Progression Of Muscle Atrophy Research Articles

Muscle atrophy is coupled with ferroptosis in the skeletal muscle of Friedreich’s ataxia

Friedreich’s ataxia (FRDA) is a rare genetic disorder caused by an expansion of GAA repeats in the frataxin gene (FXN), resulting in frataxin deficiency. As frataxin is crucial for mitochondrial function by recruiting key iron components such as iron-sulfur clusters and hemes, FRDA is characterized by mitochondrial dysfunction along with abnormal iron metabolism. Recently, ferroptosis, an iron-dependent cell death, has been suggested to play an important role in the development of FRDA, yet if and how ferroptosis impacts FRDA-related muscle weakness has not been studied. Therefore, the objective of this study was to identify the role of ferroptosis in the skeletal muscle of FRDA using whole-body frataxin deficient mice (Fxnnull::YG8s(GAA)>800, #030395, The Jackson Laboratory). FRDA mice (male, 3-months old) had a significantly smaller ratio of muscle mass to body mass, compared to strain-, age-, and sex-matched C57BL/6J control mice, suggesting that this animal model develops muscle atrophy as early as 3 months old upon frataxin deficiency. Furthermore, the FRDA muscles showed elevated non-heme iron levels, which were accompanied by altered iron regulatory protein expressions, including ferritin and ferroportin. In addition, mitochondrial function was decreased in the skeletal muscles of FRDA mice, as evidenced by significantly decreased protein levels of oxidative phosphorylation complexes (OXPHOS I, II, and III; decrease by 31%, 26%, and 15% compared to control, respectively), as well as other mitochondrial capacity markers such as nuclear factor erythroid 2-related factor 1 (Nrf1) and heme oxygenase-1 ( p=0.06). Notably, ferroptosis markers such as Nrf2 and glutathione peroxidase 4 were significantly downregulated in the skeletal muscles of FRDA mice, while apoptotic markers (p53 and caspase 3) were not altered. Collectively, ferroptosis could promote FRDA-related muscle atrophy, which is associated with abnormal mitochondrial iron status and function. Our results also suggest that ferroptosis may represent a novel target for correcting or ameliorating the development and progression of muscle atrophy in FRDA patients. Future studies are warranted to determine the molecular mechanism of ferroptosis in FRDA-related mitochondrial dysfunction and to develop therapeutic strategies for muscle weakness in FRDA or other types of ataxias. This study was supported in part by the NIH NS121394 and UMass Lowell Internal Seed Grant. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Uncovering the mechanisms of MuRF1-induced ubiquitylation and revealing similarities with MuRF2 and MuRF3

MuRF1 (Muscle-specific RING finger protein 1; gene name TRIM63) is a ubiquitin E3 ligase, associated with the progression of muscle atrophy. As a RING (Really Interesting New Gene) type E3 ligase, its unique activity of ubiquitylation is driven by a specific interaction with a UBE2 (ubiquitin conjugating enzyme). Our understanding of MuRF1 function remains unclear as candidate UBE2s have not been fully elucidated. In the present study, we screened human ubiquitin dependent UBE2s in vitro and found that MuRF1 engages in ubiquitylation with UBE2D, UBE2E, UBE2N/V families and UBE2W. MuRF1 can cause mono-ubiquitylation, K48- and K63-linked polyubiquitin chains in a UBE2 dependent manner. Moreover, we identified a two-step UBE2 dependent mechanism whereby MuRF1 is monoubiquitylated by UBE2W which acts as an anchor for UBE2N/V to generate polyubiquitin chains. With the in vitro ubiquitylation assay, we also found that MuRF2 and MuRF3 not only share the same UBE2 partners as MuRF1 but can also directly ubiquitylate the same substrates: Titin (A168-A170), Desmin, and MYLPF (Myosin Light Chain, Phosphorylatable, Fast Skeletal Muscle; also called Myosin Light Regulatory Chain 2). In summary, our work presents new insights into the mechanisms that underpin MuRF1 activity and reveals overlap in MuRF-induced ubiquitylation which could explain their partial redundancy in vivo.

