The role of TGF-ß signaling in muscle atrophy, sarcopenia and cancer cachexia.

Skeletal muscle, comprising a significant proportion (40 to 50 percent) of total body weight in humans, plays a critical role in maintaining normal physiological conditions. Muscle atrophy occurs when the rate of protein degradation exceeds protein synthesis. Sarcopenia refers to age-related muscle atrophy, while cachexia represents a more complex form of muscle wasting associated with various diseases such as cancer, heart failure, and AIDS. Recent research has highlighted the involvement of signaling pathways, including IGF1-Akt-mTOR, MuRF1-MAFbx, and FOXO, in regulating the delicate balance between muscle protein synthesis and breakdown. Myostatin, a member of the TGF-ß superfamily, negatively regulates muscle growth and promotes muscle atrophy by activating Smad2 and Smad3. It also interacts with other signaling pathways in cachexia and sarcopenia. Inhibition of myostatin has emerged as a promising therapeutic approach for sarcopenia and cachexia. Additionally, other TGF-ß family members, such as TGF-ß1, activin A, and GDF11, have been implicated in the regulation of skeletal muscle mass. Furthermore, myostatin cooperates with these family members to impair muscle differentiation and contribute to muscle loss. This review provides an overview of the significance of myostatin and other TGF-ß signaling pathway members in muscular dystrophy, sarcopenia, and cachexia. It also discusses potential novel therapeutic strategies targeting myostatin and TGF-ß signaling for the treatment of muscle atrophy.

Myokines and Biomarkers of Frailty in Older Inpatients with Undernutrition: A Prospective Study.

BACKGROUND: Population aging might increase the prevalence of undernutrition in older people, which increases the risk of frailty. Numerous studies have indicated that myokines are released by skeletal myocytes in response to muscular contractions and might be associated with frailty. This study aimed to evaluate whether myokines are biomarkers of frailty in older inpatients with undernutrition. METHODS: The frailty biomarkers were extracted from the Gene Expression Omnibus and Genecards datasets. Relevant myokines and health-related variables were assessed in 55 inpatients aged ≥ 65 years from the Peking Union Medical College Hospital prospective longitudinal frailty study. Serum was prepared for enzyme-linked immunosorbent assay using the appropriate kits. Correlations between biomarkers and frailty status were calculated by Spearman's correlation analysis. Multiple linear regression was performed to investigate the association between factors and frailty scores. RESULTS: The prevalence of frailty was 13.21%. The bioinformatics analysis indicated that leptin, adenosine 5'-monophosphate-activated protein kinase (AMPK), irisin, decorin, and myostatin were potential biomarkers of frailty. The frailty group had significantly higher concentrations of leptin, AMPK, and MSTN than the robust group (p < 0.05). AMPK was significantly positively correlated with frailty (p < 0.05). The pre-frailty and frailty groups had significantly lower concentrations of irisin than the robust group (p < 0.05), whereas the DCN concentration did not differ among the groups. Multiple linear regression suggested that the 15 factors influencing the coefficients of association, the top 50% were the ADL score, MNA-SF score, serum albumin concentration, urination function, hearing function, leptin concentration, GDS-15 score, and MSTN concentration. CONCLUSIONS: Proinflammatory myokines, particularly leptin, myostatin, and AMPK, negatively affect muscle mass and strength in older adults. ADL and nutritional status play major roles in the development of frailty. Our results confirm that identification of frailty relies upon clinical variables, myokine concentrations, and functional parameters, which might enable the identification and monitoring of frailty.

NADPH oxidase 1 promotes hepatic steatosis in obese mice and is abrogated by augmented skeletal muscle mass.

