Effect of Phonophoresis and Copaiba Oil on Oxidative Stress Biomarkers after Skeletal Muscle Injury in Rats.

The objective of this study was to analyze the effectiveness of phonophoresis with copaiba oil gel, in comparison to therapeutic pulsed ultrasound alone or topical application of copaiba oil gel, on oxidative stress after a traumatic muscle injury. Forty male Wistar rats were divided into five groups: control, muscle injury, therapeutic pulsed ultrasound (TPU), copaiba oil gel (CO) and TPU plus CO. TPU and CO application occurred at 2, 12, 24, 48, 72 and 96 h after injury. The gastrocnemius muscle was injured by mechanical trauma. Malondialdehyde (a lipoperoxidation marker) and superoxide dismutase and catalase (antioxidant enzymes) were assessed 98 h after muscle injury. All were elevated in the muscle injury group. There was a significant difference among treatment groups favoring TPU plus CO for reducing malondialdehyde levels, but all treatments reduced superoxide dismutase and catalase activity, with no between-groups difference. In conclusion, phonophoresis-the application of TPU plus CO-was superior to TPU or CO alone for reducing lipoperoxidation. Phonophoresis, TPU alone and CO were all effective in decreasing antioxidant enzyme activity after a traumatic skeletal muscle injury.

A novel quercetin/ß-cyclodextrin transdermal gel, combined or not with therapeutic ultrasound, reduces oxidative stress after skeletal muscle injury.

A gel containing the inclusion complex of quercetin and ß-cyclodextrin was developed in order to verify its effects, isolated or using phonophoresis, on oxidative biomarkers after skeletal muscle injury. 30 male rats were divided into one of five groups: Control (CTRL), Muscle Injury (MI), Therapeutic Pulsed Ultrasound (TPU), Therapeutic Pulsed Ultrasound plus Quercetin (TPU plus gel-QUE) or Quercetin gel (QUE). Quercetin gel was complexed with ß-Cyclodextrin (ß-CD) using chromatography (HPLC). TPU and quercetin application occurred with 2, 12, 24, 48, 72, 96 hours intervals after injury. Gastrocnemius muscle was injured by mechanical trauma. Lipid peroxidation, superoxide dismutase activity, and catalase activity were assessed. The inclusion complex exhibited adequate entrapment efficiency, relative density and pH. The viscosity of the complex showed a non-Newtonian pseudoplastic behavior. Quercetin/ß-cyclodextrin gel reduced lipid peroxidation, superoxide dismutase activity and catalase activity compared to muscle injury group. Similarly, phonophoresis and TPU also reduced the levels of these oxidative biomarkers. In conclusion, quercetin/ß-cyclodextrin transdermal gel reduces oxidative stress biomarkers after skeletal muscle injury irrespective of using phonophoresis.

Hemodynamic and creatine kinase changes after a 12-week equipment-based Pilates training program in hypertensive women.

Exercise therapy is a conservative strategy to manage hypertension. The aim of this study was to investigate the effects of a 12-week equipment-based Pilates training program on the hemodynamics of medication-controlled hypertensive women. Forty-five women were classified into two groups: medication-controlled hypertensive (n = 30) or control (n = 15). To be eligible for the hypertensive group, participants had to have a clinical diagnosis of hypertension, be taking medication to control their blood pressure and not to have done any exercise activity in the previous three months. In the control group, the participants needed to have normal blood pressure levels and not to have done any exercise activity in the previous three months. All women participated in a 12-week equipment-based Pilates training program (2 × 60min sessions per week). Systolic and diastolic blood pressure (SBP, DBP), heart rate (HR) and creatine kinase levels (CK) were assessed before and after the program. SBP, DBP and heart rate were also assessed before and after each session. There was no statistically significant difference after Pilates training for systolic (normotensive MD 4.1, 95%CI -9.2 to 17.5, hypertensive MD 3.8, 95%CI -5.3 to 13.1) and diastolic (normotensive MD 5.8, 95%CI -2.0 to 13.7, hypertensive MD 4.0, 95%CI -4.3 to 12.4) blood pressure. Heart rate was reduced after Pilates training in both normotensive (MD 4.5, 95%CI 1.1 to 7.8) and medication-controlled hypertensive (MD 7.9, 95%CI 4.4 to 11.4) women. Creatine kinase activity was reduced after Pilates training in medication-controlled hypertensive women (p = 0.019). Blood pressure was not altered, but heart rate and creatinine kinase activity were reduced following 12 weeks of Equipment-based Pilates training.

