Curcumin and Resveratrol Improve Muscle Function and Structure through Attenuation of Proteolytic Markers in Experimental Cancer-Induced Cachexia.

Muscle wasting and cachexia are prominent comorbidities in cancer. Treatment with polyphenolic compounds may partly revert muscle wasting. We hypothesized that treatment with curcumin or resveratrol in cancer cachectic mice may improve muscle phenotype and total body weight through attenuation of several proteolytic and signaling mechanisms in limb muscles. In gastrocnemius and soleus muscles of cancer cachectic mice (LP07 adenocarcinoma cells, N = 10/group): (1) LC-induced cachexia, (2) LC-cachexia+curcumin, and (3) LC-cachexia + resveratrol, muscle structure and damage (including blood troponin I), sirtuin-1, proteolytic markers, and signaling pathways (NF-κB and FoxO3) were explored (immunohistochemistry and immunoblotting). Compared to nontreated cachectic mice, in LC-cachexia + curcumin and LC-cachexia + resveratrol groups, body and muscle weights (gastrocnemius), limb muscle strength, muscle damage, and myofiber cross-sectional area improved, and in both muscles, sirtuin-1 increased, while proteolysis (troponin I), proteolytic markers, and signaling pathways were attenuated. Curcumin and resveratrol elicited beneficial effects on fast- and slow-twitch limb muscle phenotypes in cachectic mice through sirtuin-1 activation, attenuation of atrophy signaling pathways, and proteolysis in cancer cachectic mice. These findings have future therapeutic implications as these natural compounds, separately or in combination, may be used in clinical settings of muscle mass loss and dysfunction including cancer cachexia.

Relationship between beta-adrenergic agonists, calpain system activity and beef texture: A systematic review.

The objective was to conduct a systematic review to evaluate the effects of dietary supplementation with beta-adrenergic agonists on calpains and calpastatin activity in bovine muscle and changes in meat tenderness. A survey was conducted in June 2019 on Science Direct, Web of Science, Scopus, PubMed and Capes Periodicals, using four keyword combinations: agonist and calpain and cattle; agonist and calpain and bovine; agonist and calpain and heifers; agonist and calpain and steers. Thirteen studies were selected, 54% concluded that supplementation with beta-adrenergic agonists increases calpastatin activity, 23% observed increase in their gene expression and 23% reported no effect on activity or expression of this enzyme. Nine studies evaluated the influence of beta-adrenergic agonists supplementation on meat texture and all found an increase in shear force values. There is strong evidence that beta-adrenergic agonists may increase calpastatin activity in the muscle, causing damage to meat tenderness.

Prolonged Immobilization Exacerbates the Loss of Muscle Mass and Function Induced by Cancer-Associated Cachexia through Enhanced Proteolysis in Mice.

We hypothesized that in mice with lung cancer (LC)-induced cachexia, periods of immobilization of the hindlimb (7 and 15 days) may further aggravate the process of muscle mass loss and function. Mice were divided into seven groups (n = 10/group): (1) non-immobilized control mice, (2) 7-day unloaded mice (7-day I), (3) 15-day unloaded mice (15-day I), (4) 21-day LC-cachexia group (LC 21-days), (5) 30-day LC-cachexia group (LC 30-days), (6) 21-day LC-cachexia group besides 7 days of unloading (LC 21-days + 7-day I), (7) 30-day LC-cachexia group besides 15 days of unloading (LC 30-days + 15-day I). Physiological parameters, body weight, muscle and tumor weights, phenotype and morphometry, muscle damage (including troponin I), proteolytic and autophagy markers, and muscle regeneration markers were identified in gastrocnemius muscle. In LC-induced cachexia mice exposed to hindlimb unloading, gastrocnemius weight, limb strength, fast-twitch myofiber cross-sectional area, and muscle regeneration markers significantly decreased, while tumor weight and area, muscle damage (troponin), and proteolytic and autophagy markers increased. In gastrocnemius of cancer-cachectic mice exposed to unloading, severe muscle atrophy and impaired function was observed along with increased muscle proteolysis and autophagy, muscle damage, and impaired muscle regeneration.

