Kristy Swiderski

Antibody-directed myostatin inhibition enhances muscle mass and function in tumor-bearing mice

Cancer cachexia describes the progressive skeletal muscle wasting and weakness in many cancer patients and accounts for >20% of cancer-related deaths. We tested the hypothesis that antibody-directed myostatin inhibition would attenuate the atrophy and loss of function in muscles of tumor-bearing mice. Twelve-week-old C57BL/6 mice received a subcutaneous injection of saline (control) or Lewis lung carcinoma (LLC) tumor cells. One week later, mice received either once weekly injections of saline (control, n = 12; LLC, n = 9) or a mouse chimera of anti-human myostatin antibody (PF-354, 10 mg·kg⁻¹·wk⁻¹, LLC+PF-354, n = 11) for 5 wk. Injection of LLC cells reduced muscle mass and maximum force of tibialis anterior (TA) muscles by 8-10% (P < 0.05), but the muscle atrophy and weakness were prevented with PF-354 treatment (P > 0.05). Maximum specific (normalized) force of diaphragm muscle strips was reduced with LLC injection (P < 0.05) but was not improved with PF-354 treatment (P > 0.05). PF-354 enhanced activity of oxidative enzymes in TA and diaphragm muscles of tumor-bearing mice by 118% and 89%, respectively (P < 0.05). Compared with controls, apoptosis that was not of myofibrillar or satellite cell origin was 140% higher in TA muscle cross sections from saline-treated LLC tumor-bearing mice (P < 0.05) but was not different in PF-354-treated tumor-bearing mice (P > 0.05). Antibody-directed myostatin inhibition attenuated the skeletal muscle atrophy and loss of muscle force-producing capacity in a murine model of cancer cachexia, in part by reducing apoptosis. The improvements in limb muscle mass and function highlight the therapeutic potential of antibody-directed myostatin inhibition for cancer cachexia.

GENE AND CELL-MEDIATED THERAPIES FOR MUSCULAR DYSTROPHY

Duchenne muscular dystrophy (DMD) is a devastating muscle disorder that affects 1 in 3,500 boys. Despite years of research and considerable progress in understanding the molecular mechanism of the disease and advancement of therapeutic approaches, there is no cure for DMD. The current treatment options are limited to physiotherapy and corticosteroids, and although they provide a substantial improvement in affected children, they only slow the course of the disorder. On a more optimistic note, more recent approaches either significantly alleviate or eliminate muscular dystrophy in murine and canine models of DMD and importantly, many of them are being tested in early phase human clinical trials. This review summarizes advancements that have been made in viral and nonviral gene therapy as well as stem cell therapy for DMD with a focus on the replacement and repair of the affected dystrophin gene. Muscle Nerve 47: 649–663, 2013

Phosphorylation within the cysteine-rich region of dystrophin enhances its association with β-dystroglycan and identifies a potential novel therapeutic target for skeletal muscle wasting

Mutations in dystrophin lead to Duchenne muscular dystrophy, which is among the most common human genetic disorders. Dystrophin nucleates assembly of the dystrophin-glycoprotein complex (DGC), and a defective DGC disrupts an essential link between the intracellular cytoskeleton and the basal lamina, leading to progressive muscle wasting. In vitro studies have suggested that dystrophin phosphorylation may affect interactions with actin or syntrophin, yet whether this occurs in vivo or affects protein function remains unknown. Utilizing nanoflow liquid chromatography mass spectrometry, we identified 18 phosphorylated residues within endogenous dystrophin. Mutagenesis revealed that phosphorylation at S3059 enhances the dystrophin-dystroglycan interaction and 3D modeling utilizing the Rosetta software program provided a structural model for how phosphorylation enhances this interaction. These findings demonstrate that phosphorylation is a key mechanism regulating the interaction between dystrophin and the DGC and reveal that posttranslational modification of a single amino acid directly modulates the function of dystrophin.

