Sulforaphane improves redox homeostasis and right ventricular contractility in a model of pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is characterized by increased pulmonary vascular resistance, imposing overload on the right ventricle (RV) and imbalance of redox state. Our study investigated the influence of treatment with sulforaphane (SFN), found in cruciferous vegetables, on right ventricle (RV) remodeling and redox homeostasis in monocrotaline-induced pulmonary arterial hypertension (PAH). Male Wistar rats were separated into four groups: control (CTR); control + sulforaphane (CTR + SFN); monocrotaline (MCT); monocrotaline + sulforaphane (MCT + SFN). PAH induction was implemented by a single dose of MCT (60 mg/kg i.p.). Treatment with SFN (2.5 mg/kg/day i.p.) started on the 7th day after the MCT injection and persisted for 2 weeks. After 21 days of PAH induction, echocardiography, hemodynamic, and oxidative stress evaluation were performed. MCT group showed increase in RV hypertrophy, RV systolic area, RV systolic, mean pulmonary artery pressure (mPAP), and pulmonary vascular resistance (PVR), while exhibited a decrease in RV outflow tract AT/ET ratio, RV fractional shortening and tricuspid annular plane systolic excursion (TAPSE) compared to CTR (P<0.05). SFN-treated PAH attenuated detrimental changes in TPSE, mPAP, and PVR parameters. Catalase and GSH/GSSG ratio were diminished in MCT compared to CTR (P<0.05). SFN increased catalase and normalized GSH/GSSG ratio to control levels (P<0.05). Data express a benefit of SFN treatment on the cardiac function of rats with PAH associated with the cellular redox state.

Copaiba oil improves pulmonary nitric oxide bioavailability in monocrotaline-treated rats.

Oxidative stress is involved in increased pulmonary vascular resistance (PVR) and right ventricular (RV) hypertrophy, characteristics of pulmonary arterial hypertension (PAH). Copaiba oil, an antioxidant compound, could attenuate PAH damage. This study's aim was to determine the effects of copaiba oil on lung oxidative stress, PVR, and mean pulmonary arterial pressure (mPAP) in the monocrotaline (MCT) model of PAH. Male Wistar rats (170 g, n = 7/group) were divided into four groups: control, MCT, copaiba oil, and MCT + copaiba oil (MCT-O). PAH was induced by MCT (60 mg/kg i.p.) and, after 1 week, the treatment with copaiba oil (400 mg/kg/day gavage) was started for 14 days. Echocardiographic and hemodynamic measurements were performed. RV was collected for morphometric evaluations and lungs and the pulmonary artery were used for biochemical analysis. Copaiba oil significantly reduced RV hypertrophy, PVR, mPAP, and antioxidant enzyme activities in the MCT-O group. Moreover, increased nitric oxide synthase and decreased NADPH oxidase activities were observed in the MCT-O group. In conclusion, copaiba oil was able to improve the balance between nitric oxide and reactive oxygen species in lungs and the pulmonary artery and to reduce PVR, which could explain a decrease in RV hypertrophy in this PAH model.

Blueberry extract improves redox balance and functional parameters in the right ventricle from rats with pulmonary arterial hypertension.

PURPOSE: Pulmonary arterial hypertension (PAH) is a disease characterized by increased pulmonary vascular resistance and right ventricle (RV) failure. In this context, oxidative stress is an essential element contributing to PAH's pathophysiology. Thus, blueberry (BB), which has a high antioxidant capacity, emerges as a natural therapeutic approach in PAH. This work evaluated the effect of BB extract on redox balance in RV in a PAH's animal model. METHODS: Male Wistar rats (200 ± 20 g) (n = 72) were randomized into eight groups: control (CTR); monocrotaline (MCT); CTR and MCT treated at doses of 50, 100, and 200 mg/kg BB. PAH was induced by administration of MCT (60 mg/kg, intraperitoneal). Rats were treated with BB orally for 5 weeks (2 weeks before monocrotaline and 3 weeks after monocrotaline injection). On day 35, rats were submitted to echocardiography and catheterization, then euthanasia and RV harvesting for biochemical analyses. RESULTS: RV hypertrophy, observed in the MCT groups, was reduced with BB treatment. MCT elevated RV systolic pressure and pressure/time derivatives, while the intervention with BB decreased these parameters. PAH decreased RV output and pulmonary artery outflow acceleration/ejection time ratio, while increased RV diameters, parameters restored by BB treatment. Animals from the MCT group showed elevated lipid peroxidation and NADPH oxidase activity, outcomes attenuated in animals treated with BB, which also led to increased catalase activity. CONCLUSION: Treatment with BB partially mitigated PAH, which could be associated with improvement of RV redox state. Such findings constitute an advance in the investigation of the role of BB extract in chronic progressive cardiovascular diseases that involve the redox balance, such as PAH.

