Indian Ornamental Tarantula (Poecilotheria regalis) Venom Affects Myoblast Function and Causes Skeletal Muscle Damage.

Envenomation by the Indian ornamental tarantula (Poecilotheria regalis) is medically relevant to humans, both in its native India and worldwide, where they are kept as pets. Muscle-related symptoms such as cramps and pain are commonly reported in humans following envenomation by this species. There is no specific treatment, including antivenom, for its envenomation. Moreover, the scientific knowledge of the impact of this venom on skeletal muscle function is highly limited. Therefore, we carried out this study to better understand the myotoxic properties of Poecilotheria regalis venom by determining its effects in cultured myoblasts and in the tibialis anterior muscle in mice. While there was no effect found on undifferentiated myoblasts, the venom affected differentiated multinucleated myotubes resulting in the reduction of fusion and atrophy of myotubes. Similarly, intramuscular administration of this venom in the tibialis anterior muscle in mice resulted in extensive muscle damage on day 5. However, by day 10, the regeneration was evident, and the regeneration process continued until day 20. Nevertheless, some tissue abnormalities including reduced dystrophin expression and microthrombi presence were observed on day 20. Overall, this study demonstrates the ability of this venom to induce significant muscle damage and affect its regeneration in the early stages. These data provide novel mechanistic insights into this venom-induced muscle damage and guide future studies to isolate and characterise individual toxic component(s) that induce muscle damage and their significance in developing better therapeutics.

Current Research on Vitamin D Supplementation against Sarcopenia: A Review of Clinical Trials.

Vitamin D plays an important role in skeletal muscle function and metabolism. The aim of this review was A) to discuss the clinical evidence of vitamin D supplementation either alone or combined with other strategies in the prevention of sarcopenia in non-sarcopenic individuals and B) to critically discuss the clinical evidence on the effect of vitamin D combined with other strategies on muscle strength, mass and function in sarcopenic individuals without vitamin D deficiency. Sparse clinical data on non-sarcopenic individuals indicate that vitamin D alone has a subtle beneficial effect on knee extensor strength at doses 880-1600 IU/day without improving handgrip strength or muscle mass. When co-administered with other supplements such as protein, mixed effects appear to prevent the decline of muscle mass, possibly delaying the onset of sarcopenia in non-sarcopenic individuals, at doses of 800-1,000 IU/day over 6-12 weeks. In sarcopenic individuals, vitamin D 100-1,000 IU/day co-supplementation with protein results in increased handgrip strength between 9.8-40.5%. However, there is no strong clinical evidence that vitamin D dosage correlates with changes in muscle strength or mass. Potential sources of discrepancy among studies are discussed. Future studies with appropriate experimental design are essential to dissect the net effect of vitamin D on sarcopenia.

Current Insights into Cellular Senescence and Myotoxicity Induced by Doxorubicin: The Role of Exercise and Growth Factors.

Doxorubicin is an anti-neoplasmic drug that prevents DNA replication but induces senescence and cellular toxicity. Intensive research has focused on strategies to alleviate the doxorubicin-induced skeletal myotoxicity. The aim of the present review is to critically discuss the relevant scientific evidence about the role of exercise and growth factor administration and offer novel insights about newly developed-tools to combat the adverse drug reactions of doxorubicin treatment on skeletal muscle. In the first part, we discuss current data and mechanistic details on the impact of doxorubicin on skeletal myotoxicity. We next review key aspects about the role of regular exercise and the impact of growth factors, administered either pharmacologically or via genetic interventions. Future strategies such as combination of exercise and growth factor administration remain to be established to combat the pharmacologically-induced myotoxicity.

Pro-cachectic factors link experimental and human chronic kidney disease to skeletal muscle wasting programs.

