Vitamin C Research Today is a free monthly online journal that collates and summarizes the latest research about Vitamin C, including details on benefits, dosage, supplements, information. | ||||||||
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Preincubation with sodium ascorbate potentiates insulin-dependent PKB/Akt and c-Jun phosphorylation in L6 rat myoblasts challenged with reactive oxygen/nitrogen species.Orzechowski A, Lokociejewska M, Pawlikowska P, Kruszewski M Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw Agricultural University, Nowoursynowska 159, 02-776 Warsaw, Poland. orzechowski@alpha.sggw.waw.pl Previously, we reported that mitogenicity in L6 muscle cells was stimulated by insulin but inhibited by reactive oxygen/nitrogen species (ROS/RNS; []) and that preincubation with sodium ascorbate (ASC) protected from either the impaired DNA synthesis and/or loss of cell viability. Now, we addressed the question how ascorbate (AA) rescued DNA synthesis in L6 muscle cells being challenged with ROS/RNS. We assumed that AA might be able to influence insulin signaling. We found that insulin elevated the protein levels of both PKB/Akt kinase phosphorylated at Serine(473) (pS473-Akt), and c-Jun phosphorylated at Serine63, Serine73 (pS63, pS73-c-Jun) residues, respectively. A short-term treatment experiment (0 - 45 min) revealed that either insulin (0.1 muM) or hydrogen peroxide (0.1, 0.5 mM; H2O2) increased the pS473-Akt and pS63, pS73-c-Jun protein levels, although the effect of ROS/RNS peaked earlier (5 min) than that of insulin (45 min). Astonishingly, the elevated levels of both pS473-Akt and pS63, pS73-c-Jun in response to insulin were reduced by the concomitant treatment with H2O2 in a dose-dependent fashion. In contrast, a 4-hour preincubation with ASC (1 mM) augmented the signal from pS473-Akt and pS63, pS73-c-Jun, when both insulin and H2O2 were added. Moreover, a 24 h preincubation with ASC also elevated the pS473-Akt and pS63, pS73-c-Jun levels in response to insulin irrespective to ROS/RNS co-treatment. During chronic treatment studies, ROS/RNS stimulated neither phosphorylation of Akt nor c-Jun, indicating that ROS/RNS-dependent activation of the above-mentioned proteins was short-term and transient. Furthermore, higher levels of pS473 Akt and pS63, pS73-c-Jun after preincubation with ASC suggest that by this route AA could protect insulin-induced mitogenicity. Basal levels of Akt and its target p70(S6K) remained constant regardless of treatment. These results suggest that AA defends the insulin-stimulated mitogenicity hampered by ROS/RNS most likely by the amplification of insulin signal at the level of pS473-Akt and pS63, pS73-c-Jun, respectively. Published 20 May 2005 in Life Sci, 77(5): 496-511.
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