Exercise as a lifestyle modification is a frontline therapy for nonalcoholic fatty liver disease (NAFLD), but how components of exercise attenuate steatosis is unclear. To uncouple the effect of increased muscle mass from weight loss in obesity, myostatin knockout mice were bred on a lean and obese db/db background. Myostatin deletion increases gastrocnemius (Gastrocn.) mass and reduces hepatic steatosis and hepatic sterol regulatory element binding protein 1 (Srebp1) expression in obese mice, with no impact on adiposity or body weight. Interestingly, hypermuscularity reduces hepatic NADPH oxidase 1 (Nox1) expression but not NADPH oxidase 4 (Nox4) in db/db mice. To evaluate a deterministic function of Nox1 on steatosis, Nox1 knockout mice were bred on a lean and db/db background. NOX1 deletion significantly attenuates hepatic oxidant stress, steatosis, and Srebp1 programming in obese mice to parallel hypermuscularity, with no improvement in adiposity, glucose control, or hypertriglyceridemia to suggest off-target effects. Directly assessing the role of NOX1 on SREBP1, insulin (Ins)-mediated SREBP1 expression was significantly increased in either NOX1, NADPH oxidase organizer 1 (NOXO1), and NADPH oxidase activator 1 (NOXA1) or NOX5-transfected HepG2 cells versus ?-galactosidase control virus, indicating superoxide is the key mechanistic agent for the actions of NOX1 on SREBP1. Metabolic Nox1 regulators were evaluated using physiological, genetic, and diet-induced animal models that modulated upstream glucose and insulin signaling, identifying hyperinsulinemia as the key metabolic derangement explaining Nox1-induced steatosis in obesity. GEO data revealed that hepatic NOX1 predicts steatosis in obese humans with biopsy-proven NAFLD. Taken together, these data suggest that hypermuscularity attenuates Srebp1 expression in db/db mice through a NOX1-dependent mechanism.NEW & NOTEWORTHY This study documents a novel mechanism by which changes in body composition, notably increased muscle mass, protect against fatty liver disease. This mechanism involves NADPH oxidase 1 (NOX1), an enzyme that increases superoxide and increases insulin signaling, leading to increased fat accumulation in the liver. NOX1 may represent a new early target for preventing fatty liver to stave off later liver diseases such as cirrhosis or liver cancer.

Myostatin and CXCL11 promote nervous tissue macrophages to maintain osteoarthritis pain.

Pain is the most debilitating symptom of knee osteoarthritis (OA) that can even persist after total knee replacement. The severity and duration of pain do not correlate well with joint tissue alterations, suggesting other mechanisms may drive pain persistence in OA. Previous work identified that macrophages accumulate in the dorsal root ganglia (DRG) containing the somas of sensory neurons innervating the injured knee joint in a mouse OA model and acquire a M1-like phenotype to maintain pain. Here we aimed to unravel the mechanisms that govern DRG macrophage accumulation and programming. The accumulation of F4/80+iNOS+ (M1-like) DRG macrophages was detectable at day 3 after mono-iodoacetate (MIA)-induced OA in the mouse. Depletion of macrophages prior to induction of OA resolved pain-like behaviors by day 7 without affecting the initial development of pain-like behaviors. Analysis of DRG transcript identified CXCL11 and myostatin. CXCL11 and myostatin were increased at 3 weeks post OA induction, with CXCL11 expression partially localized in satellite glial cells and myostatin in sensory neurons. Blocking CXCL11 or myostatin prevented the persistence of OA pain, without affecting the initiation of pain. CXCL11 neutralization reduced the number of total and F4/80+iNOS+ DRG macrophages, whilst myostatin inhibition diminished the programming of F4/80+iNOS+ DRG macrophages. Intrathecal injection of recombinant CXCL11 did not induce pain-associated behaviors. In contrast, intrathecal myostatin increased the number of F4/80+iNOS+ DRG macrophages concurrent with the development of mechanical hypersensitivity that was prevented by macrophages depletion or CXCL11 blockade. Finally, myostatin inhibition during established OA, resolved pain and F4/80+iNOS+ macrophage accumulation in the DRG. In conclusion, DRG macrophages maintain OA pain, but are not required for the induction of OA pain. Myostatin is a key ligand in neuro-immune communication that drives the persistence of pain in OA through nervous tissue macrophages and represent a novel therapeutic target for the treatment of OA pain.

Myostatin expression in lung cancer induces sarcopenia and promotes cancer progression.

OBJECTIVES: Many studies have demonstrated that sarcopenia among lung cancer predicts poor prognosis due to cancer progression. However, the cytokines that link sarcopenia and lung cancer progression remain unidentified. This study aimed to investigate whether lung cancer producing myostatin, which induces skeletal muscle atrophy, leads to sarcopenia and promotes cancer progression in patients with resected lung cancer. METHODS: Tumor tissues were obtained from 148 patients who underwent curative resection for lung cancer. Tumor cells were stained with myostatin and tumor-associated macrophages (TAM) in the tumor microenvironment were stained with CD68. We assessed the association between myostatin expression and the clinicopathological features. RESULTS: High myostatin expression in lung cancer was significantly associated with low skeletal muscle mass. The 5-year overall survival and relapse-free survival were significantly worse among patients with high myostatin expression than those with low expression. A multivariate analysis showed that TAM count was positively correlated with high myostatin expression. CONCLUSION: Sarcopenia may be induced by myostatin secreted by lung cancer cells. Moreover, myostatin may promote TAM migration into the tumor microenvironment, leading to advance lung cancer. As a result, patients with high myostatin expression had poor prognosis.