Reduced lung cancer burden by selective immunomodulators elicits improvements in muscle proteolysis and strength in cachectic mice.

Identification of to what extent tumor burden influences muscle mass independently of specific treatments for cancer-cachexia remains to be elucidated. We hypothesized that reduced tumor burden by selective treatment of tumor with immunomodulators may exert beneficial effects on muscle wasting and function in mice. Body and muscle weight, grip strength, physical activity, muscle morphometry, apoptotic nuclei, troponin-I systemic levels, interleukin-6, proteolytic markers, and tyrosine release, and apoptosis markers were determined in diaphragm and gastrocnemius muscles of lung cancer (LP07 adenocarcinoma cells) mice (BALB/c) treated with monoclonal antibodies (mAbs), against immune check-points and pathways (CD-137, cytotoxic T-lymphocyte associated protein-4, programed cell death-1, and CD-19; N = 10/group). Nontreated lung cancer cachectic mice were the controls. T and B cell numbers and macrophages were counted in tumors of both mouse groups. Compared to nontreated cachectic mice, in the mAbs-treated animals, T cells increased, no differences in B cells or macrophages, the variables final body weight, body weight and grip strength gains significantly improved. In diaphragm and gastrocnemius of mAbs-treated cachectic mice, number of apoptotic nuclei, tyrosine release, proteolysis, and apoptosis markers significantly decreased compared to nontreated cachectic mice. Systemic levels of troponin-I significantly decreased in treated cachectic mice compared to nontreated animals. We conclude that reduced tumor burden as a result of selective treatment of the lung cancer cells with immunomodulators elicits per se beneficial effects on muscle mass loss through attenuation of several biological mechanisms that lead to increased protein breakdown and apoptosis, which translated into significant improvements in limb muscle strength but not in physical activity parameters.

Diabetogenic effect of pravastatin is associated with insulin resistance and myotoxicity in hypercholesterolemic mice.

BACKGROUND: HMG-CoA reductase inhibitors (statins) are cholesterol-lowering drugs widely used to treat hypercholesterolemia and prevent cardiovascular disease. Statins are generally well tolerated, but adverse reactions may occur, particularly myopathy and new onset of diabetes. The exact mechanism of statin-induced myopathy and diabetes has not been fully elucidated. We have previously shown that treatment of hypercholesterolemic (LDLr-/-) mice with pravastatin for 2 months decreased pancreatic islet insulin secretion and increased oxidative stress and cell death, but no glucose intolerance was observed. The purpose of the current work was to study long-term pravastatin effects on glucose homeostasis, insulin sensitivity, muscle protein turnover and cell viability. METHODS: LDLr-/- mice were treated with pravastatin for 3, 6 and 10 months. Glucose tolerance, insulin resistance and glucose-stimulated insulin secretion were evaluated. The rates of protein synthesis and degradation were determined in gastrocnemius muscle after 10 months of treatment. Insulin signalling, oxidative stress and cell death were analysed in vitro using C2C12 myotubes. RESULTS: After 6 and 10 months of treatment, these mice became glucose intolerant, and after 10 months, they exhibited marked insulin resistance. Reduced islet glucose-stimulated insulin secretion was observed after the 3rd month of treatment. Mice treated for 10 months showed significantly decreased body weight and increased muscle protein degradation. In addition, muscle chymotrypsin-like proteasomal activity and lysosomal cathepsin were markedly elevated. C2C12 myotubes exposed to increasing concentrations of pravastatin presented dose-dependent impairment of insulin-induced Akt phosphorylation, increased apoptotic markers (Bax protein and cleaved caspase-3) and augmented superoxide anion production. CONCLUSIONS: In addition to reduced insulin secretion, long-term pravastatin treatment induces insulin resistance and muscle wasting. These results suggest that the diabetogenic effect of statins is linked to the appearance of myotoxicity induced by oxidative stress, impaired insulin signalling, proteolysis and apoptosis.

Formoterol treatment prevents the effects of endotoxin on muscle TNF/NF-kB, Akt/mTOR, and proteolytic pathways in a rat model. Role of IGF-I and miRNA 29b.

