Muscle Ring Finger Research Articles

Hydrogen sulfide regulates muscle RING finger-1 protein S-sulfhydration at Cys44 to prevent cardiac structural damage in diabetic cardiomyopathy.

Hydrogen sulfide (H2 S) plays important roles as a gasotransmitter in pathologies. Increased expression of the E3 ubiquitin ligase, muscle RING finger-1 (MuRF1), may be involved in diabetic cardiomyopathy. Here we have investigated whether and how exogenous H2 S alleviates cardiac muscle degradation through modifications of MuRF1 S-sulfhydration in db/db mice. Neonatal rat cardiomyocytes were treated with high glucose (40mM), oleate (100μM), palmitate (400μM), and NaHS (100μM) for 72hr. MuRF1 was silenced with siRNA technology and mutation at Cys44 . Endoplasmic reticulum stress markers, MuRF1 expression, and ubiquitination level were measured. db/db mice were injected with NaHS (39μmol·kg-1 ) for 20weeks. Echocardiography, cardiac ultrastructure, cystathionine-γ-lyase, cardiac structure proteins expression, and S-sulfhydration production were measured. H2 S levels and cystathionine-γ-lyase protein expression in myocardium were decreased in db/db mice. Exogenous H2 S reversed endoplasmic reticulum stress, including impairment of the function of cardiomyocytes and structural damage in db/db mice. Exogenous H2 S could suppress the levels of myosin heavy chain 6 and myosin light chain 2 ubiquitination in cardiac tissues of db/db mice, and MuRF1 was modified by S-sulfhydration, following treatment with exogenous H2 S, to reduce the interaction between MuRF1 and myosin heavy chain 6 and myosin light chain 2. Our findings suggest that H2 S regulates MuRF1 S-sulfhydration at Cys44 to prevent myocardial degradation in the cardiac tissues of db/db mice. This article is part of a themed section on Hydrogen Sulfide in Biology & Medicine. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v177.4/issuetoc.

Knee osteoarthritis induces atrophy and neuromuscular junction remodeling in the quadriceps and tibialis anterior muscles of rats

Knee osteoarthritis (KOA) is associated with muscle weakness, but it is unclear which structures are involved in the muscle changes. This study assessed morphological alterations and the expression of genes and proteins linked to muscular atrophy and neuromuscular junctions (NMJs) in KOA, induced by anterior cruciate ligament transection (ACLT) in rats. Two groups of rats were assessed: control (without intervention) and KOA (ACLT surgery in the right knee). After 8 weeks, quadriceps, tibialis anterior (TA) and gastrocnemius muscles were analyzed (area of muscle fibers, NMJ, gene and protein expression). KOA group showed atrophy in quadriceps (15.7%) and TA (33%), with an increase in atrogin-1 and muscle RING-finger protein-1 (MuRF-1). KOA group showed quadriceps NMJ remodeling (reduction area and perimeter) and decrease in NMJ diameter in TA muscle. The expression of nicotinic acetylcholine receptor (nAChR) γ-nAChR increased and that of α-nAChR and muscle specific tyrosine kinase (MuSK) declined in the quadriceps, with a decrease in ε-nAChR in TA. MuRF-1 protein expression increased in quadriceps and TA, with no changes in neural cell adhesion molecule (NCAM). In conclusion, ACLT-induced KOA promotes NMJ remodeling and atrophy in quadriceps and TA muscles, associated with inflammatory signs and changes in muscle gene and protein expression.

SUN-026 Selective Androgen Receptor Modulator S42 Has Anabolic and Anti-Catabolic Effects on Cultured Myotubes