Effect of a resistance exercise at acute moderate altitude on muscle health biomarkers.

The intensification of the stress response during resistance training (RT) under hypoxia conditions could trigger unwanted effects that compromise muscle health and, therefore, the ability of the muscle to adapt to longer training periods. We examined the effect of acute moderate terrestrial hypoxia on metabolic, inflammation, antioxidant capacity and muscle atrophy biomarkers after a single RT session in a young male population. Twenty healthy volunteers allocated to the normoxia (N < 700 m asl) or moderate altitude (HH = 2320 m asl) group participated in this study. Before and throughout the 30 min following the RT session (3 × 10 reps, 90 s rest, 70% 1RM), venous blood samples were taken and analysed for circulating calcium, inorganic phosphate, cytokines (IL-6, IL-10 and TNF-α), total antioxidant capacity (TAC) and myostatin. Main results displayed a marked metabolic stress response after the RT in both conditions. A large to very large proportional increase in the adjusted to pre-exercise change of inflammatory and anti-inflammatory markers favoured HH (serum TNF-α [ES = 1.10; p = 0.024] and IL-10 [ES = 1.31; p = 0.009]). The exercise produced a similar moderate increment of myostatin in both groups, followed by a moderate non-significant reduction in HH throughout the recovery (ES = - 0.72; p = 0.21). The RT slightly increased the antioxidant response regardless of the environmental condition. These results revealed no clear impact of RT under acute hypoxia on the metabolic, TAC and muscle atrophy biomarkers. However, a coordinated pro/anti-inflammatory response balances the potentiated effect of RT on systemic inflammation.

Inflammation Is a Histological Characteristic of Skeletal Muscle in Chronic Limb Threatening Ischemia.

BACKGROUND: The loss of skeletal muscle is a prognostic factor in several diseases including in patients with chronic limb threatening ischemia (CLTI). Patients with CLTI also have a lower skeletal mass and area when compared to those with claudication. However, there are no currently available data regarding the histological characteristics of core muscles in patients with CLTI. This study aims to determine the differences in core skeletal muscles between patients with claudication and those with CLTI. The second aim is to evaluate the differences in myokines, which are molecules secreted by skeletal muscle, between patients with claudication and those with CLTI. METHODS: An observational, prospective study was conducted from January 2018 to July 2022 involving consecutive patients with peripheral arterial disease (PAD). The clinical characteristics were registered. In PAD patients with surgical indication for common femoral artery approach, samples of sartorius skeletal muscle (and not from the limb muscles directly involved in the ischemic process) were collected. The samples were submitted to histological characterization on hematoxylin-eosin and to immunohistochemical analysis to detect CD45+ leukocytes and CD163+ macrophages. The extent of the inflammatory cells (leukocytes and macrophages) was semiquantitatively assessed using a 0-to-4 grade scale as follows: absent (0†), mild (†), moderate (††), severe (†††), and very severe (††††). Serum levels of myokines: irisin, myostatin, IL-8, and lL-6 were determined with multiplex bead-based immunoassay. RESULTS: 119 patients (mean age: 67.58 ± 9.60 years old, 79.80% males) 64 with claudication and 54 with CLTI were enrolled in the study. No differences were registered between patients with claudication and those with CLTI on age, gender, cardiovascular risk factors, and medication, except on smoking habits. There was a significantly higher prevalence of smokers and a higher smoking load in the claudication group. Samples of sartorius skeletal muscle from 40 patients (14 with claudication and 26 with CLTI) were submitted to histological analysis. No differences were found in skeletal muscle fibers preservation, trauma, or hemorrhage (on hematoxylin-eosin staining). However, in the immunohistochemistry study, we found more inflammatory cells CD45+ leukocytes in patients with CLTI when compared to those with claudication [CD45+ ≥ moderate (††): claudication (n = 14): 4; 28.57%; CLTI (n = 25): 16; 64.00%; P = 0.034]. Patients with CLTI also had higher tissue levels of CD163+ macrophages, but this difference was not significant [CD163+ ≥ moderate (††): claudication (n = 13): 7; 53.85%; CLTI (n = 27): 21; 77.78%; P = 0.122]. The serum levels of the myokines, irisin, and myostatin were below the lower limit of detection, in the majority of patients, so no valid results were obtained. However, patients with CLTI had a higher serum level of Interleukin (IL)-6 and IL-8. CONCLUSIONS: CLTI patients exhibit increased quantities of leukocytes in their sartorius muscle, as well as elevated serum levels of myokines IL-8 and IL-6. Inflamed skeletal muscle can contribute to the loss of muscle mass and account for the lower density of skeletal muscle observed in CLTI. Additionally, inflamed skeletal muscle may contribute to the development of systemic inflammation through the secretion of pro-inflammatory cytokines into the systemic circulation. Halting the inflammatory process could eventually improve the prognosis of CLTI patients.

