title: Glucosamine-induced increase in Akt phosphorylation corresponds to increased endoplasmic reticulum stress in astroglial cells.
abstract: Increased glucose flux through the hexosamine biosynthetic pathway (HBP) is known to affect the activity of a number of signal transduction pathways and lead to insulin resistance. Although widely studied in insulin responsive tissues, the effect of increased HBP activity on largely insulin unresponsive tissues, such as the brain, remains relatively unknown. Herein, we investigate the effects of increased HBP flux on Akt activation in a human astroglial cells line using glucosamine, a compound commonly used to mimic hyperglycemic conditions by increasing HBP flux. Cellular treatment with 8 mM glucosamine resulted in a 96.8% +/- 24.6 increase in Akt phosphorylation after 5 h of treatment that remained elevated throughout the 9-h time course. Glucosamine treatment also resulted in modest increases in global levels of the O-GlcNAc protein modification. Increasing O-GlcNAc levels using the O-GlcNAcase inhibitor streptozotocin (STZ) also increased Akt phosphorylation by 96.8% +/- 11.0 after only 3 h although for a shorter duration than glucosamine; however, the more potent O-GlcNAcase inhibitors O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbamate (PUGNAc) and 1,2-dideoxy-2'-propyl-alpha-D-glucopyranoso-[2,1-d]-Delta2'-thiazoline (NAGBT) failed to mimic the increases in phospho-Akt indicating that the Akt phosphorylation is not a result of increased O-GlcNAc protein modification. Further analysis indicated that this increased phosphorylation was also not due to increased osmotic stress and was not attenuated by N-acetylcysteine eliminating the potential role of oxidative stress in the observed phospho-Akt increases. Glucosamine treatment, but not STZ treatment, did correlate with a large increase in the expression of the endoplasmic reticulum (ER) stress marker GRP 78. Altogether, these results indicate that increased HBP flux in human astroglial cells results in a rapid, short-term phosphorylation of Akt that is likely a result of increased ER stress. The mechanism by which STZ increases Akt phosphorylation, however, remains unknown.
authors.affiliation: Department of Chemistry, University of South Florida, 4202 East Fowler Ave, SCA 400, Tampa, FL 33620, USA.
authors.forename: J Aaron
authors.initials: JA
authors.lastname: Matthews
authors.forename: Jonathan L
authors.initials: JL
authors.lastname: Belof
authors.forename: Mildred
authors.initials: M
authors.lastname: Acevedo-Duncan
authors.forename: Robert L
authors.initials: RL
authors.lastname: Potter
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000117:Acetylglucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D001253:Astrocytes / Q000187:drug effects* / Q000201:enzymology*; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000473:pathology*; D000091342:Endoplasmic Reticulum Chaperone BiP; D004790:Enzyme Induction / Q000187:drug effects; D005688:Galactosamine / Q000494:pharmacology; D005944:Glucosamine / Q000494:pharmacology*; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D018832:Molecular Chaperones / Q000378:metabolism; D009997:Osmotic Pressure / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects; D010091:Oximes / Q000494:pharmacology; D048448:Phenylcarbamates / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D013311:Streptozocin / Q000494:pharmacology
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D010091:Oximes; D048448:Phenylcarbamates; C068836:N-acetylglucosaminono-1,5-lactone O-(phenylcarbamoyl)oxime; D013311:Streptozocin; D005688:Galactosamine; D051057:Proto-Oncogene Proteins c-akt; D005944:Glucosamine; D000117:Acetylglucosamine; D000111:Acetylcysteine