Nuclear morphometry and stereological analysis of gastrocnemius and pectoralis major muscle tissue from MDX mice supplemented or not with creatine for sixteen weeks

Duchenne muscular dystrophy is a genetic disease that causes a reduction in the expression of dystrophin, thus, with the reduction of the levels of this protein, a progression of muscle atrophy, reduction of strength and physical impairments is identified, which culminates in death from inability to contract the respiratory muscles. Creatine is a dietary supplement that helps maintain strength and recent findings indicate a potential anti-inflammatory effect of this amine. The present study aimed to compare the progress of muscular dystrophy in terms of inflammation, presence of intramuscular glycogen and tissue fibrosis in the gastrocnemius and pectoralis major muscles of MDX mice. We selected 20 MDX mice that were supplemented or not with creatine at a dosage of 0.3 grams per kilogram of body weight, for 16 weeks. After that, the gastrocnemius and pectoralis major were extracted for morphological and morphometric analysis. The inflammatory progress derived from Duchenne muscular dystrophy affects animals, even when supplemented, however, in a less forceful and disabling way, enabling them to have less functional damage. Nevertheless, the supplemented animals have a greater stock of intramuscular glycogen in the evaluated gastrocnemius tissue, a factor that corroborates with the survival and better quality of movement in these animals, given the availability of energy for the contraction of this musculature. Furthermore, a lower frequency and extent of tissue fibrosis is observed in supplemented animals, aspects that enable longer survival and qualify creatine, even in an experimental condition, as an auxiliary supplement in the treatment of a severe dystrophic condition, acting mainly in the maintenance of strength and reduction of local tissue fibrosis.

Influence of gonadectomy on muscle health in micro- and partial-gravity environments in rats.

Gonadal hormones, such as testosterone and estradiol, modulate muscle size and strength in males and females. However, the influence of sex hormones on muscle strength in micro- and partial-gravity environments (e.g., the Moon or Mars) is not fully understood. The purpose of this study was to determine the influence of gonadectomy (castration/ovariectomy) on progression of muscle atrophy in both micro- and partial-gravity environments in male and female rats. Male and female Fischer rats (n = 120) underwent castration/ovariectomy (CAST/OVX) or sham surgery (SHAM) at 11 wk of age. After 2 wk of recovery, rats were exposed to hindlimb unloading (0 g), partial weight bearing at 40% of normal loading (0.4 g, Martian gravity), or normal loading (1.0 g) for 28 days. In males, CAST did not exacerbate body weight loss or other metrics of musculoskeletal health. In females, OVX animals tended to have greater body weight loss and greater gastrocnemius loss. Within 7 days of exposure to either microgravity or partial gravity, females had detectable changes to estrous cycle, with greater time spent in low-estradiol phases diestrus and metestrus (∼47% in 1 g vs. 58% in 0 g and 72% in 0.4 g animals, P = 0.005). We conclude that in males testosterone deficiency at the initiation of unloading has little effect on the trajectory of muscle loss. In females, initial low estradiol status may result in greater musculoskeletal losses.NEW & NOTEWORTHY We find that removal of gonadal hormones does not exacerbate muscle loss in males or females during exposure to either simulated microgravity or partial-gravity environments. However, simulated micro- and partial gravity did affect females' estrous cycles, with more time spent in low-estrogen phases. Our findings provide important data on the influence of gonadal hormones on the trajectory of muscle loss during unloading and will help inform NASA for future crewed missions to space and other planets.

Sulforaphane ameliorates serum starvation-induced muscle atrophy via activation of the Nrf2 pathway in cultured C2C12 cells.

Oxidative stress, an imbalance of redox homeostasis, contributes to the pathogenesis and progress of muscle atrophy. However, it is debated whether oxidative stress is a cause or consequence of muscle atrophy. In this study, we investigated the relationship between menadione-induced oxidative stress and serum starvation-induced muscle atrophy in C2C12 myotubes. We found that atrophic phenotypes including myotube diameter decrease, protein ubiquitination, and the expression of atrogenes were detected under oxidative stress as well as during serum starvation. Oxidative stress during serum starvation was assessed to confirm the correlation. Both intracellular reactive oxygen species (ROS) and protein oxidation were increased in atrophic myotubes. These results indicate that menadione-induced oxidative stress triggers muscle atrophy and vice versa. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key regulator of cellular response to oxidative stress and it is considered to have a cytoprotective role in the mitigation of muscle atrophy. Transcription of heme oxygenase-1 (HO-1) and NAD(P)H quinone dehydrogenase-1, target genes of Nrf2, was decreased during serum starvation, which is related to decreased nuclear translocation of Nrf2. Pre-treatment of sulforaphane (SFN), a known Nrf2 inducer, before serum starvation showed a protective effect via Nrf2/HO-1 upregulation. SFN can liberate Nrf2 from Keap1, enabling the nuclear translocation of Nrf2. Consequently, the expression of HO-1 increased and intracellular ROS was significantly reduced by SFN pre-treatment. These results demonstrate that oxidative stress mediates the pathophysiology of muscle atrophy, which can be improved via upregulation of the Nrf2-mediated antioxidant response.

