Profile of pterostilbene-induced redox homeostasis modulation in cardiac myoblasts and heart tissue.

This study was designed to investigate the effect of pterostilbene (PTS) on cardiac oxidative stress in vitro, as this is a simple and promising methodology to study cardiac disease. Cardiac myoblasts (H9c2 cells) and homogenised cardiac tissue were incubated with the PTS and cyclodextrin (PTS + HPßCD) complex for 1 and 24 h, respectively, at concentrations of 50µM for the cells and 25 and 50µM for cardiac tissue. The PTS + HPßCD complex was used to increase the solubility of PTS in water. After the pretreatment period, cardiomyoblasts were challenged with hydrogen peroxide (6.67µM) for 10 min, while cardiac tissue was submitted to a hydroxyl radical generator system (30 min). Cellular viability, oxidative stress biomarkers (e.g. total reactive oxygen species (ROS), carbonyl assay and lipoperoxidation) and the antioxidant response (e.g. sulfhydryl and the antioxidant enzyme activities of superoxide dismutase, catalase and glutathione peroxidase) were evaluated. In cardiomyoblasts, the PTS + HPßCD complex (50µM) increased cellular viability. Moreover, the PTS + HPßCD complex also significantly increased sulfhydryl levels in the cells submitted to an oxidative challenge. In cardiac tissue, lipid peroxidation, carbonyls and ROS levels were significantly increased in the groups submitted to oxidative damage, while the PTS + HPßCD complex significantly reduced ROS levels in these groups. In addition, the PTS + HPßCD complex also provoked increased catalase activity in both experimental protocols. These data suggest that the PTS + HPßCD complex may play a cardioprotective role through a reduction of ROS levels associated with an improved antioxidant response.

Redox and metabolic strategies developed by anterior and posterior gills of the crab Neohelice granulata after short periods of hypo- or hyper-osmotic stress.

The aim of this study was to identify the response pattern of redox balance, Na+/K+ATPase activity and HSP70 expression in the posterior and anterior gills of the crab Neohelice granulata submitted to hypo- or hyper-osmotic stress for 1 h and 6 h. After 1 h of either type of osmotic stress, there was an increase in catalase activity, but a decrease in GSSG/GSH ratio (oxidized to reduced glutathione ratio) and Na+/K+ATPase activity in both gill sets. H2O2 levels decreased only in the posterior gills. H2O2 levels and Na+/K+ATPase activity remained reduced after 6 h of exposure to either type of osmotic stress in both gill sets. The GSSG/GSH ratio returned to initial levels after 6 h of hyper-osmotic stress, whereas it increased 10 times in both gill sets after hypo-osmotic stress. Furthermore, HSP70 protein expression increased in posterior gills after 6 h of hypo-osmotic stress. H2O2 levels in tank water decreased after hypo-osmotic challenge and increased after 6 h of hyper-osmotic stress, indicating increased H2O2 excretion. Therefore, N. granulata gills have redox, metabolic and molecular strategies to deal with rapid osmotic challenges, an important environmental parameter that influences juvenile and adult crab distribution and abundance within different populations.

Influence of estrogen on pulmonary arterial hypertension: role of oxidative stress.

Pulmonary arterial hypertension (PAH) is a disease that increases the pulmonary vascular resistance, causing hypertrophy and subsequent right heart failure. Oxidative stress is involved in the pathogenesis of PAH, and estrogen is considered an antioxidant. Thus, the aim of this study was to test the hypothesis that estrogen could attenuate PAH by modulating oxidative stress. Female Wistar rats were ovariectomized or suffered the surgery simulation (sham). After 7 days, subcutaneous pellets with 17ß-estradiol or sunflower oil were implanted. At this time, PAH was induced by means of a single dose of monocrotaline (MCT) (60 mg·kg(-1) i.p.). The experimental groups were as follows: (1) sham, (2) sham + MCT, (3) ovariectomy (O), (4) ovariectomy + MCT (OM), (5) ovariectomy + estrogen replacement + MCT (ORM). Hemodynamic measurements were performed 21 days after MCT or saline. Nonovariectomized animals were assessed in the stage of diestrus. Afterwards, the rats were killed to collect the heart, the lung and the liver to evaluate morphometry. Samples of the right ventricle were used to analyse the reduced glutathione : oxidized glutathione ratio. Lung congestion in the OM group, which was decreased in the ORM group, was observed. Right ventricle end-diastolic pressure was increased in the OM and the ORM groups. The glutathione ratio decreased in the groups O, OM and ORM. The data suggest that estrogen can exert great influence on the cellular redox balance. The maintenance of physiological estrogen levels may help to avoid the appearance of pulmonary oedema, characteristic of this model of PAH, and right ventricular failure.

