Activation Of Redox-sensitive Kinases Research Articles

Effect of Subtoxic DDT Exposure on Glucose Uptake and Insulin Signaling in Rat L6 Myoblast-Derived Myotubes.

Exposure to persistent organic pollutants including dichlorodiphenyltrichloroethane (DDT) induces insulin resistance. But the mechanism is not clearly known. The present study was designed to explore the effect of subtoxic DDT exposure on (1) insulin-stimulated glucose uptake, (2) malondialdehyde (MDA) level and total antioxidant content, (3) activation of redox sensitive kinases (RSKs), and (4) insulin signaling in rat L6 myoblast-derived myotubes. Exposure to 30 mg/L and 60 mg/L of DDT for 18 hours dose dependently decreased glucose uptake and antioxidant content in myotubes and increased MDA levels. The exposures did not alter tumor necrosis factor α (TNF-α) level as determined by enzyme-linked immunosorbent assay, despite decreased messenger RNA expression following DDT exposures. Phosphorylation of c-Jun N-terminal kinases and IκBα, an inhibitory component of nuclear factor κB (NFκB), was increased, suggesting activation of RSKs. The level of tyrosine phosphorylation of insulin receptor substrate 1 and serine phosphorylation of protein kinase B (Akt) on insulin stimulation decreased in myotubes with exposure to subtoxic concentrations of DDT, but there was no change in tyrosine phosphorylation level of insulin receptors. We conclude that subtoxic DDT exposure impairs insulin signaling and thereby induces insulin resistance in muscle cells. Data show that oxidative stress-induced activation of RSKs is responsible for impairment of insulin signaling on DDT exposure.

Sub-toxic exposure to lindane activates redox sensitive kinases and impairs insulin signaling in muscle cell culture: The possible mechanism of lindane-induced insulin resistance

Lindane exposure is claimed to be involved in pathogenesis of type 2 diabetes mellitus (T2DM) and insulin resistance state by an as yet unknown mechanism. The redox sensitive kinases (RSKs) and heat shock proteins (HSPs) interfere with insulin signaling and induce insulin resistance. The present study was designed to explore the mechanism of insulin resistance induced by sub-toxic lindane exposure. In an in vitro study, exposure to 60 mg/L and 120 mg/L of lindane for 18 h on rat L6 myoblasts derived myotubes significantly increased malondialdehyde level & superoxide dismutase activity, decreased total antioxidant level and insulin-induced glucose uptake in a dose dependent manner. The extent of activation of RSKs and HSP25 as measured by western blot from the extent of phosphorylation of IκBα, p38 MAPK, JNK & HSP25 in lindane-exposed myotubes was higher. HSP70 was induced and insulin signaling as measured from tyrosine phosphorylation of insulin receptor (IR) & insulin receptor substrate-1 (IRS-1) and serine phosphorylation of Akt was attenuated in comparison to those in untreated myotubes. We conclude that sub-toxic lindane exposure induces oxidative stress, activates RSKs & HSP25 and induces HSP25. These in turn, impair insulin signaling to impart insulin resistance in myotubes induced by sub-toxic lindane exposure.

OP-39 - NADPH oxidase is a target for (-)-epicatechin in the prevention of fatty acid-induced insulin resistance

While diets rich in fruit and vegetables can decrease the risk for type 2 diabetes (T2D), diets rich in fat and carbohydrates can increase it. The flavanol-3-ol (-)-epicatechin (EC) can improve insulin sensitivity both in humans and animal models of T2D. NADPH oxidases and oxidative stress can contribute to the development of insulin resistance. This study investigated the capacity of EC and EC metabolites (ECM) to downregulate NADPH oxidases and oxidative stress, and its association to an improvement of insulin sensitivity. This was studied in in vivo (high fat-fed mice) and in vitro (HepG2 cells) conditions of hepatic lipid overload. EC decreased NOX3/NOX4 liver expression and mitigated oxidative stress in high fat-fed mice. In HepG2 cells, incubation with palmitate increased: i) lipid deposition, ii) NOX3/NOX4 expression, iii) NADPH oxidase activity, and iv) oxidative stress; promoting v) the activation of redox-sensitive kinases (JNK and IKK), and vi) impaired insulin responses. Physiological concentrations of EC and ECM, and NADPH oxidase inhibitors (apocynin, VAS2870) prevented all those deleterious effects of palmitate. The obtained results points to NADPH oxidases as an important target in the capacity of EC to prevent Western style diet-induced insulin resistance.

