Altered NAD(P)H production in neonatal rat islets resistant to H(2)O(2)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Aims: We determined the involvement of NAD(P)H generation ability on the resistance of pancreatic islets B-cells to oxidative stress caused by culture exposition to H(2)O(2). Main methods: We cultured isolated neonatal Wistar rat islets for four days in medium containing 5.6 or 20 mM glucose, with or without H(2)O(2) (200 mu M), and analyzed several parameters associated with islet survival in different media. High glucose was used since it protects neonatal islets against the loss of GSIS. Key findings: While none of the culture conditions increased the rate of NAD(P)H content at 16.7 mM glucose, the islets resistant to H(2)O(2) and those exposed to 20 mM glucose showed a greater use of the pentose phosphate pathway and increased ATP synthesis from glucose. Significance: Oxidative stress contributes to the loss of glucose-induced insulin secretion (GSIS) during the onset of diabetes mellitus. Although immature rat islets have reduced GSIS compared to mature islets, they adapt better to oxidative stress and are a good model for understanding the causes involved in the destruction or survival of islet cells. These data support the idea that GSIS and resistance against oxidative stress in immature islets rely on NADH shuttle activities, with little contribution of reduced equivalents from the tricarboxylic acid cycle (TCAC). (c) 2008 Elsevier Inc. All rights reserved.8321-22709716Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Hydrogen peroxide production regulates the mitochondrial function in insulin resistant muscle cells: Effect of catalase overexpression

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The mitochondrial redox state plays a central role in the link between mitochondrial overloading and insulin resistance. However, the mechanism by which the ROS induce insulin resistance in skeletal muscle cells is not completely understood. We examined the association between mitochondrial function and H2O2 production in insulin resistant cells. Our hypothesis is that the low mitochondrial oxygen consumption leads to elevated ROS production by a mechanism associated with reduced PGC1 alpha transcription and low content of phosphorylated CREB. The cells were transfected with either the encoded sequence for catalase overexpression or the specific siRNA for catalase inhibition. After transfection, myotubes were incubated with palmitic acid (500 mu M) and the insulin response, as well as mitochondrial function and fatty acid metabolism, was determined. The low mitochondrial oxygen consumption led to elevated ROS production by a mechanism associated with beta-oxidation of fatty acids. Rotenone was observed to reduce the ratio of ROS production. The elevated H2O2 production markedly decreased the PGC1 alpha transcription, an effect that was accompanied by a reduced phosphorylation of Akt and CREB. The catalase transfection prevented the reduction in the phosphorylated level of Ala and upregulated the levels of phosphaylated CREB. The mitochondrial function was elevated and H2O2 production reduced, thus increasing the insulin sensitivity. The catalase overexpression improved mitochondrial respiration protecting the cells from fatty acid-induced, insulin resistance. This effect indicates that control of hydrogen peroxide production regulates the mitochondrial respiration preventing the insulin resistance in skeletal muscle cells by a mechanism associated with CREB phosphorylation and beta-oxidation of fatty acids. (C) 2013 Elsevier B.V. All rights reserved.18321015911604Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq