PKC-delta-dependent activation of oxidative stress in adipocytes of obese and insulin-resistant mice: role for NADPH oxidase.
Oxidative stress is thought to be one of the causative factors contributing to insulin resistance and type 2 diabetes. Previously, we showed that reactive oxygen species (ROS) production is significantly increased in adipocytes from high-fat diet-induced obese and insulin-resistant mice (HF). ROS production was also associated with the increased activity of PKC-delta. In the present studies, we hypothesized that PKC-delta contributes to ROS generation and determined their intracellular source. NADPH oxidase inhibitor diphenyleneiodonium chloride (DPI) reduced ROS levels by 50% in HF adipocytes, and inhibitors of NO synthase (L-NAME, 1 mM), xanthine oxidase (allopurinol, 100 microM), AGE formation (aminoguanidine, 10 microM), or the mitochondrial uncoupler (FCCP, 10 microM) had no effect. Rottlerin, a selective PKC-delta inhibitor, suppressed ROS levels by approximately 50%. However, neither GO-6976 nor LY-333531, effective inhibitors toward conventional PKC or PKC-beta, respectively, significantly altered ROS levels in HF adipocytes. Subsequently, adenoviral-mediated expression of wild-type PKC-delta or its dominant negative mutant (DN-PKC-delta) in HF adipocytes resulted in either a twofold increase in ROS levels or their suppression by 20%, respectively. In addition, both ROS levels and PKC-delta activity were sharply reduced by glucose depletion. Taken together, these results suggest that PKC-delta is responsible for elevated intracellular ROS production in HF adipocytes, and this is mediated by high glucose and NADPH oxidase.
Talior
Ilana
I
Dept. of Human Genetics and Molecular Medicine, Sackler School of Medicine, Tel-Aviv University, Tel-Aviv 69978, Israel.
Tennenbaum
Tamar
T
Kuroki
Toshio
T
Eldar-Finkelman
Hagit
H
D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D004041:Dietary Fats / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D007333:Insulin Resistance; D051379:Mice; D008810:Mice, Inbred C57BL; D019255:NADPH Oxidases / Q000378:metabolism*; D009765:Obesity / Q000378:metabolism*; D009861:Onium Compounds / Q000494:pharmacology; D018384:Oxidative Stress; D011493:Protein Kinase C / Q000378:metabolism*; D051745:Protein Kinase C-delta; D017382:Reactive Oxygen Species / Q000378:metabolism*
D004041:Dietary Fats; D009861:Onium Compounds; D017382:Reactive Oxygen Species; C007517:diphenyleneiodonium; D019255:NADPH Oxidases; C497259:Prkcd protein, mouse; D011493:Protein Kinase C; D051745:Protein Kinase C-delta