Denervated Muscle Atrophy Research Articles

PHD3 mediates denervation skeletal muscle atrophy through Nf-κB signal pathway.

Skeletal muscle is the largest organ of the body, the development of skeletal muscle is very important for the health of the animal body. Prolyl hydroxylases (PHDs) are the classical regulator of the hypoxia inducible factor (HIF) signal pathway, many researchers found that PHDs are involved in the muscle fiber type transformation, muscle regeneration, and myocyte differentiation. However, whether PHDs can impact the protein turnover of skeletal muscle is poorly understood. In this study, we constructed denervated muscle atrophy mouse model and found PHD3 was highly expressed in the atrophic muscles and there was a significant correlation between the expression level of PHD3 and skeletal muscle weight which was distinct from PHD1 and PHD2. Then, the similar results were getting from the different weight muscles of normal mice. To further verify the relationship between PHD3 and skeletal muscle protein turnover, we established a PHD3 interference model by injecting PHD3 sgRNA virus into tibialis anterior muscle (TA) muscle of MCK-Cre-cas9 mice and transfecting PHD3 shRNA lentivirus into primary satellite cells. It was found that the Knock-out of PHD3 in vivo led to a significant increase in muscle weight and muscle fiber area (P<.05). Besides, the activity of protein synthesis signal pathway increased significantly, while the protein degradation pathway was inhibited evidently (P<.05). In vitro, the results of 5-ethynyl-2'-deoxyuridine (EdU) and tetramethylrhodamine ethyl ester (TMRE) fluorescence detection showed that PHD3 interference could lead to a decrease in cell proliferation and an increase of cell apoptosis. After the differentiation of satellite cells, the production of puromycin in the interference group was higher than that in the control group, and the content of 3-methylhistidine in the interference group was lower than that in the control group (P<.05) which is consistent with the change of protein turnover signal pathway in the cell. Mechanistically, there is an interaction between PHD3, NF-κB, and IKBα which was detected by immunoprecipitation. With the interfering of PHD3, the expression of the inflammatory signal pathway also significantly decreased (P<.05). These results suggest that PHD3 may affect protein turnover in muscle tissue by mediating inflammatory signal pathway. Finally, we knocked out PHD3 in denervated muscle atrophy mice and LPS-induced myotubes atrophy model. Then, we found that the decrease of PHD3 protein level could alleviate the muscle weight and muscle fiber reduction induced by denervation in mice. Meanwhile, the protein level of the inflammatory signal pathway and the content of 3-methylhistidine in denervated atrophic muscle were also significantly reduced (P<.05). In vitro, PHD3 knock-out could alleviate the decrease of myotube diameter induced by LPS, and the expression of protein synthesis pathway was also significantly increased (P<.05). On the contrary, the expression level of protein degradation and inflammatory signal pathway was significantly decreased (P<.05). Through these series of studies, we found that the increased expression of PHD3 in denervated muscle might be an important regulator in inducing muscle atrophy, and this process is likely to be mediated by the inflammatory NF-κB signal pathway.

COVID-19 related muscle denervation atrophy

COVID-19 has presented with a variety of manifestations including peripheral neurological symptoms. The most commonly associated peripheral neuropathies described with COVID-19 are Guillain–Barre syndrome and its variants as well as critical illness polyneuropathy. We report in this paper the distinct MRI findings of an unusual case of peripheral neuropathy associated with COVID-19. These findings are similar to those seen in Guillain–Barre syndrome or one of its variants, although differing from the classic condition in certain key clinical and radiological features.

Hyperbaric Oxygen Therapy Attenuates Burn-Induced Denervated Muscle Atrophy.