Introduction: In an effort to develop a selective androgen receptor modulator (SARM) with beneficial activity in energy homeostasis, we previously screened 119 steroid analogs for the ability to induce uncoupling protein-1 (UCP-1) mRNA without increasing prostate-specific antigen promoter activity in human prostate cancer cell line, LNCap. As a result, we identified a novel SARM, S42, which is structural analog of testosterone. S42 dose not stimulate prostate growth but has a beneficial effect on lipid metabolism and also increased muscle weight of lavator ani in orchiectomized Sprague-Dawley rats. These findings prompted us to investigate whether S42 has a direct effect on cultured C2C12 myotubes. Method: Mouse myoblast C2C12 were seeded into 6well-plates at 2×105cells/well and maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS). When the cultured cells became 80-90% confluent, the medium was changed to DMEM supplemented with 2% horse serum (HS) and cells were further cultured for 4 days to induce differentiation into myotubes. Next, the medium was changed to DMEM containing 2% dextran-coated-stripped HS from one day before treatment with various drugs: S42, 5α-dihydrotestosterone (DHT), insulin and rapamycin were tested. Protein and mRNA expression levels were detected by Western blotting and Real time RT-PCR, respectively. Results: Muscle atrophy is mainly controlled by two specific genes encoding muscle ubiquitin ligase, atrogin1 and Muscle RING-Finger Protein1 (MuRF1). S42 significantly lowered expression levels of atrogin1 and MuRF1 in C2C12 myotubes, as determined by Real time RT-PCR. Phosphorylation of p70 S6 kinase (p70S6K), an essential factor for promoting protein synthesis in skeletal muscle, was significantly increased by S42 to almost the same extent as by insulin, but this was prevented by treatment with rapamycin, an inhibitor of mechanistic target of rapamycin complex 1 (mTORC1). However, phosphorylation of Akt upstream regulator mTORC1, was not changed by S42. S42 did not increase insulin-like growth factor 1 (IGF1) mRNA levels in C2C12 myotubes. Conclusion: These results suggest that S42 may have an anabolic effect through activation of mTORC1-p70S6K signaling, independent of IGF1-Akt signaling and may exert an anti-catabolic effect through inhibition of the degradation pathway in cultured C2C12 myotubes.

Identification and Characterization of Allograft Inflammatory Factor 1‐Like (Aif1L) in Skeletal Muscle

Skeletal muscle wasting, or atrophy, results from decreased protein synthesis and an associated increase in protein degradation. This shift towards a decrease in protein content of the cell can result from various physiological conditions, including immobilization, denervation, and aging. Muscle RING Finger 1 (MuRF1), an E3 ubiquitin ligase, has been found to be upregulated during almost all conditions of muscle wasting and is believed to play an important role in targeting proteins for degradation. However, preliminary data described in this study provides new evidence that MuRF1's role in the skeletal muscle atrophy cascade is likely more complex and it appears that MuRF1 is necessary for full transcriptional activation of a small subset of neurogenic atrophy‐induced genes, including allograft inflammatory factor 1‐like (Aif1L). Aif1L has been identified as a novel skeletal muscle gene that is upregulated in wild‐type mice, while MuRF1‐null mice show blunted upregulation in response to denervation. Aif1L is predicated to contain an EF‐hand domain, which can act as a calcium ion binding domain. Furthermore, Aif1L has also recently been implicated in regulating actomyosin contractility and filopodial extensions and may play a similar role in muscle cells. To confirm that Aif1L in expressed in muscle cells, qPCR was used to assess the expression levels in cultured muscle cells, while the cDNA of Aif1L was cloned from the C2C12 mouse myoblast cell line and fused with a green fluorescence protein (GFP) tag. C2C12 cells were then transfected with the Aif1L‐GFP expression plasmid in order to localize the Aif1L protein in proliferating myoblast cells. C2C12 cells transiently overexpressing Aif1L fused to GFP showed punctate fluorescence localized to the perinuclear region of the cytoplasm. The results presented here in combination with additional research into the transcriptional regulation and function of Aif1L will help contribute new insight into the molecular genetic mechanisms of skeletal muscle atrophy and may provide a more complete understanding of how MuRF1 may regulate the neurogenic atrophy cascade during muscle wasting.Support or Funding InformationThe work was support by University of North Florida Transformational Learning Opportunity grants and a University of North Florida Foundation Board Grant to D.W.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Dual Specificity Phosphatase and Pro Isomerase Domain Containing 1 (Dupd1) is Upregulated During Neurogenic Skeletal Muscle Atrophy and is Differentially Expressed in MuRF1‐null Mice

