Upregulation of the expression of tight and adherens junction-associated proteins during maturation of neonatal pancreatic islets in vitro

Abstract

Cell-cell contacts mediated by intercellular junctions are crucial for proper insulin secretion in the endocrine pancreas. The biochemical composition of the intercellular junctions in this organ and the role of junctional proteins in endocrine pancreatic dysfunctions are still unclear. In this study, we investigated the expression and cellular location of junctional and cytoskeletal proteins in cultured neonatal rat pancreatic islets. Neonatal B-cells had an impaired insulin secretion compared to adult cells. Cultured neonatal islets showed a time-dependent increase in the glucose-induced secretory response. The maturation of B-cells in vitro was accompanied by upregulation of the expression of some junctional proteins in islet cells. Neonatal islets cultured for only 24 h showed a low expression and a diffuse cytoplasmic location of the tight junctional proteins occludin and ZO-1 and of the adherens junctional proteins alpha- and beta-catenins, as demonstrated by immunoblotting and immunocytochemistry. Culturing islets for up to 8 days significantly increased the cell expression of these junctional proteins but not of the cytoskeletal proteins vinculin and alpha-actinin. A translocation of ZO-1 and catenins to the cell-cell contact region, as well as a higher association of F-actin with the intercellular junction, were also observed in neonatal islets following prolonged culturing. ZO-1 and beta-catenin were immunolocated in the endocrine pancreas of adult rats indicating that these junctional proteins are also expressed in this organ in situ. In conclusion, endocrine pancreatic cells express several junctional proteins that are upregulated following differentiation of the endocrine pancreas in vitro.

Claudin expression in pancreatic endocrine tumors as compared with ductal adenocarcinomas

“…Upregulation of occluding and ZO-1 has been shown during the maturation of neonatal pancreatic islet cells in vitro. Gap junctions have been involved in regulation of insulin secretion [4]. However, besides being involved in normal islet intercellular communication, it is likely that TJs also provide adaptive mechanisms intended to seal and protect islet microdomains against sudden perturbations in extracellular environment.…”

Co-expression and regulation of connexins 36 and 43 in cultured neonatal rat pancreatic islets

“…Before being used, each new batch of Cx36 and Cx43 antibodies was first tested for its specificity in rat brain and heart homogenates, respectively. To ensure equal amounts of protein were loaded in all lanes, the membranes were reprobed with an antibody against the cytoskeletal protein vinculin (Sigma), used herein as an internal standard, because it has been shown that its expression does not change over culturing in neonatal islet cells (Collares-Buzato et al 2004). …”

E-Cadherin and Cell Adhesion: a Role in Architecture and Function in the Pancreatic Islet

“…Over-expression of dominant negative hepatocyte nuclear factor-1 in mouse β-cells (the gene responsible for type 3 MODY) results in abnormal islet architecture, down-regulation of E-cadherin and reduced insulin secretion [14,15]. Similarly, the MIN6 cell line sub-clone (C3) has been identified with reduced glucose evoked insulin secretion and down-regulated E-cadherin expression [16], whilst in rat pancreatic islets, increased expression of the adherens proteins α-and β-catenin is correlated with increased glucose evoked insulin secretion [17]. It is clear that E-cadherin not only plays an important role in islet development but also in the continued function of the islet in terms of glucose responsiveness and insulin secretion.…”