Thioredoxin system activation is associated with the progression of experimental pulmonary arterial hypertension.

In addition to being an antioxidant, thioredoxin (Trx) is known to stimulate signaling pathways involved in cell proliferation and to inhibit apoptosis. The aim of this study was to explore the role of Trx in some of these pathways along the progression of monocrotaline (MCT)-induced pulmonary arterial hypertension (PAH). Male rats were first divided into two groups: monocrotaline (MCT - 60 mg/kg i.p.) and control (received saline), that were further divided into three groups: 1, 2, and 3 weeks. Animals were submitted to echocardiographic analysis. Right and left ventricles were used for the measurement of hypertrophy, through morphometric and histological analysis. The lung was prepared for biochemical and molecular analysis. One week after MCT injection, there was an increase in thioredoxin reductase (TrxR) activity, a reduction in glutathione reductase (GR) activity, and an increase in Trx-1 and vitamin D3 up-regulated protein-1 (VDUP-1) expression. Two weeks after MCT injection, there was an increase in VDUP-1, Akt and cleaved caspase-3 activation, and a decrease in Trx-1 and Nrf2 expression. PAH-induced by MCT promoted a reduction in Nrf2 and Trx-1 expression as well as an increase in Akt and VDUP-1 expression after three weeks. The increase in pulmonary vascular resistance was accompanied by increased TrxR activity, suggesting an association between the Trx system and functional changes in the progression of PAH. It seems that Trx-1 activation was an adaptive response to MCT administration to cope with pulmonary remodeling and disease progression, suggesting a potential new target for PAH therapeutics.

Pulmonary arterial hypertension induces the release of circulating extracellular vesicles with oxidative content and alters redox and mitochondrial homeostasis in the brains of rats.

Pulmonary arterial hypertension (PAH) is characterized by increased resistance of the pulmonary vasculature and afterload imposed on the right ventricle (RV). Two major contributors to the worsening of this disease are oxidative stress and mitochondrial impairment. This study aimed to explore the effects of monocrotaline (MCT)-induced PAH on redox and mitochondrial homeostasis in the RV and brain and how circulating extracellular vesicle (EV) signaling is related to these phenomena. Wistar rats were divided into control and MCT groups (60 mg/kg, intraperitoneal), and EVs were isolated from blood on the day of euthanasia (21 days after MCT injections). There was an oxidative imbalance in the RV, brain, and EVs of MCT rats. PAH impaired mitochondrial function in the RV, as seen by a decrease in the activities of mitochondrial complex II and citrate synthase and manganese superoxide dismutase (MnSOD) protein expression, but this function was preserved in the brain. The key regulators of mitochondrial biogenesis, namely, proliferator-activated receptor gamma coactivator 1-alpha and sirtuin 1, were poorly expressed in the EVs of MCT rats, and this result was positively correlated with MnSOD expression in the RV and negatively correlated with MnSOD expression in the brain. Based on these findings, we can conclude that the RV is severely impacted by the development of PAH, but this pathological injury may signal the release of circulating EVs that communicate with different organs, such as the brain, helping to prevent further damage through the upregulation of proteins involved in redox and mitochondrial function.

The progression of pulmonary arterial hypertension induced by monocrotaline is characterized by lung nitrosative and oxidative stress, and impaired pulmonary artery reactivity.