Azilsartan prevents muscle loss and fast- to slow-twitch muscle fiber shift in natural ageing sarcopenic rats.

Sarcopenia is a musculoskeletal disease that reduces muscle mass and strength in older individuals. The study investigates the effects of azilsartan (AZL) on skeletal muscle loss in natural sarcopenic rats. Male Sprague-Dawley rats aged 4-6 months and 18-21 months were selected as young-matched control and natural-aged (sarcopenic) rats, respectively. Rats were allocated into young and old control (YC and OC) and young and old AZL treatment (YT and OT) groups, which received vehicles and AZL (8 mg/kg, orally) for 6 weeks. Rats were then sacrificed after muscle function analysis. Serum and gastrocnemius (GN) muscles were isolated for further endpoints. AZL significantly improved muscle grip strength and antioxidant levels in sarcopenic rats. AZL also restored the levels of insulin, testosterone, and muscle biomarkers such as myostatin and creatinine kinase in sarcopenic rats. Furthermore, AZL treatment improved the cellular and ultrastructure of GN muscle and prevented the shift of type II (glycolytic) myofibers to type I (oxidative) myofibers. The results showed that AZL intervention restored protein synthesis in natural sarcopenic rats by increasing p-Akt-1 and decreasing muscle RING-finger protein-1 and tumor necrosis factor alpha immunoexpressions. In conclusion, the present findings showed that AZL could be an effective intervention in treating age-related muscle impairments.

Myostatin and the Heart.

Myostatin (growth differentiation factor 8) is a member of the transforming growth factor-ß superfamily. It is secreted mostly by skeletal muscles, although small amounts of myostatin are produced by the myocardium and the adipose tissue as well. Myostatin binds to activin IIB membrane receptors to activate the downstream intracellular canonical Smad2/Smad3 pathway, and additionally acts on non-Smad (non-canonical) pathways. Studies on transgenic animals have shown that overexpression of myostatin reduces the heart mass, whereas removal of myostatin has an opposite effect. In this review, we summarize the potential diagnostic and prognostic value of this protein in heart-related conditions. First, in myostatin-null mice the left ventricular internal diameters along with the diastolic and systolic volumes are larger than the respective values in wild-type mice. Myostatin is potentially secreted as part of a negative feedback loop that reduces the effects of the release of growth-promoting factors and energy reprogramming in response to hypertrophic stimuli. On the other hand, both human and animal data indicate that myostatin is involved in the development of the cardiac cachexia and heart fibrosis in the course of chronic heart failure. The understanding of the role of myostatin in such conditions might initiate a development of targeted therapies based on myostatin signaling inhibition.

Myocardial Fibrosis and Steatosis in Patients with Aortic Stenosis: Roles of Myostatin and Ceramides.

Aortic stenosis (AS) involves progressive valve obstruction and a remodeling response of the left ventriculum (LV) with systolic and diastolic dysfunction. The roles of interstitial fibrosis and myocardial steatosis in LV dysfunction in AS have not been completely characterized. We enrolled 31 patients (19 women and 12 men) with severe AS undergoing elective aortic valve replacement. The subjects were clinically evaluated, and transthoracic echocardiography was performed pre-surgery. LV septal biopsies were obtained to assess fibrosis and apoptosis and fat deposition in myocytes (perilipin 5 (PLIN5)), or in the form of adipocytes within the heart (perilipin 1 (PLIN1)), the presence of ceramides and myostatin were assessed via immunohistochemistry. After BMI adjustment, we found a positive association between fibrosis and apoptotic cardiomyocytes, as well as fibrosis and the area covered by PLIN5. Apoptosis and PLIN5 were also significantly interrelated. LV fibrosis increased with a higher medium gradient (MG) and peak gradient (PG). Ceramides and myostatin levels were higher in patients within the higher MG and PG tertiles. In the linear regression analysis, increased fibrosis correlated with increased apoptosis and myostatin, independent from confounding factors. After adjustment for age and BMI, we found a positive relationship between PLIN5 and E/A and a negative correlation between septal S', global longitudinal strain (GLS), and fibrosis. Myostatin was inversely correlated with GLS and ejection fraction. Fibrosis and myocardial steatosis altogether contribute to ventricular dysfunction in severe AS. The association of myostatin and fibrosis with systolic dysfunction, as well as between myocardial steatosis and diastolic dysfunction, highlights potential therapeutic targets.

