pancreatitis was diagnosed in all patients by demonstrating the mutation in exon 3 of the cationic trypsinogen gene (R117H). The clinical implications of genotypic analysis in hereditary pancreatitis are discussed.

Transepithelial transport processes at the intestinal mucosa in inflammatory bowel disease.

Crohn's disease (CD) and ulcerative colitis (UC) are inflammatory bowel diseases (IBD) of unknown etiology. Oral absorption studies have shown an increased intestinal permeability for various sugar molecules in patients with IBD and their healthy relatives as a possible pathogenetic factor. However, the various transport pathways through the mucosal barrier have not yet been examined. This study therefore investigated whether antigens pass the epithelial barrier by a transcellular or a paracellular pathway. Mucosa of freshly resected specimens from CD (n = 10) or UC (n = 10) patients was investigated by immunoelectron microscopy and compared with healthy mucosa. Epithelial transport was studied with the antigens ovalbumin and horseradish peroxidase after defined incubation. Labeling density of subunit c of ATP synthetase was determined in mitochondria of enterocytes of all specimens. In all specimens epithelial transport of OVA and HRP was principally transcellular through enterocytes with normal ultrastructure, although some tight junctions in CD and UC were dilated. Antigens were transported within vesicles to the basolateral membrane 2.5 min after incubation. The level of enterocytes with electron-lucent cytoplasm containing a high amount of antigens was higher in CD and UC than in healthy mucosa, depending on the grade of inflammation. ATP synthetase was significantly decreased in electron-lucent cytoplasm of CD and UC to normal ultrastructure of healthy mucosa. Our study shows that ovalbumin and horseradish peroxidase taken up by the apical membrane reach the paracellular space by vesicular transport in healthy and IBD enterocytes within a few minutes. Transcellular pathway is affected in both CD and UC, which is indicated by a high level of antigens within the cytosol. We speculate that increased intestinal permeability in IBD results substantially from enhanced transcellular transport.

Coagulation and fibrinolysis in children, adolescents, and young adults with inflammatory bowel disease.

BACKGROUND: Patients with Crohn's disease and ulcerative colitis have an increased risk of thromboembolic events. METHODS: Data were collected from 24 patients aged 4.5 to 23 years who had inflammatory bowel disease. Platelet count, antithrombin, fibrinogen, prothrombin fragment F1+2, soluble thrombomodulin, tissue plasminogen activator, D-dimer, and plasminogen activator inhibitor-1 antigen were investigated. In addition the response to activated protein C, the factor V R506Q mutation, protein C, free protein S antigen, and lipoprotein (a) were analyzed. These data were compared with medical treatment, duration, and disease activity, estimated with the Pediatric Crohn's Disease Activity Index or the Clinical Colitis Activity Index. RESULTS: Forty-five percent of our patients showed an increase in fibrinogen, 29% in prothrombin fragment F1+2, and 20% in platelet count, plasminogen activator inhibitor- antigen, and soluble thrombomodulin. Thrombomodulin was higher in active disease than in inactive disease and in Crohn's disease than in ulcerative colitis. Fibrinogen was also higher with Crohn's disease and tended to be higher in active disease than in ulcerative colitis and inactive disease. Plasminogen activator inhibitor-1 antigen was significantly higher in patients with Crohn's disease than in those with ulcerative colitis and was higher in the patient group treated with steroids. CONCLUSION: As has been shown in adults, young patients with active and inactive inflammatory bowel disease were found to have abnormal coagulation and fibrinolysis. The relevance as a thromboembolic risk factor is discussed.

False-positive gliadin and endomysium antibodies and exocrine pancreatic insufficiency as pitfalls in the differential diagnosis of duodenal Crohn's versus celiac disease.

We report a 13-yr-old boy with Crohn's disease in the upper gastrointestinal tract presenting with abdominal pain, failure to thrive, recurrent fever, iron-deficient anemia, and exocrine pancreatic insufficiency. Initially, latent celiac disease was suggested because of normal endoscopic findings, the finding of non-specific inflammation on histological evaluation of duodenal biopsies, positive IgA and IgG gliadin, as well as endomysium antibodies and exocrine pancreatic insufficiency. There was no response to a gluten-free diet. A reevaluation revealed Crohn's disease.

Ultrastructural localization of IgG and TPO in autoimmune thyrocytes referring to the transcytosis of IgG and the antigen presentation of TPO.

While autoantibodies against thyroid peroxidase (TPO) are known to produce cytotoxicity in vitro, their in vivo effects are still obscure. In addition, the mechanism of TPO autoantibody creation needs to be disclosed because the localization of TPO on thyrocytes is considered to be restricted to the apical membrane, which is not in contact with immunocompetent cells. In order to study these crucial processes in the pathogenesis of thyroid autoimmunity, the ultrastructural localization of TPO and IgG was determined and quantified in thyrocytes of normal thyroid gland and thyroid tissue of patients suffering from Graves' disease. This was done by using ultrathin frozen sections and the immunogold method. IgGs were detected in the follicular lumen, close to the apical membrane, in transport vesicles, the endoplasmic reticulum, and the Golgi apparatus of thyrocytes from patients with Graves' disease. The labeling of TPO in the basolateral membrane was distinctly lower than that of the apical membrane, but was significant in comparison to the plasma membrane labeling of fibroblasts present in the same sections. These data indicate that thyroid autoantibodies may perform their cytotoxic function in intracellular compartments besides the plasma membrane. TPO molecules on the basolateral membrane of HLA class II antigen-positive thyrocytes may initiate antigen presentation of TPO as well as the formation and uptake of TPO autoantibodies.