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Table of Contents

    Topic review

    Antimyostatin Treatment

    Subjects: Neurosciences
    View times: 8
    Submitted by: Tue Nielsen


    Myostatin, also known as growth and differentiation factor 8 (GDF-8), was identified in 1997 by McPherron and Lee.

    1. Introduction

    Muscular dystrophies consist of a broad array of inherited conditions characterized by muscular wasting and atrophy. As clinical presentations in patients may vary due to a wide spectrum of phenotype–genotype variants for a particular gene, a common treatment, not depending on correcting a single molecular defect, has emerged as an attractive target for development. For the last 20 years, one of the most promising therapeutic subjects in the field of muscular dystrophies has been myostatin. Identified for the first time in 1997, myostatin knock-out in mice caused increased muscle mass [1] and mutations in the myostatin gene (MSTN) gene have subsequently been identified in the double muscled Belgian Blue and Piedmontese cattle [2][3][4] as well as whippet racing dogs [5]. In 2004, a loss-of-function mutation of MSTN in a German boy with a hypermuscular phenotype demonstrated that the effect of myostatin is functionally conserved across different mammalian species [6]. Since myostatin loss of function did not appear to have any negative impact on viability and longevity [7][8], interest was raised towards a novel treatment by harnessing the potential of inhibiting this negative regulator of muscular growth. Numerous studies in animal models and clinical trials have tried to explore this relationship with promising results in preclinical studies, which have translated poorly in human clinical studies.

    2. Molecular Involvement of Myostatin in Mice and Humans

    Myostatin, also known as growth and differentiation factor 8 (GDF-8), was identified in 1997 by McPherron and Lee [1]. During embryogenesis, myostatin is expressed in the developing epaxial and hypaxial myotomes [9][10] and hereafter in muscular tissue postnatally, but has also been found at low expression in adipose tissue, heart and circulation throughout development [11][12]. As the mstn-gene is highly conserved among different vertebrate species [3][6][13][14][15][16], it is evident that it has an important function in muscle development and physiology, which has been preserved during the course of evolution. As a member of the TGF-β-superfamily, myostatin shows homology to other growth and differentiation factors, such as bone morphogenic proteins (BMP) and activins, which also elicit their biological function as dimers. Prepro-myostatin is synthesized as an N-terminal signal peptide followed by a propeptide domain and eventually a mature C-terminal domain [13]. During proteolytic processing, the signaling peptide is removed and the propeptide is cleaved from the mature protein. As the mature C-terminal domain dimerizes and forms disulfide bridges, it remains inactivated since noncovalent bonds between the mature dimer and the propeptide hold the mature myostatin in an inactive state [11][17][18][19]. To exert its function, the propeptide must be cleaved from the inactive complex by a family of BMP1/TLD-metalloprotease proteins [19][20]. Other than the propeptide itself, regulation of myostatin activity is also known to be mediated by follistatin [17][21], follistatin-related gene (FLRG) [11], Gasp-1 [22] and the proteoglycan protein decorin [23][24] typically by blocking the binding of myostatin to the receptors (Figure 1).

    Figure 1. An overview of various approaches used in myostatin inhibition. Various factors and approaches in myostatin inhibition as outlined in Section 2 and Section 5. Treatments applied in clinical trials have been colored yellow. The Smad2/3 intracellular signaling pathway downstream the ActRIIB leads to altered gene transcription of muscle regulatory factors.

    Once liberated from inhibitory proteins, myostatin, as well as other members of the TGF-β-family, binds to activin-receptors and, in the case of myostatin, mainly to the activin-receptor type IIB (ActRIIB) as well as the type IA receptor [17]. The activin receptors are transmembrane serine/threonine kinases that subsequently recruit and activate dimers of type I-receptors (ALK4 and ALK5) [25][26]. Depending on the receptor ligand and the composition of the receptor complex, the type I-receptor will phosphorylate and activate intracellular protein Smad2 and 3 downstream to the membrane receptors through the canonical Smad-pathway. Smad2/3 binds to Smad4 and the complex translocates to the nucleus [27], where muscle regulatory factors MyoD, Myf5 and Myogenin are repressed [28], preventing myoblast proliferation [29] and differentiation [28]. Obstruction of the myostatin pathway inhibits activation of Smad2/3, making Smad4 available in the BMP signaling pathway which promotes hypertrophy and counteracts the effects produced by myostatin [30]. Other noncanonical pathways activated by myostatin involve (among others) AMP-activated protein kinase (AMPK) and p38 mitogen-activated protein kinase (MAPK) [31][32].

    3. Myostatin in Healthy Humans and in Relation to Clinical Manifestations of Cachexia and Muscular Wasting

    Compared to healthy young men, there was no reported change in serum myostatin levels in an elderly population with mild or severe sarcopenia (as defined by muscular contractile force) [33]. Burch et al. reported that myostatin was 57% higher in a healthy cohort >25 years of age compared to a healthy cohort <25 years, with an age-dependent increase in the younger cohort but not in the older cohort [34]. A different study with more than 1100 participating men aged 20–87 years demonstrated that circulating myostatin level was dependent on age and body mass index [35]. Additionally, men had higher levels than women [34]. This is in contrast to findings of myostatin levels declining on ageing in men both measured by ELISA [36] and immunoplexed liquid chromatography with tandem mass spectrometry [37]. A smaller study of eight young and six elderly women showed higher levels of myostatin mRNA in muscle biopsies of the older group [38]. Various groups have sought to determine the use of myostatin as a potential biomarker for muscle wasting but the conclusions have been ambiguous [39][40][41].

    The effect of age on the expression of not only myostatin but also other promyogenic muscle regulatory factors (MRF) following exercise was examined by Raue et al. They found that at rest, there is a relative upregulation of both MRF and myostatin prior to exercise in elderly women compared to younger ones, but that the postexercise downregulation of myostatin is not hampered by age [38]. A study in healthy and sarcopenic elderly men demonstrated that resistance training or a combination of resistance and endurance training caused a decrease in myostatin [42][43].

    The clinical relevance of myostatin in humans was described for the first time in HIV patients, who had increased levels of myostatin compared to healthy subjects. Furthermore, the levels were even higher in the patients who met the definition of AIDS-wasting syndrome [13]. The role of myostatin in muscular atrophy and muscle wasting was also determined in mice that developed cachexia in response to myostatin overexpression [44]. Cachexia manifests as a complex metabolic syndrome due to an underlying illness characterized by muscle wasting in conditions such as chronic heart failure (HF), cancer, chronic obstructive pulmonary disease (COPD) or chronic kidney disease (CKD) [45]. The use of myostatin inhibitors in such populations with progressive muscle wasting or atrophy secondary to an underlying condition is attractive, as the preservation of muscle strength for ambulation, personal care and everyday independence is key in reducing morbidity and improving quality of life.

    In terms of cardiovascular disease, the upregulation of myostatin in the cardiomyocytes surrounding an ischemic infarction in sheep was shown in 1999 [12] and myostatin protein and mRNA in skeletal muscle and myocardium were increased in a rat-model of volume overload heart failure [46]. Lenk and colleagues also found that the protein expression of myostatin was increased in the skeletal muscle and myocardium of a murine LAD-ligation heart failure model, which corresponded to later findings in chronic heart failure patients who had elevated levels of myostatin mRNA and protein in muscle biopsies compared to healthy controls [47][48]. The relationship between myostatin levels in the circulatory system and patients suffering from chronic heart failure has been examined by various groups. Increased myostatin levels in HF-patients could be expected, since impaired cardiac output reduces oxygen supply to the vascular bed of muscle tissue and less muscle means less oxygen consumption. As various studies have detected elevated [48][49][50], equal [51] or lower [40][52] levels of the latent and inactivated myostatin complex in the circulatory system, methodological differences in the detection of myostatin (e.g., Western blotting of promyostatin versus immunoassays of full-length myostatin and Liquid chromatography–mass spectrometry (LC–MS)) may account for these fluctuating results [34]. Furthermore, myostatin levels in decompensated chronic HF patients dropped upon compensation therapy, suggesting dynamics and variability in myostatin levels, which are sensitive to therapeutic interventions [53].

    Treating cancer-associated cachexia by means of myostatin inhibition has been another field of interest. As myostatin was elevated in the gastrocnemius muscle of mice inoculated with the Yoshida AH-130 hepatoma [54], targeting the myostatin pathway seemed promising in preventing cancer cachexia. C26-tumor-bearing mice were treated with a soluble receptor of the ActRIIB (sActRIIB), which improved survival and muscle mass without reducing tumor size [55] and by treating the Lewis lung cancer-model with myostatin antibodies, muscular atrophy and loss of muscle force were attenuated [56].

    COPD has been another target of interest due to the muscle wasting, since 30–40% of all people with COPD undergo muscle wasting as a secondary complication to impaired pulmonary function [57]. The link between myostatin and chronic hypoxemia was established in rats exposed to chronic hypoxia, which induced myostatin expression in rat muscle [58], and the increased the expression of myostatin in the vastus lateralis and serum of COPD-patients compared to healthy controls has also been described [58][59]. Later, serum myostatin was found to be significantly elevated in COPD-patients compared to controls but skeletal muscle mass only correlated negatively with serum-myostatin in males [60].

    In CKD, myostatin is elevated in the serum and skeletal muscle of the rat model of CKD, (Cy/+), with increased activation of atrogenic transcription factors in EDL adding insights to the pathophysiology behind muscle wasting in this condition [61].

    4. Myostatin in Response to Exercise

    The effect of exercise on the expression of myostatin has been demonstrated numerous times. In a clinical study where subjects had immobilized a limb for two weeks following exercise rehabilitation, the casting-induced atrophy did not affect myostatin mRNA in muscle biopsies. However, exercise led to downregulated myostatin expression by approximately 48% [62]. These findings indicate that myostatin works in vivo by inhibiting hypertrophy, rather than inducing atrophy. Similar findings in exercise studies have been observed up to 24 h after exercise [63][64] and on protein-level in prediabetic patients performing moderate aerobic exercise for six months [65]. Most interestingly, “the myostatin paradox” was introduced by Kim et al., who in their exercise study discovered a positive correlation between myostatin mRNA and muscle mass [63], whereas the relationship would most intuitively be the opposite if not taking inhibitory factors into consideration. The authors speculate that high levels of myostatin transcripts in muscle might prime the muscles for additional growth.