The short external rotators dissection during the posterior approach in total hip arthroplasty did not change the blood flow.

The posterior approach in total hip arthroplasty (THA) often requires dissection of the short external rotators (SERs), which could increase the postoperative dislocation rate. The reattachment of the dissected SERs has been reported to reduce the dislocation rate, while such repair generally causes progression of muscle atrophy. 1 of the suggested causes of atrophy is reduced blood flow to the repaired SERs. The present study aimed to measure the blood flow of the SERs before dissection (pre-tenotomy) and after reattachment (post-reattachment) during the posterior approach in THA. This prospective study included 26 patients who underwent THA via the posterior approach. A laser-Doppler rheometer was used to measure the blood flow in the following SERs at the time of pre-tenotomy and post-reattachment: the piriformis muscle (PM), superior gemellus (SG), inferior gemellus (IG), obturator internus (OI), and subcutaneous tissue as a control. The average pre-tenotomy and post-reattachment blood flows (mL/minutes/100 g) were: 1.90 ± 0.28 and 1.92 ± 0.40 in the PM, 1.94 ± 0.20 and 1.99 ± 0.39 in the SG, 1.91 ± 0.21 and 1.94 ± 0.30 in the IG, 1.93 ± 0.22 and 1.98 ± 0.36 in the OI, and 1.94 ± 0.24 and 1.87 ± 0.38 in the subcutaneous tissue. The pre-tenotomy and post-reattachment blood flows did not show significant difference in any muscle. Laser-Doppler blood flow measurements showed that the blood flow is preserved, even when the SERs are dissected and reattached in THA via the posterior approach.

Increasing autophagy does not affect neurogenic muscle atrophy.

Physiological autophagy plays a crucial role in the regulation of muscle mass and metabolism, while the excessive induction or the inhibition of the autophagic flux contributes to the progression of several diseases. Autophagy can be activated by different stimuli, including cancer, exercise, caloric restriction and denervation. The latter leads to muscle atrophy through the activation of catabolic pathways, i.e. the ubiquitin-proteasome system and autophagy. However, the kinetics of autophagy activation and the upstream molecular pathways in denervated skeletal muscle have not been reported yet. In this study, we characterized the kinetics of autophagic induction, quickly triggered by denervation, and report the Akt/mTOR axis activation. Besides, with the aim to assess the relative contribution of autophagy in neurogenic muscle atrophy, we triggered autophagy with different stimuli along with denervation, and observed that four week-long autophagic induction, by either intermitted fasting or rapamycin treatment, did not significantly affect muscle mass loss. We conclude that: i) autophagy does not play a major role in inducing muscle loss following denervation; ii) nonetheless, autophagy may have a regulatory role in denervation induced muscle atrophy, since it is significantly upregulated as early as eight hours after denervation; iii) Akt/mTOR axis, AMPK and FoxO3a are activated consistently with the progression of muscle atrophy, further highlighting the complexity of the signaling response to the atrophying stimulus deriving from denervation.

