Anne Hoorens

Glucose promotes survival of rat pancreatic beta cells by activating synthesis of proteins which suppress a constitutive apoptotic program.

This study demonstrates that rat islet beta cells constitutively express an apoptotic program which is activated when mRNA or protein synthesis is blocked. Apoptotic beta cells were detectable by electron microscopy after treatment with actinomycin D or cycloheximide. With a fluorescence microscopic assay both agents were found to increase the number of apoptotic beta cells dose- and time-dependently, up to 70% after 1 wk of culture; virtually no apoptotic beta cells occurred in control preparations or in conditions leading to primary necrosis. Thus, survival of beta cells seems dependent on synthesis of proteins which suppress an endogenous suicide program. This mechanism explains earlier observed effects of glucose on survival of cultured beta cells. Glucose is known to dose-dependently increase the percentage of beta cells in active biosynthesis and the percentage that survives during culture. It is now demonstrated that the glucose-induced survival of beta cells cultured for 1 wk results from a dose-dependent reduction in the percentage of beta cells dying in apoptosis (49% at 3 mM glucose, 40% at 6 mM, 9% at 10 mM). Thus, intercellular differences in glucose sensitivity appear responsible for the heterogeneity in beta cell sensitivity to apoptotic conditions. These data indicate that glucose promotes survival of beta cells by activating synthesis of proteins which suppress apoptosis. The present model allows for further investigation of the regulation of apoptosis in beta cells and the identification of agents which induce or prevent beta cell death.

Cytokines induce deoxyribonucleic acid strand breaks and apoptosis in human pancreatic islet cells.

We have previously observed that a 6-day exposure of human pancreatic islets to a combination of cytokines (interleukin-1β 50 U/ml + tumour necrosis factor-α 1000 U/ml + interferon-γ 1000 U/ml) severely impairs β-cell functions. In the present study, we examined whether this condition affects DNA integrity and viability of human islet cells. Cells were studied after 3, 6, and 9 days of cytokine treatment by both single cell gel electrophoresis (the “comet assay,” a sensitive method for detection of DNA strand breaks) and by a cytotoxicity assay using the DNA binding dyes Hoechst 33342 and propidium iodide as indices for the number of viable, necrotic, and apoptotic cells. Cytokine treatment for 6 and 9 days resulted in a 50% increase in comet length (P < 0.01 vs. controls), indicating DNA strand breaks, as well as in a significant increase in the number of apoptotic cells (P < 0.02 vs. controls), but not in the number of necrotic cells. The arginine analogs NG-nitro-l-arginine and NG-monomethyl-l-arginine...

Acute Onset of Type I Diabetes Mellitus after Severe Echovirus 9 Infection: Putative Pathogenic Pathways

Enterovirus infections have been implicated in the development of type I diabetes mellitus. They may cause beta cell destruction either by cytolytic infection in the pancreas or indirectly by contributing to autoimmune reactivity. We sought evidence for these 2 mechanisms in a case of acute-onset diabetes mellitus that occurred during severe echovirus 9 infection. The virus was isolated and administered to cultured human beta cells. No viral proliferation was observed, and no beta cell death was induced, while parallel exposure to Coxsackie B virus serotype 3 resulted in viral proliferation and massive beta cell death. Although the viral protein 2C exhibited a sequence similar to that of the beta cell autoantigen glutamic acid decarboxylase (GAD(65)), no cross-reactive T cell responses were detected. The patient did not develop antibodies to GAD(65) either. Absence of evidence for direct cytolytic action or an indirect effect through molecular mimicry with GAD(65) in the present case raises the possibility of another indirect pathway through which enteroviruses can cause diabetes mellitus.

Undifferentiated carcinoma of the pancreas : analysis of intermediate filament profile and Ki-ras mutations provides evidence of a ductal origin