    5. Preclinical Studies of Myostatin Inhibition in Animal Models of Neuromuscular Disorders

    The potential for the pharmacological regulation of muscular growth had to be explored in animal models of muscular dystrophy, atrophy and muscular regeneration before ultimately turning towards clinical trials in human subjects. We present here an overview of the various ways in which myostatin has been targeted in animal models. As myostatin inhibition has been utilized to examine various physiological processes other than merely muscular regeneration (including cancer survival, bone- and energy metabolism), the following will focus on the bulk of scientific work that describes the effect of myostatin on muscular tissue. Table 1 presents information, if available, on the animal model and genus, the pharmacological compound, muscle morphology, fiber-type specific changes, absolute and specific force amongst glycolytic and oxidative muscles, muscular stress resistance and histopathological improvements. This review is focused in particular on treatment-mediated functional improvements of muscle function, as these are essential for any translation to human clinical trials. Histopathological recovery, muscular growth and the upregulation of desirable growth factors or genes in vitro may be of less importance, as primary outcomes are invariably functional in preclinical studies and the degree of functional improvement ultimately decides whether a treatment will advance in additional preclinical or clinical investigations. Furthermore, increasing the absolute force is of interest to patients and clinicians who are looking for improvements in the activities of daily living, while the scientist will be looking for specific force (force per cross sectional area of a muscle) as an indicator of whether the underlying deficit has been compensated for.

    Table 1. Results of previously published data from various means of myostatin inhibition in animal models.

    Species/Model Compound Muscle Morphology Fiber-Type Specific Changes Absolute Force/
    Specific Force/Glycolytic Absolute Force/
    Specific Force/Oxidative Stress-Induced Force Drop Histopathological Effect of Myostatin Inhibition Reference
    Antibodies Blocking Myostatin
    (BALB/c, C57BL/6)
    JA16, ATA-842, mRK35, YN41, muSRK-015P, GYM-mFc Fiber CSA increased in EDL [66] and Gas
    Increased weight of Gas, TA, Quad and TB, plantaris, Sol [67]
    Increased IIB fiber CSA, no effect on overall composition [68] Increased grip strength [67][69][70]           [66][67][68][69][70][71]
    Mouse/Sgcd−/−, Sgcg−/−
    JA16 EDL: Increased weight and single fiber area [72][73].
    Increase in TA, Quad, Gas [74]
      EDL: increased force EDL: No effect     No effect Sgcd−/−: No improvement in histopathology of TA, EDL, Gas and diaphragm albeit hydroxyproline reduced in TA.
    Fibrosis in diaphragm increased (Sgcd−/− [74]) and decreased (mdx [72])
    Mouse/mdx PF-354 Increase in hindlimb muscle weight of 5 weeks treatment, no effect after 8 weeks.
    No effect/reduction in CSA
      Diaphragm: No effect Diaphragm increased (young)/no effect (old)     No effect Diaphragm: Increased fiber size in young animals, decreased fiber size in old animals [75]
    Mouse/mdx, TgCTA1D286G, Sod1G93A, A17,
    (in SmnΔ7)
    TA, Gas, Quad, EDL, diaphragm weight increased.
    Increased CSA in TA, EDL.
    No effect on weight or CSA in soleus [76]
    Quad: Increased proportion of IIB fibers [77]
    Increase in IIB fiber CSA, no effect in remaining fiber-types [78]
    TA, EDL: Increased force
    Plantarflexor group increased torque [78]
    TA, EDL: No effect
    effect on plantarflexor group [78]
          Gas: reduced atrophy, preserved fiber diameter. Diaphragm integrity preserved [79].
    Reduced collagen I, III, IV deposits. No effect on intranuclear inclusion bodies [76][80].
    Increased number of tubular aggregates [77].
    C57BL/6, (Dexamethasone atrophy)
    REGN1033 Increased weight in Gas and TA. Fiber area increased in Gas No effect on fiber type composition TA: Increased force TA: No effect         [81]
    Monkey/cynomolgus MYO-029,
    Domagrozu-mab, GYM-cyfc
    Increased muscular circumference               [70][71][82]
    Myostatin Propeptide Administration or Overexpression
    Mouse/mdx Recombinant propeptide-Fc EDL: weight, CSA, single fiber area increased   EDL: Increased force EDL: Increased force     No effect Decreased pathological changes [83]
    Mouse/mdx AAV8-
    TA, Quad, Gas, Diaphragm increased   TA: Increased force TA: No effect       Larger fibers, less fibrosis [84]
    Mouse/calpatin 3-null mice (LGMD2A), Sgca−/− (LGMD2D) rAAV2/1mSeAP-propmyoD76A Increased muscle mass in calpain-3-null mice, no effect in Sgca−/−-mice   EDL: increased force (calpain-3-null mice) EDL: No effect
    (calpain-3-null mice)
    Soleus: Increased force
    (calpain-3-null mice)
    Soleus: No effect
    (calpain-3-null mice)
    Soluble Receptor (sActRIIB-Fc)
    Wild-type, C57BL/6
    Increased muscle weight.
    Fiber CSA increased in EDL [86] and in whole TA [87]
    Soleus: Type I and II-fiber CSA increased [88].
    Quad; increased size of I, IIA, IIB-fibers.
    No fiber-type switch [89]
    EDL: twitch force increased, no effect on tetanic force [86].
    Gas: no effect on max tetanic force [90]
    EDL: no effect [91][92]
    Gas: decreased [90]
    Soleus: increased force [91] Soleus: no effect force [91]     [86][87][88][89][90][91][92][93][94][95]
    Mouse/mdx RAP-031,
    Muscle weight increased
    Diaphragm and triceps myofiber increased [96].
    EDL single fiber CSA increased [97].
    No fiber-type conversion [91] EDL: increased force [97][98] EDL: increased force [97],
    No effect [98].
    EDL, TA decreased force in older animals
    Soleus decreased force Soleus decreased force No effect Diaphragm, TA: No effect on histopathology, hydroxyproline [93][97].
    Fibrosis decreased [96].
    No visible effects on H/E pathology. SDH stains without effect of treatment [99].
    eMHC: no effect [93].
    Increased muscle weight, increased fiber size Quad: oxidative fiber diameter increased.
    Diaphragm: glycolytic myofibers hypertrophy [100].
    IIB fiber hypertrophy, no fiber type switch [89][101]
    No effect [100].
    EDL, TA increased force [102]
    No effect [100] No effect [100] No effect [100]   Nemaline rod structures unchanged [100].
    Gross evaluation of diaphragm: unaffected by genotype or treatment [89].
    Fibrotic changes improved
    Anti-ActRIIB Antibody
    Mouse/SCID BYM338 Increased weight of TA, EDL, Gas.
    Soleus increased weight (in high dose) [105]
      Gas: increased force [106]           [105][106]
    Mouse/C57BL/6 (glucocorticoid-induced atrophy) BYM338 TA weight and CSA increased   TA increased force           [105]
    Follistatin Administration or Overexpression
    F66;Dysf−/−, F66;mdx
    Follistatin overexpression Muscle mass maintained in F66;mdx, decreased in F66;Dysf−/−     F66;Dysf−/−: EDL: Decreased force       F66;Dysf−/−: Exacerbation of dystrophic features.
    Increased Evans Blue Dye (EBD) uptake
    F66;mdx: Dystrophic features not exacerbated, mild improvement
    Mouse/mdx, Sod1G93A AAV-delivered follistatin i.m. Increased weight of TA, Gas, Quad, triceps   Increased grip strength         Young mdx: increased myofiber size. Satellite cell markers: no diff
    Old mdx: Fever necrotic fibers and mononuclear infiltrates
    Monkey/Cynomolgus AAV-delivered follistatin i.m Increased fiber size   Quad: Increased force         Myofiber hypertrophy [109]
    Mouse/C57BL10, mdx, ACE-083 Increased CSA, weight   TA: increased force TA: no effect         [110]
    Mouse/C57BL/6 FS-EEE-mFc and FST288-Fc Increased muscle weight               [98][111]
    Mouse/mdx FS-EEE-mFc Increased weight in gas, Quad, triceps, TA   EDL: Increased force EDL: No effect       Decreased necrosis and fibrosis in Quad, no effect in diaphragm [98]
    Liver-mediated Overexpression of Dominant-negative Myostatin (dnMSTN), sActRIIB and Myostatin Propeptide
    Mouse/MF-1 (wild-type) AAV8 over-ekspression (propeptide) Gas, TA increased mass. EDL and soleus increased CSA. Increased CSA of type I, IIA and IIB-fibers EDL: No effect EDL: No effect Soleus: increased force Soleus: No effect     [112]
    Mouse/SmaC/C AAV-mediated systemic expression (dnMSTN and sActRIIB) Increased weight in TA, Gas, Quad.
    dnMSTN-cohort: Increased CSA in EDL and TA but not in soleus
    TA: Increased IIA size
    EDL: Increased IIA and IIB size and total fiber number.
    Soleus: No effect vs. controls.
    I-fibers generally unaffected
    EDL increased vs. SMAC/C control EDL; Decreased force Soleus: Increased force Soleus: No effect     [113]
    Mouse/mdx AAV-delivered liver-specific promoter: dnMSTN, sActRIIB Increased weight in TA, Gas, Quad, EDL, Soleus
    EDL: increased CSA
    Soleus: No effect in weight [114]
    EDL: IA + IIB increased fiber size. Increased proportion of IIB fibers in EDL and Soleus.
    Soleus: Increased size and proportion of IIA-fibers
    Diaphragm: IIX fibers proportion increased, IIA fibers proportion decrease [115]
    Diaphragm: No effect in specific fiber-type size [114]
    EDL: increased force No effect (decreased force by 10 months of treatment) Soleus: increased force Soleus increased force [115]
    Soleus no difference [114].
    Diaphragm: no effect
    Dog/GRMD AAV-delivered liver-specific promoter (dnMSTN) Increased weight in Tibialis cranialis, EDL, Gas, flexor digitorum superficialis Increased size of IIA-fibers, no effect in I-fibers.
    No fiber type switch
    RNA Interference and Anti-oligonucleotides against Myostatin or ActRIIB
    Mouse/mdx Antimyostatin PMO No effect in weight of diaphragm, EDL, Gas, Soleus, TA Diaphragm: no difference in fiber-type content (I, IIA, IIX, IIB)           Diaphragm and TA: no effect on fiber diameter and collagen IV content [118]
    Mouse/mdx (female) AAV-delivered shRNA, i.m. TA: No effect on CSA, fiber number increased   TA: No effect TA: No effect         [119]
    AAV-Cas9-mediated Myostatin Gene Editing
    Mouse/C57/BL10 rAAV-SaCas9   Increased fiber area and number of fibers per area             [78]
    Myostatin Knock-out/Crossbreeding
    Mouse/Mstn−/−   Increased muscle weight vs. wild-type.
    Increased fiber number and CSA of EDL and soleus [120]
    EDL fiber-type composition:
    IIA and IIX incidence decreased, IIB increased in EDL and TA.
    Soleus CSA increased only in IIA-fibers [121]
    Increased [120]/no effect [121][122]
    Soleus: Increased Soleus: No effect EDL: Force deficit
    Soleus: No force deficit
    Decreased hydroxyproline content in EDL, no effect in soleus [120].
    Cytoplasmic inclusions of tubular aggregates in older mice [122]
    Mouse/BehC/C   Increased muscle weight   EDL: No effect [122] EDL: Decreased force [122]         [122][127]
    Mstn−/−;Sgcd −/−
      Increased mean fiber diameter and muscle weight [104][128][129]             Mstn−/−;mdx: Reduced fibrosis [128]
    Mstn −/−;Sgcd −/−:
    Hydroxyproline content decreased in EDL [74]
    Mouse/Mstn−/−; dyW/dyW   Increased muscle mass, muscle CSA and fiber CSA.
    (increased mortality)
    Decreased type I fiber composition           No effect on necrosis, inflammation or infiltrating cells. Less fat tissue. [130]

    Mainstream results from various antimyostatin treatments in animal models. Specific results that were distinct for a particular study and not general for all of the references have been titled as such. Abbreviations: AAV; adeno-associated virus, ActRIIB; activin receptor type IIB, CSA; cross-sectional area, EDL; m. extensor digitorum longus, eMHC; embryonic myosin heavy chain, Gas; m. gastrocnemius, GRMD; golden retriever muscular dystrophy i.m.; intra-muscular injection, LGMD; limb-girdle muscular dystrophy, Quad; m. quadriceps, SDH; succinate dehydrogenase, TA; m. tibialis anterior, TB; m. triceps brachii.

