Changes in cardiac heparan sulfate proteoglycan expression and streptozotocin-induced diastolic dysfunction in rats

Background: Changes in the proteoglycans glypican and syndecan-4 have been reported in several pathological conditions, but little is known about their expression in the heart during diabetes. The aim of this study was to investigate in vivo heart function changes and alterations in mRNA expression and protein levels of glypican-1 and syndecan-4 in cardiac and skeletal muscles during streptozotocin (STZ)-induced diabetes. Methods: Diabetes was induced in male Wistar rats by STZ administration. The rats were assigned to one of the following groups: control (sham injection), after 24 hours, 10 days, or 30 days of STZ administration. Echocardiography was performed in the control and STZ 10-day groups. Western and Northern blots were used to quantify protein and mRNA levels in all groups. Immunohistochemistry was performed in the control and 30-day groups to correlate the observed mRNA changes to the protein expression. Results: In vivo cardiac functional analysis performed using echocardiography in the 10-day group showed diastolic dysfunction with alterations in the peak velocity of early (E) diastolic filling and isovolumic relaxation time (IVRT) indices. These functional alterations observed in the STZ 10-day group correlated with the concomitant increase in syndecan-4 and glypican-1 protein expression. Cardiac glypican-1 mRNA and skeletal syndecan-4 mRNA and protein levels increased in the STZ 30-day group. On the other hand, the amount of glypican in skeletal muscle was lower than that in the control group. The same results were obtained from immunohistochemistry analysis. Conclusion: Our data suggest that membrane proteoglycans participate in the sequence of events triggered by diabetes and inflicted on cardiac and skeletal muscles.

Imatinib mesylate ameliorates the dystrophic phenotype in exercised mdx mice

Myofiber degeneration, inflammation, and fibrosis are remarkable features of Duchenne muscular dystrophy. We hypothesized that the administration of imatinib mesylate, an inhibitor of tyrosine kinase and TGF-beta pro-fibrogenic activity, could improve the muscular conditions in mdx mice. Four-week old mdx mice were treated and exercised for 6 weeks. Gastrocnemius and diaphragm histopathology, strength, creatine kinase, and cytokine levels were evaluated. The treated group presented increased muscular strength and decreased CK levels, injured myofibers, and inflammatory infiltrates. Pro-inflammatory cytokines and TGF-beta were also reduced, while IL-10 was increased, suggesting an immunomodulatory effect of imatinib, which can ameliorate the dystrophic phenotype in mdx mice. (C) 2009 Elsevier B.V. All rights reserved.

Chronic absence of baroreceptor inputs prevents training-induced cardiovascular adjustments in normotensive and spontaneously hypertensive rats

We investigate whether arterial baroreceptors mediate the training-induced blood pressure fall and resting bradycardia in hypertensive (SHR) and normotensive rats (WKY). Male SHR and WKY rats, submitted to sino-aortic denervation (SAD) or sham surgery (SHAM group), were allocated to training (T; 55% of maximal exercise capacity) or sedentary (S) protocols for 3 months. Rats were instrumented with arterial and venous catheters for haemodynamic measurements at rest (power spectral analysis) and baroreceptor testing. Kidney and skeletal muscles were processed for morphometric analysis of arterioles. Elevated mean arterial pressure (MAP) and heart rate (HR) in SHAM SHRS were accompanied by increased sympathetic variability and arteriolar wall/lumen ratio [+3.4-fold on low-frequency (LF) power and +70%, respectively, versus WKYS, P < 0.05]. Training caused significant HR (similar to 9% in WKY and SHR) and MAP reductions (-8% in the SHR), simultaneously with improvement of baroreceptor reflex control of HR (SHR and WKY), LF reduction (with a positive correlation between LF power and MAP levels in the SHR) and normalization of wall/lumen ratio of the skeletal muscle arterioles (SHR only). In contrast, SAD increased pressure variability in both strains of rats, causing reductions in MAP (-13%) and arteriolar wall/lumen ratio (-35%) only in the SHRS. Training effects were completely blocked by SAD in both strains; in addition, after SAD the resting MAP and HR and the wall/lumen ratio of skeletal muscle arterioles were higher in SHRT versus SHRS and similar to those of SHAM SHRS. The lack of training-induced effects in the chronic absence of baroreceptor inputs strongly suggests that baroreceptor signalling plays a decisive role in driving beneficial training-induced cardiovascular adjustments.