Background: Neuronal apoptosis and inflammation in the ventral horn of the spinal cord contribute to denervated muscle atrophy post-burn. Hyperbaric oxygen therapy (HBOT) exerts anti-inflammation and neuroprotection. Furthermore, hypoxia-inducible factor (HIF)-1α has been reported to promote inflammation and apoptosis. We investigated the therapeutic potential of HBOT and the role of HIF-1α post-burn.Methods: Sprague-Dawley rats were divided into three groups: a control group, an untreated burn group receiving burn and sham treatment, and a HBOT group receiving burn injury and HBOT. The burn injury was induced with 75ºC ± 5ºC at the right hindpaw. HBOT (100% oxygen at 2.5 atmosphere, 90 min/day) and sham HBOT (21% oxygen at 1 atmosphere, 90 min/day) was started on day 28 after burn injury and continued for 14 treatments (days 28-41). Incapacitance (hind limb weight bearing) testing was conducted before burn and weekly after burn. At day 42 post-burn, the gastrocnemius muscle and the spinal cord ventral horn were analyzed.Results: HBOT improved burn-induced weight bearing imbalance. At day 42 post-burn, less gastrocnemius muscle atrophy and fibrosis were noted in the HBOT group than in the untreated burn group. In the ventral horn, HBOT attenuated the neuronal apoptosis and glial activation post-burn. The increases in phosphorylated AKT/mTOR post-burn were reduced after HBOT. HBOT also inhibited HIF-1α signaling, as determined by immunofluorescence and western blot.Conclusions: HBOT reduces burn-induced neuronal apoptosis in the ventral horn, possibly through HIF-1α signaling.

Combining reverse end-to-side neurorrhaphy with rapamycin treatment on chronically denervated muscle in rats

This preliminary research determines whether a combination of reverse end-to-side neurorrhaphy and rapamycin treatment achieves a better functional outcome than a single application after prolonged peripheral nerve injury. We found that the tibial nerve function of the reverse end-to-side + rapamycin group recovered better, with a higher tibial function index value, higher amplitude recovery rate, and shorter latency delay rate (P < 0.05). The reverse end-to-side + rapamycin group better protected the gastrocnemius muscle with more forceful contractility, tetanic tension, and a higher myofibril cross-sectional area (P < 0.05). Combining reverse end-to-side neurorrhaphy with rapamycin treatment is a practical approach to promoting the recovery of chronically denervated muscle atrophy after peripheral nerve injury.

Ficus carica L. Attenuates Denervated Skeletal Muscle Atrophy via PPARα/NF-κB Pathway.

Treatment options for denervated skeletal muscle atrophy are limited, in part because the underlying molecular mechanisms are not well understood. Unlike previous transcriptomics studies conducted in rodent models of peripheral nerve injury, in the present study, we performed high-throughput sequencing with denervated atrophic biceps muscle and normal (non-denervated) sternocleidomastoid muscle samples obtained from four brachial plexus injury (BPI) patients. We also investigated whether Ficus carica L. (FCL.) extract can suppress denervated muscle atrophy in a mouse model, along with the mechanism of action. We identified 1471 genes that were differentially expressed between clinical specimens of atrophic and normal muscle, including 771 that were downregulated and 700 that were upregulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that the differentially expressed genes were mainly enriched in the GO terms “structural constituent of muscle,” “Z disc,” “M band,” and “striated muscle contraction,” as well as “Cell adhesion molecules,” “Glycolysis/Gluconeogenesis,” “Peroxisome proliferator-activated receptor alpha (PPARα) signaling pathway,” and “P53 signaling pathway.” In experiments using mice, the reduction in wet weight and myofiber diameter in denervated muscle was improved by FCL. extract compared to saline administration, which was accompanied by downregulation of the proinflammatory cytokines interleukin (IL)-1β and IL-6. Moreover, although both denervated groups showed increased nuclear factor (NF)-κB activation and PPARα expression, the degree of NF-κB activation was lower while PPARα and inhibitor of NF-κB IκBα expression was higher in FCL. extract-treated mice. Thus, FCL. extract suppresses denervation-induced inflammation and attenuates muscle atrophy by enhancing PPARα expression and inhibiting NF-κB activation. These findings suggest that FCL. extract has therapeutic potential for preventing denervation-induced muscle atrophy caused by peripheral nerve injury or disease.

Aspirin alleviates denervation-induced muscle atrophy via regulating the Sirt1/PGC-1α axis and STAT3 signaling.

