Activation of Akt/PKB Prevents Retinal Muller Cells from High Glucose-Induced Apoptosis
Abstract Number: 102-OR
Authors: JINZHONG ZHANG, XIA XI, MI WIDNESS, LING GAO
Institutions: Cleveland, OH
Results: The serine/threonine Akt/PKB protein kinase is central to PI3K/Akt signaling cascade that potently antagonizes apoptosis in a variety of cells. This protective effect requires glucose in response to various stimuli. Conversely, in diabetes, elevated intracellular glucose is likely responsible for the retinal cell death via apoptosis. The mechanism by which the chronic hyperglycemia causes retinal cell death is just beginning to be fleshed out. Recently, accumulating data show that retinal Muller cells are involved in the early stage of diabetic retinopathy. In the current study, we were exploring the possible role of Akt signaling transduction pathway in high glucose-induced apoptosis in retinal Muller cells. We found that culture of retinal Muller cells in 25 mM glucose for 72h resulted in several-fold decrease in Akt activity compared to that in 5.5 mM glucose, as revealed by Western blotting using anti-phospho-Akt antibody and by Akt activity assay. Inactivation of Akt induced by elevated glucose appeared to be mediated by glycolysis, since short-term incubation of cells cultured in 25 mM glucose for 72h with glycolytic inhibitors, iodoacetate and 2-deoxy-glucose, restored Akt activity. To explore the possible role of Akt in retinal cell death and the interaction of Akt survival pathway with glucose metabolism, retinal Muller cells were transfected with plasmid containing cDNA encoding the constitutively active or dominant negative form of Akt1. Overexpression of active Akt prevented retinal Muller cells from high glucose-induced apoptosis as revealed by caspase-3 activity and Annexin V binding assays. Inhibition of glycolysis by iodoacetate also blocked caspase-3 activation, and this effect was completely abolished by dominant negative Akt. These results suggest that promotion of PI3K/Akt survival pathway can prevent retinal cells from high glucose-induced apoptosis, and glycolysis is involved in this process. Mechanistic understanding of the interaction of glucose metabolism and Akt survival pathway should help to elucidate the role of cell death in the development of diabetic retinopathy.