Redox status and pro-survival/pro-apoptotic protein expression in the early cardiac hypertrophy induced by experimental hyperthyroidism.

This study was conducted to analyse the redox status and redox-sensitive proteins that may contribute to a non-genomic mechanism of cardiac hypertrophy induction by hyperthyroidism. Wistar rats, treated with L-thyroxine (T4) during 2 weeks (12 mg·l(-1) in drinking water), presented cardiac hypertrophy (68% higher than control), without signals of liver or lung congestion. Myocardial reduction of the reduced glutathione: oxidized glutathione (GSSG) ratio (45%) (redox status) and elevation in hydrogen peroxide concentration (H(2) O(2) ) (28%) were observed in hyperthyroid as compared with the control. No significant difference was found in thioredoxin (Trx), Trx reductase activity and Nrf2 (a transcriptional factor) protein expression between groups. Redox-sensitive proteins, quantified using Western blot, presented the following results: increased p-ERK: total extracellular-regulated kinase (ERK) (200%) and Bax:Bcl-2 (62%) ratios and reduced total-Akt (63%) and p-Akt (53%) expressions in the hyperthyroid rats as compared with the control. The redox imbalance, associated with increased immunocontent of a protein related to maladaptative growth (ERK) and reduced immunocontent of protein related to cytoprotection/survival (Akt), may suggest that the molecular scenario could favour the decompensation process of cardiac hypertrophy induced by experimental hyperthyroidism.

Thyroid hormone-induced haemoglobin changes and antioxidant enzymes response in erythrocytes.

Thyroid hormones modulate haemoglobin and reactive oxygen species (ROS) production, leading to antioxidant changes. This study evaluated the antioxidant response to ROS in erythrocytes in hypothyroid and hyperthyroid rats. Wistar rats were divided into four groups: control; hyperthyroid (T4-12 mg 1(-1) in drinking water); sham operated (simulation of thyroidectomy); and hypothyroid (thyroidectomized). Four weeks after, blood was collected and haemoglobin and T(4) levels, lipid peroxidation (LPO), protein oxidation, superoxide dismutase (SOD), catalase (CAT) , glutathione S-transferase (GST) and glutathione peroxidase (GPx) activities, and total radical antioxidant potential (TRAP) were measured. SOD, CAT and GST immunocontent was evaluated. Haemoglobin levels were increased in hyperthyroid erythrocytes. LPO and carbonyls were augmented (65% and 55%, respectively) in hyperthyroid and reduced (31% and 56%, respectively) in hypothyroid group. SOD and CAT activities have not changed, as well as CAT immunocontent. TRAP was diminished in both hyperthyroid and hypothyroid groups (36% and 37%, respectively). GST activity and immunocontent, as well as GPx activity, were increased in hyper and hypothyroid rats. The data suggest that thyroid hormone changes determine ROS concentration changes and decrease of some antioxidant defences that would lead to a compensatory answer of the GST and GPx enzymes, which could be consider as credible biomarkers.

Reactive oxygen and nitrogen species balance in the determination of thyroid hormones-induced cardiac hypertrophy mediated by renin-angiotensin system.