Skeletal muscle wasting is commonly associated with chronic kidney disease (CKD), resulting in increased morbidity and mortality. However, the link between kidney and muscle function remains poorly understood. Here, we took a complementary interorgan approach to investigate skeletal muscle wasting in CKD. We identified increased production and elevated blood levels of soluble pro-cachectic factors, including activin A, directly linking experimental and human CKD to skeletal muscle wasting programs. Single-cell sequencing data identified the expression of activin A in specific kidney cell populations of fibroblasts and cells of the juxtaglomerular apparatus. We propose that persistent and increased kidney production of pro-cachectic factors, combined with a lack of kidney clearance, facilitates a vicious kidney/muscle signaling cycle, leading to exacerbated blood accumulation and, thereby, skeletal muscle wasting. Systemic pharmacological blockade of activin A using soluble activin receptor type IIB ligand trap as well as muscle-specific adeno-associated virus-mediated downregulation of its receptor ACVR2A/B prevented muscle wasting in different mouse models of experimental CKD, suggesting that activin A is a key factor in CKD-induced cachexia. In summary, we uncovered a crosstalk between kidney and muscle and propose modulation of activin signaling as a potential therapeutic strategy for skeletal muscle wasting in CKD.

Platelet releasate normalises the compromised muscle regeneration in a mouse model of hyperlipidaemia.

NEW FINDINGS: What is the central question of this study? What is the impact of obesity-independent hyperlipidaemia on skeletal muscle stem cell function of ApoE-deficient (ApoE-/- ) mice? What is the main finding and its importance? Compromised muscle stem cell function accounts for the impaired muscle regeneration in hyperlipidaemic ApoE-/- mice. Importantly, impaired muscle regeneration is normalised by administration of platelet releasate. ABSTRACT: Muscle satellite cells are important stem cells for skeletal muscle regeneration and repair after injury. ApoE-deficient mice, an established mouse model of hyperlipidaemia and atherosclerosis, show evidence of oxidative stress-induced lesions and fat infiltration in skeletal muscle followed by impaired repair after injury. However, the mechanisms underpinning attenuated muscle regeneration remain to be fully defined. Key to addressing the latter is to understand the properties of muscle stem cells from ApoE-deficient mice and their myogenic potential. Muscle stem cells from ApoE-deficient mice were cultured both ex vivo (on single fibres) and in vitro (primary myoblasts) and their myogenic capacity was determined. Skeletal muscle regeneration was studied on days 5 and 10 after cardiotoxin injury. ApoE-deficient muscle stem cells showed delayed activation and differentiation on single muscle fibres ex vivo. Impaired proliferation and differentiation profiles were also evident on isolated primary muscle stem cells in culture. ApoE-deficient mice displayed impaired skeletal muscle regeneration after acute injury in vivo. Administration of platelet releasate in ApoE-deficient mice reversed the deficits of muscle regeneration after acute injury to wild-type levels. These findings indicate that muscle stem cell myogenic potential is perturbed in skeletal muscle of a mouse model of hyperlipidaemia. We propose that platelet releasate could be a therapeutic intervention for conditions with associated myopathy such as peripheral arterial disease.

A muscle growth-promoting treatment based on the attenuation of activin/myostatin signalling results in long-term testicular abnormalities.

Activin/myostatin signalling acts to induce skeletal muscle atrophy in adult mammals by inhibiting protein synthesis as well as promoting protein and organelle turnover. Numerous strategies have been successfully developed to attenuate the signalling properties of these molecules, which result in augmenting muscle growth. However, these molecules, in particular activin, play major roles in tissue homeostasis in numerous organs of the mammalian body. We have recently shown that although the attenuation of activin/myostatin results in robust muscle growth, it also has a detrimental impact on the testis. Here, we aimed to discover the long-term consequences of a brief period of exposure to muscle growth-promoting molecules in the testis. We demonstrate that muscle hypertrophy promoted by a soluble activin type IIB ligand trap (sActRIIB) is a short-lived phenomenon. In stark contrast, short-term treatment with sActRIIB results in immediate impact on the testis, which persists after the sessions of the intervention. Gene array analysis identified an expansion in aberrant gene expression over time in the testis, initiated by a brief exposure to muscle growth-promoting molecules. The impact on the testis results in decreased organ size as well as quantitative and qualitative impact on sperm. Finally, we have used a drug-repurposing strategy to exploit the gene expression data to identify a compound - N6-methyladenosine - that may protect the testis from the impact of the muscle growth-promoting regime. This work indicates the potential long-term harmful effects of strategies aimed at promoting muscle growth by attenuating activin/myostatin signalling. Furthermore, we have identified a molecule that could, in the future, be used to overcome the detrimental impact of sActRIIB treatment on the testis.