C26‐Induced Cancer Cachexia Impairs Skeletal Muscle Mitochondrial Function

Cancer cachexia is characterized by a rapid ‘loss of muscle with or without loss of fat mass'1 that can impede cancer treatments resulting in significantly increased mortality rate in afflicted individuals. Whereas impaired skeletal muscle respiratory capacity likely plays a role in cancer‐induced fatigue, few studies have evaluated mitochondrial function in models of cancer cachexia2, and none in the well‐characterized colon 26 carcinoma (C26)‐induced cancer cachexia model. The present study thus aimed to evaluate the impact of C26‐induced cancer cachexia on skeletal muscle mitochondrial respiration and content.Twelve six‐week old male CD2F1 mice were injected subcutaneously with 5 × 105 C26‐cells diluted in 100 μl of PBS per flank (C26 group), whereas 12 age‐matched mice received an equivalent PBS vehicle injection (Control group). Mice were sacrificed ~ 26 days after tumor or vehicle injection. At sacrifice, C26 mice showed a reduced body weight (~ 20%) compared to control mice (p<0.05). Muscle atrophy was observed in C26 soleus, gastrocnemii‐plantaris complex (GasPL) and tibialis anterior (−21%, −10% and −11% respectively, vs. Control; p<0.05). Soleus permeabilized muscle fiber bundles from C26 mice showed a 20% reduction in maximal mitochondrial respiration compared to Controls in the presence of 10 mM of glutamate, 5 mM of malate and 5 mM of adenosine diphosphate (p<0.05). Furthermore, a 15% lower acceptor control ratio (State 3/State 2, an index of mitochondrial coupling) was observed in the C26 vs. Controls (p<0.05). Evaluation of mitochondrial content and relative expression of the different mitochondrial components involved in the oxidative phosphorylation process is currently ongoing. Preliminary results suggest that GasPL VDAC (as an index of mitochondrial content) relative protein expression is reduced in the C26 vs. Controls (~ −20%). Similarly, reductions in representative subunits of complex I, II, III and IV (CI‐IV) expressions were observed in C26‐GasPL compared to Controls (~ −19–24%), but there was no differences in complex V (CV) relative expression (~ −8%).In conclusion, C26‐induced cancer cachexia appears to lead to a reduced mitochondrial content associated with a reduced maximal respiration capacity. Moreover, in addition to the reduced mitochondrial content, an alteration of the remaining mitochondria could be observed as mitochondrial coupling was impeded. The reduced maximal respiratory capacity appears largely to be due to reduced mitochondrial content. The temporal relationships between these changes and the progression of muscle atrophy, and whether these changes are reversible if the cancer is survived remain important areas for future investigation.Support or Funding InformationDaria Neyroud salary is supported by an Early Post‐doc fellowship obtained from the Swiss National Science Foundation. In addition, this work was supported by NIH grants obtained by Prof. Judge (R01AR060209 and 1R21CA194118‐01) and internal funding obtained by Prof. Hepple through the University of Florida Cancer Center.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Size Profile and Selective Protein Packaging of Exosomes Released from Atrophying Muscle Cells

Skeletal muscle atrophy occurs in a variety of conditions and can result in decreased quality of life and mortality. Previous work from our lab established that certain microRNAs in muscle cells play a role in the progression of muscle atrophy and the intracellular level of these microRNAs are altered during atrophy, at least in part, due to incorporation into small vesicles (termed exosomes) released into the extracellular environment. Currently, little information exists about muscle released exosomes. Potentially these vesicles could be taken up by other tissues and identify a mechanism by which muscle signals other tissues during chronic conditions in which atrophy is occurring. However, to know what signaling pathways these exosomes may potentially be involved in, it is important to know what potential signaling molecules are present in exosomes released from muscle cells during atrophy. PURPOSE: To identify if exosomes released from muscle cells during atrophy contain different internal cargo proteins than exosomes from healthy muscle cells. METHODS: C2C12 cells were treated with dexamethasone (DEX; 1μM) for 6 hours in serum free media, media was collected, and exosomes were isolated from the media. LC-MS proteomic analysis was performed on proteins isolated from exosomes, and analyzed using Ingenuity Pathway Analysis software. Nanoparticle tracking analysis (Nanosight) was performed on a separate set of exomes measure vesicle size and number. RESULTS: Compared to control cells, the exosomes released during DEX-induced muscle atrophy contained 135 proteins increased greater than two-fold and 159 proteins decreased greater than two. Nanoparticle tracking analysis revealed no change in the number of exosomes released during atrophy (6.77x108 vs 7.06 x 108 vesicles/mL). However, while there was no change in the total number of exosomes the size profiles of the exosomes released during atrophy was significantly different (p<0.05). CONCLUSIONS: Skeletal muscle atrophy results in both a selective packaging of proteins into exosomes and unique size profile of exosomes released from muscles, but does not alter the total number of exosomes released. These novel findings could have broad implications for the development of biomarkers and signaling during skeletal muscle atrophy.

