title: Preconditioning with millimolar concentrations of vitamin C or N-acetylcysteine protects L6 muscle cells insulin-stimulated viability and DNA synthesis under oxidative stress.
abstract: The effect of reactive oxygen/nitrogen species (ROS/RNS)(hydrogen peroxide -- H(2)O(2), superoxide anion radical O(2)*- and hydroxyl radical *OH -- the reaction products of hypoxanthine/xanthine oxidase system), nitric oxide (NO* from sodium nitroprusside -- SNP), and peroxynitrite (ONOO(-) from 3-morpholinosydnonimine -- SIN-1) on insulin mitogenic effect was studied in L6 muscle cells after one day pretreatment with/or without antioxidants. ROS/RNS inhibited insulin-induced mitogenicity (DNA synthesis). Insulin (0.1 microM), however, markedly improved mitogenicity in the muscle cells treated with increased concentrations (0.1, 0.5, 1 mM) of donors of H(2)O(2), O(2)*-, *OH, ONOO(-) and NO*. Cell viability assessed by morphological criteria was also monitored. Massive apoptosis was induced by 1 mM of donors of H(2)O(2) and ONOO(-), while NO* additionally induced necrotic cell death. Taken together, these results have shown that ROS/RNS provide a good explanation for the developing resistance to the growth promoting activity of insulin in myoblasts under conditions of oxidative or nitrosative stress. Cell viability showed that neither donor induced cell death when given below 0.5 mM. In order to confirm the deleterious effects of ROS/RNS prior to the subsequent treatment with ROS/RNS plus insulin one day pretreatment with selected antioxidants (sodium ascorbate - ASC (0.01, 0.1, 1 mM), or N-acetylcysteine - NAC (0.1, 1, 10 mM) was carried out. Surprisingly, at a low dose (micromolar) antioxidants did not abrogate and even worsened the concentration-dependent effects of ROS/RNS. In contrast, pretreatment with millimolar dose of ASC or NAC maintained an elevated mitogenicity in response to insulin irrespective of the ROS/RNS donor type used.
authors.affiliation: Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw Agricultural University, 02-787 Warsaw, Poland. orzechowski@alpha.sggw.waw.pl
authors.forename: Arkadiusz
authors.initials: A
authors.lastname: Orzechowski
authors.forename: Małgorzata
authors.initials: M
authors.lastname: Łokociejewska
authors.forename: Patrycja
authors.initials: P
authors.lastname: Muras
authors.forename: Jean-Francois
authors.initials: JF
authors.lastname: Hocquette
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology*; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001205:Ascorbic Acid / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003470:Culture Media; D004247:DNA / Q000096:biosynthesis*; D053938:DNA Fragmentation / Q000187:drug effects; D016166:Free Radical Scavengers / Q000494:pharmacology*; D005609:Free Radicals / Q000378:metabolism; D006651:Histocytochemistry; D007328:Insulin / Q000494:pharmacology*; D008931:Mitochondria, Muscle / Q000187:drug effects / Q000378:metabolism; D008934:Mitogens / Q000494:pharmacology; D018482:Muscle, Skeletal / Q000187:drug effects*; D009707:Nucleosomes / Q000737:chemistry; D018384:Oxidative Stress / Q000187:drug effects*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000975:Antioxidants; D003470:Culture Media; D016166:Free Radical Scavengers; D005609:Free Radicals; D007328:Insulin; D008934:Mitogens; D009707:Nucleosomes; D017382:Reactive Oxygen Species; D004247:DNA; D001205:Ascorbic Acid; D000111:Acetylcysteine