    The pharmacological approaches to inhibiting myostatin activity in vivo have included: (a) systemic administration of antibodies against myostatin; (b) overexpression or administration of the myostatin propeptide; (c) systemic administration of the activin-IIB-receptor itself; (d) administration of antibodies directed against ActRIIB; (e) overexpression or administration of follistatin; (f) liver-mediated overexpression of a soluble receptor (sActRIIB), dominant-negative myostatin (dnMSTN) or the propeptide; (g) RNA interference and antioligonucleotides against myostatin or ActRIIB or; (h) AAV-Cas9 mediated myostatin gene editing. Finally, we have also included works on the effects of transgenic knock-out models and crossbreeding with the preexisting models of muscular dystrophy (i).

    5.1. Antibodies against Myostatin

    Bogdanovich et al. were the first to successfully treat the commonly used mouse model of Duchenne Muscular Dystrophy (DMD), the mdx, with antibodies directed towards myostatin (monoclonal antibodies, JA16) [72]. The results were promising, as the diaphragm and the skeletal muscle, which in the mdx reproduces the pathological features seen in muscles of DMD-patients most accurately [131], showed fewer degenerative features compared to controls. Meanwhile, m. extensor digitorum longus (EDL) had increased weight, cross-sectional area (CSA) and absolute force but failed to show improvement on specific force and stretch resistance. Similar results with increased muscle weight and absolute force but lack of improvement in specific force and resistance were seen in the Sgcg−/− model of limb-girdle muscular dystrophy (LGMD) 2C in a design of similar age and treatment length to the previously mentioned study [73]. The Sgcd−/− mouse model of LGMD2F also treated with JA16 antibodies was not able to improve fibrosis in either young or older Sgcd−/− animals (4 and 20 weeks old at treatment start, respectively) with older animals even showing signs of worsening of fibrosis [74]. Interestingly, a 5-week treatment period of very young (16 days old) mdx-animals showed positive effects on the diaphragm, as specific force increased while absolute force was unaffected, fiber size increased and connective tissue infiltration of the diaphragm was reduced [75], indicating that early initiation of treatment is crucial for a positive effect. Another monoclonal antibody developed by Pfizer, mRK-35, was also able to increase absolute but not specific force in mdx mice [71] and the TgActa1D286G mouse model of nemaline myopathy [77]. Treatment of the Sod1G93A mouse and rat models of amyotrophic lateral sclerosis (ALS) with RK35 improved grip strength compared to placebo controls but did not delay disease onset or extend survival of either model [79]. Later, Muramatsu and colleagues introduced the concept of “sweeping antibody technology” with the GYM329-antibody designed to bind and clear latent myostatin from the circulatory system, which increased muscle mass in three mice models and cynomolgus monkeys and also improved grip strength in the mice [70]. As opposed to other antimyostatin antibodies, GYM329 did not bind GDF-11 and this specificity appears to induce an enhanced effect on muscle mass in treated animals. Especially in older animals, where other myostatin inhibition treatments fail or struggle to achieve an effect, GYM329 appeared superior. Other models of neuromuscular disorders such as the SmnΔ7 mouse of spinal muscle atrophy (SMA) had increased absolute muscle torque but not specific torque after treatment with the muSRK-015P antibody versus myostatin (combined with salvation of Smn2-gene mRNA) [132]. In a study of micro-gravity-induced muscular atrophy, mice were held at the International Space Station and treated with YN41 for 6 weeks, inducing improved grip strength compared to controls, as well as increased muscle mass [67].

    5.2. Myostatin Propeptide Administration or Overexpression

    As previously mentioned, the myostatin propeptide functions as an inhibitor of myostatin, as it binds myostatin in an inactive complex. Propeptide-based inhibition by intraperitoneal injection for three months resulted in increased body mass, EDL mass, absolute and specific force in EDL. There was no effect on stretch-resistance but the histopathological phenotype of the diaphragm improved compared to untreated mdx [83]. Bartoli et al. treated calpain-3-null and Sgca−/−-mouse models of LGMD2A and 2D, respectively, by local and systemic overexpression of the propeptide but were only able to improve the calpain-3-null mice [85].

    5.3. Systemic Administration of the Soluble Receptor ActRIIB

    In order to increase the specific targeting of myostatin and reduce binding of the variety of other ligands that also bind to and activate ActRIIB, another approach based on the systemic administration of a soluble activin type IIB receptor, sActRIIB, has been widely utilized. The compound RAP-031, developed by Acceleron, is a fusion protein consisting of the extracellular domain of the ActRIIB linked to the Fc-portion of murine IgG to delay systemic clearance. Applying this approach, Pistilli and colleagues demonstrated an increase in both absolute and specific force of EDL in mdx mice [97], a functional outcome, which unfortunately has been difficult to replicate in both wild-type [86][92], mdx [91][93][98], and nemaline myopathy mouse models [100]. The treatment of mice with muscular atrophy due to spinal cord injury with RAP-031 did not alleviate the atrophy [133]. A hypoxia model in wild-type mice showed improved resistance to eccentric lengthening but no other studies using the soluble receptor have shown improvements to stretch resistance [134]. The specific hypertrophy of fibers with a IIB fiber-type composition was observed in two models of myotubularin-deficient mice [89][101] but also in other fibers of wild-type animals [88][89].

    5.4. Administration of Antibodies Directed against ActRIIB

    Blocking the ActRIIB itself by antibodies has not been widely used as another means of myostatin inhibition. Novartis developed BYM338 (bimagrumab, which would progress into clinical trials as mentioned below) and described the receptor-specificity in cell cultures and myoblasts while also showing the effects on body and muscle mass in both SCID-mice and a glucocorticoid atrophy model [105].

    5.5. Follistatin Administration or Overexpression

    Like the myostatin propeptide, follistatin is able to inhibit not only myostatin but also shows affinity for other TGF-β-family members (such as BMPs and activins) [22][135]. Transgenic overexpression of follistatin primarily showed increased muscle weight and fiber diameter [17]. Transgenic mice overexpressing a follistatin-derived myostatin inhibitor crossed with the mdx ameliorated the dystrophic features in terms of grip strength and pathohistological features [136]. When transgenic overexpression of follistatin (F66-mice) is crossed with the dysferlinopathy LGMD2B model Dysf−/−, the positive effect on muscle weight in F66;Dysf−/−-mice declines with age and the specific force of EDL is reduced, compared to F66-mice, exacerbating the dystrophic phenotype [103]. Furthermore, ActRIIB-FC-administration in Dysf−/−-mice ameliorated histopathological changes, but increased creatine kinase (CK, a marker of muscular damage and membrane integrity) levels. The authors conclude that follistatin overexpression accelerated the degenerative features in the dysferlinopathy model, as the dystrophin-deficient mdx was not exacerbated [103], and suggest that muscle hypertrophy may have pernicious effects depending on the disease context.

    Another approach using a follistatin-based fusion protein ACE-083 by local intramuscular injections increased CSA, weight and absolute, but not specific, force of injected muscle tibialis anterior (TA) in the Trembler-J mouse model of Charcot–Marie–Tooth disease and mdx [110].

    As the systemic clearance of follistatin is rather quick, systemic versus local administration poses a challenge. Thus, the pharmacokinetic properties were edited and a long-acting follistatin-based molecule (FS-EEE-hFc) was engineered by Shen and colleagues [137] and applied by intravenous and subcutaneous administration to wild-type and mdx-animals [98]. The subcutaneous treatment of young (4 weeks) mdx-mice for 12 weeks also undergoing an exhaustion-exercise regime showed increased muscle weight and absolute but not specific force increments [98].

    5.6. Liver-Mediated Overexpression of Dominant-Negative Myostatin (dnMSTN), sActRIIB and Myostatin Propeptide

    Using the same approach as mentioned earlier with adeno-associated virus 8 (AAV8)-delivered myostatin inhibitors, Morine and colleagues treated the mdx with AAV-vectors, which brought liver-mediated transcripts of sActRIIB [114] or dnMSTN [115] into circulation. The sActRIIB treatment did increase the muscle mass, fiber size and absolute force of the EDL, while CK decreased. However, there were no positive effects in soleus or specific force [114]. The dnMSTN paper showed that the treatment in mdx-mice was predominantly observed in the fast fibers (IIA, IIX and IIB) of both the EDL and soleus, while soleus increased both absolute and specific force and CK decreased [115]. A similar study in the D2.mdx only reported beneficial effects on absolute force in EDL [116].

    Similar to the treatment regimes in the mdx, Liu and colleagues treated the C/C mouse model of spinal muscle atrophy (SMA) with AAV8-vectors containing transcripts for dnMSTN and sActRIIB, respectively [113]. Both treatments increased the size of type IIA and IIB-fibers, leaving type I-fibers unaffected (IIX was not measured). While specific force was unaffected by treatment, absolute force increased in EDL (both treatments) and soleus (only sActRIIB).

    Another approach was used in wild-type MF-1 mice, where propeptide coupled to an immunoglobulin Fc molecule was delivered by means of AAV8 vectors to hepatocytes, ensuring an intrinsic production of the inhibitor [112]. In contrast to exogenic injections of the propeptide, Foster and colleagues treated mice from six weeks of age and found an increased absolute force in oxidative muscle soleus but not in EDL. Both EDL and soleus increased CSA, as well as subanalyses of fiber-types I, IIA and IIB. In a similar design, mdx-mice were treated at the age of three months, which increased body mass, grip strength, muscle mass and fiber radius [84]. The absolute twitch and tetanic force production improved but specific force did not.