Chemical sympathectomy further increases muscle protein degradation of acutely diabetic rats

The present work investigated the role of the sympathetic nervous system (SINS) in the control of protein degradation in skeletal muscles from rats with streptozotocin (STZ)-induced diabetes. Diabetes (1, 3, and 5 days after STZ) induced a significant increase in the norepinephrine content of soleus and EDL muscles, but it did not affect plasma catecholamine levels. Chemical sympathectomy induced by guanethidine (100 mg/kg body weight, for 1 or 2 days) reduced muscle norepinephrine content to negligible levels (less than 5%), decreased plasma epinephrine concentration, and further increased the high rate of protein degradation in muscles from acutely diabetic rats. The rise in the rate of proteolysis (nmol.mg wet wt(-1).2h(-1)) in soleus from 1-day diabetic sympathectomized rats was associated with increased activities of lysosomal (0.127 +/- 0.008 vs. 0.086 +/- 0.013 in diabetic control) and ubiquitin (Ub)-proteasome-dependent proteolytic pathways (0.154 +/- 0,007 vs. 0.121 +/- 0.006 in diabetic control). Increases in Ca2+-depenclent (0.180 +/- 0.007 vs. 0.121 +/- 0.011 in diabetic control) and Ub-proteasome-dependent proteolytic systems (0.092 +/- 0.003 vs. 0.060 +/- 0.002 in diabetic control) were observed in EDL from 1-day diabetic sympathectomized rats. The lower phosphorylation levels of AKT and Foxo3a in EDL muscles from 3-day diabetic rats were further decreased by sympathectomy. The data suggest that the SNS exerts acute inhibitory control of skeletal muscle proteolysis during the early stages of diabetes in rats, probably involving the AKT/Foxo signaling pathway.

Is gender crucial for cardiovascular adjustments induced by exercise training in female spontaneously hypertensive rats?

Evidence of mild hypertension in women and female rats and our preliminary observation showing that training is not effective to reduce pressure in female as it does in male spontaneously hypertensive rats (SHR) prompt us to investigate the effects of gender on hemodynamic pattern and microcirculatory changes induced by exercise training. Female SHR and normotensive controls (Wistar- Kyoto rats) were submitted to training (55% VO2 peak; 3 months) or kept sedentary and instrumented for pressure and hindlimb flow measurements at rest and during exercise. Heart, kidney, and skeletal muscles (locomotor/ nonlocomotor) were processed for morphometric analysis of arterioles, capillaries, and venules. High pressure in female SHR was accompanied by an increased arteriolar wall: lumen ratio in the kidney (+30%; P < 0.01) but an unchanged ratio in the skeletal muscles and myocardium. Female SHR submitted to training did not exhibit further changes on the arteriolar wall: lumen ratio and pressure, showing additionally increased hindlimb resistance at rest (+29%; P < 0.05). On the other hand, female SHR submitted to training exhibited increased capillary and venular densities in locomotor muscles (+50% and 2.3- fold versus sedentary SHR, respectively) and normalized hindlimb flow during exercise hyperemia. Left ventricle pressure and weight were higher in SHR versus WKY rats, but heart performance (positive dP/dt(max) and negative dP/dt(max)) was not changed by hypertension or training, suggesting a compensated heart function in female SHR. In conclusion, the absence of training- induced structural changes on skeletal muscle and myocardium arterioles differed from changes observed previously in male SHR, suggesting a gender effect. This effect might contribute to the lack of pressure fall in trained female SHRs.

Tyrosine hydroxylase immunoreactivity as indicator of sympathetic activity: simultaneous evaluation in different tissues of hypertensive rats