BackgroundOur prior studies have shown that inflammation may play an important triggering role during the process of denervated muscle atrophy. The nonsteroidal anti-inflammatory drug aspirin exhibits the effect of anti-inflammatory factors. This study will investigate the protective effect of aspirin on denervated muscle atrophy and the underlying mechanism.MethodsMouse models of denervated muscle atrophy were established. The protective effect of aspirin (20 mg/kg/d, i.p.) on denervated muscle atrophy was analyzed using the wet weight ratio of tibialis anterior (TA) muscle and muscle fiber cross-sectional area (CSA). The levels of inflammatory factors were detected using quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. Sirtuins1 (SIRT1)/Peroxisome Proliferator-Activated Receptor γ Co-Activator 1α (PGC-1α) and Signal transducer and activator of transcription 3 (STAT3) signaling pathway and the muscle fiber type related proteins in TA muscle after denervation were analyzed by western blot assay.ResultsIntraperitoneal injection of aspirin (20 mg/kg/d) effectively alleviated denervation-induced muscle atrophy. This mainly manifested as follows: The wet weight ratio of TA muscle and muscle fiber CSA of mice treated with aspirin were significantly greater compared with mice treated with normal saline. The level of myosin heavy chain (MHC) increased, and the levels of muscle specific E3 ubiquitin ligase Muscle-specific RING finger-1 (MuRF-1) and muscle atrophy F-box (MAFbx) were decreased. Mitochondrial vacuolation and autophagy were inhibited, as evidenced by reduced level of autophagy related proteins PINK1, BNIP3, LC3B and Atg7 in mice treated with aspirin compared with mice treated with saline. In addition, aspirin treatment inhibited the slow-to-fast twitch muscle fiber conversion, which were related with triggering the expression of Sirt1 and PGC-1α. Moreover, aspirin reduced the levels of inflammatory factors interleukin-6, interleukin-1β and tumor necrosis factor-α and decreased the activation of STAT3 signaling pathway.ConclusionsThis is the first study to find that aspirin can alleviate denervation-induced muscle atrophy and inhibit the type I-to-type II muscle fiber conversion and mitophagy possibly through regulating the STAT3 inflammatory signaling pathway and Sirt1/PGC-1α signal axis. This study expands our knowledge regarding the pharmacological function of aspirin and provides a novel strategy for prevention and treatment of denervated muscle atrophy.

Therapeutic benefit of Muse cells in a mouse model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive motor neuron loss. Muse cells are endogenous reparative pluripotent-like stem cells distributed in various tissues. They can selectively home to damaged sites after intravenous injection by sensing sphingosine-1-phosphate produced by damaged cells, then exert pleiotropic effects, including tissue protection and spontaneous differentiation into tissue-constituent cells. In G93A-transgenic ALS mice, intravenous injection of 5.0 × 104 cells revealed successful homing of human-Muse cells to the lumbar spinal cords, mainly at the pia-mater and underneath white matter, and exhibited glia-like morphology and GFAP expression. In contrast, such homing or differentiation were not recognized in human mesenchymal stem cells but were instead distributed mainly in the lung. Relative to the vehicle groups, the Muse group significantly improved scores in the rotarod, hanging-wire and muscle strength of lower limbs, recovered the number of motor neurons, and alleviated denervation and myofiber atrophy in lower limb muscles. These results suggest that Muse cells homed in a lesion site-dependent manner and protected the spinal cord against motor neuron death. Muse cells might also be a promising cell source for the treatment of ALS patients.

Isoquercitrin Delays Denervated Soleus Muscle Atrophy by Inhibiting Oxidative Stress and Inflammation.

Although denervated muscle atrophy is common, the underlying molecular mechanism remains unelucidated. We have previously found that oxidative stress and inflammatory response may be early events that trigger denervated muscle atrophy. Isoquercitrin is a biologically active flavonoid with antioxidative and anti-inflammatory properties. The present study investigated the effect of isoquercitrin on denervated soleus muscle atrophy and its possible molecular mechanisms. We found that isoquercitrin was effective in alleviating soleus muscle mass loss following denervation in a dose-dependent manner. Isoquercitrin demonstrated the optimal protective effect at 20 mg/kg/d, which was the dose used in subsequent experiments. To further explore the protective effect of isoquercitrin on denervated soleus muscle atrophy, we analyzed muscle proteolysis via the ubiquitin-proteasome pathway, mitophagy, and muscle fiber type conversion. Isoquercitrin significantly inhibited the denervation-induced overexpression of two muscle-specific ubiquitin ligases—muscle RING finger 1 (MuRF1) and muscle atrophy F-box (MAFbx), and reduced the degradation of myosin heavy chains (MyHCs) in the target muscle. Following isoquercitrin treatment, mitochondrial vacuolation and autophagy were inhibited, as evidenced by reduced level of autophagy-related proteins (ATG7, BNIP3, LC3B, and PINK1); slow-to-fast fiber type conversion in the target muscle was delayed via triggering expression of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α); and the production of reactive oxygen species (ROS) in the target muscle was reduced, which might be associated with the upregulation of antioxidant factors (SOD1, SOD2, NRF2, NQO1, and HO1) and the downregulation of ROS production-related factors (Nox2, Nox4, and DUOX1). Furthermore, isoquercitrin treatment reduced the levels of inflammatory factors—interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α)—in the target muscle and inactivated the JAK/STAT3 signaling pathway. Overall, isoquercitrin may alleviate soleus muscle atrophy and mitophagy and reverse the slow-to-fast fiber type conversion following denervation via inhibition of oxidative stress and inflammatory response. Our study findings enrich the knowledge regarding the molecular regulatory mechanisms of denervated muscle atrophy and provide a scientific basis for isoquercitrin as a protective drug for the prevention and treatment of denervated muscle atrophy.