The time-course of pulmonary arterial hypertension in the monocrotaline (MCT) model was investigated. Male rats were divided into two groups: MCT (received a 60 mg/kg i.p. injection) and control (received saline). The MCT and control groups were further divided into three cohorts, based on the follow-up interval: 1, 2, and 3 weeks. Right ventricle (RV) catheterization was performed and RV hypertrophy (RVH) was estimated. The lungs were used for biochemical, histological, molecular, and immunohistochemical analysis, while pulmonary artery rings were used for vascular reactivity. MCT promoted lung perivascular edema, inflammatory cells exudation, greater neutrophils and lymphocytes profile, and arteriolar wall thickness, compared to CTR group. Increases in pulmonary artery pressure and in RVH were observed in the MCT 2- and 3-week groups. The first week was marked by the presence of nitrosative stress (50% moderate and 33% accentuated staining by nitrotyrosine). These alterations lead to an adaptation of NO production by NO synthase activity after 2 weeks. Oxidative stress was evident in the third week, probably by an imbalance between endothelin-1 receptors, resulting in extracellular matrix remodeling, endothelial dysfunction, and RVH. Also, it was found a reduced pulmonary arterial vasodilatory response to acetylcholine after 2 (55%) and 3 (45%) weeks in MCT groups. The relevance of this study is precisely to show that nitrosative and oxidative stress predominate in distinct time windows of the disease progression.

Thyroid hormone treatment improved the response to maximum exercise test and preserved the ventricular geometry in myocardial infarcted rats.

NEW FINDINGS: What is the central question of this study? Does thyroid hormone treatment given after myocardial infarction preserve left ventricular function and treadmill exercise performance, and improve parameters of oxidative stress in the right ventricle and lungs of Wistar rats? What is the main finding and its importance? Thyroid hormone treatment improved the performance of the maximum exercise test in infarcted rats and induced effects in the heart and lungs that were similar to those observed with exercise training. This suggests there is a significant value of thyroid hormones for preserving exercise tolerance after myocardial infarction. ABSTRACT: Left ventricular myocardial infarction (MI) provokes damage in the heart and in other tissues, such as right ventricle and lungs. The present study elucidated whether thyroid hormone treatment (THT) may present positive effects in heart and lungs after MI, and whether or not these effects are similar to those of exercise training (ET). Male Wistar rats were divided into four groups: sham operated (SHAM), infarcted (MI), infarcted + exercise training (MIE), and infarcted + thyroid hormones (MIH). A maximum exercise test, left ventricle echocardiography, pulmonary histology, and oxidative stress in the right ventricle and lung were evaluated. THT and ET both reduced left ventricular dilatation and end-diastolic wall stress indexes to a similar extent. MI accentuated the content of macrophages and inflammatory infiltrate in the lungs, which was partially prevented in the MIH and MIE groups. THT and ET presented similar effects in the heart and lungs, and both improved the performance of the maximum exercise test in infarcted animals.

Pterostilbene improves cardiac function in a rat model of right heart failure through modulation of calcium handling proteins and oxidative stress.

This study explored the effect of pterostilbene (PTS) complexed with hydroxypropyl-ß-cyclodextrin (HPßCD) on right heart function, glutathione and glutaredoxin systems, and the expression of redox-sensitive proteins involved with regulation calcium levels in the experimental model of pulmonary arterial hypertension (PAH) induced by monocrotaline (MCT). After 7 days of PAH induction, rats received daily doses of the PTS:HPßCD complex (corresponding to 25, 50, or 100 mg·kg-1 of PTS) or vehicle (control group, CTR0) (an aqueous solution containing HPßCD; CTR0 and MCT0 (MCT group that did not receive PTS treatment)) via oral administration for 2 weeks. The results showed that the PTS:HPßCD complex increased the content of reduced glutathione and the activity of glutathione-S-transferase and glutaredoxin in the right ventricle (RV) of MCT-treated rats in a dose-dependent manner. Additionally, at higher doses, it also prevented the reduction of stroke volume and cardiac output, prevented myocardial performance index (MPI) increase, reduced lipoperoxidation, reduced total phospholamban, and increased the expression of sarcoplasmic reticulum calcium ATPase in the RV of MCT-treated rats. These results demonstrate that the PTS:HPßCD complex has a dose-dependent antioxidant mechanism that results in improved cardiac function in experimental right heart failure. Our results open a field of possibilities to PTS administration as new therapeutic approach to conventional therapy for right ventricular dysfunction. Novelty Pterostilbene complexed with hydroxypropyl-ß-cyclodextrin could be a new therapeutic approach. Pterostilbene complexed with hydroxypropyl-ß-cyclodextrin reestablishes redox homeostasis through glutathione metabolism modulation, leading to an improved MPI in pulmonary arterial hypertension-provoked right heart failure.