Inflammatory diseases are associated with muscle wasting as a result of an increase in proteolysis. The purpose of this study was to elucidate whether administration of a ß2 adrenergic agonist, formoterol, was able to prevent the acute effects of sepsis induced by liposaccharide (LPS) injection on rat gastrocnemius muscle and to evaluate the possible roles of corticosterone, IGF-I, miR-23a, and miR-29b. For this purpose, male Wistar rats were injected with LPS and/or formoterol. Formoterol treatment decreased LPS-induced increase in serum corticosterone, TNFα upregulation, and NF-κB(p65) and Forkhead box protein O1 activation in the gastrocnemius. Atrogin-1, muscle RING-finger protein-1, microtubule-associated protein-1 light chain 3b (LC3b), and the lipidation of LC3b-I to LC3b-II were increased by LPS, and formoterol blocked these effects. Serum IGF-I and its mRNA levels in the gastrocnemius were decreased, whereas mecano growth factor and IGF binding protein 3 mRNA levels were increased in the rats injected with LPS but not in the rats that received LPS and formoterol. Similarly, LPS decreased Akt and mammalian target of rapamycin phosphorylation, and formoterol blocked these decreases. Finally, miR-29b expression in the gastrocnemius was upregulated by endotoxin injection, whereas miR-23a was not significantly different. Formoterol treatment did not significantly modify LPS-induced increase in muscle miR-29b. Furthermore, in control rats formoterol increased the expression of this miRNA. We conclude that formoterol decreases endotoxin-induced inflammation and proteolysis in rat skeletal muscle. Those responses can be a direct effect of ß2 adrenergic receptor stimulation or/and of blocking the effects of LPS on corticosterone and IGF-I. Muscle miR-23a and -29b do not seem to play an important role in those responses.

Isoflavin-ß modifies muscle oxidative stress and prevents a thyrotoxicosis-induced loss of muscle mass in rats.

INTRODUCTION: We sought to verify whether isoflavin-beta (Iso-ß), a mixture of isoflavones with antioxidant properties, could prevent thyrotoxicosis-induced loss of muscle mass and the participation of oxidative stress (OS) in the mechanisms of this prevention. METHODS: Two experimental periods of thyrotoxicosis induction were used in Wistar rats: 3 and 5 days to assess Iso-ß effects before and after thyrotoxicosis-induced muscle wasting. After euthanasia, peritoneal fat and gastrocnemius muscle were collected, weighed, and muscle OS was assessed. RESULTS: Iso-ß prevented the loss of gastrocnemius mass in thyrotoxic rats through the prevention of muscle OS generation during thyrotoxicosis, increasing muscle total antioxidant capacity and decreasing mitochondrial cytochrome c oxidase activity, lipid peroxidation, and protein carbonyl content. CONCLUSION: Iso-ß decreased oxidative modification of proteins, which is known to exert a major role during proteolysis induction and is present in thyrotoxic myopathy, highlighting the potential action of Iso-ß in this complication of the disease. Muscle Nerve 56: 975-981, 2017.

Insulin treatment reverses the increase in atrogin-1 expression in atrophied skeletal muscles of diabetic rats with acute joint inflammation.