Detergent-insoluble inclusion constitutes the first pathology in PFN1 transgenic rats.

Mutation of profilin 1 (PFN1) can cause amyotrophic lateral sclerosis (ALS). To assess how PFN1 mutation causes the disease, we created transgenic rats with human genomic DNA that harbors both the coding and the regulatory sequences of the human PFN1 gene. Selected transgenic lines expressed human PFN1 with or without the pathogenic mutation C71G at a moderate and a comparable level and in the similar pattern of spatial and temporal expression to rat endogenous PFN1. The artificial effects of arbitrary transgene expression commonly observed in cDNA transgenic animals were minimized in PFN1 transgenic rats. Expression of the mutant, but not the wild type, human PFN1 in rats recapitulated the cardinal features of ALS including the progressive loss of motor neurons and the subsequent denervation atrophy of skeletal muscles. Detergent-insoluble PFN1 inclusions were detected as the first pathology in otherwise asymptomatic transgenic rats expressing mutant human PFN1. The findings suggest that protein aggregation is involved in the neurodegeneration of ALS associated with PFN1 mutation. The resulting rat model is useful to mechanistic study on the ALS.

Modulation of HOXA9 after skeletal muscle denervation and reinnervation.

Homeobox A9 (HOXA9), the expression of which is promoted by mixed lineage leukemia 1 (MLL1) and WD-40 repeat protein 5 (WDR5), is a homeodomain-containing transcription factor that plays an essential role in regulating stem cell activity. HOXA9 has been found to inhibit skeletal muscle regeneration and delay recovery after muscle wounding in aged mice, but little is known about its role in denervated/reinnervated muscles. We performed detailed time-dependent expression analyses of HOXA9 and its promoters, MLL1 and WDR5, in rat gastrocnemius muscles after the following three types of sciatic nerve surgeries: nerve transection (denervation), end-to-end repair (repair), and sham operation (sham). Then, the specific mechanisms of HOXA9 were detected in vitro by transfecting primary satellite cells with empty pIRES2-DsRed2, pIRES2-DsRed2-HOXA9, empty pPLK/GFP-Puro, and pPLK/GFP-Puro-HOXA9 small hairpin RNA (shRNA) plasmids. We found, for the first time, that HOXA9 protein expression simultaneously increased with increasing denervated muscle atrophy severity and that upregulated MLL1 and WDR5 expression was partly associated with denervation. Indeed, in vitro experiments revealed that HOXA9 inhibited myogenic differentiation, affected the best known atrophic signaling pathways, and promoted apoptosis but did not eliminate the differentiation potential of primary satellite cells. HOXA9 may promote denervated muscle atrophy by regulating the activity of satellite cells.

Prevention of denervated muscle atrophy with accelerated nerve-regeneration by babysitter procedure in rat facial nerve paralysis model.

The "babysitter" procedure is a reconstruction technique for facial nerve complete paralysis and uses the movement source from the healthy facial nerve with a cross-nerve graft. First, an end-to-side neurorrhaphy is performed between the affected facial nerve trunk and hypoglossal nerve for continuously delivering stimuli to the mimetic muscles for preventing the atrophy of mimetic muscles. Despite favorable clinical results, histological and physiological mechanisms remain unknown. This study attempted to establish a model for the "babysitter" procedure and find its efficacy in rats with facial nerve complete paralysis. A total of 16 Lewis rats were used and divided into 2 groups; cross nerve graft (n = 8) and babysitter groups (n = 8). The facial nerve trunk was transected in both groups. Babysitter group underwent a two-stage procedure. Cross nerve graft group underwent only the transfer of nerve graft from the healthy side to affected side. The animals were assessed physiologically by compound muscle action potential (CMAP), and the regenerated nerve tissues were evaluated histopathologically at 13 weeks after surgery. Facial nucleus stained with retrograde tracers proved the re-innervation of affected facial muscle by the babysitter procedure. In CMAP, the amplitude of babysitter group was significantly higher than that of the cross-facial nerve graft group (p < .05). Histological examination found a significant difference in myelin g-ratio between two groups (p < .05). This study investigated the "babysitter" procedure for rat facial nerve palsy. Babysitter procedure shortened the denervation period without mimic muscle atrophy.

