Role of insulin signaling in pancreatic beta cells and measurement of insulin release from islets of Langerhans using a microfluidic chip

Insulin resistance in peripheral tissues and defective insulin secretion from pancreatic β-cells leads to non-insulin dependent diabetes mellitus, more commonly known as type 2 diabetes. It has been proposed that a defect in the insulin signal transduction pathway could lead to both hallmarks of type 2 diabetes. To determine how a defect in the insulin signal transduction pathway would affect normal β-cell function, intracellular Ca 2+ changes and secretion were measured from genetically-modified pancreatic β-cells upon stimulation with various secretagogues.; Application of glucose, glyceraldehyde and arginine initiates secretion from insulin receptor substrate-1 (IRS-1) wildtype (WT) and knockout (KO) pancreatic β-cells as measured by amperometry with microelectrodes; however, the amount of secretory events from IRS-1 KO β-cells is significantly less. Additionally, IRS-1 deletion disrupts intracellular Ca2+ handling as KO β-cells have an increase in latency to Ca2+ changes and a decrease in duration of stimulated Ca2+ levels compared to WT control cells. Islets of Langerhans, microorgans composed of 2000–5000 cells of which the majority are β-cells, display irregular Ca2+ oscillations in the presence of stimulatory glucose levels. An insulin mimetic, which stimulates glucose uptake in peripheral tissues by activating the insulin signaling pathway, also initiates secretion from β-cells through an IRS-1 dependent process. Thus, the insulin signaling pathway appears to contribute to Ca2+ homeostasis in pancreatic β-cells and disruption of this pathway can lead to defective insulin secretion. These results provide evidence for the insulin signal transduction pathway as a link between insulin resistance and defective insulin secretion.; To monitor insulin secretion from islets of Langerhans, a microfluidic chip is used to house an islet and perform a capillary electrophoresis competitive immunosassay. Insulin is electrokinetically sampled through a 4 cm heated reaction channel where it mixes with fluorescein isothiocyanate-labeled insulin (FITC-insulin) and anti-insulin immunoglobulin (Ab). The resulting mixture is injected every 15 s onto a 1.5 cm separation channel and separation of FITC-insulin:Ab complex and free FITC-insulin occurs in 5 s. The method allows the kinetics of first and second phase insulin release to be measured from single islets with high temporal resolution.