Diminution in sperm quantity and quality in mouse models of Duchenne Muscular Dystrophy induced by a myostatin-based muscle growth-promoting intervention.

Duchenne Muscular Dystrophy is a devastating disease caused by the absence of a functional rod-shaped cytoplasmic protein called dystrophin. Several avenues are being developed aimed to restore dystrophin expression in boys affected by this X-linked disease. However, its complete cure is likely to need combinational approaches which may include regimes aimed at restoring muscle mass. Augmenting muscle growth through the manipulation of the Myostatin/Activin signalling axis has received much attention. However, we have recently shown that while manipulation of this axis in wild type mice using the sActRIIB ligand trap indeed results in muscle growth, it also had a detrimental impact on the testis. Here we examined the impact of administering a powerful Myostatin/Activin antagonist in two mouse models of Duchenne Muscular Dystrophy. We report that whilst the impact on muscle growth was not always positive, both models showed attenuated testis development. Sperm number, motility and ultrastructure were significantly affected by the sActRIIB treatment. Our report suggests that interventions based on Myostatin/Activin should investigate off-target effects on tissues as well as muscle.

Inhibition of Activin/Myostatin signalling induces skeletal muscle hypertrophy but impairs mouse testicular development.

Numerous approaches are being developed to promote post-natal muscle growth based on attenuating Myostatin/Activin signalling for clinical uses such as the treatment neuromuscular diseases, cancer cachexia and sarcopenia. However there have been concerns about the effects of inhibiting Activin on tissues other than skeletal muscle. We intraperitoneally injected mice with the Activin ligand trap, sActRIIB, in young, adult and a progeric mouse model. Treatment at any stage in the life of the mouse rapidly increased muscle mass. However at all stages of life the treatment decreased the weights of the testis. Not only were the testis smaller, but they contained fewer sperm compared to untreated mice. We found that the hypertrophic muscle phenotype was lost after the cessation of sActRIIB treatment but abnormal testis phenotype persisted. In summary, attenuation of Myostatin/Activin signalling inhibited testis development. Future use of molecules based on a similar mode of action to promote muscle growth should be carefully profiled for adverse side-effects on the testis. However the effectiveness of sActRIIB as a modulator of Activin function provides a possible therapeutic strategy to alleviate testicular seminoma development.

Quantitative Redox Biology of Exercise.

Biology is rich in claims that reactive oxygen and nitrogen species are involved in every biological process and disease. However, many quantitative aspects of redox biology remain elusive. The important quantitative parameters you need to address the feasibility of redox reactions in vivo are: rate of formation and consumption of a reactive oxygen and nitrogen species, half-life, diffusibility and membrane permeability. In the first part, we explain the basic chemical kinetics concepts and algebraic equations required to perform "street fighting" quantitative analysis. In the second part, we provide key numbers to help thinking about sizes, concentrations, rates and other important quantities that describe the major oxidants (superoxide, hydrogen peroxide, nitric oxide) and antioxidants (vitamin C, vitamin E, glutathione). In the third part, we present the quantitative effect of exercise on superoxide, hydrogen peroxide and nitric oxide concentration in mitochondria and whole muscle and calculate how much hydrogen peroxide concentration needs to increase to transduce signalling. By taking into consideration the quantitative aspects of redox biology we can: i) refine the broad understanding of this research area, ii) design better future studies and facilitate comparisons among studies, and iii) define more efficiently the "borders" between cellular signaling and stress.

Evaluating Trastuzumab in the treatment of HER2 positive breast cancer.