P2X7 activation enhances skeletal muscle metabolism and regeneration in SOD1G93A mouse model of amyotrophic lateral sclerosis.

Muscle weakness plays an important role in neuromuscular disorders comprising amyotrophic lateral sclerosis (ALS). However, it is not established whether muscle denervation originates from the motor neurons, the muscles or more likely both. Previous studies have shown that the expression of the SOD1G93A mutation in skeletal muscles causes denervation of the neuromuscular junctions, inability to regenerate and consequent atrophy, all clear symptoms of ALS. In this work, we used SOD1G93A mice, a model that best mimics some pathological features of both familial and sporadic ALS, and we investigated some biological effects induced by the activation of the P2X7 receptor in the skeletal muscles. The P2X7, belonging to the ionotropic family of purinergic receptors for extracellular ATP, is abundantly expressed in the healthy skeletal muscles, where it controls cell duplication, differentiation, regeneration or death. In particular, we evaluated whether an in vivo treatment in SOD1G93A mice with the P2X7 specific agonist 2′(3′)‐O‐(4‐Benzoylbenzoyl) adenosine5′‐triphosphate (BzATP) just before the onset of a pathological neuromuscular phenotype could exert beneficial effects in the skeletal muscles. Our findings indicate that stimulation of P2X7 improves the innervation and metabolism of myofibers, moreover elicits the proliferation/differentiation of satellite cells, thus preventing the denervation atrophy of skeletal muscles in SOD1G93A mice. Overall, this study suggests that a P2X7‐targeted and site‐specific modulation might be a strategy to interfere with the complex multifactorial and multisystem nature of ALS.

UCHL1 regulates muscle fibers and mTORC1 activity in skeletal muscle

AimsSkeletal muscle wasting is associated with many chronic diseases. Effective prevention and treatment of muscle wasting remain as a challenging task due to incomplete understanding of mechanisms by which muscle mass is maintained and regulated. This study investigated the functional role of Ubiquitin C-terminal hydrolase L1 (UCHL1) in skeletal muscle. Main methodsMice with skeletal muscle specific gene knockout of UCHL1 and C2C12 myoblast cells with UCHL1 knockdown were used. Muscle fiber types and size were measured using tissue or cell staining. The mammalian target of rapamycin complex 1 (mTORC1) and mTORC2 activities were assessed with the phosphorylation of their downstream targets. Key findingsIn mouse skeletal muscle, UCHL1 was primarily expressed in slow twitch muscle fibers. Mice with skeletal muscle specific knockout (skmKO) of UCHL1 exhibited enlarged muscle fiber sizes in slow twitch soleus but not fast twitch extensor digitorum longus (EDL) muscle. Meanwhile, UCHL1 skmKO enhanced mTORC1 activity and reduced mTORC2 activity in soleus but not in EDL. Consistently, in C2C12 cells, UCHL1 knockdown increased the myotube size, enhanced mTORC1 activity, and reduced mTORC2 activities as compared with control cells. UCHL1 knockdown did not change the major proteins of mTOR complex but decreased the protein turnover of PRAS40, an inhibitory factor of mTORC1. SignificanceThese data revealed a novel function of UCHL1 in regulation of mTORC1 activity and skeletal muscle growth in slow twitch skeletal muscle. Given the upregulation of UCHL1 in denervation and spinal muscle atrophy, our finding advances understanding of regulators that are involved in muscle wasting.

Exosomes from human adipose-derived stem cells promote sciatic nerve regeneration via optimizing Schwann cell function.