The natural course of nonoperatively treated rotator cuff tears: an 8.8-year follow-up of tear anatomy and clinical outcome in 49 patients

The natural course of nonoperatively treated rotator cuff tears is not fully understood. We explored the long-term development of tear anatomy and assessed functional outcomes. Eighty-nine small to medium-sized full-thickness tears of the rotator cuff, all primarily treated by physiotherapy, were identified retrospectively. Twenty-three tears needed surgical treatment later on, and 17 patients were unable to meet for follow-up. The remaining 49 still unrepaired tears were re-examined after 8.8 (8.2-11.0) years with sonography. Re-examination by magnetic resonance imaging was possible for 37 patients. Shoulder function was assessed with shoulder scores. Primary outcome measures were progression of tear size, muscle atrophy, and fatty degeneration and the Constant score (CS). Mean tear size increased by 8.3 mm in the anterior-posterior plane (P = .001) and by 4.5 mm in the medial-lateral plane (P = .001). Increase of tear size was -5 to +9.9 mm in 33 patients, 10 to 19.9 mm in 8 patients, and ≥20 mm in 8 patients. The CS was 81 points for tear increases <20 mm and 58.5 points for increases ≥20 mm (P = .008). Muscle atrophy and fatty degeneration progressed in 18 and 15 of the 37 patients, respectively. In tears with no progression of atrophy, the CS was 82 points compared with 75.5 points in tears with progression (P = .04). Anatomic tear deterioration was found in the majority of patients, but it was often moderate. Large tear size increases and progression of muscle atrophy were correlated to a poorer functional outcome.

Differential expression of HDAC and HAT genes in atrophying skeletal muscle.

Histone deacetylase (HDAC) proteins, which counter the activity of histone acetyltransferases (HATs), are necessary for normal muscle atrophy in response to several pathophysiological conditions. Despite this, it remains unknown whether a common or unique transcriptional profile of HDAC and HAT genes exist during the progression of muscle atrophy. Muscles were harvested from cast immobilized, denervated, or nutrient deprived animals for quantitative reverse transcriptase-polymerase chain reaction analysis of HDAC and HAT gene expression. The mRNA levels of Hdac2, Hdac4, Hdac6, Sirt1, p300, Cbp, and Pcaf increased, and Hdac7 decreased in skeletal muscle in each experimental model of muscle atrophy. Hdac1 and Hdac3 were increased only in cast immobilized and denervated muscles. While specific HDACs and HATs are increased in multiple models of muscle atrophy, increased expression of class I HDACs was unique to muscle disuse, reinforcing that specific HDAC inhibitors may be more effective than pan-HDAC inhibitors at countering muscle atrophy.

Mitochondrial Network Genes in the Skeletal Muscle of Amyotrophic Lateral Sclerosis Patients

Recent evidence suggested that muscle degeneration might lead and/or contribute to neurodegeneration, thus it possibly play a key role in the etiopathogenesis and progression of amyotrophic lateral sclerosis (ALS). To test this hypothesis, this study attempted to categorize functionally relevant genes within the genome-wide expression profile of human ALS skeletal muscle, using microarray technology and gene regulatory network analysis. The correlation network structures significantly change between patients and controls, indicating an increased inter-gene connection in patients compared to controls. The gene network observed in the ALS group seems to reflect the perturbation of muscle homeostasis and metabolic balance occurring in affected individuals. In particular, the network observed in the ALS muscles includes genes (PRKR1A, FOXO1, TRIM32, ACTN3, among others), whose functions connect the sarcomere integrity to mitochondrial oxidative metabolism. Overall, the analytical approach used in this study offer the possibility to observe higher levels of correlation (i.e. common expression trends) among genes, whose function seems to be aberrantly activated during the progression of muscle atrophy.

Accumulation of severely atrophic myofibers marks the acceleration of sarcopenia in slow and fast twitch muscles