[Adipomyokines in children with obesity].

BACKGROUND: Adipomyokines are synthesized and secreted into the bloodstream by cells of both muscle and adipose tissue. They can have both a negative metabolic effect, acting as pro-inflammatory adipokines in obesity, and a positive one, increasing in response to physical exertion in the form of myokines. AIM: To study the features of adipocytokine secretion in children with constitutionally exogenous obesity. MATERIALS AND METHODS: The study included 80 patients: 60 adolescents aged 15 [13; 16] years with constitutionally exogenous obesity SDS BMI: 3.0 [2.6; 3.3] and 20 control group children aged 16 [15; 17] years without excess weight SDS BMI: -0.3 [-1.25; 0.33]. Commercial enzyme immunoassay kits were used to determine the level of adipomyokines. The compositional composition of the body was evaluated by bioimpedance analysis (InBody 770 analyzer, South Korea) in the morning, on an empty stomach. Statistical processing was carried out using STATISTICA v.12.0 (StatSoft Inc., USA). The results are presented in the form of median (Me) and quartiles (Q1; Q3) corresponding to 25 and 75 percentiles. The critical significance level (p) was assumed to be <0.05. RESULTS: Levels of IL-6 and irisin are statistically significantly higher in obese adolescents compared to the control group: 0.55 [0.226; 1.35] pg/ml vs 0.202 [0.128; 0.652] pg/ml (p=0.041) and 11.16 [6.6; 22.76] mcg/ml vs 7.36 [6.48; 9.68] mcg/ml (p=0.043), respectively. Concentrations of IL-6, myostatin and decorin increase with an increase in the degree of obesity: grade I vs III: 0.226 [0.224; 0.398] vs 0.80 [0.36; 1.81] pg/ml (p=0,0197), 25,85 [21,53; 28,23] vs 31.41 [24.36; 35.06] ng/ml (p=0.03), 4065.3 [3244.9; 5245.5] vs 5322.5 [4199.8; 7702.4] pg/ml (p=0.0376), respectively. In obese children, IL-6 levels positively correlate with BMI, SDS BMI and the amount of adipose tissue, and myostatin - with BMI and SDS BMI. The concentration of irisin in the blood serum is significantly higher in obese girls than in obese boys and healthy girls. Obese patients, compared with lean peers, are characterized by a statistically significantly higher content of both fat and lean mass. With the progression of obesity, there is a statistically significant increase in the ratio of fat to lean mass (I degree - 0.66 [0.56; 0.7], III - 0.78 [0.68; 0.98] (p=0.0073). CONCLUSION: Patients with obesity and normal body weight have different levels of adipomyokines. An increase in the level of IL-6 with the progression of obesity is directly related to an increase in the content of adipose tissue. Further study of the features of adipocytokine secretion, their relationship with the features of the body composition and metabolic complications in obesity is required.

Inhibitor Design Strategy for Myostatin: Dynamics and Interaction Networks Define the Affinity and Release Mechanisms of the Inhibited Complexes.

Myostatin, an important negative regulator of muscle mass, is a therapeutic target for muscle atrophic disorders such as muscular dystrophy. Thus, the inhibition of myostatin presents a strategy to treat these disorders. It has long been established that the myostatin prodomain is a strong inhibitor of the mature myostatin, and the minimum peptide of the prodomain-corresponding to the α1-helix of its lasso-region-responsible for the inhibitory efficiency was defined and characterized as well. Here we show that the minimum peptide segment based on the growth differentiation factor 11 (GDF11), which we found to be more helical in its stand-alone solvated stfate than the similar segment of myostatin, is a promising new base scaffold for inhibitor design. The proposed inhibitory peptides in their solvated state and in complex with the mature myostatin were analyzed by in silico molecule modeling supplemented with the electronic circular dichroism spectroscopy measurements. We defined the Gaussian-Mahalanobis mean score to measure the fraction of dihedral angle-pairs close to the desired helical region of the Ramachandran-plot, carried out RING analysis of the peptide-protein interaction networks and characterized the internal motions of the complexes using our rigid-body segmentation protocol. We identified a variant-11m2-that is sufficiently ordered both in solvent and within the inhibitory complex, forms a high number of contacts with the binding-pocket and induces such changes in its internal dynamics that lead to a rigidified, permanently locked conformation that traps this peptide in the binding site. We also showed that the naturally evolved α1-helix has been optimized to simultaneously fulfill two very different roles: to function as a strong binder as well as a good leaving group. It forms an outstanding number of non-covalent interactions with the mature core of myostatin and maintains the most ordered conformation within the complex, while it induces independent movement of the gate-keeper ß-hairpin segment assisting the dissociation and also results in the least-ordered solvated form which provides extra stability for the dissociated state and discourages rebinding.