Role of reactive oxygen species (ROS)/nitric oxide (NO) balance and renin-angiotensin system in mediating cardiac hypertrophy in hyperthyroidism was evaluated in an in vivo and in vitro experimental model. Male Wistar rats were divided into four groups: control, thyroid hormone, vitamin E (or Trolox, its hydrosoluble analogue), thyroid hormone+vitamin E. Angiotensin II receptor (AT1/AT2) gene expression, immunocontent of AT1/AT2 receptors, angiotensinogen, NADPH oxidase (Nox2), and nitric oxide synthase isoforms, as well as ROS concentration (hydrogen peroxide and superoxide anion) were quantified in myocardium. Thyroid hormone increased ROS and NO metabolites, iNOS, nNOS and eNOS isoforms and it was accompanied by cardiac hypertrophy. AT1/AT2 expression and the immunocontent of angiotensinogen and Nox2 were enhanced by thyroid hormone. Antioxidants reduced ROS levels, Nox2, AT1/AT2, NOS isoforms and cardiac hypertrophy. In conclusion, ROS/NO balance may play a role in the control of thyroid hormone-induced cardiac hypertrophy mediated by renin-angiotensin system.

The role of redox signaling in cardiac hypertrophy induced by experimental hyperthyroidism.

This study was conducted to test whether oxidative stress activates the intracellular protein kinase B (AKT1) signaling pathway, which culminates with cardiac hypertrophy in experimental hyperthyroidism. Male Wistar rats were divided into four groups: control, vitamin E, thyroxine (T(4)), and T(4)+vitamin E. Hyperthyroidism was induced by T(4) administration (12 mg/l in drinking water for 28 days). Vitamin E treatment was given during the same period via s.c. injections (20 mg/kg per day). Morphometric and hemodynamic parameters were evaluated at the end of the 4-week treatment period. Protein oxidation, redox state (reduced glutathione, GSH/glutathione dissulfide, GSSG), vitamin C, total radical-trapping antioxidant potential (TRAP), hydrogen peroxide (H2O2), and nitric oxide metabolites (NO(X)) were measured in heart homogenates. The p-AKT1/AKT1 ratio, p-glycogen-synthase kinase (GSK)3B/GSK3B ratio, FOS, and JUN myocardial protein expression were also quantified by western blot after 4 weeks. Increases in biochemical parameters, such as protein oxidation (41%), H2O2 (62%), and NO(X) (218%), and increase in the left ventricular end-diastolic pressure were observed in the T(4) group. T(4) treatment also caused a decrease in GSH/GSSG ratio (83%), vitamin C (34%), and TRAP (55%). These alterations were attenuated by vitamin E administration to the hyperthyroid rats. Expression of p-AKT1/AKT1, p-GSK3B/GSK3B, FOS, and JUN were elevated in the T(4) group (by 69, 37, 130, and 33% respectively), whereas vitamin E administration promoted a significant reduction in their expression. These results indicate that oxidative stress plays an important role in cardiac hypertrophy, and suggest redox activation of AKT1 and JUN/FOS signaling pathways with H2O2 acting as a possible intracellular mediator in this adaptive response to experimental hyperthyroidism.

Oxidative stress activates insulin-like growth factor I receptor protein expression, mediating cardiac hypertrophy induced by thyroxine.

Thyroxine can cause cardiac hypertrophy by activating growth factors, such as IGF-I (insulin-like growth factor-I). Since oxidative stress is enhanced in the hyperthyroidism, it would control protein expression involved in this hypertrophy. Male Wistar rats were divided into four groups: (I) control, (II) vitamin E-supplemented (20 mg/kg/day subcutaneous), (III) hyperthyroid (thyroxine 12 mg/l, in drinking water), and (IV) hyperthyroid + vitamin E. After 4 weeks, the contractility and relaxation indexes of left ventricle (LV), and cardiac mass were increased by 54%, 60%, and 60%, respectively, in hyperthyroid group. An increase in lipid peroxidation (around 40%), and a decrease in total glutathione (by 20%) was induced by thyroxine and avoided by vitamin E administration. Superoxide dismutase (SOD) and glutathione-S-transferase (GST) activities were increased (by 83% and 54%, respectively) in hyperthyroid, and vitamin E avoided changes in SOD. Protein expression of SOD, GST, and IGF-I receptor (IGF-IR) were increased (by 87%, 84%, and 60%, respectively) by thyroxine, and vitamin E promoted a significant reduction in SOD and IGF-IR expression (by 36% and 17%, respectively). These results indicate that oxidative stress is involved in cardiac hypertrophy, and suggest a role for IGF-IR as a mediator of this adaptive response in experimental hyperthyroidism.