The age-related decline in muscle mass, known as sarcopenia, exhibits a marked acceleration in advanced age. Although many studies have remarked upon the accumulation of very small myofibers, particularly at advanced stages of sarcopenia, the significance of this phenomenon in the acceleration of sarcopenia has never been examined. Furthermore, although mitochondrial dysfunction characterized by a lack of cytochrome oxidase (COX) activity has been implicated in myofiber atrophy in sarcopenia, the contribution of this phenotype to the accumulation of severely atrophied fibers in aged muscles has never been determined. To this end, we examined the fiber size distribution in the slow twitch soleus (Sol) and fast twitch gastrocnemius (Gas) muscles between young adulthood (YA) and senescence (SEN). We also quantified the abundance of COX deficient myocytes and their size attributes to gain insight into the contribution of this phenotype to myofiber atrophy with aging. Our data showed that the progression of muscle atrophy, particularly its striking acceleration between late middle age and SEN, was paralleled by an accumulation of severely atrophic myofibers (≤ 1000 μm 2 in size) in both Sol and Gas. On the other hand, we observed no COX deficient myofibers in Sol, despite nearly 20% of the myofibers being severely atrophic. Similarly, only 0.17 ± 0.06% of all fibers in Gas were COX deficient, and their size was generally larger (2375 ± 319 μm 2) than the severely atrophied myofibers noted above. Collectively, our results suggest that similar processes likely contribute to the acceleration of sarcopenia in both slow twitch and fast twitch muscles, and that COX deficiency is not a major contributor to this phenomenon.

Muscle pathology in upper motor neuron paraplegia

During the last 30 years acceptance of the muscle biopsy by paraplegic patients have accumulated a large body of knowledge on effects of spinal cord injury (SCI) on the skeletal muscle microstructure. Light and electron microscopy analyses combined with computerized methods provided a large body of knowledge on the progression of muscle atrophy and on the effects of rehabilitation strategies. Beside a summary of normal human muscle results, we will here discuss the time-course of muscle events related to SCI affecting the upper motor neuron. Furthermore the concomitant events on the microcirculation of skeletal muscle will be described in relation to the frequent complications of deep venous thrombosis, decubitus ulcers and paraosteoarthrosis.

Muscle pathology in upper motor neuron paraplegia

During the last 30 years acceptance of the muscle biopsy by paraplegic patients have accumulated a large body of knowledge on effects of spinal cord injury (SCI) on the skeletal muscle microstructure. Light and electron microscopy analyses combined with computerized methods provided a large body of knowledge on the progression of muscle atrophy and on the effects of rehabilitation strategies. Beside a summary of normal human muscle results, we will here discuss the time-course of muscle events related to SCI affecting the upper motor neuron. Furthermore the concomitant events on the microcirculation of skeletal muscle will be described in relation to the frequent complications of deep venous thrombosis, decubitus ulcers and paraosteoarthrosis.

Inhibitory Effect of a Combination of Thermotherapy with Exercise Therapy on Progression of Muscle Atrophy

[Purpose] This study was performed to investigate the inhibitory effect of a short-term thermal load in combination with treadmill running or neuromuscular electrical stimulation on the progression of disuse muscle atrophy and its causative mechanism. [Subjects and Methods] The ankle joints of Wistar rats were fixed at maximum plantar flexion, and the fixation was removed for intervention once a day. The rats were divided into 6 groups for intervention: control, fixation, treadmill running, thermal treadmill running, neuromuscular electrical stimulation, and thermal neuromuscular electrical stimulation groups. [Results] The progression of disuse muscle atrophy was inhibited only in the group given a thermal load before treadmill running. Regarding the atrophy mechanism, the expression of heat shock protein 72, which promotes protein synthesis and repair, was enhanced, while the expression of protein ligase E3, which promotes protein degradation, was inhibited. [Conclusion] These findings suggest that thermotherapy prior to exercise therapy promotes the effect of exercise therapy in patients with disuse muscle atrophy and elderly persons.

Ouabain exacerbates botulinum neurotoxin-induced muscle paralysis via progression of muscle atrophy in mice

Botulinum neurotoxin serotype A (BoNT/A) inhibits acetylcholine release at the neuromuscular junction in isolated muscles, and ouabain can partially block its effect. However, it is not clear whether ouabain attenuates BoNT/A-induced neuromuscular paralysis in vivo. In this work, we investigated the effects of ouabain on BoNT/A-induced neuromuscular paralysis in mice. Ouabain was administered to mice intraperitoneally immediately after a single injection of BoNT/A into skeletal muscle. The effects of ouabain on BoNT/A-induced muscle paralysis were assessed by quantitative monitoring of muscle tension and digit abduction via the digit abduction scoring (DAS) assay. A single administration of ouabain significantly prolonged BoNT/A-induced neuromuscular paralysis. Moreover, consecutive daily injection of ouabain exacerbated BoNT/A-induced neuromuscular paralysis, and led to a significant decrease in both twitch and tetanic forces as assayed in isolated BoNT/A-injected muscles. We next looked at the effects of ouabain on BoNT/A-induced muscle atrophy. Administration of ouabain led to a decrease in the myofibrillar cross-sectional area (CSAs) by 14 post-BoNT/A injection. In addition, repeated administration of ouabain increased mRNA expression levels of ubiquitin ligases, which are markers of muscle atrophy, in BoNT/A-injected muscle. These results suggest that ouabain exacerbates BoNT/A-induced neuromuscular paralysis via a marked progression of BoNT/A-induced muscle atrophy.