Blueberry extract decreases oxidative stress and improves functional parameters in lungs from rats with pulmonary arterial hypertension.

OBJECTIVES: Pulmonary arterial hypertension (PAH) is a condition characterized by an increased resistance of pulmonary vasculature, culminating in an increase in pulmonary pressure. This process involves disturbances in lung redox homeostasis, causing progressive right heart failure. In this context, the use of natural antioxidants, such as those found in blueberries, may represent a therapeutic approach. The aim of this study was to evaluate the effect of blueberry extract (BB) on functional parameters and oxidative stress levels in rat lungs with induced PAH. METHODS: Forty-eight male Wistar rats (weighing 200 ± 20 g) were randomized into five groups: control, monocrotaline, monocrotaline + BB 50, monocrotaline + BB 100, and monocrotaline + BB 200. PAH was induced by the administration of monocrotaline (60 mg/kg, intraperitoneal). Rats were treated with BB at doses of 50, 100, and 200 mg/kg via gavage for 5 wk (2 wk before monocrotaline and 3 wk after monocrotaline injection). At day 35, rats were submitted to echocardiography and catheterization. They were then sacrificed and lungs were harvested for biochemical analyses. RESULTS: BB increased the E/A ratio of blood flow across the tricuspid valve and tricuspid annular phase systolic excursion, as wells as decreased the mean pulmonary artery pressure of animals compared with the PAH group. Moreover, BB decreased total reactive species concentration and lipid oxidation, reduced activity of nicotinamide adenine dinucleotide phosphate oxidase and expression of xanthine oxidase, increased the activity of superoxide dismutase and restored sulfhydryl content in the animal lungs compared with those in the PAH group. Additionally, BB restored expression of the antioxidant transcriptional factor Nrf2 in the lungs of the animal subjects. Finally, BB normalized the endothelin receptor (ETA/ETB) expression ratio in the animal lungs, which were increased in the PAH group. CONCLUSION: Intervention with BB mitigated functional PAH outcomes through improvement of the pulmonary redox state. Our results provide a basis for future research on natural antioxidant interventions as a novel treatment strategy in PAH.

Copaiba Oil Attenuates Right Ventricular Remodeling by Decreasing Myocardial Apoptotic Signaling in Monocrotaline-Induced Rats.

There is an increase in oxidative stress and apoptosis signaling during the transition from hypertrophy to right ventricular (RV) failure caused by pulmonary arterial hypertension (PAH) induced by monocrotaline (MCT). In this study, it was evaluated the action of copaiba oil on the modulation of proteins involved in RV apoptosis signaling in rats with PAH. Male Wistar rats (±170 g, n = 7/group) were divided into 4 groups: control, MCT, copaiba oil, and MCT + copaiba oil. PAH was induced by MCT (60 mg/kg intraperitoneally) and, 7 days later, treatment with copaiba oil (400 mg/kg by gavage) was given for 14 days. Echocardiographic and hemodynamic measurements were performed, and the RV was collected for morphometric evaluations, oxidative stress, apoptosis, and cell survival signaling, and eNOS protein expression. Copaiba oil reduced RV hypertrophy (24%), improved RV systolic function, and reduced RV end-diastolic pressure, increased total sulfhydryl levels and eNOS protein expression, reduced lipid and protein oxidation, and the expression of proteins involved in apoptosis signaling in the RV of MCT + copaiba oil as compared to MCT group. In conclusion, copaiba oil reduced oxidative stress, and apoptosis signaling in RV of rats with PAH, which may be associated with an improvement in cardiac function caused by this compound.

Stilbenoid pterostilbene complexed with cyclodextrin preserves left ventricular function after myocardial infarction in rats: possible involvement of thiol proteins and modulation of phosphorylated GSK-3ß.