Topical application of (S)-(-)-limonene is as effective as phonophoresis for improving oxidative parameters of injured skeletal muscle in rats.

The aim of this study is to investigate the effects of limonene, alone or associated with therapeutic ultrasound, on oxidative stress following skeletal muscle injury. Thirty male Wistar rats were divided into 5 groups: CTR-control, MI-muscle injury without treatment, TPU-therapeutic pulsed ultrasound alone, TPU + LIM-phonophoresis with 5% limonene, and LIM-5% limonene applied topically. Muscle injury was induced by a mechanical abrupt impact over gastrocnemius muscle. The animals were treated in the following intervals: 2, 12, 24, 48, 72, and 96 h after injury. Blood and gastrocnemius samples were collected 98 h after lesion for data analysis. Creatine kinase (CK) and lactate dehydrogenase (LDH) activity, lipid peroxidation (TBARS) levels, catalase (CAT), and superoxide dismutase (SOD) activity were assessed. CK (p = 0.01), SOD activity (p < 0.01), and TBARS levels (p < 0.01) were increased after injury. There was no effect on LDH levels in any group. Phonophoresis (TABRS p < 0.01; SOD p = 0.01), TPU alone (TBARS p < 0.01; SOD p = 0.01), and LIM alone (TBARS p < 0.01; SOD p < 0.01) reduced TBARS levels and SOD activity after muscle injury. There was no change for CAT activity after injury. Only phonophoresis reduced CK activity after injury (p < 0.01). There was no difference between phonophoresis, TPU alone and LIM alone groups for TBARS, SOD, CAT, and LDH. Limonene alone and TPU alone were effective in reducing oxidative stress parameters after skeletal muscle injury. Only phonophoresis decreased CK activity. Skeletal muscle injury increases reactive oxidative species (ROS) levels and muscle proteins activity as creatine kinase (CK) and lactate dehydrogenase (LDH). Five percent limonene, alone or associated with therapeutic pulsed ultrasound, exhibited reduction of CK, superoxide dismutase (SOD) and catalase (CAT) activity, and lipid peroxidation markers (TBARS). Graphical abstract.

Effect of Pulsed Therapeutic Ultrasound and Diosmin on Skeletal Muscle Oxidative Parameters.

Cyclodextrins (CDs) have been widely used as a promising alternative in the formation of inclusion complexes with poorly soluble molecules. From this perspective, the present study aimed to study the inclusion complexes of diosmin in ß-cyclodextrin, chemically quantify the diosmin-in-gel preparation and analyze the stability of the gels. Furthermore, we evaluated the effect of therapeutic pulsed ultrasound (TPU) in association with the gel-diosmin complex on the parameters of muscle damage and oxidative stress in rats. Serum creatine kinase (CK) levels were used as an indicator of skeletal muscle injury. Lipid peroxidation (thiobarbituric acid-reactive substances [TBARS]) and superoxide dismutase and catalase activities were used as indicators of oxidative stress. The results obtained indicated that the inclusion complex obtained by co-evaporation had the highest complexation efficiency and stability; there was no change in the features of diosmin on incorporation into the Carbopol gel. Additionally, a significant (p <0.05) decrease was observed in CK levels (TPU plus gel-diosmin: 178.4 ± 85.3 U/L) relative to the untreated group (527.8 ± 46.1 U/L). Levels of TBARS were lower in the TPU plus gel-diosmin group (0.008 ± 0.0004 nmol malondialdehyde/mg protein, p <0.05) compared with the untreated group (0.081 ± 0.011 nmol malondialdehyde/mg protein, p <0.05, n = 6). Catalase activity did not statistically significantly differ between the treatment groups, and superoxide dismutase activity was lower in the diosmin-treated group (0.320 ± 0.11 U/mg protein) compared with the untreated group (0.983 ± 0.40 U/mg protein). These results suggest that TPU in association with the diosmin-gel complex is effective in reducing muscle damage and oxidative stress after mechanical trauma.