    5.7. RNA Interference and Antioligonucleotides against Myostatin or ActRIIB

    Myostatin has also been sought downregulated by means of RNA interference. Dumonceaux et al. combined short hairpin RNA (shRNA) interference of ActRIIB mRNA with AAV mediated exon-skipping of dystrophin. The number of fibers increased in TA, but force production was unchanged in mice that received myostatin interference solely compared to untreated mdx [119].

    In contrast to AAV-mediated gene therapy, antisense oligomers (AOs) hold no risk of uncontrolled genome insertion and levels of exon skipping can be regulated or aborted over time. Antisense phosphorodiamidate morpholine oligomers (PMOs) causing exon-skipping of myostatin increased TA weight and CSA locally in mdx-mice [138]. A follow up study combining systemic treatment with two different PMOs that restored dystrophin and inhibited myostatin, respectively, was promising but the mdx mice receiving the myostatin-inhibiting PMO did not benefit from this treatment alone [118]. A similar study demonstrated similar increases in muscle mass in PMO-skipped myostatin, but also demonstrated that skipping varied among muscles, with the highest level of skipping in the soleus. These studies emphasize the importance of the design of the PMO, as well as the variable results obtained in healthy and mdx animals, suggesting that histopathology plays a role in efficiency of the treatment [118][138].

    5.8. AAV-Cas9-Mediated Myostatin Gene Knock-Down

    Recently, it was demonstrated that myostatin knock-out by the means of AAV-Sa-Cas9 gene editing delivered by intramuscular injections increased fiber area and number of fibers per area in aged wild-type mice [78]. However, functional outcomes were not described.

    5.9. Crossbreeding Transgenic Myostatin Knock-Out Animals

    The murine hypermuscular myostatin knockout (Mstn−/−, also denominated ‘the myostatin-null’) described in 1997, has subsequently been further examined and crossed with various mouse models of neuromuscular diseases. The myostatin-null itself has been described numerous times [1][122][126] with increased muscle and body mass. Force measurements have shown both positive and no effect on absolute force in the myostatin knock-outs but decreased specific force has generally been reported [120][121][122]. An increased proportion of fast fiber-types has been the common observation [122][123][124][125][126], in line with the findings in studies of pharmacological myostatin inhibition (see above). Another model of myostatin malfunction includes the Compact-mouse (also known as the Berlin High Line BEHC/C), which contains a 12-bp deletion in the propeptide domain of promyostatin (MstnCmpt-dl1Abc) but leaves the biologically active growth-factor domain of myostatin unaffected [139]. Kocsis and colleagues later found that the Compact genetic background itself, in addition to the promyostatin genetic deletion, determines the phenotype [127] and the use of this model has been rather limited.

    A third mouse model is the lean myostatin mouse (Mstnln/ln), which has an induced loss-of-function mutation leading to a peptide without the ligand, thus a complete lack of myostatin. This model has similarly increased muscularity but has had most of its use in the field of metabolic research [140].

    Crossing myostatin-null with other models of muscular dystrophy has occasionally been the preceding study to pharmacological interventions. Crossing myostatin-null with mdx [128][129] or caveolin-3-deficient mice with transgenic mice overexpressing the myostatin prodomain (“MSTNPro”) [104] did ameliorate the pathological features by increasing body weight, fiber numbers and improving grip strength. However, the crossing of myostatin-null mice with the dyW/dyW laminin-deficient mouse model of congenital muscular dystrophy failed to improve the dystrophic phenotype and postnatal lethality was even increased [130].

    In addition, a recent study crossing a follistatin overexpressing mouse strain with the calpain 3 knock-out mouse model for LGMD2A led to increased glycolytic muscle mass, but caused the loss of AMP-activated protein kinase signaling, important for contraction-induced glycolysis and poor exercise tolerance [141].