Oxidative stress alters signalling pathways for survival and cell death favouring the adverse remodelling of postmyocardial remnant cardiomyocytes, promoting functional impairment. The administration of pterostilbene (PTS), a phytophenol with antioxidant potential, can promote cardioprotection and represents a therapeutic alternative in acute myocardial infarction (AMI). The present study aims to explore the effects of oral administration of PTS complexed with hydroxypropyl-ß-cyclodextrin HPßCD (PTS:HPßCD complex) on the glutathione cycle, thiol protein activities and signalling pathways involving the protein kinase B (AKT) and glycogen synthase kinase-3ß (GSK-3ß) proteins in the left ventricle (LV) of infarcted rats. Animals were submitted to acute myocardial infarction through surgical ligation of the descending anterior branch of the left coronary artery and received over 8 days, by gavage, PTS:HPßCD complex at dose of 100 mg kg-1 day-1 (AMI + PTS group) or vehicle (aqueous solution with HPßCD) divided into Sham-operated (SHAM) and infarcted (AMI) groups. The results showed that the PBS: HPßCD complex decreased lipid peroxidation, prevented the decrease in thioredoxin reductase (TRxR) activity, and increased the activity of glutathione-S-transferase (GST) and glutaredoxin (GRx). Additionally, the expression of nuclear factor-erythroid two (Nrf2) and p-GSK-3ß was increased, whereas the p-GSK-3ß/GSK-3ß ratio was reduced in the LV of the infarcted animals. Overall, the PTS:HPßCD complex modulates activity of thiol-dependent enzymes and induces to the expression of antioxidant proteins, improving systolic function and mitigating the adverse cardiac remodelling post infarction.

Sulforaphane effects on oxidative stress parameters in culture of adult cardiomyocytes.

The aim of this study was to analyse the effect of sulforaphane (SFN) in cultures of adult cardiomyocytes, evaluating oxidative stress at different times. Cells were isolated, cultured, and divided into 4 groups: Control, SFN (5µM), H2O2 (5µM), and SFN+H2O2 (5µM both), and subdivided into groups undergoing 1 or 24 h of SFN incubation. After 1 h of incubation, reactive oxygen species production was 40% lower in the SFN group than the Control, and lipid peroxidation was 63% higher in the H2O2 group than the Control, and it was reduced in both of the SFN groups. The SOD activity was 59% higher in groups incubated for 24 h than in those incubated for 1 h. Protein expression of SOD-1 and SOD-2 was higher in the 24-h groups compared to the 1-h groups (55% and 24%, respectively). The Nrf2 protein expression in the 1-h groups was 17% higher than in the 24-h groups, and the SFN + H2O2 group had 40% more Nrf2 than the Control in the 1-h groups. Unlike Nrf2, the PGC-1α expression was 69% higher in the 24-h groups in relation to the 1-h groups. Regarding the 24-h groups, the SFN and SFN+H2O2 groups were higher than the Control (32% and 33%, respectively), and the SFN+H2O2 group was increased (21%) compared to H2O2. SFN had a protective action against oxidative damage, but had no effect on the antioxidant enzymes analyzed. The different responses in the expression of Nrf2 and PGC-1α in relation to the incubation times, draws attention to the importance of establishing a timeline of the action of SFN, since there appears to be a temporal difference in its mechanism in adult cardiomyocytes.

Pterostilbene reduces oxidative stress, prevents hypertrophy and preserves systolic function of right ventricle in cor pulmonale model.

BACKGROUND AND PURPOSE: In cor pulmonale, the increased afterload imposed on the right ventricle (RV) generates a maladaptive response, impairing the contractile cardiac function. Oxidative mechanisms play an important role in the pathophysiology and progression of this disease. The administration of pterostilbene (PTS), a phytophenol with antioxidant potential, may represent a therapeutic option. In the present study, we evaluated the effect of PTS complexed with hydroxypropyl-ß-cyclodextrin (HPßCD) on hypertrophy, contractile function and oxidative parameters in the RV of rats with pulmonary hypertension, induced by the administration of monocrotaline (MCT). EXPERIMENTAL APPROACH: The rats received daily doses of the PTS : HPßCD complex at 25, 50 or 100 mg·kg-1 , p.o., for 14 days. The diastolic function, E/A ratio, and systolic function, shortening fraction, fractional area change (FAC) and tricuspid annular plane systolic excursion (TAPSE) of the RV were determined by echocardiography. KEY RESULTS: The PTS : HPßCD complex reduced the production of NADPH oxidase-dependent superoxide anions and oxidative stress in the RV of MCT-treated rats in a dose-dependent manner. At higher doses it prevented the reduction in FAC and TAPSE in MCT-treated animals. CONCLUSIONS AND IMPLICATIONS: The PTS : HPßCD complex prevented the maladaptative remodelling and protected systolic function in the RV of rats with pulmonary hypertension. These cardioprotective mechanisms may be related, in part, to the antioxidant potential of PTS, favoured by the increased p.o. bioavailability promoted by the presence of HPßCD in the complex.

