title: Long-term effects of tumor necrosis factor-alpha treatment on insulin signaling pathway in HepG2 cells and HepG2 cells overexpressing constitutively active Akt/PKB.
abstract: Tumor necrosis factor-alpha (TNF-alpha) mediated attenuation of insulin signaling pathway is an important cause in several disorders like obesity, obesity linked diabetes mellitus. TNF-alpha actions vary depending upon concentration and time of exposure in various cells. In the present study, the effects of long-term TNF-alpha (1 ng/ml) exposure on the components of insulin signaling pathway in HepG2 and HepG2 cells overexpressing constitutively active Akt1/PKB-alpha (HepG2-CA-Akt/PKB) have been investigated. In parental HepG2 cells, TNF-alpha treatment for 24 h reduced the phosphorylation of Akt1/PKB-alpha and GSK-3beta and under these conditions cells also showed reduced insulin responsiveness in terms of Akt1/PKB-alpha and GSK-3beta phosphorylation. TNF-alpha pre-incubated HepG2-CA-Akt/PKB cells showed lower reduction in Akt1/PKB-alpha and GSK-3beta phosphorylation and insulin responsiveness after 24 h as compared to parental HepG2 cells. We report that the long-term TNF-alpha pre-incubation in both parental HepG2 and HepG2-CA-Akt/PKB-alpha cells leads to the reduction in the levels of IRS-1 without altering the levels of IRS-2. In order to understand the reason for the differential insulin resistance in both the cell types, the effect of long-term TNF-alpha treatment on the proteins upstream to Akt/PKB was investigated. TNF-alpha pre-incubation also showed reduced insulin-stimulated Tyr phosphorylation of insulin receptor (IR-beta) in both the cell types, moreover hyperphosphorylation of IRS-1 at Ser 312 residue was observed in TNF-alpha pre-incubated cells. As hyperphosphorylation of IRS-1 at Ser 312 can induce its degradation, it is possible that reduced insulin responsiveness after long-term TNF-alpha pre-incubation observed in this study is due to the decrease in IRS-1 levels.
authors.affiliation: Department of Biochemistry, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
authors.forename: Dhananjay
authors.initials: D
authors.lastname: Gupta
authors.forename: Shailly
authors.initials: S
authors.lastname: Varma
authors.forename: Ramji L
authors.initials: RL
authors.lastname: Khandelwal
mesh_terms: D002460:Cell Line; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D006868:Hydrolysis; D007328:Insulin / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins; D007333:Insulin Resistance; D010750:Phosphoproteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: C526292:IRS1 protein, human; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D010750:Phosphoproteins; D014409:Tumor Necrosis Factor-alpha; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3