Peripheral markers of oxidative stress in chronic mercuric chloride intoxication.

The present study was designed to evaluate the time course changes in peripheral markers of oxidative stress in a chronic HgCl2 intoxication model. Twenty male adult Wistar rats were treated subcutaneously daily for 30 days and divided into two groups of 10 animals each: Hg, which received HgCl2 (0.16 mg kg(-1) day(-1)), and control, receiving the same volume of saline solution. Blood was collected at the first, second and fourth weeks of Hg administration to evaluate lipid peroxidation (LPO), total radical trapping antioxidant potential (TRAP), and superoxide dismutase (Cu,Zn-SOD), glutathione peroxidase (GPx), glutathione-S-transferase (GST), and catalase (CAT). HgCl2 administration induced a rise (by 26%) in LPO compared to control (143 +/- 10 cps/mg hemoglobin) in the second week and no difference was found at the end of the treatment. At that time, GST and GPx were higher (14 and 24%, respectively) in the Hg group, and Cu,Zn-SOD was lower (54%) compared to control. At the end of the treatment, Cu,Zn-SOD and CAT were higher (43 and 10%, respectively) in the Hg group compared to control (4.6 +/- 0.3 U/mg protein; 37 +/- 0.9 pmol/mg protein, respectively). TRAP was lower (69%) in the first week compared to control (43.8 +/- 1.9 mM Trolox). These data provide evidence that HgCl2 administration is accompanied by systemic oxidative damage in the initial phase of the process, which leads to adaptive changes in the antioxidant reserve, thus decreasing the oxidative injury at the end of 30 days of HgCl2 administration. These results suggest that a preventive treatment with antioxidants would help to avoid oxidative damage in subjects with chronic intoxication.

Peripheral markers of oxidative stress in chronic mercuric chloride intoxication

The present study was designed to evaluate the time course changes in peripheral markers of oxidative stress in a chronic HgCl2 intoxication model. Twenty male adult Wistar rats were treated subcutaneously daily for 30 days and divided into two groups of 10 animals each: Hg, which received HgCl2 (0.16 mg kg-1 day-1), and control, receiving the same volume of saline solution. Blood was collected at the first, second and fourth weeks of Hg administration to evaluate lipid peroxidation (LPO), total radical trapping antioxidant potential (TRAP), and superoxide dismutase (Cu,Zn-SOD), glutathione peroxidase (GPx), glutathione-S-transferase (GST), and catalase (CAT). HgCl2 administration induced a rise (by 26 percent) in LPO compared to control (143 ± 10 cps/mg hemoglobin) in the second week and no difference was found at the end of the treatment. At that time, GST and GPx were higher (14 and 24 percent, respectively) in the Hg group, and Cu,Zn-SOD was lower (54 percent) compared to control. At the end of the treatment, Cu,Zn-SOD and CAT were higher (43 and 10 percent, respectively) in the Hg group compared to control (4.6 ± 0.3 U/mg protein; 37 ± 0.9 pmol/mg protein, respectively). TRAP was lower (69 percent) in the first week compared to control (43.8 ± 1.9 mM Trolox). These data provide evidence that HgCl2 administration is accompanied by systemic oxidative damage in the initial phase of the process, which leads to adaptive changes in the antioxidant reserve, thus decreasing the oxidative injury at the end of 30 days of HgCl2 administration. These results suggest that a preventive treatment with antioxidants would help to avoid oxidative damage in subjects with chronic intoxication.