Marcadores bioquímicos propuestos para el estudio de la sarcopenia / Proposed biochemical markers for the study of sarcopenia

La sarcopenia asociada a la edad es una condición clínica caracterizada por una disminución en la fuerza, calidad y cantidad de masa muscular así como también en la función muscular. Un biomarcador se define como una característica que es medible objetivamente y evaluable como indicador de un proceso biológico normal, patológico o respuesta terapéutica a una intervención farmacológica. Los marcadores bioquímicos propuestos para el estudio de la sarcopenia pueden ser categorizados en dos grupos. El primero de ellos evalúa el estatus musculoesquelético; este panel de marcadores está formado por miostatina/folistatina, procolágeno aminoterminal tipo III e índice de sarcopenia. El segundo grupo de marcadores bioquímicos evalúa factores causales, para lo cual se sugiere medir el factor de crecimiento insulino-símil tipo 1 (IGF-1), dehidroepiandrosterona (DHEAS), cortisol, facto-res inflamatorios [proteína C reactiva (PCR), interleuquina 6 (IL-6) y factor de necrosis tu-moral (TNF-a)]. Las recomendaciones realiza-das están basadas en la evidencia científica disponible en la actualidad y la disponibilidad de la metodología apropiada para cada uno de los biomarcadores. (AU)

Sarcopenia is a progressive and generalized skeletal muscle disorder defined by decrease in the strength, quality and quantity of muscle mass as well as in muscle function. A biomarker is defined as a feature objectively measured and evaluated as an indicator of a normal biologic process, a pathogenic process or a pharmacologic response to therapeutic intervention. The biochemical markers proposed for the study of sarcopenia may be classified in two groups. The first group evaluates the musculoskeletal status, made up by myostatin/follistatin, N-terminal Type III Procollagen and the sarcopenia index. The second evaluates causal factors, where the measurement of the following is suggested: hormones insulin-like growth factor-1 (IGF-I), dehydroepiandrosterone sulphate (DHEAS), cortisol, inflammatory factors [C-reactive protein (CRP), interleukin-6 (IL-6), and tumor necrosis factor-a (TNF-a)]. The recommendations made are based on scientific evidence currently available and the appropriate methodology availability for each biomarker. (AU)

Effects of Lenvatinib on Skeletal Muscle Volume and Cardiac Function in Patients with Hepatocellular Carcinoma.

INTRODUCTION: Previously, we reported that the tyrosine kinase inhibitor (TKI) sorafenib decreases serum levels of carnitine and reduces skeletal muscle volume. Moreover, others reported that TKIs might lead to cardiomyopathy or heart failure. Therefore, this study aimed to evaluate the effects of lenvatinib (LEN) on skeletal muscle volume and cardiac function in patients with hepatocellular carcinoma (HCC). METHODS: This retrospective study included 58 adult Japanese patients with chronic liver diseases and HCC treated with LEN. Blood samples were collected before and after 4 weeks of treatment, and serum carnitine fraction and myostatin levels were measured. Before and after 4-6 weeks of treatment, the skeletal muscle index (SMI) was evaluated from computed tomography images and cardiac function was assessed by ultrasound cardiography. RESULTS: After treatment, SMI, serum levels of total carnitine, and global longitudinal strain were significantly lower, but serum levels of myostatin were significantly higher. Left ventricular ejection fraction showed no significant change. CONCLUSION: In patients with HCC, LEN decreases serum levels of carnitine, skeletal muscle volume, and worsens cardiac function.

Red blood cell extracellular vesicles deliver therapeutic siRNAs to skeletal muscles for treatment of cancer cachexia.

Cancer cachexia is a multifactorial syndrome characterized by a significant loss of skeletal muscle, which negatively affects the quality of life. Inhibition of myostatin (Mstn), a negative regulator of skeletal muscle growth and differentiation, has been proven to preserve muscle mass in muscle atrophy diseases, including cachexia. However, myostatin inhibitors have repeatedly failed clinical trials because of modest therapeutic effects and side effects due to the poor efficiency and toxicity of existing delivery methods. Here, we describe a novel method for delivering Mstn siRNA to skeletal muscles using red blood cell-derived extracellular vesicles (RBCEVs) in a cancer cachectic mouse model. Our data show that RBCEVs are taken up by myofibers via intramuscular administration. Repeated intramuscular administrations with RBCEVs allowed the delivery of siRNAs, thereby inhibiting Mstn, increasing muscle growth, and preventing cachexia in cancer-bearing mice. We observed the same therapeutic effects when delivering siRNAs against malonyl-CoA decarboxylase, an enzyme driving dysfunctional fatty acid metabolism in skeletal muscles during cancer cachexia. We demonstrate that intramuscular siRNA delivery by RBCEVs is safe and non-inflammatory. Hence, this method is useful to reduce the therapeutic dose of siRNAs, to avoid toxicity and off-target effects caused by systemic administration of naked siRNAs at high doses.