The transmembrane oncoprotein HER2 is encoded by ERBB2 gene and overexpressed in around 20% of invasive breast cancers. It can be specifically targeted by Trastuzumab (Herceptin®), a humanised IgG1 antibody. Trastuzumab has been regarded as one of the most effective therapeutic drugs targeted to HER2 positive cancers. However, there are drawbacks, notably cardiotoxicity and resistance, which have raised awareness in clinical use. Therefore, understanding the mechanism of action is vital to establish improved therapeutic strategies. Here we evaluate Trastuzumab application in the treatment of HER2 positive breast cancer, focusing on its mechanistic actions and clinical effectiveness. Alternative therapies targeting the HER2 receptor and its downstream anomalies will also be discussed, as these could highlight further targets that could be key to improving clinical outcomes.

Loss of CD36 protects against diet-induced obesity but results in impaired muscle stem cell function, delayed muscle regeneration and hepatic steatosis.

AIM: The prevalence of obesity is a major risk factor for cardiovascular and metabolic diseases including impaired skeletal muscle regeneration. Since skeletal muscle regenerative capacity is regulated by satellite cells, we aimed to investigate whether a high-fat diet impairs satellite cell function and whether this is linked to fatty acid uptake via CD36. We also aimed to determine whether loss of CD36 impacts on muscle redox homeostasis and skeletal muscle regenerative capacity. METHODS: We studied the impact of a high-fat diet and CD36 deficiency on murine skeletal muscle morphology, redox homeostasis, satellite cell function, bioenergetics and lipid accumulation in the liver. We also determined the effect of CD36 deficiency on skeletal muscle regeneration. RESULTS: High-fat diet increased body weight, intramuscular lipid accumulation and oxidative stress in wild-type mice that were significantly mitigated in CD36-deficient mice. High-fat diet and CD36 deficiency independently attenuated satellite cell function on single fibres and myogenic capacity on primary satellite cells. CD36 deficiency resulted in delayed skeletal muscle regeneration following acute injury with cardiotoxin. CD36-deficient and wild-type primary satellite cells had distinct bioenergetic profiles in response to palmitate. High-fat diet induced hepatic steatosis in both genotypes that was more pronounced in the CD36-deficient mice. CONCLUSION: This study demonstrates that CD36 deficiency protects against diet-induced obesity, intramuscular lipid deposition and oxidative stress but results in impaired muscle satellite cell function, delayed muscle regeneration and hepatic steatosis. CD36 is a key mediator of fatty acid uptake in skeletal muscle, linking obesity with satellite cell function and muscle regeneration.

Optimising platelet secretomes to deliver robust tissue-specific regeneration.

Promoting cell proliferation is the cornerstone of most tissue regeneration therapies. As platelet-based applications promote cell division and can be customised for tissue-specific efficacy, this makes them strong candidates for developing novel regenerative therapies. Therefore, the aim of this study was to determine if platelet releasate could be optimised to promote cellular proliferation and differentiation of specific tissues. Growth factors in platelet releasate were profiled for physiological and supraphysiological platelet concentrations. We analysed the effect of physiological and supraphysiological releasate on C2C12 skeletal myoblasts, H9C2 rat cardiomyocytes, human dermal fibroblasts (HDF), HaCaT keratinocytes, and chondrocytes. Cellular proliferation and differentiation were assessed through proliferation assays, mRNA, and protein expression. We show that supraphysiological releasate is not simply a concentrated version of physiological releasate. Physiological releasate promoted C2C12, HDF, and chondrocyte proliferation with no effect on H9C2 or HaCaT cells. Supraphysiological releasate induced stronger proliferation in C2C12 and HDF cells compared with physiological releasate. Importantly, supraphysiological releasate induced proliferation of H9C2 cells. The proliferative effects of skeletal and cardiac muscle cells were in part driven by vascular endothelial growth factor alpha. Furthermore, supraphysiological releasate induced differentiation of H9C2 and C2C12, HDF, and keratinocytes. This study provides insights into the ability of releasate to promote muscle, heart, skin, and cartilage cell proliferation and differentiation and highlights the importance of optimising releasate composition for tissue-specific regeneration.

The effect of high-fat diet on the morphological properties of the forelimb musculature in hypertrophic myostatin null mice.