Therapeutic ultrasound associated with copaiba oil reduces pain and improves range of motion in patients with knee osteoarthritis / Ultrassom terapêutico associado ao óleo de copaíba reduz a dor e melhora amplitude de movimento de pacientes com osteoartrite de joelho

Abstract Introduction: Osteoarthritis is a disease that affects millions of Brazilians.Therapeutic ultrasound has been used in its treatment, either alone or associated with drugs. Objective: The aim of this study was to evaluate the effects of ultrasound (US) associated with Copaiba oil (CO) on knee osteoarthritis. Methods: Patients were divided into three different groups: US, US+CO, CO.Ten treatment sessions were held twice a week, 30 minutes each.Pain intensity was assessed through the Visual Analog Scale (VAS) and Range of Motion (ROM) by goniometry, and muscle strength was assessed by means of the Medical Research Council Scale. Statistical analysis was performed by Cohen's d test, student's t test and ANOVA, considering p<0.05 as significant. Results: Pain reduced in all groups.The US+CO group (d = -3.50) presented larger effect size when compared to the other groups. Regarding ROM, the largest effect size was observed in the US+CO group for flexion (d = 0.86) and extension (d = 0.97) in comparison with the remainder groups. Muscle strength increased in the US (d = 1.54) and US+CO (d = 1.60) groups for flexion.Regarding extension, the US group presented the largest effect size (d = 1.80). Conclusion: Therapeutic ultrasound associated with copaiba oil is a practical and effective therapy for the treatment of inflammatory diseases such as osteoarthritis.

Resumo Introdução: A osteoartrite é uma doença que afeta milhões de brasileiros. O ultrassom terapêutico tem sido utilizado em seu tratamento tanto sozinho, quanto associado a fármacos. Objetivo: Avaliar o efeito do ultrassom (US) associado ao óleo de copaíba (OC) em pacientes com osteoartrite de joelho. Métodos: Os pacientes foram distribuídos em 3 grupos distintos: US, US+OC e OC. Foram realizadas 10 sessões de tratamento, duas vezes por semana durante 30 minutos. A intensidade da dor foi avaliada pela Escala Visual Analógica da Dor (EVA), amplitude de movimento ADM pela goniometria e força muscular pelo Score Medical Research Council. A análise estatística foi feita pelo Teste T de Student e ANOVA e a magnitude do efeito (d), considerando p<0,05 como valores significativos. Resultados: A dor foi atenuada em todos os grupos, sendo apresentada uma magnitude maior do efeito para o grupo US+OC (d = -3,50) quando comparado aos demais grupos. Em relação a ADM a magnitude do efeito foi maior no grupo US+OC (d= 0,86) para a flexão e extensão (d = 0,97) quando comparados com os outros grupos. Na variável força muscular os grupos US (d= 1,54) e US+OC (d = 1,60) foram mais eficazes no movimento de flexão e na extensão, o grupo US exibiu o maior tamanho de efeito (d = 1,80) quando comparados aos demais grupos. Conclusão: O ultrassom terapêutico associado ao óleo de copaíba é uma terapia efetiva e prática para o tratamento de doenças inflamatórias, tais como a osteoartrite.

Dissection of the Effects of Quercetin on Mouse Myocardium.