BACKGROUND: The aim of this study was to evaluate the changes in biomarkers of skeletal muscle proteolysis (atrogin-1, muscle RING finger-1 protein [MuRF-1]) and inflammation (nuclear factor kappa-B) in skeletal muscles of rats under two catabolic conditions, diabetes mellitus (DM) and acute joint inflammation, and the effects of insulin therapy. MATERIALS AND METHODS: Male Wistar rats were divided into groups without diabetes - normal (N), saline (NS), or ι-carrageenan (NCa) injection into the tibiotarsal joint - and groups with diabetes - diabetes (D), plus insulin (DI), saline (DS), or ι-carrageenan (DCa) injection into the tibiotarsal joint, or ι-carrageenan injection and treatment with insulin (DCaI). Three days after ι-carrageenan injection (17 days after diabetes induction), tibialis anterior (TA) and soleus (SO) skeletal muscles were used for analysis. RESULTS: DM alone caused a significant decrease in the mass of TA and SO muscles, even with low levels of atrogenes (atrogin-1, MuRF-1), which could be interpreted as an adaptive mechanism to spare muscle proteins under this catabolic condition. The loss of muscle mass was exacerbated when ι-carrageenan was administered in the joints of diabetic rats, in association with increased expression of atrogin-1, MuRF-1, and nuclear factor kappa-B. Treatment with insulin prevented the increase in atrogin-1 (TA, SO) and the loss of muscle mass (SO) in diabetic-carrageenan rats; in comparison with TA, SO muscle was more responsive to the anabolic actions of insulin. CONCLUSION: Acute joint inflammation overcame the adaptive mechanism in diabetic rats to prevent excessive loss of muscle mass, worsening the catabolic state. The treatment of diabetic-carrageenan rats with insulin prevented the loss of skeletal muscle mass mainly via atrogin-1 inhibition. Under the condition of DM and inflammation, muscles with the prevalence of slow-twitch, type 1 fibers were more responsive to insulin treatment, recovering the ability to grow.

Involvement of cAMP/EPAC/Akt signaling in the antiproteolytic effects of pentoxifylline on skeletal muscles of diabetic rats.

Advances in the knowledge of the mechanisms controlling protein breakdown in skeletal muscles have allowed the exploration of new options for treating muscle-wasting conditions. Pentoxifylline (PTX), a nonselective phosphodiesterase (PDE) inhibitor, attenuates the loss of muscle mass during catabolic conditions, mainly via inhibiting protein breakdown. The aim of this study was to explore the mechanisms by which PTX inhibits proteolysis in the soleus and extensor digitorum longus (EDL) muscles of streptozotocin-induced diabetic rats. The levels of atrogin-1 and muscle RING finger-1 were decreased, as were the activities of caspase-3 (EDL) and calpains (soleus and EDL), in diabetic rats treated with PTX, which at least partly explains the drop in the ubiquitin conjugate (EDL) levels and in proteasome activity (soleus and EDL). Treatment with PTX decreased PDE activity and increased cAMP content in muscles of diabetic rats; moreover, it also increased both the protein levels of exchange protein directly activated by cAMP (EPAC, a cAMP effector) and the phosphorylation of Akt. The loss of muscle mass was practically prevented in diabetic rats treated with PTX. These findings advance our understanding of the mechanisms underlying the antiproteolytic effects of PTX and suggest the use of PDE inhibitors as a strategy to activate cAMP signaling, which is emerging as a promising target for treating muscle mass loss during atrophic conditions. NEW & NOTEWORTHY cAMP signaling has been explored as a strategy to attenuate skeletal muscle atrophies. Therefore, in addition to ß2AR agonists, phosphodiesterase inhibitors such as pentoxifylline (PTX) can be an interesting option. This study advances the understanding of the mechanisms related to the antiproteolytic effects of PTX on skeletal muscles of diabetic rats, which involve the activation of both exchange protein directly activated by cAMP and Akt effectors, inhibiting the expression of atrogenes and calpain/caspase-3-proteolytic machinery.

Resistance exercise attenuates skeletal muscle oxidative stress, systemic pro-inflammatory state, and cachexia in Walker-256 tumor-bearing rats.

The aim of this study was to investigate the effects of resistance exercise training (RET) on oxidative stress, systemic inflammatory markers, and muscle wasting in Walker-256 tumor-bearing rats. Male (Wistar) rats were divided into 4 groups: sedentary controls (n = 9), tumor-bearing (n = 9), exercised (n = 9), and tumor-bearing exercised (n = 10). Exercised and tumor-bearing exercised rats were exposed to resistance exercise of climbing a ladder apparatus with weights tied to their tails for 6 weeks. The physical activity of control and tumor-bearing rats was confined to the space of the cage. After this period, tumor-bearing and tumor-bearing exercised animals were inoculated subcutaneously with Walker-256 tumor cells (11.0 × 107 cells in 0.5 mL of phosphate-buffered saline) while control and exercised rats were injected with vehicle. Following inoculation, rats maintained resistance exercise training (exercised and tumor-bearing exercised) or sedentary behavior (control and tumor-bearing) for 12 more days, after which they were euthanized. Results showed muscle wasting in the tumor-bearing group, with body weight loss, increased systemic leukocytes, and inflammatory interleukins as well as muscular oxidative stress and reduced mTOR signaling. In contrast, RET in the tumor-bearing exercised group was able to mitigate the reduced body weight and muscle wasting with the attenuation of muscle oxidative stress and systemic inflammatory markers. RET also prevented loss of muscle strength associated with tumor development. RET, however, did not prevent the muscle proteolysis signaling via FBXO32 gene messenger RNA expression in the tumor-bearing group. In conclusion, RET performed prior tumor implantation prevents cachexia development by attenuating tumor-induced systemic pro-inflammatory condition with muscle oxidative stress and muscle damage.

