Analysis of synaptic-like microvesicle exocytosis of B-cells using a live imaging technique.

Pancreatic β-cells play central roles in blood glucose homeostasis. Beside insulin, these cells release neurotransmitters and other signaling molecules stored in synaptic-like microvesicles (SLMVs). We monitored SLMV exocytosis by transfecting a synaptophysin-pHluorin construct and by visualizing the cells by Total Internal Reflection Fluorescence (TIRF) microscopy. SLMV fusion was elicited by 20 mM glucose and by depolarizing K(+) concentrations with kinetics comparable to insulin secretion. SLMV exocytosis was prevented by Tetanus and Botulinum-C neurotoxins indicating that the fusion machinery of these organelles includes VAMP-2/-3 and Syntaxin-1, respectively. Sequential visualization of SLMVs by TIRF and epifluorescence microscopy showed that after fusion the vesicle components are rapidly internalized and the organelles re-acidified. Analysis of single fusion episodes revealed the existence of two categories of events. While under basal conditions transient fusion events prevailed, long-lasting episodes were more frequent upon secretagogue exposure. Our observations unveiled similarities between the mechanism of exocytosis of insulin granules and SLMVs. Thus, diabetic conditions characterized by defective insulin secretion are most probably associated also with inappropriate release of molecules stored in SLMVs. The assessment of the contribution of SLMV exocytosis to the manifestation of the disease will be facilitated by the use of the imaging approach described in this study.

Hirschsprung's disease: the "Swiss roll" technique revisited.

PURPOSE: During pull-through for Hirschsprung's disease (HSCR), the assessment of innervation is mainly based on the presence of ganglion cells when conventional Hematoxylin and Eosin (HE) staining is used. In hypoganglionosis, the evaluation is difficult. We adapted a standardized methodology for the examination of resected bowel after HSCR surgery, using the technique described by Moolenbeek on rodent intestine and later by Meier-Ruge in children. We have analysed the entire innervation of surgically resected bowels and compared the results with the follow up of patients. METHODS: Three longitudinal strips of colon were harvested from the mesenteric, anti-mesenteric and intermediate part in the whole length of resected colon of six patients with HSCR. Each strip was divided into two parts. One of the contiguous strips was assessed with HE and Hematoxylin-Phloxin-Safran, and the other one with acetylcholinesterase (AChE) histochemistry. We analyzed the distribution of ganglion cells and nerve arrangement along the strips with both techniques and compared the results obtained in the three different regions of the bowel. RESULTS: There was no significant difference in the pattern of innervation circumferentially. There was a correlation between a progressive increase of AChE activity and nerve hypertrophy and a decrease of ganglion cells from the proximal to the distal part of the resected colon in the submucosa and the myenteric plexus. Nerve hypertrophy and AChE-positive reaction in the mucosa were found at the resection border in patients who presented postoperative complications. CONCLUSIONS: Simultaneous assessment of nerve cells, nerve fibers and AChE activity is important in the evaluation of the innervation of the bowel segment proximal to the aganglionic zone. The method described is feasible and can be adapted to older children and adults with larger bowels. These results point out the importance of assessing nerve fibers in intraoperative biopsies during pull-through procedures to prevent uncomplete surgical bowel resection.

Partial eating disorders among adolescents: a review.

PURPOSE: Many adolescents do not fulfill all the DSM-IV criteria's for anorexia nervosa and bulimia, but do nevertheless suffer from partial eating disorders (EDs). This review focuses on the definition, epidemiology and clinical aspects of these disorders. METHODS: Search on Medline & PsycINFO, review of websites, screening of bibliographies of articles and book chapters. RESULTS: There is still no consensus on the definition of these disorders, which cover a wide range of severity. Affected adolescents often suffer from physical and psychological problems owing to co-morbidity or as a consequence of their eating patterns: chronic constipation, dyspeptic symptoms, nausea, abdominal pain, fatigue, headaches, hypotension, menstrual dysfunction as well as dysthymia, depressive and anxiety disorders, or substance misuse and abuse. In comparison with those who are unaffected, adolescents with partial ED are at higher risk of evolving into full ED. However, most of them evolve into spontaneous remission. Adolescents with partial ED engaged, over a period of several months, in potentially unhealthy weight-control practices, suffering from intense fear of gaining weight and a disturbed body weight/image should be offered therapeutic support. CONCLUSION: Future research should focus on the exact delineation of various subtypes of clinical presentations in partial ED and on evidence-based treatment and follow-up of these various situations.