MG53 Regulates Membrane Budding and Exocytosis in Muscle Cells

Membrane recycling and remodeling contribute to multiple cellular functions, including cell fusion events during myogenesis. We have identified a tripartite motif (TRIM72) family member protein named MG53 and defined its role in mediating the dynamic process of membrane fusion and exocytosis in striated muscle. MG53 is a muscle-specific protein that contains a TRIM motif at the amino terminus and a SPRY motif at the carboxyl terminus. Live cell imaging of green fluorescent protein-MG53 fusion construct in cultured myoblasts showed that although MG53 contains no transmembrane segment it is tightly associated with intracellular vesicles and sarcolemmal membrane. RNA interference-mediated knockdown of MG53 expression impeded myoblast differentiation, whereas overexpression of MG53 enhanced vesicle trafficking to and budding from sarcolemmal membrane. Co-expression studies indicated that MG53 activity is regulated by a functional interaction with caveolin-3. Our data reveal a new function for TRIM family proteins in regulating membrane trafficking and fusion in striated muscles.

Stable muscle atrophy in long-term paraplegics with complete upper motor neuron lesion from 3- to 20-year SCI

Unrandomized trial. To investigate the structural and functional relationships and the progression of muscle atrophy up to 20 years of spastic paraplegia. Clinical follow-up in Vienna, Austria; muscle biopsies analyzed by light microscopy in Padova and by electron microscopy (EM) in Chieti, Italy. Force was measured as knee extension torque; trophism by computer tomography scan; tissue composition and fiber morphology by histopathology and EM. In the long-term group of patients (17.0+/-2.6 years), force and size of thigh muscles were only slightly different from those of mid-term subjects (2.2+/-0.5 years). Histology and ultrastructure confirm that the difference in average size of muscle fibers between long-term and mid-term paralyzed leg muscles is actually very small. In addition, muscle fibers maintain the striated appearance characteristic of normal skeletal fibers even after 14-20 years of paralysis. Ultrastructural alterations of the activating and metabolic machineries, and the presence of fibers with lower motor neuron denervation features, may explain the low-force output and the reduced endurance of paretic muscles. The stable muscle atrophy that characterizes long-lasting spastic paraplegia suggests that there are no upper-time limits to begin a training program based on functional electrical stimulation.

Mergla K + channel induces skeletal muscle atrophy by activating the ubiquitin proteasome pathway

Skeletal muscle atrophy results from an imbalance in protein degradation and protein synthesis and occurs in response to injury, various disease states, disuse, and normal aging. Current treatments for this debilitating condition are inadequate. More information about mechanisms involved in the onset and progression of muscle atrophy is necessary for development of more effective therapies. Here we show that expression of the mouse ether-a-go-go related gene (Merg1a) K+ channel is up-regulated in skeletal muscle of mice experiencing atrophy as a result of both malignant tumor expression and disuse. Further, ectopic expression of Merg1a in vivo induces atrophy in healthy wt-bearing mice, while expression of a dysfunctional Merg1a mutant suppresses atrophy in hindlimb-suspended mice. Treatment of hindlimb-suspended mice with astemizole, a known Merg1a channel blocker, inhibits atrophy in these animals. Importantly, in vivo expression of Merg1a in mouse skeletal muscle activates the ubiquitin proteasome pathway that is responsible for the majority of protein degradation that causes muscle atrophy, yet expression of a dysfunctional Merg1a mutant decreases levels of ubiquitin-proteasome proteolysis. Thus, expression of Merg1a likely initiates atrophy by activating ubiquitin-proteasome proteolysis. This gene and its product are potential targets for prevention and treatment of muscle atrophy.