Obesity is a worldwide nutritional disorder affecting body performance, including skeletal muscle. Inhibition of myostatin not only increases the muscle mass but also it reduces body fat accumulation. We examined the effect of high-fat diet on the phenotypic properties of forelimb muscles from myostatin null mice. Male wild-type and myostatin null mice were fed on either a normal diet or a high-fat diet (45% fat) for 10 weeks. Musculus triceps brachii Caput longum; M. triceps brachii Caput laterale; M. triceps brachii Caput mediale; M. extensor carpi ulnaris and M. flexor carpi ulnaris were processed for fiber type composition using immunohistochemistry and morphometric analysis. Although the muscle mass revealed no change under a high-fat diet, there were morphometric alterations in the absence of myostatin. We show that high-fat diet reduces the cross-sectional area of the fast (IIB and IIX) fibers in M. triceps brachii Caput longum and M. triceps brachii Caput laterale of both genotypes. In contrast, increases of fast fiber areas were observed in both M. extensor carpi ulnaris of wild-type and M. flexor carpi ulnaris of myostatin null mice. Meanwhile, a high-fat diet increased the area of the fast IIA fibers in wild-type mice; myostatin null mice display a muscle-dependent alteration in the area of the same fiber type. The combined high-fat diet and myostatin deletion shows no effect on the area of slow type I fibers. Although a high-fat diet causes a reduction in the area of the peripheral IIB fibers in both genotypes, only myostatin null mice show an increase in the area of the central IIB fibers. We provide evidence that a high-fat diet induces a muscle-dependent fast to slow myofiber shift in the absence of myostatin. The data suggest that the morphological alterations of muscle fibers under a combined high-fat diet and myostatin deletion reflect a functional adaptation of the muscle to utilize the high energy intake.

Current Insights into the Potential Misuse of Platelet-based Applications for Doping in Sports.

Platelet-based applications are currently used for the delivery of growth factors and other biomolecules as autologous biomaterials in regenerative medicine and cosmetic therapies. Many studies have revealed that platelet-based applications such as platelet-rich plasma and platelet releasate exhibit beneficial biological effects after a sports injury or trauma when administered locally by intramuscular injections. At present, treatment of the public, patients and athletes with platelet-based applications is permitted and regulated by the Food and Drug Administration and the World Anti-Doping Agency. Since 2011 the use of autologous platelet-rich plasma is permitted in competitive sports by the World Anti-Doping Agency, due to the lack of evidence in performance enhancement and anabolic effects. However, accumulating research has recently shed light on the role of platelet-derived growth factors in wound healing, skeletal myogenesis, muscle stem cell function and tissue regeneration. Although any ergogenic potential of platelet-rich plasma and platelet releasate on intact skeletal muscle and human sports performance remain to be established, novel evidence suggests that platelet-derived growth factors can modulate muscle, tendon, ligament, protein synthesis/degradation, vascularization, energy utilization and regenerative capacity in various experimental settings. Since platelet-based applications are currently not prohibited, they constitute a tool for potential abuse and doping in sports. The aim of this review is to critically discuss and assimilate current insights and biological evidence that set the ground for exploitation and misuse in competitive sports, and develop strategies to combat these activities.

Compression of morbidity in a progeroid mouse model through the attenuation of myostatin/activin signalling.