Sulforaphane effects on postinfarction cardiac remodeling in rats: modulation of redox-sensitive prosurvival and proapoptotic proteins.

This study investigated whether sulforaphane (SFN), a compound found in cruciferous vegetables, could attenuate the progression of post-myocardial infarction (MI) cardiac remodeling. Male Wistar rats (350 g) were allocated to four groups: SHAM (n=8), SHAM+SFN (n=7), MI (n=8) and MI+SFN (n=5). On the third day after surgery, cardiac function was assessed and SFN treatment (5 mg/kg/day) was started. At the end of 25 days of treatment, cardiac function was assessed and heart was collected to measure collagen content, oxidative stress and protein kinase. MI and MI+SFN groups presented cardiac dysfunction, without signs of congestion. Sulforaphane reduced fibrosis (2.1-fold) in infarcted rats, which was associated with a slight attenuation in the cardiac remodeling process. Both infarcted groups presented increases in the oxidative markers xanthine oxidase and 4-hydroxinonenal, as well as a parallel increase in the antioxidant enzymes glutathione peroxidase and superoxide dismutase. Moreover, sulforaphane stimulated the cytoprotective heme oxygenase-1 (HO-1) (38%). Oxidative markers correlated with ERK 1/2 activation. In the MI+SFN group, up-regulation of ERK 1/2 (34%) and Akt (35%), as well as down-regulation of p38 (52%), was observed. This change in the prosurvival kinase balance in the MI+SFN group was related to a down-regulation of apoptosis pathways (Bax/Bcl-2/caspase-3). Sulforaphane was unable to modulate autophagy. Taken together, sulforaphane increased HO-1, which may generate a redox environment in the cardiac tissue favorable to activation of prosurvival and deactivation of prodeath pathways. In conclusion, this natural compound contributes to attenuation of the fibrotic process, which may contribute to mitigation against the progression of cardiac remodeling postinfarction.

Effect of Deep Intramuscular Stimulation and Transcranial Magnetic Stimulation on Neurophysiological Biomarkers in Chronic Myofascial Pain Syndrome.

OBJECTIVE: The aim was to assess the neuromodulation techniques effects (repetitive transcranial magnetic stimulation [rTMS] and deep intramuscular stimulation therapy [DIMST]) on pain intensity, peripheral, and neurophysiological biomarkers chronic myofascial pain syndrome (MPS) patients. DESIGN: Randomized, double blind, factorial design, and controlled placebo-sham clinical trial. SETTING: Clinical trial in the Laboratory of Pain and Neuromodulation at Hospital de Clínicas de Porto Alegre (NCT02381171). SUBJECTS: We recruited women aged between 19- and 75-year old, with MPS diagnosis. METHODS: Patients were randomized into four groups: rTMS + DIMST, rTMS + sham-DIMST, sham-rTMS + DIMST, sham-rTMS + sham-DIMST; and received 10 sessions for 20 minutes each one (rTMS and DIMST). Pain was assessed by visual analogue scale (VAS); neurophysiological parameters were assessed by transcranial magnetic stimulation; biochemical parameters were: BDNF, S100ß, lactate dehydrogenase, inflammatory (TNF-α, IL6, and IL10), and oxidative stress parameters. RESULTS: We observed the pain relief assessed by VAS immediately assessed before and after the intervention (P < 0.05, F(1,3)= 3.494 and F(1,3)= 4.656, respectively); in the sham-rTMS + DIMST group and both three active groups in relation to sham-rTMS + sham-DIMST group, respectively. There was an increase in the MEP after rTMS + sham-DIMST (P < 0.05). However, there was no change in all-peripheral parameters analyzed across the treatment (P > 0.05). CONCLUSION: Our findings add additional evidence about rTMS and DIMST in relieving pain in MPS patients without synergistic effect. No peripheral biomarkers reflected the analgesic effect of both techniques; including those related to cellular damage. Additionally, one neurophysiological parameter (increased MEP amplitude) needs to be investigated.