Quercetin is a plant flavonoid with several biological activities. This study aimed to describe quercetin effects on contractile and electrophysiological properties of the cardiac muscle as well as on calcium handling. Quercetin elicited positive inotropism that was significantly reduced by propranolol indicating an involvement of the sympathetic nervous system. In cardiomyocytes, 30 µM quercetin increased ICa,L at 0 mV from -0.95 ± 0.01 A/F to -1.21 ± 0.08 A/F. The membrane potential at which 50% of the channels are activated (V0.5 ) shifted towards more negative potentials from -13.06 ± 1.52 mV to -19.26 ± 1.72 mV and did not alter the slope factor. Furthermore, quercetin increased [Ca2+ ]i transient by 28% when compared to control. Quercetin accelerated [Ca2+ ]i transient decay time, which could be attributed to SERCA activation. In resting cardiomyocytes, quercetin did not change amplitude or frequency of Ca2+ sparks. In isolated heart, quercetin increased heart rate and decreased PRi, QTc and duration of the QRS complex. Thus, we showed that quercetin activates ß-adrenoceptors, leading to increased L-type Ca2+ current and cell-wide intracellular Ca2+ transient without visible changes in Ca2+ sparks.

Geraniol blocks calcium and potassium channels in the mammalian myocardium: useful effects to treat arrhythmias.

Geraniol is a monoterpene present in several essential oils, and it is known to have a plethora of pharmacological activities. In this study, we explored the contractile and electrophysiological properties of geraniol and its antiarrhythmic effects in the heart. The geraniol effects on atrial contractility, L-type Ca(2+) current, K(+) currents, action potential (AP) parameters, ECG profile and on the arrhythmia induced by ouabain were evaluated. In the atrium, geraniol reduced the contractile force (~98%, EC = 1,510 ± 160 µM) and diminished the positive inotropism of CaCl2 and BAY K8644. In cardiomyocytes, the IC a,L was reduced by 50.7% (n = 5) after perfusion with 300 µM geraniol. Moreover, geraniol prolonged the AP duration (APD) measured at 50% (n = 5) after repolarization, without changing the resting potential. The increased APD could be attributed to the blockade of the transient outward K(+) current (Ito ) (59.7%, n = 4), the non-inactivation K(+) current (Iss ) (39.2%, n = 4) and the inward rectifier K(+) current (IK 1 ) (33.7%, n = 4). In isolated hearts, geraniol increased PRi and QTi without affecting the QRS complex (n = 6), and it reduced both the left ventricular pressure (83%) and heart rate (16.5%). Geraniol delayed the time to onset of ouabain-induced arrhythmias by 128%, preventing 30% of the increase in resting tension (n = 6). Geraniol exerts its negative inotropic and chronotropic responses in the heart by decreasing both L-type Ca(2+) and voltage-gated K(+) currents, ultimately acting against ouabain-induced arrhythmias.

Aqueous fraction from Costus spiralis (Jacq.) Roscoe leaf reduces contractility by impairing the calcium inward current in the mammalian myocardium.

