Oxidative Stress Is a Mediator of Glucose Toxicity in Insulin-secreting Pancreatic Islet Cell Lines

Abstract

Pancreatic ␤ cells secrete insulin in response to changes in the extracellular glucose. However, prolonged exposure to elevated glucose exerts toxic effects on ␤ cells and results in ␤ cell dysfunction and ultimately ␤ cell death (glucose toxicity). To investigate the mechanism of how increased extracellular glucose is toxic to ␤ cells, we used two model systems where glucose metabolism was increased in ␤ cell lines by enhancing glucokinase (GK) activity and exposing cells to physiologically relevant increases in extracellular glucose (3.3-20 mM). Exposure of cells with enhanced GK activity to 20 mM glucose accelerated glycolysis, but reduced cellular NAD(P)H and ATP, caused accumulation of intracellular reactive oxygen species (ROS) and oxidative damage to mitochondria and DNA, and promoted apoptotic cell death. These changes required both enhanced GK activity and exposure to elevated extracellular glucose. A ROS scavenger partially prevented the toxic effects of increased glucose metabolism. These results indicate that increased glucose metabolism in ␤ cells generates oxidative stress and impairs cell function and survival; this may be a mechanism of glucose toxicity in ␤ cells. The level of ␤ cell GK may also be critical in this process.Pancreatic ␤ cells play a crucial role in maintaining glucose homeostasis through secretion of insulin in response to changes in the extracellular glucose. Glucose-stimulated insulin secretion (GSIS) 1 is regulated by the rate of glucose metabolism within ␤ cells (1, 2), and a key event in this process is the phosphorylation of glucose by glucokinase (GK). GK, also referred to as hexokinase IV, is a member of the mammalian hexokinase (HK) family that catalyzes the initial step in glucose metabolism by most metabolic pathways (e.g. glycolysis, pentose phosphate cycle and glycogen synthesis) (3). In pancreatic ␤ cells, GK assumes the rate-limiting role for glucose metabolism (4). GK has a K m for glucose of ϳ5 mM, a value that is within the physiological range of blood glucose levels and almost two orders of magnitude higher than any other HK (3-5). Furthermore, the activity of GK is insensitive to feedback inhibition by physiological concentrations of its product, glucose 6-phosphate (Glc-6-P) (3). These characteristics of GK enable ␤ cells to increase glucose metabolism in proportion to elevations in extracellular glucose. Subsequent to glucose phosphorylation, glucose metabolism involves both cytosolic and mitochondrial processes and generates signals leading to insulin secretion (2).In contrast to the ability of glucose to acutely stimulate insulin secretion, chronic exposure of ␤ cells to increased glucose concentrations results in ␤ cell dysfunction and ultimately ␤ cell death, a phenomenon termed ␤ cell glucose toxicity (glucotoxicity) (6 -8). During the progression of type 2 diabetes, glucose toxicity is likely an important factor that contributes to progressive ␤ cell failure and development of overt diabetes. The molecular mechanisms of how chronic exposure to elevated...

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