    The entry is from 10.3390/cells10030533


    1. McPherron, A.C.; Lawler, A.M.; Lee, S.-J. Regulation of Skeletal Muscle Mass in Mice by a New TGF-p Superfamily Member. Nature 1997, 387, 83–90.
    2. Grobet, L.; Martin, L.J.; Poncelet, D.; Pirottin, D.; Brouwers, B.; Riquet, J.; Schoeberlein, A.; Dunner, S.; Ménissier, F.; Massabanda, J.; et al. A Deletion in the Bovine Myostatin Gene Causes the Double-Muscled Phenotype in Cattle. Nat. Genet. 1997, 17, 71–74.
    3. Kambadur, R.; Sharma, M.; Smith, T.P.; Bass, J.J. Mutations in Myostatin (GDF8) in Double-Muscled Belgian Blue and Piedmontese Cattle. Genome Res. 1997, 7, 910–916.
    4. McPherron, A.C.; Lee, S.J. Double Muscling in Cattle Due to Mutations in the Myostatin Gene. Proc. Natl. Acad. Sci. USA 1997, 94, 12457–12461.
    5. Mosher, D.S.; Quignon, P.; Bustamante, C.D.; Sutter, N.B.; Mellersh, C.S.; Parker, H.G.; Ostrander, E.A. A Mutation in the Myostatin Gene Increases Muscle Mass and Enhances Racing Performance in Heterozygote Dogs. PLoS Genet. 2007, 3.
    6. Schuelke, M.; Wagner, K.R.; Stolz, L.E.; Hübner, C.; Riebel, T.; Kömen, W.; Braun, T.; Tobin, J.F.; Lee, S.-J. Myostatin Mutation Associated with Gross Muscle Hypertrophy in a Child. N. Engl. J. Med. 2004, 350, 2682–2688.
    7. Mendias, C.L.; Bakhurin, K.I.; Gumucio, J.P.; Shallal-Ayzin, M.V.; Davis, C.S.; Faulkner, J.A. Haploinsufficiency of Myostatin Protects against Aging-Related Declines in Muscle Function and Enhances the Longevity of Mice. Aging Cell 2015, 14, 704–706.
    8. Morissette, M.R.; Stricker, J.C.; Rosenberg, M.A.; Buranasombati, C.; Levitan, E.B.; Mittleman, M.A.; Rosenzweig, A. Effects of Myostatin Deletion in Aging Mice. Aging Cell 2009, 8, 573–583.
    9. Amthor, H.; Huang, R.; McKinnell, I.; Christ, B.; Kambadur, R.; Sharma, M.; Patel, K. The Regulation and Action of Myostatin as a Negative Regulator of Muscle Development during Avian Embryogenesis. Dev. Biol. 2002, 251, 241–257.
    10. Manceau, M.; Gros, J.; Savage, K.; Thomé, V.; McPherron, A.; Paterson, B.; Marcelle, C. Myostatin Promotes the Terminal Differentiation of Embryonic Muscle Progenitors. Genes Dev. 2008, 22, 668–681.
    11. Hill, J.J.; Davies, M.V.; Pearson, A.A.; Wang, J.H.; Hewick, R.M.; Wolfman, N.M.; Qiu, Y. The Myostatin Propeptide and the Follistatin-Related Gene Are Inhibitory Binding Proteins of Myostatin in Normal Serum. J. Biol. Chem. 2002, 277, 40735–40741.
    12. Sharma, M.; Kambadur, R.; Matthews, K.G.; Somers, W.G.; Devlin, G.P.; Conaglen, J.V.; Fowke, P.J.; Bass, J.J. Myostatin, a Transforming Growth Factor-β Superfamily Member, Is Expressed in Heart Muscle and Is Upregulated in Cardiomyocytes after Infarct. J. Cell. Physiol. 1999, 180, 1–9.
    13. Gonzalez-Cadavid, N.F.; Taylor, W.E.; Yarasheski, K.; Sinha-Hikim, I.; Ma, K.; Ezzat, S.; Shen, R.; Lalani, R.; Asa, S.; Mamita, M.; et al. Organization of the Human Myostatin Gene and Expression in Healthy Men and HIV-Infected Men with Muscle Wasting. Proc. Natl. Acad. Sci. USA 1998, 95, 14938–14943.
    14. Rodgers, B.D.; Weber, G.M. Sequence Conservation among Fish Myostatin Orthologues and the Characterization of Two Additional CDNA Clones from Morone Saxatilis and Morone Americana. Comp. Biochem. Physiol. Bbiochem. Mol. Biol. 2001, 129, 597–603.
    15. Smith, T.P.; Lopez-Corrales, N.L.; Kappes, S.M.; Sonstegard, T.S. Myostatin Maps to the Interval Containing the Bovine Mh Locus. Mamm. Genome 1997, 8, 742–744.
    16. Stavaux, D.; Art, T.; McEntee, K.; Reznick, M.; Lekeux, P. Muscle Fibre Type and Size, and Muscle Capillary Density in Young Double-Muscled Blue Belgian Cattle. Zent. Vet. A 1994, 41, 229–236.
    17. Lee, S.-J.; McPherron, A.C. Regulation of Myostatin Activity and Muscle Growth. Proc. Natl. Acad. Sci. USA 2001, 98, 9306–9311.
    18. Thies, R.S.; Chen, T.; Davies, M.V.; Tomkinson, K.N.; Pearson, A.A.; Shakey, Q.A.; Wolfman, N.M. GDF-8 Propeptide Binds to GDF-8 and Antagonizes Biological Activity by Inhibiting GDF-8 Receptor Binding. Growth Factors 2001, 18, 251–259.
    19. Wolfman, N.M.; McPherron, A.C.; Pappano, W.N.; Davies, M.V.; Song, K.; Tomkinson, K.N.; Wright, J.F.; Zhao, L.; Sebald, S.M.; Greenspan, D.S.; et al. Activation of Latent Myostatin by the BMP-1/Tolloid Family of Metalloproteinases. Proc. Natl. Acad. Sci. USA 2003, 100, 15842–15846.
    20. Lee, S.-J. Genetic Analysis of the Role of Proteolysis in the Activation of Latent Myostatin. PLoS ONE 2008, 3.
    21. Amthor, H.; Nicholas, G.; McKinnell, I.; Kemp, C.F.; Sharma, M.; Kambadur, R.; Patel, K. Follistatin Complexes Myostatin and Antagonises Myostatin-Mediated Inhibition of Myogenesis. Dev. Biol. 2004, 270, 19–30.
    22. Hill, J.J.; Qiu, Y.; Hewick, R.M.; Wolfman, N.M. Regulation of Myostatin in Vivo by Growth and Differentiation Factor-Associated Serum Protein-1: A Novel Protein with Protease Inhibitor and Follistatin Domains. Mol. Endocrinol. 2003, 17, 1144–1154.
    23. Miura, T.; Kishioka, Y.; Wakamatsu, J.; Hattori, A.; Hennebry, A.; Berry, C.J.; Sharma, M.; Kambadur, R.; Nishimura, T. Decorin Binds Myostatin and Modulates Its Activity to Muscle Cells. Biochem. Biophys. Res. Commun. 2006, 340, 675–680.
    24. Zhu, J.; Li, Y.; Shen, W.; Qiao, C.; Ambrosio, F.; Lavasani, M.; Nozaki, M.; Branca, M.F.; Huard, J. Relationships between Transforming Growth Factor-Β1, Myostatin, and Decorin Implications for Skeletal Muscle Fibrosis. J. Biol. Chem. 2007, 282, 25852–25863.
    25. Kemaladewi, D.U.; de Gorter, D.J.J.; Aartsma-Rus, A.; van Ommen, G.-J.; ten Dijke, P.; ’t Hoen, P.A.C.; Hoogaars, W.M. Cell-Type Specific Regulation of Myostatin Signaling. FASEB J. 2012, 26, 1462–1472.
    26. Rebbapragada, A.; Benchabane, H.; Wrana, J.L.; Celeste, A.J.; Attisano, L. Myostatin Signals through a Transforming Growth Factor β-Like Signaling Pathway To Block Adipogenesis. Mol. Cell Biol. 2003, 23, 7230–7242.
    27. Derynck, R.; Zhang, Y.E. Smad-Dependent and Smad-Independent Pathways in TGF-Beta Family Signalling. Nature 2003, 425, 577–584.
    28. Langley, B.; Thomas, M.; Bishop, A.; Sharma, M.; Gilmour, S.; Kambadur, R. Myostatin Inhibits Myoblast Differentiation by Down-Regulating MyoD Expression. J. Biol. Chem. 2002, 277, 49831–49840.
    29. Thomas, M.; Langley, B.; Berry, C.; Sharma, M.; Kirk, S.; Bass, J.; Kambadur, R. Myostatin, a Negative Regulator of Muscle Growth, Functions by Inhibiting Myoblast Proliferation. J. Biol. Chem. 2000, 275, 40235–40243.
    30. Sartori, R.; Schirwis, E.; Blaauw, B.; Bortolanza, S.; Zhao, J.; Enzo, E.; Stantzou, A.; Mouisel, E.; Toniolo, L.; Ferry, A.; et al. BMP Signaling Controls Muscle Mass. Nat. Genet. 2013, 45, 1309–1318.
    31. Biesemann, N.; Mendler, L.; Wietelmann, A.; Hermann, S.; Schäfers, M.; Krüger, M.; Boettger, T.; Borchardt, T.; Braun, T. Myostatin Regulates Energy Homeostasis in the Heart and Prevents Heart Failure. Circ. Res. 2014, 115, 296–310.
    32. Philip, B.; Lu, Z.; Gao, Y. Regulation of GDF-8 Signaling by the P38 MAPK. Cell Signal. 2005, 17, 365–375.
    33. Ratkevicius, A.; Joyson, A.; Selmer, I.; Dhanani, T.; Grierson, C.; Tommasi, A.M.; DeVries, A.; Rauchhaus, P.; Crowther, D.; Alesci, S.; et al. Serum Concentrations of Myostatin and Myostatin-Interacting Proteins Do Not Differ between Young and Sarcopenic Elderly Men. J. Gerontol. A Biol. Sci. Med. Sci. 2011, 66, 620–626.
    34. Burch, P.M.; Pogoryelova, O.; Palandra, J.; Goldstein, R.; Bennett, D.; Fitz, L.; Guglieri, M.; Bettolo, C.M.; Straub, V.; Evangelista, T.; et al. Reduced Serum Myostatin Concentrations Associated with Genetic Muscle Disease Progression. J. Neurol. 2017, 264, 541–553.
    35. Szulc, P.; Schoppet, M.; Goettsch, C.; Rauner, M.; Dschietzig, T.; Chapurlat, R.; Hofbauer, L.C. Endocrine and Clinical Correlates of Myostatin Serum Concentration in Men--the STRAMBO Study. J. Clin. Endocrinol Metab. 2012, 97, 3700–3708.
    36. Lakshman, K.M.; Bhasin, S.; Corcoran, C.; Collins-Racie, L.A.; Tchistiakova, L.; Forlow, S.B.; St Ledger, K.; Burczynski, M.E.; Dorner, A.J.; Lavallie, E.R. Measurement of Myostatin Concentrations in Human Serum: Circulating Concentrations in Young and Older Men and Effects of Testosterone Administration. Mol. Cell Endocrinol. 2009, 302, 26–32.
    37. Schafer, M.J.; Atkinson, E.J.; Vanderboom, P.M.; Kotajarvi, B.; White, T.A.; Moore, M.M.; Bruce, C.J.; Greason, K.L.; Suri, R.M.; Khosla, S.; et al. Quantification of GDF11 and Myostatin in Human Aging and Cardiovascular Disease. Cell Metab. 2016, 23, 1207–1215.
    38. Raue, U.; Slivka, D.; Jemiolo, B.; Hollon, C.; Trappe, S. Myogenic Gene Expression at Rest and after a Bout of Resistance Exercise in Young (18–30 Yr) and Old (80–89 Yr) Women. J. Appl. Physiol. 2006, 101, 53–59.
    39. Chew, J.; Tay, L.; Lim, J.P.; Leung, B.P.; Yeo, A.; Yew, S.; Ding, Y.Y.; Lim, W.S. Serum Myostatin and IGF-1 as Gender-Specific Biomarkers of Frailty and Low Muscle Mass in Community-Dwelling Older Adults. J. Nutr. Health Aging 2019, 23, 979–986.