Burgi K, Cavalleri MT, Alves AS, Britto LRG, Antunes VR, Michelini LC. Tyrosine hydroxylase immunoreactivity as indicator of sympathetic activity: simultaneous evaluation in different tissues of hypertensive rats. Am J Physiol Regul Integr Comp Physiol 300: R264-R271, 2011. First published December 9, 2010; doi: 10.1152/ajpregu.00687.2009.-Vasomotor control by the sympathetic nervous system presents substantial heterogeneity within different tissues, providing appropriate homeostatic responses to maintain basal/stimulated cardiovascular function both at normal and pathological conditions. The availability of a reproducible technique for simultaneous measurement of sympathetic drive to different tissues is of great interest to uncover regional patterns of sympathetic nerve activity (SNA). We propose the association of tyrosine hydroxylase immunoreactivity (THir) with image analysis to quantify norepinephrine (NE) content within nerve terminals in arteries/arterioles as a good index for regional sympathetic outflow. THir was measured in fixed arterioles of kidney, heart, and skeletal muscle of WistarKyoto rats (WKY) and spontaneously hypertensive rats (SHR) (123 +/- 2 and 181 +/- 4 mmHg, 300 +/- 8 and 352 +/- 8 beats/min, respectively). There was a differential THir distribution in both groups: higher THir was observed in the kidney and skeletal muscle (similar to 3-4-fold vs. heart arterioles) of WKY; in SHR, THir was increased in the kidney and heart (2.4- and 5.3-fold vs. WKY, respectively) with no change in the skeletal muscle arterioles. Observed THir changes were confirmed by either: 1) determination of NE content (high-performance liquid chromatography) in fresh tissues (SHR vs. WKY): +34% and +17% in kidney and heart, respectively, with no change in the skeletal muscle; 2) direct recording of renal (RSNA) and lumbar SNA (LSNA) in anesthetized rats, showing increased RSNA but unchanged LSNA in SHR vs. WKY. THir in skeletal muscle arterioles, NE content in femoral artery, and LSNA were simultaneously reduced by exercise training in the WKY group. Results indicate that THir is a valuable technique to simultaneously evaluate regional patterns of sympathetic activity.

Complexation of the anti-Trypanosoma cruzi drug benznidazole improves solubility and efficacy

The ruthenium complex,trans-[Ru(Bz)(NH3)(4)SO2](CF3SO3)(2) 1, Bz = benznidazole (N-benzyl-2-(2-nitro-1H-imidazol-1-yl)acetamide), is more hydrosoluble and more active (IC50try/1 h = 79 +/- 3 mu M) than free benznidazole 2 (IC50try/1 h > 1 mM). 1 also exhibits low acute toxicity in vitro (IC50macrophages > 1 mM) and in vivo (400 mu mol/kg < LD50 < 600 mu mol/kg) and the formation of hydroxylamine is more favorable in 1 than in 2 by 9.6 kcal/mol. In murine acute models of Chagas` disease, 1 was more active than 2 even when only one dose was administrated. Moreover, 1 at a thousand-fold smaller concentration than the considered optimal dose for 2 (385 mu mol/kg/day = 100 mg/kg/day), proved to be sufficient to protect all infected mice, eliminating the amastigotes in their hearts and skeletal muscles as observed in H&E micrographics.

Electromyographic activity of masticatory muscles in women with osteoporosis

The purpose of this study was to analyze the electromyographic (EMG) activity and the maximal molar bite force in women diagnosed with osteoporosis in the maxillary and mandibular regions, considering the habits and conditions that lead to development of generalized skeletal bone loss, including on face bones, can disturb the functional harmony of the stomatognathic system. Twenty-seven women with mandibular and maxillary osteoporosis and 27 healthy controls volunteered to participate in the study. A 5-channel electromyographer was used. Muscle activity was evaluated by means of EMG recordings of the masticatory musculature (masseter and temporalis muscles, bilaterally) during the following clinical conditions: rest (5 s); right and left lateral excursions (5 s); protrusion (5 s); maximal dental clenching on Parafilm™ (4 s) and maximal voluntary contraction (4 s). This latter clinical condition was used as the normalization factor of the sample data. It was observed that individuals with osteoporosis presented greater EMG activity when maintaining mandible posture conditions and less activity during dental clenching and when obtaining maximal molar bite force. It may be concluded that facial osteoporosis can interfere on the patterns of masticatory muscle activation and maximal bite force of the stomatognathic system.

Vibration-induced extra torque during electrically-evoked contractions of the human calf muscles

Background: High-frequency trains of electrical stimulation applied over the lower limb muscles can generate forces higher than would be expected from a peripheral mechanism (i.e. by direct activation of motor axons). This phenomenon is presumably originated within the central nervous system by synaptic input from Ia afferents to motoneurons and is consistent with the development of plateau potentials. The first objective of this work was to investigate if vibration (sinusoidal or random) applied to the Achilles tendon is also able to generate large magnitude extra torques in the triceps surae muscle group. The second objective was to verify if the extra torques that were found were accompanied by increases in motoneuron excitability. Methods: Subjects (n = 6) were seated on a chair and the right foot was strapped to a pedal attached to a torque meter. The isometric ankle torque was measured in response to different patterns of coupled electrical (20-Hz, rectangular 1-ms pulses) and mechanical stimuli (either 100-Hz sinusoid or gaussian white noise) applied to the triceps surae muscle group. In an additional investigation, M(max) and F-waves were elicited at different times before or after the vibratory stimulation. Results: The vibratory bursts could generate substantial self-sustained extra torques, either with or without the background 20-Hz electrical stimulation applied simultaneously with the vibration. The extra torque generation was accompanied by increased motoneuron excitability, since an increase in the peak-to-peak amplitude of soleus F waves was observed. The delivery of electrical stimulation following the vibration was essential to keep the maintained extra torques and increased F-waves. Conclusions: These results show that vibratory stimuli applied with a background electrical stimulation generate considerable force levels (up to about 50% MVC) due to the spinal recruitment of motoneurons. The association of vibration and electrical stimulation could be beneficial for many therapeutic interventions and vibration-based exercise programs. The command for the vibration-induced extra torques presumably activates spinal motoneurons following the size principle, which is a desirable feature for stimulation paradigms.