Undifferentiated carcinomas and osteoclast‐like giant cell tumours of the pancreas commonly contain foci of neoplastic ductal glands. To test the hypothesis that undifferentiated carcinomas and osteoclast‐like giant cell tumours have a ductal origin, the immunocytochemical cytokeratin pattern and the frequency and type of Ki‐ras mutations at colon 12 were studied in a series of 17 undifferentiated carcinomas and two osteoclast‐like giant cell tumours. The cytokeratin features of undifferentiated carcinomas and osteoclast‐like giant cell tumours were compared with those found in 10 ductal adenocarcinomas, 20 acinar cell carcinomas, 25 neuroendocrine tumours, and 15 solid‐pseudopapillary tumours. All undifferentiated carcinomas and osteoclast‐like giant cell tumours stained with at least one cytokeratin antibody, and 13/19 of them with antibodies against cytokeratins 7, 8, 18, and 19. The latter cytokeratins were expressed in all ductal adenocarcinomas, but only in 15/20 acinar cell carcinomas, 2/25 neuroendocrine tumours, and 1/15 solid‐pseudopapillary tumours. In addition to cytokeratin, 15/19 undifferentiated carcinomas/osteoclast‐like giant cell tumours were positive for vimentin. Ki‐ras mutations at codon 12 were found in 10 undifferentiated carcinomas and one osteoclast‐like giant cell tumour from which DNA could be successfully amplified. The Ki‐ras mutation patterns were analysed in six tumours and corresponded to those typical of ductal adenocarcinomas. In tumours with ductal and anaplastic components, both components revealed identical mutation patterns. From these findings, it is concluded that both undifferentiated carcinomas and osteoclast‐like giant cell tumours belong to the pancreatic tumours that show a ductal phenotype. Since undifferentiated carcinomas and osteoclast‐like giant cell tumours share the same cytokeratin and Ki‐ras features, they are probably derived from the same cell lineage.

Nicotinamide protects human beta cells against chemically-induced necrosis, but not against cytokine-induced apoptosis

Summary Nicotinamide intervention trials are presently undertaken to prevent Type I (insulin-dependent) diabetes in high risk subjects. They are based on studies in rodents reporting nicotinamide protection against beta-cell injury in vitro and in vivo. This study examines whether nicotinamide can protect human beta cells in vitro. At concentrations (2 and 5 mmol/l) to protect rat beta cells against necrosis by streptozotocin or hydrogen peroxide, nicotinamide prevents hydrogen peroxide-induced necrosis of human beta cells. As with rat beta cells, nicotinamide fails to protect human beta cells against apoptosis induced by a combination of the cytokines interleukin-1β, interferon-γ and tumour necrosis factor-α. In rat beta cells, nicotinamide (2 to 20 mmol/l) was also found to induce apoptosis, in particular during the days following its protection against necrosis; this cytotoxic effect was not observed with human beta cells. These data demonstrate that nicotinamide can protect human beta cells against radical-induced necrosis, but not against cytokine-induced apoptosis. This effect is not associated with a delayed apoptosis as in rat beta cells. [Diabetologia (1999) 42: 55–59]

The Hippo pathway effector YAP controls mouse hepatic stellate cell activation

BACKGROUND & AIMSnHepatic stellate cell activation is a wound-healing response to liver injury. However, continued activation of stellate cells during chronic liver damage causes excessive matrix deposition and the formation of pathological scar tissue leading to fibrosis and ultimately cirrhosis. The importance of sustained stellate cell activation for this pathological process is well recognized, and several signalling pathways that can promote stellate cell activation have been identified, such as the TGFβ-, PDGF-, and LPS-dependent pathways. However, the mechanisms that trigger and drive the early steps in activation are not well understood.nnnMETHODS AND RESULTSnWe identified the Hippo pathway and its effector YAP as a key pathway that controls stellate cell activation. YAP is a transcriptional co-activator and we found that it drives the earliest changes in gene expression during stellate cell activation. Activation of stellate cells in vivo by CCl4 administration to mice or activation in vitro caused rapid activation of YAP as revealed by its nuclear translocation and by the induction of YAP target genes. YAP was also activated in stellate cells of human fibrotic livers as evidenced by its nuclear localization. Importantly, knockdown of YAP expression or pharmacological inhibition of YAP prevented hepatic stellate cell activation in vitro and pharmacological inhibition of YAP impeded fibrogenesis in mice.nnnCONCLUSIONSnYAP activation is a critical driver of hepatic stellate cell activation and inhibition of YAP presents a novel approach for the treatment of liver fibrosis.