    40. Christensen, H.M.; Kistorp, C.; Schou, M.; Keller, N.; Zerahn, B.; Frystyk, J.; Schwarz, P.; Faber, J. Prevalence of Cachexia in Chronic Heart Failure and Characteristics of Body Composition and Metabolic Status. Endocrine 2013, 43, 626–634.
    41. Peng, L.-N.; Lee, W.-J.; Liu, L.-K.; Lin, M.-H.; Chen, L.-K. Healthy Community-Living Older Men Differ from Women in Associations between Myostatin Levels and Skeletal Muscle Mass. J. Cachexia Sarcopenia Muscle 2018, 9, 635–642.
    42. Bagheri, R.; Moghadam, B.H.; Church, D.D.; Tinsley, G.M.; Eskandari, M.; Moghadam, B.H.; Motevalli, M.S.; Baker, J.S.; Robergs, R.A.; Wong, A. The Effects of Concurrent Training Order on Body Composition and Serum Concentrations of Follistatin, Myostatin and GDF11 in Sarcopenic Elderly Men. Exp. Gerontol. 2020, 133, 110869.
    43. Negaresh, R.; Ranjbar, R.; Baker, J.; Habibi, A.; Mokhtarzade, M.; Gharibvand, M.; Fokin, A. Skeletal Muscle Hypertrophy, Insulin-like Growth Factor 1, Myostatin and Follistatin in Healthy and Sarcopenic Elderly Men: The Effect of Whole-Body Resistance Training. Int. J. Prev. Med. 2019, 10, 29.
    44. Zimmers, T.A.; Davies, M.V.; Koniaris, L.G.; Haynes, P.; Esquela, A.F.; Tomkinson, K.N.; McPherron, A.C.; Wolfman, N.M.; Lee, S.-J. Induction of Cachexia in Mice by Systemically Administered Myostatin. Science 2002, 296, 1486–1488.
    45. Lenk, K.; Schuler, G.; Adams, V. Skeletal Muscle Wasting in Cachexia and Sarcopenia: Molecular Pathophysiology and Impact of Exercise Training. J. Cachexia Sarcopenia Muscle 2010, 1, 9–21.
    46. Shyu, K.G.; Lu, M.J.; Wang, B.W.; Sun, H.Y.; Chang, H. Myostatin Expression in Ventricular Myocardium in a Rat Model of Volume-Overload Heart Failure. Eur. J. Clin. Investig. 2006, 36, 713–719.
    47. Lenk, K.; Schur, R.; Linke, A.; Erbs, S.; Matsumoto, Y.; Adams, V.; Schuler, G. Impact of Exercise Training on Myostatin Expression in the Myocardium and Skeletal Muscle in a Chronic Heart Failure Model. Eur. J. Heart Fail. 2009, 11, 342–348.
    48. Lenk, K.; Erbs, S.; Höllriegel, R.; Beck, E.; Linke, A.; Gielen, S.; Winkler, S.M.; Sandri, M.; Hambrecht, R.; Schuler, G.; et al. Exercise Training Leads to a Reduction of Elevated Myostatin Levels in Patients with Chronic Heart Failure. Eur. J. Prev. Cardiol. 2012, 19, 404–411.
    49. George, I.; Bish, L.T.; Kamalakkannan, G.; Petrilli, C.M.; Oz, M.C.; Naka, Y.; Lee Sweeney, H.; Maybaum, S. Myostatin Activation in Patients with Advanced Heart Failure and after Mechanical Unloading. Eur. J. Heart Fail. 2010, 12, 444–453.
    50. Gruson, D.; Ahn, S.A.; Ketelslegers, J.-M.; Rousseau, M.F. Increased Plasma Myostatin in Heart Failure. Eur. J. Heart Fail. 2011, 13, 734–736.
    51. Zamora, E.; Simó, R.; Lupón, J.; Galán, A.; Urrutia, A.; González, B.; Mas, D.; Valle, V. Serum Myostatin Levels in Chronic Heart Failure. Rev. Esp. Cardiol. 2010, 63, 992–996.
    52. Furihata, T.; Kinugawa, S.; Fukushima, A.; Takada, S.; Homma, T.; Masaki, Y.; Abe, T.; Yokota, T.; Oba, K.; Okita, K.; et al. Serum Myostatin Levels Are Independently Associated with Skeletal Muscle Wasting in Patients with Heart Failure. Int. J. Cardiol. 2016, 220, 483–487.
    53. Wintgens, K.F.; Dschietzig, T.; Stoeva, S.; Paulsson, M.; Armbruster, F.P. Plasma Myostatin Measured by a Competitive ELISA Using a Highly Specific Antiserum. Clin. Chim. Acta 2012, 413, 1288–1294.
    54. Costelli, P.; Muscaritoli, M.; Bonetto, A.; Penna, F.; Reffo, P.; Bossola, M.; Bonelli, G.; Doglietto, G.B.; Baccino, F.M.; Fanelli, F.R. Muscle Myostatin Signalling Is Enhanced in Experimental Cancer Cachexia. Eur. J. Clin. Investig. 2008, 38, 531–538.
    55. Zhou, X.; Wang, J.L.; Lu, J.; Song, Y.; Kwak, K.S.; Jiao, Q.; Rosenfeld, R.; Chen, Q.; Boone, T.; Simonet, W.S.; et al. Reversal of Cancer Cachexia and Muscle Wasting by ActRIIB Antagonism Leads to Prolonged Survival. Cell 2010, 142, 531–543.
    56. Murphy, K.T.; Chee, A.; Gleeson, B.G.; Naim, T.; Swiderski, K.; Koopman, R.; Lynch, G.S. Antibody-Directed Myostatin Inhibition Enhances Muscle Mass and Function in Tumor-Bearing Mice. Am. J. Physiol. Regul. Integr. Comp. Physiol. 2011, 301, R716–726.
    57. Sharma, M.; McFarlane, C.; Kambadur, R.; Kukreti, H.; Bonala, S.; Srinivasan, S. Myostatin: Expanding Horizons: Myostatin. Iubmb Life 2015, 67, 589–600.
    58. Hayot, M.; Rodriguez, J.; Vernus, B.; Carnac, G.; Jean, E.; Allen, D.; Goret, L.; Obert, P.; Candau, R.; Bonnieu, A. Myostatin Up-Regulation Is Associated with the Skeletal Muscle Response to Hypoxic Stimuli. Mol. Cell. Endocrinol. 2011, 332, 38–47.
    59. Plant, P.J.; Brooks, D.; Faughnan, M.; Bayley, T.; Bain, J.; Singer, L.; Correa, J.; Pearce, D.; Binnie, M.; Batt, J. Cellular Markers of Muscle Atrophy in Chronic Obstructive Pulmonary Disease. Am. J. Respir. Cell Mol. Biol. 2010, 42, 461–471.
    60. Ju, C.-R.; Chen, R.-C. Serum Myostatin Levels and Skeletal Muscle Wasting in Chronic Obstructive Pulmonary Disease. Respir. Med. 2012, 106, 102–108.
    61. Avin, K.G.; Chen, N.X.; Organ, J.M.; Zarse, C.; O’Neill, K.; Conway, R.G.; Konrad, R.J.; Bacallao, R.L.; Allen, M.R.; Moe, S.M. Skeletal Muscle Regeneration and Oxidative Stress Are Altered in Chronic Kidney Disease. PLoS ONE 2016, 11, e0159411.
    62. Jones, S.W.; Hill, R.J.; Krasney, P.A.; O’Conner, B.; Peirce, N.; Greenhaff, P.L. Disuse Atrophy and Exercise Rehabilitation in Humans Profoundly Affects the Expression of Genes Associated with the Regulation of Skeletal Muscle Mass. FASEB J. 2004, 18, 1025–1027.
    63. Kim, J.; Cross, J.M.; Bamman, M.M. Impact of Resistance Loading on Myostatin Expression and Cell Cycle Regulation in Young and Older Men and Women. Am. J. Physiol.-Endocrinol. Metab. 2005, 288, E1110–E1119.
    64. Louis, E.; Raue, U.; Yang, Y.; Jemiolo, B.; Trappe, S. Time Course of Proteolytic, Cytokine, and Myostatin Gene Expression after Acute Exercise in Human Skeletal Muscle. J. Appl. Physiol. 2007, 103, 1744–1751.
    65. Hittel, D.S.; Axelson, M.; Sarna, N.; Shearer, J.; Huffman, K.M.; Kraus, W.E. Myostatin Decreases with Aerobic Exercise and Associates with Insulin Resistance. Med. Sci. Sports Exerc. 2010, 42, 2023–2029.
    66. Whittemore, L.-A.; Song, K.; Li, X.; Aghajanian, J.; Davies, M.; Girgenrath, S.; Hill, J.J.; Jalenak, M.; Kelley, P.; Knight, A.; et al. Inhibition of Myostatin in Adult Mice Increases Skeletal Muscle Mass and Strength. Biochem. Biophys. Res. Commun. 2003, 300, 965–971.
    67. Smith, R.C.; Cramer, M.S.; Mitchell, P.J.; Lucchesi, J.; Ortega, A.M.; Livingston, E.W.; Ballard, D.; Zhang, L.; Hanson, J.; Barton, K.; et al. Inhibition of Myostatin Prevents Microgravity-Induced Loss of Skeletal Muscle Mass and Strength. PLoS ONE 2020, 15.
    68. Pirruccello-Straub, M.; Jackson, J.; Wawersik, S.; Webster, M.T.; Salta, L.; Long, K.; McConaughy, W.; Capili, A.; Boston, C.; Carven, G.J.; et al. Blocking Extracellular Activation of Myostatin as a Strategy for Treating Muscle Wasting. Sci. Rep. 2018, 8.
    69. Camporez, J.-P.G.; Petersen, M.C.; Abudukadier, A.; Moreira, G.V.; Jurczak, M.J.; Friedman, G.; Haqq, C.M.; Petersen, K.F.; Shulman, G.I. Anti-Myostatin Antibody Increases Muscle Mass and Strength and Improves Insulin Sensitivity in Old Mice. Proc. Natl. Acad. Sci. USA 2016, 113, 2212–2217.
    70. Muramatsu, H.; Kuramochi, T.; Katada, H.; Ueyama, A.; Ruike, Y.; Ohmine, K.; Shida-Kawazoe, M.; Miyano-Nishizawa, R.; Shimizu, Y.; Okuda, M.; et al. Novel Myostatin-Specific Antibody Enhances Muscle Strength in Muscle Disease Models. Sci. Rep. 2021, 11, 2160.
    71. St. Andre, M.; Johnson, M.; Bansal, P.N.; Wellen, J.; Robertson, A.; Opsahl, A.; Burch, P.M.; Bialek, P.; Morris, C.; Owens, J. A Mouse Anti-Myostatin Antibody Increases Muscle Mass and Improves Muscle Strength and Contractility in the Mdx Mouse Model of Duchenne Muscular Dystrophy and Its Humanized Equivalent, Domagrozumab (PF-06252616), Increases Muscle Volume in Cynomolgus Monkeys. Skelet. Muscle 2017, 7, 25.
    72. Bogdanovich, S.; Krag, T.O.B.; Barton, E.R.; Morris, L.D.; Whittemore, L.-A.; Ahima, R.S.; Khurana, T.S. Functional Improvement of Dystrophic Muscle by Myostatin Blockade. Nature 2002, 420, 418–421.
    73. Bogdanovich, S.; McNally, E.M.; Khurana, T.S. Myostatin Blockade Improves Function but Not Histopathology in a Murine Model of Limb-Girdle Muscular Dystrophy 2C. Muscle Nerve 2008, 37, 308–316.
    74. Parsons, S.A.; Millay, D.P.; Sargent, M.A.; McNally, E.M.; Molkentin, J.D. Age-Dependent Effect of Myostatin Blockade on Disease Severity in a Murine Model of Limb-Girdle Muscular Dystrophy. Am. J. Pathol. 2006, 168, 1975–1985.
