Igor H. Sampaio

Melatonin prevents mitochondrial dysfunction and insulin resistance in rat skeletal muscle

Melatonin has a number of beneficial metabolic actions and reduced levels of melatonin may contribute to type 2 diabetes. The present study investigated the metabolic pathways involved in the effects of melatonin on mitochondrial function and insulin resistance in rat skeletal muscle. The effect of melatonin was tested both in vitro in isolated rats skeletal muscle cells and in vivo using pinealectomized rats (PNX). Insulin resistance was induced in vitro by treating primary rat skeletal muscle cells with palmitic acid for 24 hr. Insulin‐stimulated glucose uptake was reduced by palmitic acid followed by decreased phosphorylation of AKT which was prevented my melatonin. Palmitic acid reduced mitochondrial respiration, genes involved in mitochondrial biogenesis and the levels of tricarboxylic acid cycle intermediates whereas melatonin counteracted all these parameters in insulin‐resistant cells. Melatonin treatment increases CAMKII and p‐CREB but had no effect on p‐AMPK. Silencing of CREB protein by siRNA reduced mitochondrial respiration mimicking the effect of palmitic acid and prevented melatonin‐induced increase in p‐AKT in palmitic acid‐treated cells. PNX rats exhibited mild glucose intolerance, decreased energy expenditure and decreased p‐AKT, mitochondrial respiration, and p‐CREB and PGC‐1 alpha levels in skeletal muscle which were restored by melatonin treatment in PNX rats. In summary, we showed that melatonin could prevent mitochondrial dysfunction and insulin resistance via activation of CREB‐PGC‐1 alpha pathway. Thus, the present work shows that melatonin play an important role in skeletal muscle mitochondrial function which could explain some of the beneficial effects of melatonin in insulin resistance states.

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

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 PGC1α 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 (500μM) 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 β-oxidation of fatty acids. Rotenone was observed to reduce the ratio of ROS production. The elevated H2O2 production markedly decreased the PGC1α 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 Akt and upregulated the levels of phosphorylated 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 β-oxidation of fatty acids.

Long-chain acyl-CoA synthetase 6 regulates lipid synthesis and mitochondrial oxidative capacity in human and rat skeletal muscle.

Long‐chain acyl‐CoA synthetase 6 (ACSL6) mRNA is present in human and rat skeletal muscle, and is modulated by nutritional status: exercise and fasting decrease ACSL6 mRNA, whereas acute lipid ingestion increase its expression. ACSL6 genic inhibition in rat primary myotubes decreased lipid accumulation, as well as activated the higher mitochondrial oxidative capacity programme and fatty acid oxidation through the AMPK/PGC1‐α pathway. ACSL6 overexpression in human primary myotubes increased phospholipid species and decreased oxidative metabolism.

Regulação do metabolismo de glicose e ácido graxo no músculo esquelético durante exercício físico

The glucose-fatty acid cycle explains the preference for fatty acid during moderate and long duration physical exercise. In contrast, there is a high glucose availability and oxidation rate in response to intense physical exercise. The reactive oxygen species (ROS) production during physical exercise suggests that the redox balance is important to regulate of lipids/carbohydrate metabolism. ROS reduces the activity of the Krebs cycle, and increases the activity of mitochondrial uncoupling proteins. The opposite effects happen during moderate physical activity. Thus, some issues is highlighted in the present review: Why does skeletal muscle prefer lipids in the basal and during moderate physical activity? Why does glucose-fatty acid fail to carry out their effects during intense physical exercise? How skeletal muscles regulate the lipids and carbohydrate metabolism during the contraction-relaxation cycle?

Oncophosphosignaling Favors a Glycolytic Phenotype in Human Drug Resistant Leukemia.

In chemoresistant leukemia cells (Lucena‐1), the low molecular weight protein tyrosine phosphatase (LMWPTP) is about 20‐fold more active than in their susceptible counterpart (K562). We found this phosphatase ensures the activated statuses of Src and Bcr‐Abl. Since, phosphorylation and dephosphorylation of proteins represent a key post‐translational regulation of several enzymes, we also explored the kinome. We hereby show that LMWPTP superactivation, together with kinome reprogramming, cooperate towards glucose addiction. Resistant leukemia cells present lower levels of oxidative metabolism, in part due to downexpression of the following mitochondrial proteins: pyruvate dehydrogenase subunit alpha 1, succinate dehydrogenase, and voltage‐dependent anion channel. Those cells displayed higher expression levels of glucose transporter 1 and higher production of lactate. In addition, Lucena‐1 siRNA LMWPTP cells showed lower expression levels of glucose transporter 1 and lower activity of lactate dehydrogenase. On the other hand, K562 cells overexpressing LMWPTP presented higher expression/activity of both proteins. In this study, we show that LMWPTP is a pivotal mediator of metabolic reprogramming that confers survival advantages to leukemia cells against death stimuli. J. Cell. Biochem. 118: 3846–3854, 2017.

Effects of intermittent dietary supplementation with conjugated linoleic acid and fish oil (EPA/DHA) on body metabolism and mitochondrial energetics in mice

Understanding the mitochondrial processes that contribute to body energy metabolism may provide an attractive therapeutic target for obesity and co-morbidities. Here we investigated whether intermittent dietary supplementation with conjugated linoleic (CLA, 18:2n-6), docosahexaenoic (22:6n-3, DHA) and eicosapentaenoic (20:5n-3, EPA) acids, either alone or in combination, changes body metabolism associated with mitochondrial functions in the brain, liver, skeletal muscle and brown adipose tissue (BAT). Male C57Bl/6 mice were divided into groups: CLA (50% cis-9, trans-11; 50% trans-10, cis-12), EPA/DHA (64% EPA; 28% DHA), CLA plus EPA/DHA or control (linoleic acid). Each mouse received 3 g/kg b.w. of the stated oil by gavage on alternating days for 60 days. Dietary supplementation with CLA or EPA/DHA increased body VO2 consumption, VCO2 production and energy expenditure, being fish oil (FO) the most potent even in combination with CLA. Individually, both oils reduced mitochondrial density in BAT. CLA supplementation alone also a) elevated the expression of uncoupling proteins in soleus, liver and hippocampus and the uncoupling activity in the last two, ad this effect was associated with reduced hydrogen peroxide production in hippocampus; b) increased proteins related to mitochondrial fission in liver. EPA/DHA supplementation alone also a) induced mitochondrial biogenesis in liver, soleus and hippocampus associated with increased expression of PGC1-α; b) induced proteins related to mitochondrial fusion in the liver, and fission and fusion in the hippocampus. Therefore, this study shows changes on mitochondrial mechanisms induced by CLA and/or EPA/DHA that can be associated with elevated body energy expenditure.