Molecular mechanisms of cancer cachexia-related loss of skeletal muscle mass: data analysis from preclinical and clinical studies.

Cancer cachexia is a systemic hypoanabolic and catabolic syndrome that diminishes the quality of life of cancer patients, decreases the efficiency of therapeutic strategies and ultimately contributes to decrease their lifespan. The depletion of skeletal muscle compartment, which represents the primary site of protein loss during cancer cachexia, is of very poor prognostic in cancer patients. In this review, we provide an extensive and comparative analysis of the molecular mechanisms involved in the regulation of skeletal muscle mass in human cachectic cancer patients and in animal models of cancer cachexia. We summarize data from preclinical and clinical studies investigating how the protein turnover is regulated in cachectic skeletal muscle and question to what extent the transcriptional and translational capacities, as well as the proteolytic capacity (ubiquitin-proteasome system, autophagy-lysosome system and calpains) of skeletal muscle are involved in the cachectic syndrome in human and animals. We also wonder how regulatory mechanisms such as insulin/IGF1-AKT-mTOR pathway, endoplasmic reticulum stress and unfolded protein response, oxidative stress, inflammation (cytokines and downstream IL1ß/TNFα-NF-κB and IL6-JAK-STAT3 pathways), TGF-ß signalling pathways (myostatin/activin A-SMAD2/3 and BMP-SMAD1/5/8 pathways), as well as glucocorticoid signalling, modulate skeletal muscle proteostasis in cachectic cancer patients and animals. Finally, a brief description of the effects of various therapeutic strategies in preclinical models is also provided. Differences in the molecular and biochemical responses of skeletal muscle to cancer cachexia between human and animals (protein turnover rates, regulation of ubiquitin-proteasome system and myostatin/activin A-SMAD2/3 signalling pathways) are highlighted and discussed. Identifying the various and intertwined mechanisms that are deregulated during cancer cachexia and understanding why they are decontrolled will provide therapeutic targets for the treatment of skeletal muscle wasting in cancer patients.

Myostatin inhibitory peptides in sports drug testing.

Across species, skeletal muscle mass is negatively regulated by the TGF-ß cytokine myostatin (MSTN). Inhibitors of this growth factor and its signaling pathways are therefore not only promising therapeutics for muscular diseases but also potential performance-enhancing agents in sports. Within this study, protein precipitation and liquid chromatography high-resolution tandem mass spectrometry (LC-HRMS/MS) were employed to develop a detection method for six novel MSTN inhibitory peptides derived from the regulatory MSTN propeptide and the natural MSTN inhibitor follistatin (FST) from doping control serum samples. The approach was comprehensively characterized and found to allow for a specific detection down to concentrations of 3-9 ng/mL. Moreover, several potential metabolites of the drug candidates referred to as DF-3, DF-25, and Peptide 7 were identified as valuable complementary analytical targets for doping control analytical assays. Overall, the acquired data pave the way for an implementation of MSTN inhibitory peptides into routine sports drug testing. Even though no drug candidate has obtained clinical approval yet, a proactive development of detection assays is of utmost importance to deter athletes from misusing such compounds, which are readily available for research purposes and on the black market.

The Preventive Effect of Specific Collagen Peptides against Dexamethasone-Induced Muscle Atrophy in Mice.

Muscle atrophy, also known as muscle wasting, is the thinning of muscle mass due to muscle disuse, aging, or diseases such as cancer or neurological problems. Muscle atrophy is closely related to the quality of life and has high morbidity and mortality. However, therapeutic options for muscle atrophy are limited, so studies to develop therapeutic agents for muscle loss are always required. For this study, we investigated how orally administered specific collagen peptides (CP) affect muscle atrophy and elucidated its molecular mechanism using an in vivo model. We treated mice with dexamethasone (DEX) to induce a muscular atrophy phenotype and then administered CP (0.25 and 0.5 g/kg) for four weeks. In a microcomputed tomography analysis, CP (0.5 g/kg) intake significantly increased the volume of calf muscles in mice with DEX-induced muscle atrophy. In addition, the administration of CP (0.25 and 0.5 g/kg) restored the weight of the gluteus maximus and the fiber cross-sectional area (CSA) of the pectoralis major and calf muscles, which were reduced by DEX. CP significantly inhibited the mRNA expression of myostatin and the phosphorylation of Smad2, but it did not affect TGF-ß, BDNF, or FNDC5 gene expression. In addition, AKT/mTOR, a central pathway for muscle protein synthesis and related to myostatin signaling, was enhanced in the groups that were administered CP. Finally, CP decreased serum albumin levels and increased TNF-α gene expression. Collectively, our in vivo results demonstrate that CP can alleviate muscle wasting through a multitude of mechanisms. Therefore, we propose CP as a supplement or treatment to prevent muscle atrophy.