    75. Murphy, K.T.; Ryall, J.G.; Snell, S.M.; Nair, L.; Koopman, R.; Krasney, P.A.; Ibebunjo, C.; Holden, K.S.; Loria, P.M.; Salatto, C.T.; et al. Antibody-Directed Myostatin Inhibition Improves Diaphragm Pathology in Young but Not Adult Dystrophic Mdx Mice. Am. J. Pathol. 2010, 176, 2425–2434.
    76. Harish, P.; Malerba, A.; Lu-Nguyen, N.; Forrest, L.; Cappellari, O.; Roth, F.; Trollet, C.; Popplewell, L.; Dickson, G. Inhibition of Myostatin Improves Muscle Atrophy in Oculopharyngeal Muscular Dystrophy (OPMD). J. Cachexia Sarcopenia Muscle 2019, 10, 1016–1026.
    77. Tinklenberg, J.A.; Siebers, E.M.; Beatka, M.J.; Meng, H.; Yang, L.; Zhang, Z.; Ross, J.A.; Ochala, J.; Morris, C.; Owens, J.M.; et al. Myostatin Inhibition Using MRK35 Produces Skeletal Muscle Growth and Tubular Aggregate Formation in Wild Type and TgACTA1D286G Nemaline Myopathy Mice. Hum. Mol. Genet. 2018, 27, 638–648.
    78. Weng, S.; Gao, F.; Wang, J.; Li, X.; Chu, B.; Wang, J.; Yang, G. Improvement of Muscular Atrophy by AAV-SaCas9-Mediated Myostatin Gene Editing in Aged Mice. Cancer Gene 2020, 27, 960–975.
    79. Holzbaur, E.L.F.; Howland, D.S.; Weber, N.; Wallace, K.; She, Y.; Kwak, S.; Tchistiakova, L.A.; Murphy, E.; Hinson, J.; Karim, R.; et al. Myostatin Inhibition Slows Muscle Atrophy in Rodent Models of Amyotrophic Lateral Sclerosis. Neurobiol. Dis. 2006, 23, 697–707.
    80. Harish, P.; Forrest, L.; Herath, S.; Dickson, G.; Malerba, A.; Popplewell, L. Inhibition of Myostatin Reduces Collagen Deposition in a Mouse Model of Oculopharyngeal Muscular Dystrophy (OPMD) With Established Disease. Front. Physiol. 2020, 11, 184.
    81. Latres, E.; Pangilinan, J.; Miloscio, L.; Bauerlein, R.; Na, E.; Potocky, T.B.; Huang, Y.; Eckersdorff, M.; Rafique, A.; Mastaitis, J.; et al. Myostatin Blockade with a Fully Human Monoclonal Antibody Induces Muscle Hypertrophy and Reverses Muscle Atrophy in Young and Aged Mice. Skelet Muscle 2015, 5.
    82. Singh, P.; Rong, H.; Gordi, T.; Bosley, J.; Bhattacharya, I. Translational Pharmacokinetic/Pharmacodynamic Analysis of MYO-029 Antibody for Muscular Dystrophy. Clin. Transl. Sci. 2016, 9, 302–310.
    83. Bogdanovich, S.; Perkins, K.J.; Krag, T.O.B.; Whittemore, L.-A.; Khurana, T.S. Myostatin Propeptide-Mediated Amelioration of Dystrophic Pathophysiology. FASEB J. 2005, 19, 543–549.
    84. Qiao, C.; Li, J.; Jiang, J.; Zhu, X.; Wang, B.; Li, J.; Xiao, X. Myostatin Propeptide Gene Delivery by Adeno-Associated Virus Serotype 8 Vectors Enhances Muscle Growth and Ameliorates Dystrophic Phenotypes in Mdx Mice. Hum. Gene Ther. 2008, 19, 241–254.
    85. Bartoli, M.; Poupiot, J.; Vulin, A.; Fougerousse, F.; Arandel, L.; Daniele, N.; Roudaut, C.; Noulet, F.; Garcia, L.; Danos, O.; et al. AAV-Mediated Delivery of a Mutated Myostatin Propeptide Ameliorates Calpain 3 but Not α -Sarcoglycan Deficiency. Gene Ther. 2007, 14, 733–740.
    86. Akpan, I.; Goncalves, M.D.; Dhir, R.; Yin, X.; Pistilli, E.E.; Bogdanovich, S.; Khurana, T.S.; Ucran, J.; Lachey, J.; Ahima, R.S. The Effects of a Soluble Activin Type IIB Receptor on Obesity and Insulin Sensitivity. Int. J. Obes. 2009, 33, 1265–1273.
    87. Chiu, C.-S.; Peekhaus, N.; Weber, H.; Adamski, S.; Murray, E.M.; Zhang, H.Z.; Zhao, J.Z.; Ernst, R.; Lineberger, J.; Huang, L.; et al. Increased Muscle Force Production and Bone Mineral Density in ActRIIB-Fc-Treated Mature Rodents. J. Gerontol. Ser. A Biol. Sci. Med. Sci. 2013, 68, 1181–1192.
    88. Cadena, S.M.; Tomkinson, K.N.; Monnell, T.E.; Spaits, M.S.; Kumar, R.; Underwood, K.W.; Pearsall, R.S.; Lachey, J.L. Administration of a Soluble Activin Type IIB Receptor Promotes Skeletal Muscle Growth Independent of Fiber Type. J. Appl. Physiol. 2010, 109, 635–642.
    89. Lawlor, M.W.; Read, B.P.; Edelstein, R.; Yang, N.; Pierson, C.R.; Stein, M.J.; Wermer-Colan, A.; Buj-Bello, A.; Lachey, J.L.; Seehra, J.S.; et al. Inhibition of Activin Receptor Type IIB Increases Strength and Lifespan in Myotubularin-Deficient Mice. Am. J. Pathol. 2011, 178, 784–793.
    90. Béchir, N.; Pecchi, É.; Relizani, K.; Vilmen, C.; Le Fur, Y.; Bernard, M.; Amthor, H.; Bendahan, D.; Giannesini, B. Mitochondrial Impairment Induced by Postnatal ActRIIB Blockade Does Not Alter Function and Energy Status in Exercising Mouse Glycolytic Muscle in Vivo. Am. J. Physiol.-Endocrinol. Metab. 2016, 310, E539–E549.
    91. Relizani, K.; Mouisel, E.; Giannesini, B.; Hourdé, C.; Patel, K.; Gonzalez, S.M.; Jülich, K.; Vignaud, A.; Piétri-Rouxel, F.; Fortin, D.; et al. Blockade of ActRIIB Signaling Triggers Muscle Fatigability and Metabolic Myopathy. Mol. Ther. 2014, 22, 1423–1433.
    92. Nagy, J.A.; Kapur, K.; Taylor, R.S.; Sanchez, B.; Rutkove, S.B. Electrical Impedance Myography as a Biomarker of Myostatin Inhibition with ActRIIB-MFc: A Study in Wild-Type Mice. Future Sci. OA 2018, 4, FSO308.
    93. Hoogaars, W.M.H.; Mouisel, E.; Pasternack, A.; Hulmi, J.J.; Relizani, K.; Schuelke, M.; Schirwis, E.; Garcia, L.; Ritvos, O.; Ferry, A.; et al. Combined Effect of AAV-U7-Induced Dystrophin Exon Skipping and Soluble Activin Type IIB Receptor in Mdx Mice. Hum. Gene Ther. 2012, 23, 1269–1279.
    94. Béchir, N.; Pecchi, É.; Vilmen, C.; Bernard, M.; Bendahan, D.; Giannesini, B. Activin Type IIB Receptor Blockade Does Not Limit Adenosine Triphosphate Supply in Mouse Skeletal Muscle in Vivo. Muscle Nerve 2018, 58, 834–842.
    95. Tauer, J.T.; Rauch, F. Novel ActRIIB Ligand Trap Increases Muscle Mass and Improves Bone Geometry in a Mouse Model of Severe Osteogenesis Imperfecta. Bone 2019, 128, 115036.
    96. Li, Z.B.; Zhang, J.; Wagner, K.R. Inhibition of Myostatin Reverses Muscle Fibrosis through Apoptosis. J. Cell Sci. 2012, 125, 3957–3965.
    97. Pistilli, E.E.; Bogdanovich, S.; Goncalves, M.D.; Ahima, R.S.; Lachey, J.; Seehra, J.; Khurana, T. Targeting the Activin Type IIB Receptor to Improve Muscle Mass and Function in the Mdx Mouse Model of Duchenne Muscular Dystrophy. Am. J. Pathol. 2011, 178, 1287–1297.
    98. Iskenderian, A.; Liu, N.; Deng, Q.; Huang, Y.; Shen, C.; Palmieri, K.; Crooker, R.; Lundberg, D.; Kastrapeli, N.; Pescatore, B.; et al. Myostatin and Activin Blockade by Engineered Follistatin Results in Hypertrophy and Improves Dystrophic Pathology in Mdx Mouse More than Myostatin Blockade Alone. Skelet. Muscle 2018, 8, 34.
    99. Hulmi, J.J.; Oliveira, B.M.; Silvennoinen, M.; Hoogaars, W.M.H.; Pasternack, A.; Kainulainen, H.; Ritvos, O. Exercise Restores Decreased Physical Activity Levels and Increases Markers of Autophagy and Oxidative Capacity in Myostatin/Activin-Blocked Mdx Mice. Am. J. Physiol.-Endocrinol. Metab. 2013, 305, E171–E182.
    100. Tinklenberg, J.; Meng, H.; Yang, L.; Liu, F.; Hoffmann, R.G.; Dasgupta, M.; Allen, K.P.; Beggs, A.H.; Hardeman, E.C.; Pearsall, R.S.; et al. Treatment with ActRIIB-MFc Produces Myofiber Growth and Improves Lifespan in the Acta1 H40Y Murine Model of Nemaline Myopathy. Am. J. Pathol. 2016, 186, 1568–1581.
    101. Lawlor, M.W.; Viola, M.G.; Meng, H.; Edelstein, R.V.; Liu, F.; Yan, K.; Luna, E.J.; Lerch-Gaggl, A.; Hoffmann, R.G.; Pierson, C.R.; et al. Differential Muscle Hypertrophy Is Associated with Satellite Cell Numbers and Akt Pathway Activation Following Activin Type IIB Receptor Inhibition in Mtm1 p.R69C Mice. Am. J. Pathol. 2014, 184, 1831–1842.
    102. Bondulich, M.K.; Jolinon, N.; Osborne, G.F.; Smith, E.J.; Rattray, I.; Neueder, A.; Sathasivam, K.; Ahmed, M.; Ali, N.; Benjamin, A.C.; et al. Myostatin Inhibition Prevents Skeletal Muscle Pathophysiology in Huntington’s Disease Mice. Sci. Rep. 2017, 7, 14275.
    103. Lee, Y.-S.; Lehar, A.; Sebald, S.; Liu, M.; Swaggart, K.A.; Talbot, C.C.; Pytel, P.; Barton, E.R.; McNally, E.M.; Lee, S.-J. Muscle Hypertrophy Induced by Myostatin Inhibition Accelerates Degeneration in Dysferlinopathy. Hum. Mol. Genet. 2015, 24, 5711–5719.
    104. Ohsawa, Y.; Hagiwara, H.; Nakatani, M.; Yasue, A.; Moriyama, K.; Murakami, T.; Tsuchida, K.; Noji, S.; Sunada, Y. Muscular Atrophy of Caveolin-3–Deficient Mice Is Rescued by Myostatin Inhibition. J. Clin. Investig. 2006, 116, 2924–2934.
    105. Lach-Trifilieff, E.; Minetti, G.C.; Sheppard, K.; Ibebunjo, C.; Feige, J.N.; Hartmann, S.; Brachat, S.; Rivet, H.; Koelbing, C.; Morvan, F.; et al. An Antibody Blocking Activin Type II Receptors Induces Strong Skeletal Muscle Hypertrophy and Protects from Atrophy. Mol. Cell Biol. 2014, 34, 606–618.
    106. Morvan, F.; Rondeau, J.-M.; Zou, C.; Minetti, G.; Scheufler, C.; Scharenberg, M.; Jacobi, C.; Brebbia, P.; Ritter, V.; Toussaint, G.; et al. Blockade of Activin Type II Receptors with a Dual Anti-ActRIIA/IIB Antibody Is Critical to Promote Maximal Skeletal Muscle Hypertrophy. Proc. Natl. Acad. Sci. USA 2017, 114, 12448–12453.
    107. Haidet, A.M.; Rizo, L.; Handy, C.; Umapathi, P.; Eagle, A.; Shilling, C.; Boue, D.; Martin, P.T.; Sahenk, Z.; Mendell, J.R.; et al. Long-Term Enhancement of Skeletal Muscle Mass and Strength by Single Gene Administration of Myostatin Inhibitors. Proc. Natl. Acad. Sci. USA 2008, 105, 4318–4322.