Human adipose-derived stem cells (ASCs) have a potential for the treatment of peripheral nerve injury. Recent studies demonstrated that stem cells can mediate therapeutic effect by secreting exosomes. We aimed to investigate the effect of human ASCs derived exosomes (ASC-Exos) on peripheral nerve regeneration in vitro and in vivo. Our results showed after being internalized by Schwann cells (SCs), ASC-Exos significantly promoted SC proliferation, migration, myelination, and secretion of neurotrophic factors by upregulating corresponding genes in vitro. We next evaluated the efficacy of ASC-Exo therapy in a rat sciatic nerve transection model with a 10-mm gap. Axon regeneration, myelination, and restoration of denervation muscle atrophy in ASC-Exos treated group was significantly improved compared to vehicle control. This study demonstrates that ASC-Exos effectively promote peripheral nerve regeneration via optimizing SC function and thereby represent a novel therapeutic strategy for regenerative medicine and nerve tissue engineering.

A Potassium Channel Blocker, 4‐Aminopyridine, Attenuates Peripheral Nerve Injury‐induced Muscle Atrophy, Improves Intrinsic Muscle Force, and Suppresses the Expression of Muscle Atrophy‐related Transcription Factors in Mice

4‐Aminopyridine (4‐AP), a broad‐spectrum potassium channel blocker and FDA‐approved drug for the symptomatic treatment of multiple sclerosis, is used to improve neuromuscular function in patients with diverse demyelinating disorders. The neurological benefits of 4‐AP are believed to result from increases in action potential duration, calcium influx, neurotransmitter release and synaptic transmission. Traumatic peripheral nerve injury (TPNI) represents a major health problem that often leads to significant functional impairment and permanent disability from the loss in axonal continuity, neuronal cell death, nerve demyelination, nerve conduction defects, muscle denervation, and muscle atrophy. We have recently repurposed the use of 4‐AP and demonstrated that systemic 4‐AP administration enhances global functional recovery of the affected limb, promotes remyelination of the nerve and improves the nerve conduction velocity in a mouse model of TPNI. While muscle atrophy occurs very rapidly following nerve injury, the effect 4‐AP on muscle atrophy and muscle contractile function is largely unknown. This study was designed to explore the possible beneficial effects of 4‐AP treatment in muscle atrophy, intrinsic muscle function, and muscle regeneration following acute sciatic nerve crush injury. Mice were assigned to moderate sciatic nerve crush injury and no‐injury groups and followed for 3, 7 and 14 days with/without 4‐AP (10 mg/daily, intraperitoneal injection) or saline treatment. Morphological, functional and transcriptional properties of skeletal muscle were assessed. In addition to improving in vivo global motor function as early as post‐operative day 3, 4‐AP treatment significantly attenuated muscle atrophy of the injured limb with increased muscle mass and muscle fiber area. 4‐AP also improved ex vivo intrinsic muscle contractile force 7 days post‐injury. Most importantly, the reduced muscle atrophy with 4‐AP treatment was concurrent with significantly reduced expression of atrophy‐related genes (myogenin, MuRF‐1, FoxO1, FoxO3) and increased expression of Pax7+ satellite cells and proliferating Ki67+ cells. These findings provide new insights into the beneficial effects of 4‐AP in nerve injury‐induced muscle atrophy and dysfunction and open a new window for further investigation.Support or Funding InformationThis work was supported by grants from the NIH (K08 AR060164‐01A) and DOD (W81XWH‐16‐1‐0725) in addition to institutional support from the University of Rochester and Pennsylvania State University Medical Centers.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

"Isolated" Suprascapular Neuropathy: Compression, Traction, or Inflammation?

Several hypotheses have been proposed for the pathophysiology of suprascapular nerve (SSN) palsy, including compression, traction, and nerve inflammation. To provide insight into the pathophysiology of isolated nontraumatic SSN palsy by performing critical reinterpretations of electrodiagnostic (EDX) studies and magnetic resonance (MR) images of patients with such diagnosis. We retrospectively reviewed all patients referred to our institution for the past 20 yr with a diagnosis of nontraumatic isolated suprascapular neuropathy who had an upper extremity EDX study and a shoulder or brachial plexus MR scan. Patient charts were reviewed to analyze their initial clinical examination, and their original EDX study and MR images were reinterpreted by an experienced neurologist and a musculoskeletal radiologist, respectively, both blinded from the authors' hypothesis and from each other's findings. Fifty-nine patients were included. Fifty of them (85%) presented with at least 1 finding that was inconsistent with an isolated SSN palsy. Forty patients (68%) had signs on physical examination beyond the SSN distribution. Thirty-one patients (53%) had abnormalities on their EDX studies not related to the SSN. Twenty-two patients (37%) had denervation atrophy in other muscles than the spinati, or neural hyperintensity in other nerves than the SSN on their MR scans, without any evidence of SSN extrinsic compression. The great majority of patients with presumed isolated SSN palsy had clinical, electrophysiological, and/or imaging evidence of a more diffuse pattern of neuromuscular involvement. These data strongly support an inflammatory pathophysiology in many cases of "isolated" SSN palsy.