Myocardial antioxidant enzyme activities and concentration and glutathione metabolism in experimental hyperthyroidism.

Hyperthyroidism was induced in rats by l-thyroxine administration (12 mg/L in drinking water, 4 weeks). Animals were assessed hemodynamically, and heart, lung, and liver morphometry were performed. Lipid peroxidation (LPO) and protein oxidation (carbonyls) were measured in heart homogenates. It was quantified glutathione (GSH) metabolism, and antioxidant enzyme activities its and protein expression (by Western blot). At the end of treatment, it was observed cardiac hypertrophy, elevation of left ventricular systolic and end diastolic pressures, lung and liver congestion. LPO and carbonyls were increased in the hyperthyroid group, and GSH was decreased by 46% in the fourth week. Myocardial oxidative stress time course analysis revealed that it was increased in the second week of treatment. Antioxidant enzyme activities elevation was accompanied by protein expression induction in the hyperthyroid group in the fourth week. These results imply that hyperthyroidism generates myocardial dysfunction associated with oxidative stress inducing antioxidant enzyme activities and protein expression.

Myocardial antioxidant and oxidative stress changes due to sex hormones.

The purpose of the present study was to examine myocardial antioxidant and oxidative stress changes in male and female rats in the presence of physiological sex hormone concentrations and after castration. Twenty-four 9-week-old Wistar rats were divided into four groups of 6 animals each: 1) sham-operated females, 2) castrated females, 3) sham-operated males, and 4) castrated males. When testosterone and estrogen levels were measured by radioimmunoassay, significant differences were observed between the castrated and control groups (both males and females), demonstrating the success of castration. Progesterone and catalase levels did not change in any group. Control male rats had higher levels of glutathione peroxidase (50%) and lower levels of superoxide dismutase (SOD, 14%) than females. Control females presented increased levels of SOD as compared to the other groups. After castration, SOD activity decreased by 29% in the female group and by 14% in the male group as compared to their respective controls. Lipid peroxidation (LPO) was assessed to evaluate oxidative damage to cardiac membranes by two different methods, i.e., TBARS and chemiluminescence. LPO was higher in male controls compared to female controls when evaluated by both methods, TBARS (360%) and chemiluminescence (46%). Castration induced a 200% increase in myocardial damage in females as determined by TBARS and a 20% increase as determined by chemiluminescence. In males, castration did not change LPO levels. These data suggest that estrogen may have an antioxidant role in heart muscle, while testosterone does not.

Myocardial antioxidant and oxidative stress changes due to sex hormones

The purpose of the present study was to examine myocardial antioxidant and oxidative stress changes in male and female rats in the presence of physiological sex hormone concentrations and after castration. Twenty-four 9-week-old Wistar rats were divided into four groups of 6 animals each: 1) sham-operated females, 2) castrated females, 3) sham-operated males, and 4) castrated males. When testosterone and estrogen levels were measured by radioimmunoassay, significant differences were observed between the castrated and control groups (both males and females), demonstrating the success of castration. Progesterone and catalase levels did not change in any group. Control male rats had higher levels of glutathione peroxidase (50 percent) and lower levels of superoxide dismutase (SOD, 14 percent) than females. Control females presented increased levels of SOD as compared to the other groups. After castration, SOD activity decreased by 29 percent in the female group and by 14 percent in the male group as compared to their respective controls. Lipid peroxidation (LPO) was assessed to evaluate oxidative damage to cardiac membranes by two different methods, i.e., TBARS and chemiluminescence. LPO was higher in male controls compared to female controls when evaluated by both methods, TBARS (360 percent) and chemiluminescence (46 percent). Castration induced a 200 percent increase in myocardial damage in females as determined by TBARS and a 20 percent increase as determined by chemiluminescence. In males, castration did not change LPO levels. These data suggest that estrogen may have an antioxidant role in heart muscle, while testosterone does not