ETHNOPHARMACOLOGICAL RELEVANCE: Brazilian folk medicine uses infusion of Costus spiralis leaf to help people to treat arterial hypertension and syndromes of cardiac hyperexcitability. AIM OF THE STUDY: Evaluate the aqueous fraction (AqF) effect on atrial contractility and investigate its mechanism of action. MATERIALS AND METHODS: The AqF effect on the cardiac contractility was studied on isolated electrically driven guinea pig left atria. Atropine and tetraethylammonium (TEA) were employed to investigate whether potassium contributes for the inotropic mechanism of the AqF. The role of calcium in this effect was also studied. This was done by analysing the AqF effect on the Bowditch's phenomenon, as well as by studying whether it could interfere with the concentration-effect curve for CaCl(2), isoproterenol, and BAY K8644. Mice isolated cardiomyocytes were submitted to a whole-cell patch-clamp technique in order to evaluate whether the L-type calcium current participates on the AqF effect. Furthermore, the intracellular calcium transient was studied by confocal fluorescence microscopy. RESULTS: AqF depressed the atrial contractile force. It was the most potent fraction from C. spiralis leaf (EC(50)=305 ± 41 mg/l) (crude extract: EC(50)=712 ± 41; ethyl acetate: EC(50)=788 ± 121; chloroform: EC(50)=8,948 ± 1,346 mg/l). Sodium and potassium content in the AqF was 0.15 mM and 1.91 mM, respectively. Phytochemical analysis revealed phenols, tannins, flavones, xanthones, flavonoids, flavonols, flavononols, flavonones, and saponins. Experiments with atropine and TEA showed that potassium does not participate of the inotropic mechanism of AqF. However, this fraction decreased the force overshoot characteristic of the Bowditch's phenomenon, and shifted the concentration-response curve for CaCl(2) (EC(50) from 1.12 ± 0.07 to 7.23 ± 0.47 mM) indicating that calcium currents participate on its mechanism of action. Results obtained with isoproterenol (1-1,000 pM) and BAY K8644 (5-2000nM) showed that AqF abolished the inotropic effect of these substances. On cardiomyocytes, 48mg/l AqF reduced (∼23%) the L-type calcium current density from -6.3 ± 0.3 to -4.9 ± 0.2 A/F (n=5 cells, p<0.05) and reduced the intracellular calcium transient (∼20%, 4.7 ± 1.2 a.u., n=42 cells to 3.7 ± 1.00 a.u., n=35 cells, p<0.05). However, the decay time of the fluorescence was not changed (control: 860 ± 32 ms, n=42 cells; AqF: 876 ± 26 ms, n=35 cells, p>0.05). CONCLUSIONS: The AqF of C. spiralis leaf depresses myocardial contractility by reducing the L-type calcium current and by decreasing the intracellular calcium transient. Despite the lack of data on the therapeutic dose of AqF used in folk medicine, our results support, at least in part, the traditional use of this plant to treat cardiac disorders.

R(+)-pulegone impairs Ca²+ homeostasis and causes negative inotropism in mammalian myocardium.

The present study aimed to investigate the inotropic effects of R(+)-pulegone, a monoterpene found in plant species belonging to the genus Mentha, on the mammalian heart. In electrically stimulated guinea pig atria, R(+)-pulegone reduced the contractile force (~83%) and decreased the contraction time measured at 50% of the maximum force amplitude (CT(50)) from 45.8 ± 6.2 ms to 36.9 ± 6.2 ms, suggesting that R(+)-pulegone may have an effect on Ca(2+) homeostasis. Nifedipine (40 µM), taken as a positive control, showed a very similar profile. To explore the hypothesis that R(+)-pulegone is somehow affecting Ca(2+) handling, we determined concentration-response curves for both CaCl(2) and BAY K8644. R(+)-pulegone shifted these curves rightward. Using isolated mouse ventricular cardiomyocytes, we measured whole-cell L-type Ca(2+) current and observed an I(Ca,L) peak reduction of 13.7 ± 2.5% and 40.2 ± 2.9% after a 3-min perfusion with 0.11 and 1.1mM of R(+)-pulegone, respectively. In addition, the intracellular Ca(2+) transient was decreased (72.9%) by 3.2mM R(+)-pulegone, with no significant changes in [Ca(2+)](i) transient decay kinetics. Moreover, R(+)-pulegone at 1.1mM prolonged the action potential duration at 10, 50, and 90% of repolarisation. The lengthening of the action potential duration may be attributed to the substantial blockade of the outward K(+) currents caused by 1.1mM of R(+)-pulegone (90.5% at 60 mV). These findings suggest that R(+)-pulegone exerts its negative inotropic effect on mammalian heart mainly by decreasing the L-type Ca(2+) current and the global intracellular Ca(2+) transient.