Influence of Laser Photobiomodulation on Collagen IV During Skeletal Muscle Tissue Remodeling After Injury in Rats

Objective: The aim of the present study was to determine the effect of GaAlAs low-level laser therapy (LLLT) on collagen IV remodeling of the tibialis anterior (TA) muscle in rats after cryolesion. Background: Considerable interest exists in skeletal muscle regeneration in situations such as repair after exercise-induced muscle injury, after muscle transplantation, in muscular dystrophy, exercise-induced muscle injury, and the recovery of strength after atrophy due to disuse. A number of studies have demonstrated the potential of LLLT in facilitating the muscle-healing process; however, no consensus is found in the literature regarding the best laser-irradiation parameters. Methods: Adult male Wistar rats (n = 45) were used and randomly divided into three groups: control (n = 5); nontreated cryolesioned group (n = 20), and LLLT-cryolesioned group (n = 20). The cryolesioned groups were analyzed at 1, 7, 14, and 21 days after the injury procedure. Laser irradiation was performed 3 times per week on the injured region by using the GaAlAs laser (660 nm; beam spot of 0.04 cm(2), output power of 20 mW, power density of 500 mW/cm(2), and energy density of 5 J/cm(2), for 10 sec). The muscles were removed, frozen, cryosectioned, and then stained with hematoxylin-eosin for the visualization of general morphology or used for immunohistochemical analysis of collagen IV. Results: It was demonstrated that LLLT promotes an increase in collagen IV immunolabeling in skeletal muscle in the first 7 days after acute trauma caused by cryoinjury, but does not modify the duration of the tissue-repair process. Even with LLLT, the injured muscle tissue needs similar to 21 days to achieve the same state of organization as that in the noninjured muscle. Conclusion: The collagen IV content is modulated in regenerating skeletal muscle under LLLT, which might be associated with better tissue outcome, although the histologic analysis did not detect tissue improvement in the LLLT group.

beta-Hydroxy-beta-methylbutyrate (HM beta) supplementation stimulates skeletal muscle hypertrophy in rats via the mTOR pathway

beta-Hydroxy-beta-methylbutyrate (HM beta) supplementation is used to treat cancer, sepsis and exercise-induced muscle damage. However, its effects on animal and human health and the consequences of this treatment in other tissues (e. g., fat and liver) have not been examined. The purpose of this study was to evaluate the effects of HM beta supplementation on skeletal muscle hypertrophy and the expression of proteins involved in insulin signalling. Rats were treated with HM beta (320 mg/kg body weight) or saline for one month. The skeletal muscle hypertrophy and insulin signalling were evaluated by western blotting, and hormonal concentrations were evaluated using ELISAs. HM beta supplementation induced muscle hypertrophy in the extensor digitorum longus (EDL) and soleus muscles and increased serum insulin levels, the expression of the mammalian target of rapamycin (mTOR) and phosphorylation of p70S6K in the EDL muscle. Expression of the insulin receptor was increased only in liver. Thus, our results suggest that HM beta supplementation can be used to increase muscle mass without adverse health effects.

Human multipotent adipose-derived stem cells restore dystrophin expression of Duchenne skeletal-muscle cells in vitro

Background information. DMD (Duchenne muscular dystrophy) is a devastating X-linked disorder characterized by progressive muscle degeneration and weakness. The use of cell therapy for the repair of defective muscle is being pursued as a possible treatment for DMD. Mesenchymal stem cells have the potential to differentiate and display a myogenic phenotype in vitro. Since liposuctioned human fat is available in large quantities, it may be an ideal source of stem cells for therapeutic applications. ASCs (adipose-derived stem cells) are able to restore dystrophin expression in the muscles of mdx (X-linked muscular dystrophy) mice. However, the outcome when these cells interact with human dystrophic muscle is still unknown. Results. We show here that ASCs participate in myotube formation when cultured together with differentiating human DMD myoblasts, resulting in the restoration of dystrophin expression. Similarly, dystrophin was induced when ASCs were co-cultivated with DMD myotubes. Experiments with GFP (green fluorescent protein)-positive ASCs and DAPI (4,6-diamidino-2-phenylindole)-stained DMD myoblasts indicated that ASCs participate in human myogenesis through cellular fusion. Conclusions. These results show that ASCs have the potential to interact with dystrophic muscle cells, restoring dystrophin expression of DMD cells in vitro. The possibility of using adipose tissue as a source of stem cell therapies for muscular diseases is extremely exciting.