Evaluation of Associations of Growth Differentiation Factor-11, Growth Differentiation Factor-8, and Their Binding Proteins Follistatin and Follistatin-Like Protein-3 With Dementia and Cognition.

BACKGROUND: Studies using heterochronic parabiosis discovered that circulating factors mediate brain aging in animal models. METHODS: We assessed growth differentiation factors (GDF)-11 and GDF-8 using mass spectrometry and inhibitors follistatin and follistatin-like protein-3 (FSTL-3) with ELISA in the Cardiovascular Health Study (CHS; N = 1 506) and the Health, Aging and Body Composition (Health ABC) Study (N = 1 237). CLL-11 and beta-2 microglobulin (ß2M) were measured with ELISA in a subset of 400 individuals in Health ABC. Associations were assessed with cognitive function, brain magnetic resonance imaging (MRI) findings (CHS only), and incident dementia using correlations, linear regression, and Cox proportional hazards models. RESULTS: In CHS, levels of GDF-11, GDF-8, and follistatin were not correlated cross-sectionally with the 3MSE or DSST, brain MRI findings of white matter hyperintensity, atrophy, or small infarcts, nor were they associated with incident dementia. FSTL-3 was modestly correlated with poorer cognitive function, greater white matter hyperintensities, and atrophy on MRI, as well as with incident dementia with an adjusted hazard ratio (HR) of 1.72 (95% CI = 1.13, 2.61) per doubling of FSTL-3. FSTL-3 was not associated with cognition or dementia in Health ABC, but GDF-8 was associated with both. The adjusted HR for incident dementia was 1.50 (95% CI = 1.07, 2.10) per doubling of GDF-8. CONCLUSIONS: Total GDF-11 level was not related to cognition or dementia in older adults. Associations of GDF-8 with cognitive outcomes in Health ABC were not expected, but consistent with animal models. Associations of FSTL-3 with cognition, brain abnormalities, and incident dementia in CHS implicate TGFß superfamily inhibition in the pathogenesis of dementia.

Myostatin-mediated regulation of skeletal muscle damage post-acute Aeromonas hydrophila infection in Nile tilapia (Oreochromis niloticus L.).

This study focuses on the relationship between myostatin (MyoS), myogenin (MyoG), and the growth hormone/insulin-like growth factor-1 (GH/IGF-1) axis for muscle growth and histopathological changes in muscle after an Aeromonas hydrophila infection. A total number of 90 Nile tilapia (55.85 g) were randomly allocated into two equal groups of three replicates each. The first group was an uninfected control group that was injected intraperitoneally (ip) with 0.2 ml phosphate buffer saline (PBS), while the second group was injected ip with 0.2 ml (1.3 × 108 CFU/ml) Aeromonas hydrophila culture suspension. Sections of white muscle and liver tissues were taken from each group 24 h, 48 h, 72 h, and 1 week after infection for molecular analysis and histopathological examination. The results revealed that with time progression, the severity of muscle lesions increased from edema between bundles and mononuclear inflammatory cell infiltration 24 h post-challenge to severe atrophy of muscle bundles with irregular and curved fibers with hyalinosis of the fibers 1 week postinfection. The molecular analysis showed that bacterial infection was able to induce the muscle expression levels of GH with reduced ILGF-1, MyoS, and MyoG at 24 h postinfection. However, time progression postinfection reversed these findings through elevated muscle expression levels of MyoS with regressed expression levels of muscle GH, ILGF-1, and MyoG. There have been no previous reports on the molecular expression analysis of the aforementioned genes and muscle histopathological changes in Nile tilapia following acute Aeromonas hydrophila infection. Our findings, collectively, revealed that the up-and down-regulation of the myostatin signaling is likely to be involved in the postinfection-induced muscle wasting through the negative regulation of genes involved in muscle growth, such as GH, ILGF-1, and myogenin, in response to acute Aeromonas hydrophila infection in Nile tilapia, Oreochromis niloticus.