The Sulfonylurea Glimepiride Regulates Intracellular Routing of the Insulin-Receptor Complexes through Their Interaction with Specific Protein Kinase C Isoforms

Abstract

Sulfonylureas may stimulate glucose metabolism by protein kinase C (PKC) activation. Because interaction of insulin receptors with PKC plays an important role in controlling the intracellular sorting of the insulin-receptor complex, we investigated the possibility that the sulfonylurea glimepiride may influence intracellular routing of insulin and its receptor through a mechanism involving PKC, and that changes in these processes may be associated with improved insulin action. Using human hepatoma Hep-G2 cells, we found that glimepiride did not affect insulin binding, insulin receptor isoform expression, and insulininduced receptor internalization. By contrast, glimepiride significantly increased intracellular dissociation of the insulin-receptor complex, degradation of insulin, recycling of internalized insulin receptors, release of internalized radioactivity, and prevented insulin-induced receptor down-regulation. Association of PKC-␤II and -⑀ with insulin receptors was increased in glimepiride-treated cells. Selective depletion of cellular PKC-␤II and -⑀ by exposure to 12-O-tetradecanoylphorbol-13-acetate (TPA) or treatment of cells with PKC-␤II inhibitor G06976 reversed the effect of glimepiride on intracellular insulin-receptor processing. Glimepiride increased the effects of insulin on glucose incorporation into glycogen by enhancing both sensitivity and maximal efficacy of insulin. Exposing cells to TPA or G06976 inhibitor reversed these effects. Results indicate that glimepiride increases intracellular sorting of the insulin-receptor complex toward the degradative route, which is associated with both an increased association of the insulin receptor with PKCs and improved insulin action. These data suggest a novel mechanism of action of sulfonylurea, which may have a therapeutic impact on the treatment of type 2 diabetes.

The Mode of Action of the Antidiabetic Drug Glimepiride-Beyond Insulin Secretion

“…In accordance with findings with Hep-G2 cells, this could be attributed to the accelerated recycling of the insulin receptor during its endocytic route and the prevention of insulin-induced receptor down-regulation involving enhanced association of the insulin receptor with the protein kinase C [32]. However, in Hep-G2 cells as well as in skeletal muscles of insulinresistant KK-Ay mice glimepiride did not affect the binding characteristics of the insulin receptor [32,34].…”

“…The putative insulinsensitizing activity was investigated using glycogen synthesis as parameter. In human hepatoma Hep-G2 cells glimepiride (20 µM) increased glycogen synthesis by 30 to 40% vs. control in the presence of increasing concentrations of insulin, but not in its absence [32]. Similarly, in cultured human myotubes glimepiride (0.1 µM), but not glibenclamide, stimulated the insulin-induced glycogen synthesis by 50 to 70% during 4 h incubation [33].…”

“…In contrast, the third pathway, the Src homology collagen (Shc)/mitogen-activated protein kinase (MAPK)-pathway, is primarily engaged in insulin regulation of the general gene expression and, in cooperation with the IRS/PI3K-pathway, determines the differentiation of fibroblasts into adipocytes. Importantly, glimepiride does not activate the insulin receptor tyrosine kinase in cultured Hep-G2 cells [32] and in skeletal muscles of insulin-resistant KK-Ay mice [34]. Nevertheless, it potently activated the IRS/PI3K-pathway as reflected in tyrosine phosphorylation of the IRS-1, accompanied by increased PI3K and downstream protein kinase B activities in isolated rat adipocytes in concentrationdependent fashion (1-10 µM) to up to 35% of the maximal insulin effect [56].…”

Effect of Inhibiting Vacuolar Acidification on Insulin Signaling in Hepatocytes

“…Thus, transformed lymphoblasts from subjects with type 2 diabetes mellitus showed impaired intracellular dissociation of insulin-IRK complexes, insulin degradation, and recycling of the IRK when compared with cells from normal subjects, raising the possibility that these defects might underly the insulin resistance seen in these patients (46). Also it has recently been observed that the sulfonylurea glimepiride improved insulin action in hepatoma cells and correspondingly increased the intracellular dissociation of insulin-IRK complexes, the degradation of insulin, and the recycling of internalized IRK in these cells (47). It is tempting to consider that a defect in the vacuolar system responsible for mediating insulin action (endosomes) and insulin processing and secretion (trans Golgi vesicles) may explain both the insulin resistance and impaired ␤ cell function of type 2 diabetes mellitus.…”

Protein Kinase C Function in Muscle, Liver, and β-Cells and Its Therapeutic Implications for Type 2 Diabetes

“…While direct phosphorylation by PKC does not appear to modulate tyrosine kinase activity (41), an interaction of PKCε with the receptor in liver may affect its ability to phosphorylate its substrates (42). Alternatively, such an association may affect insulin receptor trafficking in hepatocytes (43) and hence both the clearance of insulin by the liver (44) and degradation of the receptor itself (14). …”