Histone deacetylase 4 protects from denervation and skeletal muscle atrophy in a murine model of amyotrophic lateral sclerosis

BackgroundHistone deacetylase 4 (HDAC4) has been proposed as a target for Amyotrophic Lateral Sclerosis (ALS) because it mediates nerve-skeletal muscle interaction and since its expression in skeletal muscle correlates with the severity of the disease. However, our recent studies on the skeletal muscle response upon long-term denervation highlighted the importance of HDAC4 in maintaining muscle integrity.MethodsTo fully identify the yet uncharacterized HDAC4 functions in ALS, we genetically deleted HDAC4 in skeletal muscles of a mouse model of ALS. Body weight, skeletal muscle, innervation and spinal cord were analyzed over time by morphological and molecular analyses. Transcriptome analysis was also performed to delineate the signaling modulated by HDAC4 in skeletal muscle of a mouse model of ALS.FindingsHDAC4 deletion in skeletal muscle caused earlier ALS onset, characterized by body weight loss, muscle denervation and atrophy, and compromised muscle performance, although the main catabolic pathways were not activated. Transcriptome analysis identified the gene networks modulated by HDAC4 in ALS, revealing UCP1 as a top regulator that may be implicated in worsening ALS features.InterpretationHDAC4 plays an important role in preserving innervations and skeletal muscle in ALS, likely by modulating the UCP1 gene network. Our study highlights a possible risk in considering HDAC inhibitors for the treatment of ALS.FundThis work was supported by FIRB grant (RBFR12BUMH) from Ministry of Education, Universities and Research, by Fondazione Veronesi, by Sapienza research project 2017 (RM11715C78539BD8) and Polish National Science Center grant (UMO-2016/21/B/NZ3/03638).

Effect of electroacupuncture on denervated skeletal muscle atrophy and expression of IGF-1/PI3K/AKT in rats

To observe the effects of electroacupuncture (EA) on denervated skeletal muscle atrophy and expression of IGF-1/PI3K/AKT signaling pathway in rats, and to explore the possible mechanisms of it. Thirty-six male SD rats were randomly divided into a control group, a model group and an EA group. Denervated muscle atrophy model was made by cutting off the left hindlimb sciatic nerve in the model group and the EA group. In the EA group, EA, dense-disperse wave, 2 Hz/33 Hz and 1.5-2 mA, was connected at "Zusanli" (ST 36) and "Shangjuxu" (ST 37). The treatment was given for 30 min every time, once a day and a total of 20 times were needed, there was 2 days interval every 5 times. The rats in the control group and model group were fixed in the same cage every day but not applied EA. Sciatic nerve function index (SFI) and ratio of muscle wet weight (RWW) were measured after treatment. HE staining was used to determine the ratio of cross-sectional area (RCA) and the ratio of fiber diameter (RFD). TUNEL and double immunofluorescence staining were used to evaluate the myocyte apoptosis, proliferation and differentiation. The mRNA expressions of IGF-1 and AKT were detected by real-time PCR. The total protein of AKT, p-AKT (Ser473) and p-AKT (Thr308) were detected by western blot. Compared with the control group, SFI, RWW, RCA and RFD of the model group were significantly decreased and apoptotic index of myocytes were significantly increased (all P<0.01). Compared with the model group, the above indexes of the EA group were improved (all P<0.05), and the expressions of PAX3 and PAX7 related to proliferation and differentiation of muscle satellite cells were significantly increased (both P<0.01), p-AKT (Ser473) protein level was increased (P<0.05). EA can improve the atrophy of denervated muscles, reduce myocyte apoptosis, and promote the proliferation and differentiation of muscle satellite cells. The mechanism may be related to the increase of p-AKT (Ser473) level and activation of AKT signaling pathway.