BACKGROUND: One of the principles underpinning our understanding of ageing is that DNA damage induces a stress response that shifts cellular resources from growth towards maintenance. A contrasting and seemingly irreconcilable view is that prompting growth of, for example, skeletal muscle confers systemic benefit. METHODS: To investigate the robustness of these axioms, we induced muscle growth in a murine progeroid model through the use of activin receptor IIB ligand trap that dampens myostatin/activin signalling. Progeric mice were then investigated for neurological and muscle function as well as cellular profiling of the muscle, kidney, liver, and bone. RESULTS: We show that muscle of Ercc1Δ/- progeroid mice undergoes severe wasting (decreases in hind limb muscle mass of 40-60% compared with normal mass), which is largely protected by attenuating myostatin/activin signalling using soluble activin receptor type IIB (sActRIIB) (increase of 30-62% compared with untreated progeric). sActRIIB-treated progeroid mice maintained muscle activity (distance travel per hour: 5.6 m in untreated mice vs. 13.7 m in treated) and increased specific force (19.3 mN/mg in untreated vs. 24.0 mN/mg in treated). sActRIIb treatment of progeroid mice also improved satellite cell function especially their ability to proliferate on their native substrate (2.5 cells per fibre in untreated progeroids vs. 5.4 in sActRIIB-treated progeroids after 72 h in culture). Besides direct protective effects on muscle, we show systemic improvements to other organs including the structure and function of the kidneys; there was a major decrease in the protein content in urine (albumin/creatinine of 4.9 sActRIIB treated vs. 15.7 in untreated), which is likely to be a result in the normalization of podocyte foot processes, which constitute the filtration apparatus (glomerular basement membrane thickness reduced from 224 to 177 nm following sActRIIB treatment). Treatment of the progeric mice with the activin ligand trap protected against the development of liver abnormalities including polyploidy (18.3% untreated vs. 8.1% treated) and osteoporosis (trabecular bone volume; 0.30 mm3 in treated progeroid mice vs. 0.14 mm3 in untreated mice, cortical bone volume; 0.30 mm3 in treated progeroid mice vs. 0.22 mm3 in untreated mice). The onset of neurological abnormalities was delayed (by ~5 weeks) and their severity reduced, overall sustaining health without affecting lifespan. CONCLUSIONS: This study questions the notion that tissue growth and maintaining tissue function during ageing are incompatible mechanisms. It highlights the need for future investigations to assess the potential of therapies based on myostatin/activin blockade to compress morbidity and promote healthy ageing.

Regulation of the dystrophin-associated glycoprotein complex composition by the metabolic properties of muscle fibres.

The dystrophin-glycoprotein complex (DGC) links the muscle cytoskeleton to the extracellular matrix and is responsible for force transduction and protects the muscle fibres from contraction induced damage. Mutations in components of the DGC are responsible for muscular dystrophies and congenital myopathies. Expression of DGC components have been shown to be altered in many myopathies. In contrast we have very little evidence of whether adaptive changes in muscle impact on DGC expression. In this study we investigated connection between muscle fibre phenotype and the DGC. Our study reveals that the levels of DGC proteins at the sarcolemma differ in highly glycolytic muscle compared to wild-type and that these changes can be normalised by the super-imposition of an oxidative metabolic programme. Importantly we show that the metabolic properties of the muscle do not impact on the total amount of DGC components at the protein level. Our work shows that the metabolic property of a muscle fibre is a key factor in regulating the expression of DGC proteins at the sarcolemma.

The inhibitory subunit of cardiac troponin (cTnI) is modified by arginine methylation in the human heart.

BACKGROUND: The inhibitory subunit of cardiac troponin (cTnI) is a gold standard cardiac biomarker and also an essential protein in cardiomyocyte excitation-contraction coupling. The interactions of cTnI with other proteins are fine-tuned by post-translational modification of cTnI. Mutations in cTnI can lead to hypertrophic cardiomyopathy. METHODS AND RESULTS: Here we report, for the first time, that cTnI is modified by arginine methylation in human myocardium. Using Western blot, we observed reduced levels of cTnI arginine methylation in human hypertrophic cardiomyopathy compared to dilated cardiomyopathy biopsies. Similarly, using a rat model of cardiac hypertrophy we observed reduced levels of cTnI arginine methylation compared to sham controls. Using mass spectrometry, we identified cTnI methylation sites at R74/R79 and R146/R148 in human cardiac samples. R146 and R148 lie at the boundary between the critical cTnI inhibitory and switch peptides; PRMT1 methylated an extended inhibitory peptide at R146 and R148 in vitro. Mutations at R145 that have been associated with hypertrophic cardiomyopathy hampered R146/R148 methylation by PRMT1 in vitro. H9c2 cardiac-like cells transfected with plasmids encoding for a methylation-deficient R146A/R148A cTnI protein developed cell hypertrophy, with a 32% increase in cell size after 72 h, compared to control cells. DISCUSSION: Our results provide evidence for a novel and significant cTnI post-translational modification. Our work opens the door to translational investigations of cTnI arginine methylation as a biomarker of disease, which can include e.g. cardiomyopathies, myocardial infarction and heart failure, and offers a novel way to investigate the effect of cTnI mutations in the inhibitory/switch peptides.

