Intercellular contacts affect secretion and biosynthesis of pancreatic islet cells.

Cell protein biosynthesis is regulated by different factors, but implication of intercellular contacts on alpha and beta cell protein biosyntheses activity has not been yet investigated. Islet cell biosynthetic activity is essential in regulating not only the hormonal reserve within cells but also in renewing all the proteins involved in the control of secretion. Here we aimed to assess whether intercellular interactions affected similarly secretion and protein biosynthesis of rat alpha and beta cells. Insulin and glucagon secretion were analyzed by ELISA or reverse hemolytic plaque assay, and protein biosynthesis evaluated at single cell level using bioorthogonal noncanonical amino acid tagging. Regarding beta cells, we showed a positive correlation between insulin secretion and protein biosynthesis. We also observed that homologous contacts increased both activities at low or moderate glucose concentrations. By contrast, at high glucose concentration, homologous contacts increased insulin secretion and not protein biosynthesis. In addition, heterogeneous contacts between beta and alpha cells had no impact on insulin secretion and protein biosynthesis. Regarding alpha cells, we showed that when they were in contact with beta cells, they increased their glucagon secretion in response to a drop of glucose concentration, but, on the other hand, they decreased their protein biosynthesis under any glucose concentrations. Altogether, these results emphasize the role of intercellular contacts on the function of islet cells, showing that intercellular contacts increased protein biosynthesis in beta cells, except at high glucose, and decreased protein biosynthesis in alpha cells even when glucagon secretion is stimulated.

Cadherin engagement improves insulin secretion of single human ß-cells.

The aim of this study was to assess whether cadherin-mediated adhesion of human islet cells was affected by insulin secretagogues and explore the role of cadherins in the secretory activity of ß-cells. Experiments were carried out with single islet cells adherent to chimeric proteins made of functional E-, N-, or P-cadherin ectodomains fused to the Fc fragment of immunoglobulin (E-cad/Fc, N-cad/Fc, and P-cad/Fc) and immobilized on an inert substrate. We observed that cadherin expression in islet cells was not affected by insulin secretagogues. Adhesion tests showed that islet cells attached to N-cad/Fc and E-cad/Fc acquired, in a time- and secretagogue-dependent manner, a spreading form that was inhibited by blocking cadherin antibodies. By reverse hemolytic plaque assay, we showed that glucose-stimulated insulin secretion of single ß-cells was increased by N-cad/Fc and E-cad/Fc adhesion compared with control. In the presence of E-cad/Fc and after glucose stimulation, we showed that total insulin secretion was six times higher in spreading ß-cells compared with round ß-cells. Furthermore, cadherin-mediated adhesion induced an asymmetric distribution of cortical actin in ß-cells. Our results demonstrate that adhesion of ß-cells to E- and N-cadherins is regulated by insulin secretagogues and that E- and N-cadherin engagement promotes stimulated insulin secretion.

Expression and secretion of alpha1-proteinase inhibitor are regulated by proinflammatory cytokines in human pancreatic islet cells.

AIMS/HYPOTHESIS: Alpha1-proteinase inhibitor (alpha1-PI) has been considered a key player in inflammatory processes. In humans, the main production site of alpha1-PI is the liver, but other tissues, including pancreatic islets, also synthesise this molecule. The aims of this study were to assess the islet cell types that produce alpha1-PI, to determine whether alpha1-PI is actually secreted by islet cells, and to assess how its production and/or secretion are regulated. METHODS: Expression of alpha1-PI in human islet cells was assessed by immunofluorescence, electron microscopy and western blotting. Release of alpha1-PI was analysed by reverse haemolytic plaque assay and ELISA. The effects of cytokines on alpha1-PI synthesis and secretion were tested. RESULTS: Immunofluorescence showed that alpha and delta cells do express alpha1-PI, whereas beta cells do not. By electron microscopy, we demonstrated a colocalisation of alpha1-PI with glucagon and somatostatin within secretory granules. Immunolabelling also revealed localisation of alpha1-PI within the Golgi apparatus, related vesicles and lysosomal structures. The expression of alpha1-PI in islet cells was also demonstrated by western blotting and ELISA of protein extracts. ELISA and reverse haemolytic plaque assay showed that alpha1-PI is secreted into the culture medium. Treatment of islet cells with IL-1beta and oncostatin M for 4 days increased the production and release of alpha1-PI. CONCLUSIONS/INTERPRETATION: Our results demonstrate that alpha1-PI is expressed by the alpha and delta cells of human islets, and that proinflammatory cytokines enhance the production and release of this inhibitor.