    108. Miller, T.M.; Kim, S.H.; Yamanaka, K.; Hester, M.; Umapathi, P.; Arnson, H.; Rizo, L.; Mendell, J.R.; Gage, F.H.; Cleveland, D.W.; et al. Gene Transfer Demonstrates That Muscle Is Not a Primary Target for Non-Cell-Autonomous Toxicity in Familial Amyotrophic Lateral Sclerosis. Proc. Natl. Acad. Sci. USA 2006, 103, 19546–19551.
    109. Kota, J.; Handy, C.R.; Haidet, A.M.; Montgomery, C.L.; Eagle, A.; Rodino-Klapac, L.R.; Tucker, D.; Shilling, C.J.; Therlfall, W.R.; Walker, C.M.; et al. Follistatin Gene Delivery Enhances Muscle Growth and Strength in Nonhuman Primates. Sci. Transl. Med. 2009, 1, 6ra15.
    110. Pearsall, R.S.; Davies, M.V.; Cannell, M.; Li, J.; Widrick, J.; Mulivor, A.W.; Wallner, S.; Troy, M.E.; Spaits, M.; Liharska, K.; et al. Follistatin-Based Ligand Trap ACE-083 Induces Localized Hypertrophy of Skeletal Muscle with Functional Improvement in Models of Neuromuscular Disease. Sci. Rep. 2019, 9, 1–14.
    111. Castonguay, R.; Lachey, J.; Wallner, S.; Strand, J.; Liharska, K.; Watanabe, A.E.; Cannell, M.; Davies, M.V.; Sako, D.; Troy, M.E.; et al. Follistatin-288-Fc Fusion Protein Promotes Localized Growth of Skeletal Muscle. J. Pharm. Exp. 2018, 365, 435–445.
    112. Foster, K.; Graham, I.R.; Otto, A.; Foster, H.; Trollet, C.; Yaworsky, P.J.; Walsh, F.S.; Bickham, D.; Curtin, N.A.; Kawar, S.L.; et al. Adeno-Associated Virus-8-Mediated Intravenous Transfer of Myostatin Propeptide Leads to Systemic Functional Improvements of Slow but Not Fast Muscle. Rejuvenation Res. 2009, 12, 85–94.
    113. Liu, M.; Hammers, D.W.; Barton, E.R.; Sweeney, H.L. Activin Receptor Type IIB Inhibition Improves Muscle Phenotype and Function in a Mouse Model of Spinal Muscular Atrophy. PLoS ONE 2016, 11, e0166803.
    114. Morine, K.J.; Bish, L.T.; Selsby, J.T.; Gazzara, J.A.; Pendrak, K.; Sleeper, M.M.; Barton, E.R.; Lee, S.-J.; Sweeney, H.L. Activin IIB Receptor Blockade Attenuates Dystrophic Pathology in a Mouse Model of Duchenne Muscular Dystrophy. Muscle Nerve 2010, 42, 722–730.
    115. Morine, K.J.; Bish, L.T.; Pendrak, K.; Sleeper, M.M.; Barton, E.R.; Sweeney, H.L. Systemic Myostatin Inhibition via Liver-Targeted Gene Transfer in Normal and Dystrophic Mice. PLoS ONE 2010, 5, e9176.
    116. Hammers, D.W.; Hart, C.C.; Patsalos, A.; Matheny, M.K.; Wright, L.A.; Nagy, L.; Sweeney, H.L. Glucocorticoids Counteract Hypertrophic Effects of Myostatin Inhibition in Dystrophic Muscle. JCI Insight 2020, 5.
    117. Bish, L.T.; Sleeper, M.M.; Forbes, S.C.; Morine, K.J.; Reynolds, C.; Singletary, G.E.; Trafny, D.; Pham, J.; Bogan, J.; Kornegay, J.N.; et al. Long-Term Systemic Myostatin Inhibition via Liver-Targeted Gene Transfer in Golden Retriever Muscular Dystrophy. Hum. Gene Ther. 2011, 22, 1499–1509.
    118. Lu-Nguyen, N.; Malerba, A.; Popplewell, L.; Schnell, F.; Hanson, G.; Dickson, G. Systemic Antisense Therapeutics for Dystrophin and Myostatin Exon Splice Modulation Improve Muscle Pathology of Adult Mdx Mice. Mol. Nucleic Acids 2017, 6, 15–28.
    119. Dumonceaux, J.; Marie, S.; Beley, C.; Trollet, C.; Vignaud, A.; Ferry, A.; Butler-Browne, G.; Garcia, L. Combination of Myostatin Pathway Interference and Dystrophin Rescue Enhances Tetanic and Specific Force in Dystrophic Mdx Mice. Mol. Ther. 2010, 18, 881–887.
    120. Mendias, C.L.; Marcin, J.E.; Calerdon, D.R.; Faulkner, J.A. Contractile Properties of EDL and Soleus Muscles of Myostatin-Deficient Mice. J. Appl. Physiol. (1985) 2006, 101, 898–905.
    121. Qaisar, R.; Renaud, G.; Morine, K.; Barton, E.R.; Sweeney, H.L.; Larsson, L. Is Functional Hypertrophy and Specific Force Coupled with the Addition of Myonuclei at the Single Muscle Fiber Level? FASEB J. 2011, 26, 1077–1085.
    122. Amthor, H.; Macharia, R.; Navarrete, R.; Schuelke, M.; Brown, S.C.; Otto, A.; Voit, T.; Muntoni, F.; Vrbóva, G.; Partridge, T.; et al. Lack of Myostatin Results in Excessive Muscle Growth but Impaired Force Generation. Proc. Natl. Acad. Sci. USA 2007, 104, 1835–1840.
    123. Girgenrath, S.; Song, K.; Whittemore, L.-A. Loss of Myostatin Expression Alters Fiber-Type Distribution and Expression of Myosin Heavy Chain Isoforms in Slow- and Fast-Type Skeletal Muscle. Muscle Nerve 2005, 31, 34–40.
    124. Hennebry, A.; Berry, C.; Siriett, V.; O’Callaghan, P.; Chau, L.; Watson, T.; Sharma, M.; Kambadur, R. Myostatin Regulates Fiber-Type Composition of Skeletal Muscle by Regulating MEF2 and MyoD Gene Expression. Am. J. Physiol.-Cell Physiol. 2009, 296, C525–C534.
    125. Hennebry, A.; Oldham, J.; Shavlakadze, T.; Grounds, M.D.; Sheard, P.; Fiorotto, M.L.; Falconer, S.; Smith, H.K.; Berry, C.; Jeanplong, F.; et al. IGF1 Stimulates Greater Muscle Hypertrophy in the Absence of Myostatin in Male Mice. J. Endocrinol. 2017, 234, 187–200.
    126. Matsakas, A.; Mouisel, E.; Amthor, H.; Patel, K. Myostatin Knockout Mice Increase Oxidative Muscle Phenotype as an Adaptive Response to Exercise. J. Muscle Res. Cell Motil. 2010, 31, 111–125.
    127. Kocsis, T.; Trencsenyi, G.; Szabo, K.; Baan, J.A.; Muller, G.; Mendler, L.; Garai, I.; Reinauer, H.; Deak, F.; Dux, L.; et al. Myostatin Propeptide Mutation of the Hypermuscular Compact Mice Decreases the Formation of Myostatin and Improves Insulin Sensitivity. Am. J. Physiol.-Endocrinol. Metab. 2016, 312, E150–E160.
    128. Wagner, K.R.; McPherron, A.C.; Winik, N.; Lee, S.-J. Loss of Myostatin Attenuates Severity of Muscular Dystrophy in Mdx Mice. Ann. Neurol. 2002, 52, 832–836.
    129. Wagner, K.R.; Liu, X.; Chang, X.; Allen, R.E. Muscle Regeneration in the Prolonged Absence of Myostatin. Proc. Natl. Acad. Sci. USA 2005, 102, 2519–2524.
    130. Li, Z.; Shelton, G.D.; Engvall, E. Elimination of Myostatin Does Not Combat Muscular Dystrophy in Dy Mice but Increases Postnatal Lethality. Am. J. Pathol. 2005, 166, 491–497.
    131. Stedman, H.H.; Sweeney, H.L.; Shrager, J.B.; Maguire, H.C.; Panettieri, R.A.; Petrof, B.; Narusawa, M.; Leferovich, J.M.; Sladky, J.T.; Kelly, A.M. The Mdx Mouse Diaphragm Reproduces the Degenerative Changes of Duchenne Muscular Dystrophy. Nature 1991, 352, 536–539.
    132. Long, K.K.; O’Shea, K.M.; Khairallah, R.J.; Howell, K.; Paushkin, S.; Chen, K.S.; Cote, S.M.; Webster, M.T.; Stains, J.P.; Treece, E.; et al. Specific Inhibition of Myostatin Activation Is Beneficial in Mouse Models of SMA Therapy. Hum. Mol. Genet. 2019, 28, 1076–1089.
    133. Graham, Z.A.; Collier, L.; Peng, Y.; Saéz, J.C.; Bauman, W.A.; Qin, W.; Cardozo, C.P. A Soluble Activin Receptor IIB Fails to Prevent Muscle Atrophy in a Mouse Model of Spinal Cord Injury. J. Neurotrauma 2016, 33, 1128–1135.
    134. Pistilli, E.E.; Bogdanovich, S.; Mosqueira, M.; Lachey, J.; Seehra, J.; Khurana, T.S. Pretreatment with a Soluble Activin Type IIB Receptor/Fc Fusion Protein Improves Hypoxia-Induced Muscle Dysfunction. Am. J. Physiol.-Regul. Integr. Comp. Physiol. 2010, 298, R96–R103.
    135. Tortoriello, D.V.; Sidis, Y.; Holtzman, D.A.; Holmes, W.E.; Schneyer, A.L. Human Follistatin-Related Protein: A Structural Homologue of Follistatin with Nuclear Localization. Endocrinology 2001, 142, 3426–3434.
    136. Nakatani, M.; Takehara, Y.; Sugino, H.; Matsumoto, M.; Hashimoto, O.; Hasegawa, Y.; Murakami, T.; Uezumi, A.; Takeda, S.; Noji, S.; et al. Transgenic Expression of a Myostatin Inhibitor Derived from Follistatin Increases Skeletal Muscle Mass and Ameliorates Dystrophic Pathology in Mdx Mice. FASEB J. 2008, 22, 477–487.
    137. Shen, C.; Iskenderian, A.; Lundberg, D.; He, T.; Palmieri, K.; Crooker, R.; Deng, Q.; Traylor, M.; Gu, S.; Rong, H.; et al. Protein Engineering on Human Recombinant Follistatin: Enhancing Pharmacokinetic Characteristics for Therapeutic Application. J. Pharm. Exp. 2018, 366, 291–302.
    138. Malerba, A.; Kang, J.K.; McClorey, G.; Saleh, A.F.; Popplewell, L.; Gait, M.J.; Wood, M.J.; Dickson, G. Dual Myostatin and Dystrophin Exon Skipping by Morpholino Nucleic Acid Oligomers Conjugated to a Cell-Penetrating Peptide Is a Promising Therapeutic Strategy for the Treatment of Duchenne Muscular Dystrophy. Mol. Nucleic. Acids 2012, 1, e62.
    139. Varga, L.; Szabo, G.; Darvasi, A.; Muller, G.; Sass, M.; Soller, M. Inheritance and Mapping of Compact (Cmpt), a New Mutation Causing Hypermuscularity in Mice. Genetics 1997, 147, 755–764.
    140. Wilkes, J.J.; Lloyd, D.J.; Gekakis, N. Loss-of-Function Mutation in Myostatin Reduces Tumor Necrosis Factor Alpha Production and Protects Liver against Obesity-Induced Insulin Resistance. Diabetes 2009, 58, 1133–1143.
    141. Kramerova, I.; Marinov, M.; Owens, J.; Lee, S.-J.; Becerra, D.; Spencer, M.J. Myostatin Inhibition Promotes Fast Fibre Hypertrophy but Causes Loss of AMP-Activated Protein Kinase Signalling and Poor Exercise Tolerance in a Model of Limb-Girdle Muscular Dystrophy R1/2A. J. Physiol. 2020, 598, 3927–3939.