title: Hexosamines and TGF-beta1 use similar signaling pathways to mediate matrix protein synthesis in mesangial cells.
abstract: Hyperglycemia-induced alterations in mesangial (MES) cell function and extracellular matrix (ECM) protein accumulation are seen in diabetic glomerulopathy. Transforming growth factor-beta1 (TGF-beta1) mediates high-glucose-induced matrix production in the kidney. Recent studies demonstrated that some of the effects of high glucose on cellular metabolism are mediated by the hexosamine biosynthesis pathway (HBP) in which fructose-6-phosphate is converted to glucosamine (GlcN) 6-phosphate. We previously showed that the high-glucose-mediated fibronectin and laminin synthesis in MES cells is mediated by the HBP and that GlcN is more potent than glucose in inducing TGF-beta1 promoter luciferase activity. In this study, we investigated the hypothesis that the effects of glucose on MES matrix production occur via hexosamine regulation of TGF-beta1. Culturing simian virus (SV)-40-transformed rat kidney MES cells in 25 mM glucose (HG) for 48 h increases cellular fibronectin and laminin levels about twofold on Western blots compared with low glucose (5 mM). GlcN (1.5 mM) or TGF-beta1 (2.5-5 ng/ml) for 24-48 h also increases ECM synthesis. However, the effects of HG or GlcN with TGF-beta1 are not additive. The presence of anti-TGF-beta1 antibodies (20 microg/ml) blocks both TGF-beta1- and GlcN-induced fibronectin synthesis. TGF-beta1 increased ECM levels via PKA (laminin and fibronectin) and PKC (fibronectin) pathways. Similarly, TGF-beta1 and hexosamines led to nonadditive increases in phosphorylation of the cAMP responsive element binding transcription factor. These results suggest that the effects of excess glucose on MES ECM synthesis occur via HBP-mediated regulation of TGF-beta1.
authors.affiliation: Department of Internal Medicine, Division of Nephrology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA. ecrook@dmc.org
authors.forename: Lalit P
authors.initials: LP
authors.lastname: Singh
authors.forename: Kenneith
authors.initials: K
authors.lastname: Green
authors.forename: Michelle
authors.initials: M
authors.lastname: Alexander
authors.forename: Shira
authors.initials: S
authors.lastname: Bassly
authors.forename: Errol D
authors.initials: ED
authors.lastname: Crook
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002461:Cell Line, Transformed; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D005944:Glucosamine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D019298:Glucose-6-Phosphate / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D006595:Hexosamines / Q000378:metabolism* / Q000494:pharmacology; D007797:Laminin / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D016212:Transforming Growth Factor beta / Q000276:immunology / Q000378:metabolism* / Q000494:pharmacology; D053773:Transforming Growth Factor beta1
chemical_list: D000906:Antibodies; D017362:Cyclic AMP Response Element-Binding Protein; D004791:Enzyme Inhibitors; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D006595:Hexosamines; D007797:Laminin; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C001293:glucosamine 6-phosphate; D019298:Glucose-6-Phosphate; D000255:Adenosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D005947:Glucose; D005944:Glucosamine