(-)-Epicatechin and its metabolites prevent palmitate-induced NADPH oxidase upregulation, oxidative stress and insulin resistance in HepG2 cells

While diets rich in fruit and vegetables can decrease the risk for type 2 diabetes (T2D), diets rich in fat and carbohydrates can increase it. The flavanol-3-ol (-)-epicatechin (EC) can improve insulin sensitivity both in humans and animal models of T2D. NADPH oxidases and oxidative stress can contribute to the development of insulin resistance. This study investigated the capacity of EC and EC metabolites (ECM) to downregulate NADPH oxidases and oxidative stress, and its association to an improvement of insulin sensitivity. This was studied in in vivo (high fat-fed mice) and in vitro (HepG2 cells) conditions of hepatic lipid overload. EC decreased NOX3/NOX4 liver expression and mitigated oxidative stress in high fat-fed mice. In HepG2 cells, incubation with palmitate increased: i) lipid deposition, ii) NOX3/NOX4 expression, iii) NADPH oxidase activity, and iv) oxidative stress; promoting v) the activation of redox-sensitive kinases (JNK and IKK), and vi) impaired insulin responses. Physiological concentrations of EC and ECM, and NADPH oxidase inhibitors (apocynin, VAS2870) prevented all those deleterious effects of palmitate. The obtained results points to NADPH oxidases as an important target in the capacity of EC to improve insulin sensitivity in conditions of liver lipid overload, as those associated with Western-style diets.

Abstract 19439: Melatonin Blunts Pressure-overload Left Ventricular Hypertrophy and Improves Cardiac Performance in Pathologic and PPhysiologic Cardiac Hypertrophy

Introduction: Reactive oxygen species (ROS) has been established as key mediators in cardiac hypertrophy and function through activation of redox-sensitive kinases, by stimulating NADPH oxidase. Melatonin has been described as having potent antioxidant properties in different tissues. Hypothesis: The hormone melatonin, would prevent pressure-overload (pathologic) hypertrophy, Methods: Mice (C57Bl6) were subjected to transverse aortic constriction (TAC), and treated with melatonin (MEL, 10 mg/Kg/day) in water (“TAC+MEL”) or placebo (TAC). A second group underwent intense treadmill exercise for 8 weeks and also received melatonin (EX-MEL) or water (EX). A sham operated group served as control (“sham”). After 7 weeks of TAC and 8 weeks of exercise, cardiac function was assessed by echocardiography and pressure –volumetric analysis. Cardiac ROS activity (TBARS, NRF2, Keap-1) was determined. Results: TAC promoted LVH in mice (LVMI: 6.38±0.38 mg/mm tibia length (sham), 8.11±0.63 (TAC) which was blunted by melatonin (LVMI: 6.34±0.31 (TAC+MEL) mg/mm, P<0.05). Increased LVH was associated with decreased endocardial shortening measured by echocardiography, from 61±2 (sham) to 54±1.8 % (TAC + placebo), P <0.05. This decrease in fractional shortening was less pronounced in the TAC+ MEL group (58±2.9%) as well as in PV loop analysis. The antioxidant action of MEL was evidenced by the decrease in myocardial lipid peroxidation (TBARS) from 0.35±0.02 (TAC) to 0.25±0.02 (TAC+MEL) nmol/mg of protein, P<0.05. Adult mice isolated myocyte studies revealed that Ang 2 (20nM) increased sarcomere shortening and calcium transient, both of which were blunted when cells were pre-incubated with melatonin (p< 0.005, Ang2 vs MEL + Ang2). In the exercise group, melatonin had only a small, non-significant reduction in LVMI, but a marked improvement in cardiac performance (PV loop analysis). Conclusion: Melatonin prevents pressure-overload LVH and improves contractility due to its antioxidant action. In physiologic LVH (exercise), melatonin does not affect hypertrophy, but exerts a positive effect on cardiac performance. These effects seem to be, at least in part, mediated through melatonin interfering with the Keap-1- Nrf2 antioxidant pathway.

Curcumin Attenuates Oxidative Stress and Activation of Redox-Sensitive Kinases in High Fructose- and High-Fat-Fed Male Wistar Rats.

The present study was carried out to investigate the effects of curcumin on oxidative stress and redox-sensitive kinases in high fructose- and high-fat-fed rats. Sixty rats were randomly divided into six groups with ten animals each. Rats were fed with a standard rodent diet, high fructose diet (60%), and high-fat diet (30%). Curcumin was administered to control, high fructose and high fat diet groups for ten weeks. At the end of the study, body weight and blood glucose levels were measured. The antioxidant enzymes GSH (reduced glutathione), GPx (glutathione peroxidase), and catalase activities were estimated in the blood. MDA, TAS, and TOS were estimated in the plasma, liver, and kidney. Curcumin treatment decreased body weight and blood glucose levels in the rats fed with fructose and high-fat diet. Antioxidant enzymes and plasma TAS were significantly improved by curcumin treatment in high fructose-fed rats, whereas in high-fat-fed rats, there was an increase only in the GPx activity. Curcumin significantly attenuated the elevation of plasma MDA and TOS in both diet groups. Hepatic MDA and TOS were found to be decreased upon curcumin supplementation in both diet groups, whereas a decrease in the renal MDA levels was observed only in fructose-treated rats, not in fat-fed rats. Curcumin treatment elevated liver TAS in rats fed only with the fructose-rich diet. Curcumin showed a significant decrease in the oxidative stress index (OSI) in plasma, liver, and kidney tissues in both diet groups. ERK phosphorylation was significantly decreased in both diet groups by curcumin treatment. Similarly, curcumin reduced the phosphorylation of p38 MAPK only in the high fructose-fed rats, not in the high-fat-fed rats. No significant changes were found in JNK phosphorylation in both diet groups. Thus, curcumin may be effective in the management of diet-induced oxidative stress and could be explored as a therapeutic adjuvant against complications associated with obesity and diabetes.