Predictors of surgery-induced muscle proteolysis in patients undergoing cardiac surgery.

BACKGROUND: Muscle proteolysis due to post-operative hypercatabolism is responsible for the functional decline observed in patients undergoing cardiac surgery. The aim of this study was to explore the factors underlying increased muscle proteolysis by measuring the urinary 3-methylhistidine/creatinine ratio (3-MH/Cr) in patients who had recently undergone cardiac surgery. METHODS AND RESULTS: Sixty-nine patients undergoing elective cardiac surgery participated in this study. The 24-h urinary 3-MH/Cr was collected for 3 days after surgery. Serum levels of metabolic markers, amino acids, and skeletal muscle strength were measured before and after surgery. Cumulative 3-MH/Cr during 3 days after surgery (cum3-MH/Cr) was 676.7±169.0nmol/gCr, and was positively associated with the decrease in muscle strength. In multivariate analysis, factors associated with an increased cum3-MH/Cr were preoperative grip strength (ß=-0.309, p=0.003), body mass index (ß=-0.299, p=0.001), hemoglobin (ß=-0.243, p=0.007), cardiopulmonary bypass time (ß=0.184, p=0.049), and immediate post-operative interleukin-6 (ß=0.295, p=0.002). CONCLUSIONS: Our findings suggest that post-operative muscle proteolysis is facilitated by preoperative catabolic accelerators in patients undergoing cardiac surgery. The factors of muscle proteolysis immediately after surgery may be a novel therapeutic target in rehabilitation intervention.

Effects of concentric and eccentric muscle actions on serum myostatin and follistatin-like related gene levels.

The present study determined the effects of concentric and eccentric muscle actions on the contents of serum myostatin and follistatin-like related gene (FLRG). Eight untrained males performed one exercise bout with each leg, separated by three weeks. One bout consisted of 7 sets of 10 repetitions of eccentric muscle actions of the knee extensors at 150% of the concentric 1-RM while the other bout consisted of 7 sets of 10 repetitions of concentric muscle actions at 75% 1-RM. The legs used and the bouts performed were randomized. Five days prior to each exercise bout, baseline measurements were taken for muscle strength. For both bouts, a venous blood sample was obtained immediately prior to exercise and again at 6, 24, and 48 hr post-exercise. Data were analyzed with 2 X 4 (bout x test) ANOVA (p < 0.05). Increases in serum myostatin and FLRG occurred with each exercise bout and, excluding 48 hr post-exercise, were significantly correlated to one another (p < 0.05). After eccentric exercise, peak increases of 68% and 50% (p < 0.05) were observed for myostatin and FLRG, respectively. Similar increases of 54% and 44% (p < 0.05) were observed after concentric muscle actions. There was no significant difference in expression of myostatin or FLRG as a function of muscle action type. Our results suggest that a single bout of exercise with either eccentric or concentric muscle actions appear to elicit a similar increase in serum myostatin and FLRG. Therefore, the type of muscle action may not be as much a mitigating factor for increasing serum myostatin and FLRG rather than the muscle action per se. Key PointsEccentric muscle actions do not preferentially increase serum myostatin.Increases in serum myostatin in response to eccentric muscle actions are associated with increase in serum FLRG.Increases in serum myostatin and FLRG in response to eccentric muscle actions are not correlated to serum cortisol.