Experimental chronic low-frequency resistance training produces skeletal muscle hypertrophy in the absence of muscle damage and metabolic stress markers

Volitional animal resistance training constitutes an important approach to modeling human resistance training. However, the lack of standardization protocol poses a frequent impediment to the production of skeletal muscle hypertrophy and the study of related physiological variables (i.e., cellular damage/inflammation or metabolic stress). Therefore, the purposes of the present study were: (1) to test whether a long-term and low frequency experimental resistance training program is capable of producing absolute increases in muscle mass; (2) to examine whether cellular damage/inflammation or metabolic stress is involved in the process of hypertrophy. In order to test this hypothesis, animals were assigned to a sedentary control (C, n = 8) or a resistance trained group (RT, n = 7). Trained rats performed 2 exercise sessions per week (16 repetitions per day) during 12 weeks. Our results demonstrated that the resistance training strategy employed was capable of producing absolute mass gain in both soleus and plantaris muscles (12%, p<0.05). Furthermore, muscle tumor necrosis factor (TNF-alpha) protein expression (soleus muscle) was reduced by 24% (p<0.01) in trained group when compared to sedentary one. Finally, serum creatine kinase (CK) activity and serum lactate concentrations were not affected in either group. Such information may have practical applications if reproduced in situations where skeletal muscle hypertrophy is desired but high mechanical stimuli of skeletal muscle and inflammation are not. Copyright (C) 2010 John Wiley & Sons, Ltd.

Exercise training changes IL-10/TNF-alpha ratio in the skeletal muscle of post-MI rats

Heart failure (HF) is associated with changes in the skeletal muscle (SM) which might be a consequence of the unbalanced local expression of pro- (TNF-alpha) and anti- (IL-10) inflammatory cytokines, leading to inflammation-induced myopathy, and SM wasting. This local effect of HF on SM may, on the other hand, contribute to systemic inflammation, as this tissue actively secretes cytokines. Since increasing evidence points out to an anti-inflammatory effect of exercise training, the goal of the present study was to investigate its effect in rats with HF after post-myocardial infarction (MI), with special regard to the expression of TNF-alpha and IL-10 in the soleus and extensor digitorum longus (EDL), muscles with different fiber composition. Wistar rats underwent left thoracotomy with ligation of the left coronary artery, and were randomly assigned to either a sedentary (Sham-operated and MI sedentary) or trained (Sham-operated and MI trained) group. Animals in the trained groups ran on a treadmill (0% grade at 13-20 m/min) for 60 min/day, 5 days/week, for 8-10 weeks. The training protocol was able to reverse the changes induced by MI, decreasing TNF-alpha protein (26%, P < 0.05) and mRNA (58%, P < 0.05) levels in the soleus, when compared with the sedentary MI group. Training also increased soleus IL-10 expression (2.6-fold, P < 0.001) in post-MI HF rats. As a consequence, the IL-10/TNF-alpha ratio was increased. This ""anti-inflammatory effect"" was more pronounced in the soleus than in the EDL, suggesting a fiber composition dependent response. (C) 2009 Elsevier Ltd. All rights reserved.

Changes in postnatal skeletal muscle development induced by alternative immobilization model in female rat

The objective of this study was to adapt a model of hind limb immobilization to newly weaned female rats and to determine the morphology of shortened soleus and plantaris muscles. Female Wistar rats were divided into three groups: control zero (n = 3) and control and free (n = 8), animals aged 21 and 31 days, respectively, submitted to no intervention, and immobilized (n = 25), animals aged 21 days submitted to immobilization for 10 days and sacrificed at 31 days of age. The device used for immobilization had advantages such as easy connection, good fit, and low cost. The immobilized rats showed a reduction in muscle fiber area and in connective tissue. The adaptation of this immobilization model originally used for adult rats was an excellent alternative for newly weaned rats and was also efficient in inducing significant hind limb disuse.