Mechanisms underpinning the permanent muscle damage induced by snake venom metalloprotease.

Snakebite is a major neglected tropical health issue that affects over 5 million people worldwide resulting in around 1.8 million envenomations and 100,000 deaths each year. Snakebite envenomation also causes innumerable morbidities, specifically loss of limbs as a result of excessive tissue/muscle damage. Snake venom metalloproteases (SVMPs) are a predominant component of viper venoms, and are involved in the degradation of basement membrane proteins (particularly collagen) surrounding the tissues around the bite site. Although their collagenolytic properties have been established, the molecular mechanisms through which SVMPs induce permanent muscle damage are poorly understood. Here, we demonstrate the purification and characterisation of an SVMP from a viper (Crotalus atrox) venom. Mass spectrometry analysis confirmed that this protein is most likely to be a group III metalloprotease (showing high similarity to VAP2A) and has been referred to as CAMP (Crotalus atrox metalloprotease). CAMP displays both collagenolytic and fibrinogenolytic activities and inhibits CRP-XL-induced platelet aggregation. To determine its effects on muscle damage, CAMP was administered into the tibialis anterior muscle of mice and its actions were compared with cardiotoxin I (a three-finger toxin) from an elapid snake (Naja pallida) venom. Extensive immunohistochemistry analyses revealed that CAMP significantly damages skeletal muscles by attacking the collagen scaffold and other important basement membrane proteins, and prevents their regeneration through disrupting the functions of satellite cells. In contrast, cardiotoxin I destroys skeletal muscle by damaging the plasma membrane, but does not impact regeneration due to its inability to affect the extracellular matrix. Overall, this study provides novel insights into the mechanisms through which SVMPs induce permanent muscle damage.

Platelet releasate promotes skeletal myogenesis by increasing muscle stem cell commitment to differentiation and accelerates muscle regeneration following acute injury.

AIM: The use of platelets as biomaterials has gained intense research interest. However, the mechanisms regarding platelet-mediated skeletal myogenesis remain to be established. The aim of this study was to determine the role of platelet releasate in skeletal myogenesis and muscle stem cell fate in vitro and ex vivo respectively. METHODS: We analysed the effect of platelet releasate on proliferation and differentiation of C2C12 myoblasts by means of cell proliferation assays, immunohistochemistry, gene expression and cell bioenergetics. We expanded in vitro findings on single muscle fibres by determining the effect of platelet releasate on murine skeletal muscle stem cells using protein expression profiles for key myogenic regulatory factors. RESULTS: TRAP6 and collagen used for releasate preparation had a more pronounced effect on myoblast proliferation vs thrombin and sonicated platelets (P < 0.05). In addition, platelet concentration positively correlated with myoblast proliferation. Platelet releasate increased myoblast and muscle stem cell proliferation in a dose-dependent manner, which was mitigated by VEGFR and PDGFR inhibition. Inhibition of VEGFR and PDGFR ablated MyoD expression on proliferating muscle stem cells, compromising their commitment to differentiation in muscle fibres (P < 0.001). Platelet releasate was detrimental to myoblast fusion and affected differentiation of myoblasts in a temporal manner. Most importantly, we show that platelet releasate promotes skeletal myogenesis through the PDGF/VEGF-Cyclin D1-MyoD-Scrib-Myogenin axis and accelerates skeletal muscle regeneration after acute injury. CONCLUSION: This study provides novel mechanistic insights on the role of platelet releasate in skeletal myogenesis and set the physiological basis for exploiting platelets as biomaterials in regenerative medicine.