L'hémisphérotomie péri-insulaire: technique chirurgicale, monitoring EEG intraopératoire et résultats sur le contrôle de l'épilepsie [Periinsular hemispherotomy: surgical technique, intraoperative EEG monitoring and results on seizure outcome]

Peri-insular hemispherotomy is a surgical technique used in the treatment of drug-resistant epilepsy of hemispheric origin. It is based on the exposure of insula and semi-circular sulci, providing access to the lateral ventricle through a supra- and infra-insular window. From inside the ventricle, a parasagittal callosotomy is performed. The basal and medial portion of the frontal lobe is isolated. Projections to the anterior commissure are interrupted at the time of amygdala resection. The hippocampal tail and fimbria-fornix are disrupted posteriorly. We report our experience of 18 cases treated with this approach. More than half of them presented with congenital epilepsy. Neuronavigation was useful in precisely determining the center and extent of the craniotomy, as well as the direction of tractotomies and callosotomy, allowing minimal exposure and blood loss. Intra-operative monitoring by scalp EEG on the contralateral hemisphere was used to follow the progression of the number of interictal spikes during the disconnection procedure. Approximately 90% of patients were in Engel's Class I. We observed one case who presented with transient postoperative neurological deterioration probably due to CSF overdrainage and documented one case of incomplete disconnection in a patient presenting with hemimegalencephaly who needed a second operation. We observed a good correlation between a significant decrease in the number of spikes at the end of the procedure and seizure outcome. Peri-insular hemispherotomy provides a functional disconnection of the hemisphere with minimal resection of cerebral tissue. It is an efficient technique with a low complication rate. Intra-operative EEG monitoring might be used as a predictive factor of completeness of the disconnection and consequently, seizure outcome.

Portal vein embolization with N-butyl cyanoacrylate before partial hepatectomy in patients with hepatocellular carcinoma and underlying cirrhosis or advanced fibrosis.

PURPOSE: To describe the safety, complications, and liver regeneration associated with the left liver after embolization of the right portal vein (PV) in patients with hepatocellular carcinoma (HCC) developed in the setting of advanced liver fibrosis and cirrhosis. MATERIALS AND METHODS: Forty patients (31 men, nine women; mean age, 62 years) with HCC underwent PV embolization over a 4-year period. Embolization was performed from a left PV percutaneous access with use of n-butyl cyanoacrylate (NBCA) mixed with iodized oil. Computed tomography (CT) volumetry was performed before and 1 month after PV embolization to measure the left lobe volume as well as the functional liver ratio defined by the ratio between the left lobe and the total liver volume minus tumoral volume. PV pressure and liver enzyme levels were compared before and 1 month after the procedure and complications were registered. Factors potentially affecting regeneration (age, sex, diabetes, chemoembolization, functional liver ratio before PV embolization, and Knodell histologic score) were evaluated by one-way and stepwise regression analysis. RESULTS: PV embolization could be achieved successfully in all cases. Two patients had partial PV thrombosis on the 1-month follow-up CT and two patients developed transient ascites after PV embolization. The left lobe volume increase was 41% +/- 32% after PV embolization and the functional liver ratio increased from 28% +/- 10% to 36% +/- 10% (P < .0001). Hypertrophy of the left lobe was greater in patients with a low functional liver ratio before PV embolization and those with an F3 fibrosis score. Other factors had no influence on left lobe regeneration. CONCLUSION: PV embolization with use of NBCA is feasible in patients with advanced fibrosis and cirrhosis. Hypertrophy of the left lobe of the liver after PV embolization has a statistically significant correlation with lower functional liver ratio and lower degrees of fibrosis.

Percutaneous extraction of retained biliary T-tubes: a new technique.

Retained T-tubes are rare complications after biliary surgery. The authors present three cases of retained T-tubes in patients with transplanted liver that could not be removed by a standard manual traction. The authors describe a new simple percutaneous method that allows removal of these T-tubes without complication.