Therapeutic potential of heat shock protein induction for muscular dystrophy and other muscle wasting conditions

Duchenne muscular dystrophy is the most common and severe of the muscular dystrophies, a group of inherited myopathies caused by different genetic mutations leading to aberrant expression or complete absence of cytoskeletal proteins. Dystrophic muscles are prone to injury, and regenerate poorly after damage. Remorseless cycles of muscle fibre breakdown and incomplete repair lead to progressive and severe muscle wasting, weakness and premature death. Many other conditions are similarly characterized by muscle wasting, including sarcopenia, cancer cachexia, sepsis, denervation, burns, and chronic obstructive pulmonary disease. Muscle trauma and loss of mass and physical capacity can significantly compromise quality of life for patients. Exercise and nutritional interventions are unlikely to halt or reverse the conditions, and strategies promoting muscle anabolism have limited clinical acceptance. Heat shock proteins (HSPs) are molecular chaperones that help proteins fold back to their original conformation and restore function. Since many muscle wasting conditions have pathophysiologies where inflammation, atrophy and weakness are indicated, increasing HSP expression in skeletal muscle may have therapeutic potential. This review will provide evidence supporting HSP induction for muscular dystrophy and other muscle wasting conditions. This article is part of the theme issue ‘Heat shock proteins as modulators and therapeutic targets of chronic disease: an integrated perspective’.

Skeletal muscle-specific overexpression of IGFBP-2 promotes a slower muscle phenotype in healthy but not dystrophic mdx mice and does not affect the dystrophic pathology

OBJECTIVE The insulin-like growth factor binding proteins (IGFBPs) are thought to modulate cell size and homeostasis via IGF-I-dependent and -independent pathways. There is a considerable dearth of information regarding the function of IGFBPs in skeletal muscle, particularly their role in the pathophysiology of Duchenne muscular dystrophy (DMD). In this study we tested the hypothesis that intramuscular IGFBP-2 overexpression would ameliorate the pathology in mdx dystrophic mice. DESIGN 4week old male C57Bl/10 and mdx mice received a single intramuscular injection of AAV6-empty or AAV6-IGFBP-2 vector into the tibialis anterior muscle. At 8weeks post-injection the effect of IGFBP-2 overexpression on the structure and function of the injected muscle was assessed. RESULTS AAV6-mediated IGFBP-2 overexpression in the tibialis anterior (TA) muscles of 4-week-old C57BL/10 and mdx mice reduced the mass of injected muscle after 8weeks, inducing a slower muscle phenotype in C57BL/10 but not mdx mice. Analysis of inflammatory and fibrotic gene expression revealed no changes between control and IGFBP-2 injected muscles in dystrophic (mdx) mice. CONCLUSIONS Together these results indicate that the IGFBP-2-induced promotion of a slower muscle phenotype is impaired in muscles of dystrophin-deficient mdx mice, which contributes to the inability of IGFBP-2 to ameliorate the dystrophic pathology. The findings implicate the dystrophin-glycoprotein complex (DGC) in the signaling required for this adaptation.

Muscle-specific deletion of SOCS3 increases the early inflammatory response but does not affect regeneration after myotoxic injury

Background Muscles of old animals are injured more easily and regenerate poorly, attributed in part to increased levels of circulating pro-inflammatory cytokines. The Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling cascade is a key mediator of inflammatory cytokine action, and signaling via this pathway is increased in muscles with aging. As a negative regulator of JAK/STAT signaling, a key mediator of myogenic proliferation and differentiation, altered expression of suppressor of cytokine signaling (SOCS3) is likely to have important consequences for muscle regeneration. To model this scenario, we investigated the effect of SOCS3 deletion within mature muscle fibers on injury and repair. We tested the hypothesis that reduced SOCS3 function would alter the inflammatory response and impair muscle regeneration after myotoxic injury. Methods Mice with a specific deletion of SOCS3 within mature skeletal muscle fibers were used to assess the effect of SOCS3 deletion on muscle injury and repair. Twelve-week-old or 24-month-old SOCS3 muscle-specific knockout (SOCS3 MKO) mice and littermate controls were either left uninjured or injured with a single injection of notexin (10 μg/ml) into the right tibialis anterior (TA) muscle. At 1, 2, 3, 5, 7, or 14 days post-injury, the right TA muscle was excised and subjected to histological, western immunoblotting, and gene expression analyses. Force production and fatigue were assessed in uninjured muscles and at 7 days post-notexin injury. Results In uninjured muscles, SOCS3 deletion decreased force production during fatigue but had no effect on the gross or histological appearance of the TA muscles. After notexin injury, deletion of SOCS3 increased STAT3 phosphorylation at day 1 and increased the mRNA expression of the inflammatory cytokine TNF-α, and the inflammatory cell markers F4/80 and CD68 at day 2. Gene expression analysis of the regeneration markers Pax7, MyoD, and Myogenin indicated SOCS3 deletion had no effect on the progression of muscle repair after notexin injury. Inflammation and regeneration were also unchanged in the muscles of 24-month-old SOCS3 MKO mice compared with control. Conclusions Loss of SOCS3 expression in mature muscle fibers increased the inflammatory response to myotoxic injury but did not impair muscle regeneration in either adult or old mice. Therefore, reduced SOCS3 expression in muscle fibers is unlikely to underlie impaired muscle regeneration. Further investigation into the role of SOCS3 in other cell types involved in muscle repair is warranted. Electronic supplementary material The online version of this article (doi:10.1186/s13395-016-0108-4) contains supplementary material, which is available to authorized users.