Prognostic value of the lymph node ratio in node positive colon cancer

Surgery is the primary treatment of non-metastatic colon cancer. En bloc removal of the colon with its associated mesenteric lymph nodes is essential. However, the number of lymph nodes reported with colectomy varies widely and may be a result of variation in the actual number of regional lymph nodes, surgical technique, or the thoroughness of the pathologist in finding lymph nodes. The number of lymph node metastases is an important negative prognostic factor and is used in stratification schemes for clinical trials.1nnRecent studies have emphasised the fact that examining a greater number of nodes increases the likelihood of correct staging and is associated with better survival, after controlling …

Laparoscopic resection of gastric gastrointestinal stromal tumors (GIST) is safe and effective, irrespective of tumor size

BackgroundFeasibility and long-term safety of laparoscopic removal of gastric gastrointestinal stromal tumors (GISTs) of the stomach is well established for lesions smaller than 2xa0cm. Our specific aim was to explore whether laparoscopic treatment is equally applicable for gastric GISTs larger than 2xa0cm.MethodsBetween 1997 and 2010, 31 consecutive patients presenting with a primary gastric GIST were scheduled for laparoscopic resection, irrespective of tumor size. Prerequisites for laparoscopic approach were the absence of metastases and the presence of a well-defined tumor on CT scanning without involvement of adjacent organs, the esophagogastric junction, or the pylorus of the stomach. Data were retrieved retrospectively from a prospectively collected database, including information on patient demographics, surgical procedure, complications, hospital stay, and recurrence. Diagnosis of GIST was based on microscopic analysis, including immunohistochemistry with a panel of antibodies: CD117, CD34, DOG1, S100, desmin, and smooth muscle actin.ResultsAll 31 laparoscopic resections were carried out successfully. The most common symptoms were melena, anemia, and abdominal pain. In one case we performed a laparoscopic approach for a GIST with acute bleeding. Tumor size was smaller than 2xa0cm in 5 patients and larger than 2xa0cm in 26 patients. The median tumor size was 4.4xa0cm (rangexa0=xa00.4–11.0xa0cm). Median blood loss was identical in both groups (20xa0ml), but duration of operation (60 vs. 103xa0min) and duration of hospital stay (6 vs. 8xa0days) were lower when tumor size was less than 2xa0cm. Only one patient (with tumor size <2xa0cm) experienced a postoperative hemorrhage. After a median follow-up of 52xa0months, there were no recurrences or metastases.ConclusionThe low morbidity rates and the long-term disease-free interval of 100% observed in our cohort indicate that laparoscopic resection is safe and effective in treating gastric GISTs, even for tumors larger than 2xa0cm.

Clinical and diagnostic characteristics of complex III deficiency due to mutations in the BCS1L gene.

We investigated two siblings of a Spanish family presenting with congenital lactic acidosis. They had severe failure to thrive, liver dysfunction, and renal tubulopathy. An isolated biochemical complex III deficiency was detected in liver. A search for mutations in the human bc1 synthesis like (BCS1L) gene was undertaken. Direct sequencing revealed a missense mutation R45C and a nonsense mutation R56X, both located in exon 1 of BCS1L. The missense mutation in combination with a loss of function of the second allele is responsible for the isolated complex III deficiency in this family.

Laparoscopic versus open resection of gastrointestinal stromal tumors of the stomach

BackgroundGastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract. Surgical treatment is the only chance of cure for patients with a primary localized GIST. A laparoscopic approach has been considered reasonable for these tumors of gastric origin. The current study compares the outcome of laparoscopic versus open resection of gastric GISTs and compares our series with the few published studies comparing the open versus the laparoscopic approach.MethodsFrom a prospectively collected database, we found 53 primary gastric GIST resections that were performed in our department. Laparoscopic (LAP) resections were performed in 37 patients and traditional (OPEN) resections in 16 patients. Clinical and pathologic characteristics and surgical outcomes were analyzed according to surgical procedure.ResultsPatients who underwent LAP or OPEN resection of gastric GISTs did not differ with respect to age at operation, gender, clinical presentation, and tumor size. Operative time was significantly lower for LAP than for OPEN resection, with a mean duration of 45 and 132.5xa0min, respectively (pxa0<xa00.001). LAP resection yielded a significantly shorter length of stay (median 7 vs. 14xa0days; pxa0=xa00.007) and lower 30-day morbidity rate (2.7xa0% vs. 18.9xa0%; pxa0=xa00.077). The operative mortality was 12.5xa0% after OPEN resection and there was no operative mortality after LAP (pxa0=xa00.087). The recurrence rate was significantly lower after LAP surgery (0xa0% vs. 37.5xa0%; pxa0<xa00.001). All patients in the LAP group are alive without recurrence, and 25xa0% (4/16) of the OPEN group are alive with recurrence but in complete remission under imatinib mesylate treatment. Two patients of the open group died due to progression of GIST (pxa0=xa00.087).ConclusionsCompared to open resection, laparoscopic resection of gastric stromal tumors is associated with a shorter operation time, a shorter hospital stay, and a lower recurrence rate.