Mutant UBQLN2P497H in motor neurons leads to ALS-like phenotypes and defective autophagy in rats

Mutations in ubiquilin2 (UBQLN2) have been linked to abnormal protein aggregation in amyotrophic lateral sclerosis (ALS). The mechanisms underlying UBQLN2-related neurodegenerative diseases remain unclear. Using a tetracycline-regulated gene expression system, the ALS-linked UBQLN2P497H mutant was selectively expressed in either the spinal motor neurons or astrocytes in rats. We found that selectively expressing mutant UBQLN2P497H in the spinal motor neurons caused several core features of ALS, including the progressive degeneration of motor neurons, the denervation atrophy of skeletal muscles, and the abnormal protein accumulation. Furthermore, mutant UBQLN2P497H accumulation was associated with an age-dependent decrease in several core autophagy-related proteins. ALS-like phenotypes were not observed when mutant UBQLN2P497H was overexpressed in the astrocytes, however, even though the expression of the mutant UBQLN2P497H protein was higher in these rats. Our results suggest that selectively expressing mutant UBQLN2P497H in motor neurons is sufficient to trigger the development of ALS in rats. Our results further indicate that the compromised autophagy-lysosomal pathway plays a critical role in the pathogenesis of UBQLN2-related neurodegenerative diseases.

Study on denervated rat skeletal muscle by 7.0 T 31P-MR phosphorus spectroscopy

Objective To investigate the changes of 31P-MRS in denervated skeletal muscle at 7.0 T MR system. Methods In the experiment group, a total of 18 male Sprague-Dawley rats aged 6-8 weeks old and weighing 200-250 g were obtained. The right posterior femoral nerve were transected, and the proximal stumps were ligated by using 5-0 nylon stitches to preclude spontaneous repair. A sham surgery (incision and exploration of the nerve) was performed at the same time (n=6). Before rat model established and at varying times after initial surgery (3 d, and 1, 2, 4, 6, and 8 weeks). 31P-MR spectra of rat quadriceps femoris were acquired on 7.0 T Agilent small animal MR imaging system. We quantified the phosphocreatine (PCr), adenosine triphosphate(β-ATP), and inorganic phosphate (Pi) using Creatine phosphate disodium salt(10 mmol/L) as an external standard. The ratios, include PCr/Pi, Pi/β-ATP, PCr/β-ATP, PCr/(PCr+Pi) and Pi/(PCr+Pi) were quantified in jMRUI. All data were analyzed by one way analysis of variance (ANOVA) with posttest inter-group comparisons using Bonferroni test. Comparative analysis methods between the experimental and control group were performed via independent samples t test. P<0.05 was considered as statistically significance. Results The average measured concentrations of β-ATP, PCr, and Pi in control groups were (6.654±0.178) μmol/g, (25.656±0.738) μmol/g, and (1.594±0.096) μmol/g, respectively. There were significant statistically differences in β-ATP, PCr, Pi, PCr/Pi, PCr/PCr+Pi,Pi/PCr+Pi between the study and control group(P<0.05) at any measurement time point after denervation. Pi/β-ATP,PCr/β-ATP and pHi in the experimental group significantly statistically differed from those of the control group (all P<0.05)at each time point except on day 3 after operation. The total concentrations of β-ATP and PCr were reduced by 11.5% and 19.7% respectively on day 3. Thereafter, β-ATP and PCr declined rapidly by 63.1% and 68.8% at week 4 respectively, then decreased slowely by 74.0%和82.3% till week 10. The change of PCr/Pi is similar to β-ATP and PCr, but more remarkable. Pi, Pi/PCr+Pi, Pi/ATP showed a progressively increase till week 10. The intra-cellular pH (pHi) of normal rat muscles was 7.033±0.017, While the pHi gently in experiment rat muscles decreased during the entire experiment. Conclusion 31P-MRS with 7.0 T can quantify the temporal changes of energy metabolism and pHi in normal and denervated rat muscles. It shows that the dysfunction of energy metabolism are progressive with time and that they begin within a short period following the nerve section, the change of β-ATP, PCr, PCr/Pi take place primarily within 4 weeks after denervation. β-ATP, PCr, and PCr/Pi may be potential biomarkers of energy metabolism in the evaluation of denervated muscle atrophy. Key words: Magnetic resonance spectroscopy; Muscular disorders, atrophic; Denervation; Energy metabolism; Intracellular pH