Role of autocrine/paracrine mechanisms in response to myocardial strain

Myocardial strain triggers an autocrine/paracrine mechanism known to participate in myocardial hypertrophy development. After the onset of stretch, there is a rapid augmentation in developed tension due to an increase in myofilament calcium sensitivity (the Frank Starling mechanism) followed by a gradual increase in tension over the next 10-15min. This second phase is called the slow force response (SFR) to stretch and is known to be the result of an increase in calcium transient amplitude. In the present review, we will discuss what is known thus far about the SFR, which is the in vitro equivalent of the Anrep effect and the mechanical counterpart of the autocrine/paracrine mechanism elicited by myocardial stretch. The chain of events triggered by myocardial stretch comprises: (1) release of angiotensin II, (2) release/formation of endothelin, (3) NADPH oxidase activation and transactivation of the EGFR, (4) mitochondrial reactive oxygen species production, (5) activation of redox-sensitive kinases, (6) NHE-1 hyperactivity, (7) increase in intracellular Na(+) concentration, and (8) increase in Ca(2+) transient amplitude through the Na(+)/Ca(2+) exchanger. The evidence for each step of the intracellular signaling pathway leading to the development of SFR and their relationship with the mechanisms proposed for cardiac hypertrophy development will be analyzed.

Role of NADPH oxidase in H9c2 cardiac muscle cells exposed to simulated ischaemia‐reperfusion

Oxidative stress is associated with several cardiovascular pathologies, including hypertension, cardiac hypertrophy and heart failure. Although oxidative stress is also increased after ischaemia-reperfusion (I/R), little is known about the role and the activation mechanisms, in cardiac myocytes under these conditions, of NADPH oxidase, a superoxide-producing enzyme. We found that rat cardiac muscle cells (H9c2) subjected to an in vitro simulated ischaemia (substrate-free medium plus hypoxia) followed by 'reperfusion', displayed increased reactive oxygen species (ROS) production attributable to a parallel increase of NADPH oxidase activity. Our investigation on mechanisms responsible for NADPH oxidase activation showed a contribution of both the increase of NOX2 expression and p47(phox) translocation to the membrane. We also found that the increase of NADPH oxidase activity was associated with higher levels of lipid peroxidation, the activation of redox-sensitive kinases, in particular ERK and JNK, and with cell death. Diphenyleneiodonium (DPI), a flavoprotein inhibitor used as NADPH oxidase inhibitor, prevented I/R-induced ROS formation in treated cells, together with the related lipoperoxidative damage, and JNK phosphorylation without affecting ERK activation, resulting in protection against cell death. Our results provide evidence that NADPH oxidase is a key enzyme involved in I/R-induced oxidant generation and suggest it can be a possible target in cardioprotective strategies against I/R injury, a condition of great importance in human pathology.

NADPH oxidase is related with lipid peroxidation and redox-sensitive kinase activation in human failing hearts

NADPH oxidase is related with lipid peroxidation and redox-sensitive kinase activation in human failing hearts

WITHDRAWN: NADPH oxidase is related with lipid peroxidation and redox-sensitive kinase activation in human failing hearts

WITHDRAWN: NADPH oxidase is related with lipid peroxidation and redox-sensitive kinase activation in human failing hearts

Activation of NADPH oxidase during progression of cardiac hypertrophy to failure.

Increased reactive oxygen species (ROS) production is implicated in the pathophysiology of left ventricular (LV) hypertrophy and heart failure. However, the enzymatic sources of myocardial ROS production are unclear. We examined the expression and activity of phagocyte-type NADPH oxidase in LV myocardium in an experimental guinea pig model of progressive pressure-overload LV hypertrophy. Concomitant with the development of LV hypertrophy, NADPH-dependent O2- production in LV homogenates, measured by lucigenin (5 micro mol/L) chemiluminescence or cytochrome c reduction assays, significantly and progressively increased (by approximately 40% at the stage of LV decompensation; P<0.05). O2- production was fully inhibited by diphenyleneiodonium (100 micromol/L). Immunoblotting revealed a progressive increase in expression of the NADPH oxidase subunits p22(phox), gp91(phox), p67(phox), and p47(phox) in the LV hypertrophy group, whereas immunolabeling studies indicated the presence of oxidase subunits in cardiomyocytes and endothelial cells. In parallel with the increase in O2- production, there was a significant increase in activation of extracellular signal-regulated kinase 1/2, extracellular signal-regulated kinase 5, c-Jun NH2-terminal kinase 1/2, and p38 mitogen-activated protein kinase. These data indicate that an NADPH oxidase expressed in cardiomyocytes is a major source of ROS generation in pressure overload LV hypertrophy and may contribute to pathophysiological changes such as the activation of redox-sensitive kinases and progression to heart failure.