Therapeutic potential of orphan drugs for the rare skeletal muscle diseases

Introduction: Rare diseases affect ∼350 million people worldwide yet 95% of these diseases do not have a single FDA-approved drug for treatment. The rare skeletal muscle diseases include numerous severe muscle wasting disorders, including spinal muscular atrophy, inclusion body myopathies, glycogen storage disorders and the many muscular dystrophies. While the underlying genetic defects responsible for each of these disorders is usually unique and not always confined to skeletal muscle, muscle fiber atrophy and weakness are common to all of them. Areas covered: Progress in the development of therapies with ‘orphan drug’ status is detailed for some of the rare skeletal muscle diseases. These therapies include drugs at varying stages and progression through the development pipeline such as membrane sealants, anti-inflammatory medications, enzyme replacers, growth promoting agents, anti-fibrotics, as well as gene- and cell-based approaches. Expert opinion: Multiple drugs have received orphan designation for rare muscle diseases and several have entered clinical trials and progressed through the development pipeline. Many drugs enter trials without having undergone sufficiently rigorous preclinical testing in relation to their effects on skeletal muscle structure–function; a shortcoming that could compromise their efficacy for treating the rare muscle diseases. More rigorous preclinical physiological assessments are important for identifying which drugs should advance to clinical trials.

Muscle-specific deletion of SOCS3 does not reduce the anabolic response to leucine in a mouse model of acute inflammation

HighlightsSOCS3 deletion in skeletal muscle does not alter LPS‐induced inflammation.SOCS3 does not mediate the anabolic response to leucine in skeletal muscle.Leucine augments the LPS‐induced expression of IL‐6 and STAT3 phosphorylation. Abstract Excessive inflammation reduces skeletal muscle protein synthesis leading to wasting and weakness. The janus kinase/signal transducers and activators of transcription‐3 (JAK/STAT3) pathway is important for the regulation of inflammatory signaling. As such, suppressor of cytokine signaling‐3 (SOCS3), the negative regulator of JAK/STAT signaling, is thought to be important in the control of muscle homeostasis. We hypothesized that muscle‐specific deletion of SOCS3 would impair the anabolic response to leucine during an inflammatory insult. Twelve week old (n = 8 per group) SOCS3 muscle‐specific knockout mice (SOCS3‐MKO) and littermate controls (WT) were injected with lipopolysaccharide (LPS, 1 mg/kg) or saline and were studied during fasted conditions or after receiving 0.5 g/kg leucine 3 h after the injection of LPS. Markers of inflammation, anabolic signaling, and protein synthesis were measured 4 h after LPS injection. LPS injection robustly increased mRNA expression of inflammatory molecules (Socs3, Socs1, Il‐6, Ccl2, Tnf&agr; and Cd68). In muscles from SOCS3‐MKO mice, the Socs3 mRNA response to LPS was significantly blunted (˜6‐fold) while STAT3 Tyr705 phosphorylation was exacerbated (18‐fold). Leucine administration increased protein synthesis in both WT (˜1.6‐fold) and SOCS3‐MKO mice (˜1.5‐fold) compared to basal levels. LPS administration blunted this effect, but there were no differences between WT and SOCS3‐MKO mice. Muscle‐specific SOCS3 deletion did not alter the response of AKT, mTOR, S6 or 4EBP1 under any treatment conditions. Therefore, SOCS3 does not appear to mediate the early inflammatory or leucine‐induced changes in protein synthesis in skeletal muscle.