Skeletal muscle atrophy is caused by a decrease in muscle size and strength and results from a range of physiological conditions, including denervation, immobilization, corticosteroid exposure and aging. Dual Specificity Phosphatase and Pro Isomerase Domain Containing 1 (Dupd1) has been identified as a novel atrophy‐induced gene in skeletal muscle and has been found to be differentially regulated in wild‐type mice compared to Muscle RING Finger 1 (MuRF1) knockout mice in response to denervation. Muscle RING Finger 1 (MuRF1), an E3 ubiquitin ligase, has previously been shown to be upregulated under almost all conditions of skeletal muscle wasting. Preliminary data described in this study provides evidence that MuRF1 may play a more complex role in the muscle atrophy cascade than previously believed and appears to be necessary for full activation of a subset of neurogenic atrophy‐induced genes, including Dupd1. A recent study that utilized microarray analysis of skeletal muscle isolated from wild‐type and MuRF1‐null mice revealed that Dupd1 expression is significantly induced at 3 days and 14 days post‐denervation in wild‐type mice, while Dupd1 expression is dramatically blunted in the MuRF1‐null mice at both 3 days and 14 days post‐denervation. qPCR analysis revealed that Dupd1 is significantly upregulated during differentiation of cultured muscle cells. To determine how Dupd1 might be transcriptionally regulated in skeletal muscle, fragments of the promoter region of Dupd1 were cloned, fused to a reporter gene and found to be highly inducible in response to ectopic expression of MyoD and myogenin. Furthermore, site‐directed mutagenesis of conserved E‐box elements in the proximal promoter of Dupd1 rendered the Dupd1 reporter genes incapable of being induced by overexpression of myogenic regulatory factors. The Dupd1 gene codes for a dual specificity phosphatase (DUSP) that can recognize and dephosphorylate tyrosine, serine and threonine amino acids of substrate proteins. Finally, we have found that Dupd1, an atypical DUSP, can modulate the ERK branch of the MAP Kinase signaling in muscle cells and inhibits muscle cell differentiation when ectopically expressed in proliferating muscle cells. Elucidating the role of Dupd1 in muscle will help contribute new insight into the molecular mechanisms for skeletal muscle atrophy.Support or Funding InformationThe work was support by University of North Florida Transformational Learning Opportunity grants and a University of North Florida Foundation Board Grant to D.W.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Identification and Characterization of 1700029J07RIK, a Novel Gene Expressed in Skeletal Muscle

Skeletal muscle atrophy is the loss of muscle mass that results from a wide range of conditions including denervation, corticosteroid exposure, and aging. 1700029J07RIK is a putative novel gene suggested to be involved in skeletal muscle atrophy. The 1700029J07RIK gene was found to be induced following denervation in mice and is differentially expressed in Muscle RING Finger 1 (MuRF1) knockout mice compared to wild‐type mice. A previous microarray analysis of muscle tissue isolated from wild‐type control mice and MuRF1 knockout mice revealed that the expression of the 1700029J07RIK gene is unchanged at 3 days post‐denervation, while expression is significantly induced in wild‐type mice at 14 days post denervation but remains unchanged in the MuRF1‐null animals. It has been well established that muscle cells go through a distinct developmental timeline in which they proliferate as single cell myoblasts and then differentiate and fuse into multinucleated myotubes. Quantitative PCR was used to confirm that the 1700029J07RIK gene is expressed in proliferating cells and increases in expression as muscle cells differentiate. To characterize the transcriptional regulation of 1700029J07RIK, promoter fragments were cloned into a reporter plasmid, transfected into muscle cells and found to have significant transcriptional activity, while ectopic expression of myogenic regulatory factors appear to modulate the activity of the 1700029J07RIK reporter genes. In addition, the 1700029J07RIK cDNA was cloned from muscle cells and fused with green fluorescent protein and visualized by confocal fluorescent microscopy revealing an interesting perinuclear and cytoplasmic localization of the 1700029J07RIK protein in muscle cells. Finally, it was observed that overexpression of 1700029J07RIK resulted in inhibition of muscle cell differentiation and attenuation of the ERK branch of the MAP Kinase signaling pathway. While there is virtually nothing known about this novel gene in the context of skeletal muscle, the results presented here suggest 1700029J07RIK may play a role in muscle cell differentiation and in the skeletal muscle atrophy cascade.Support or Funding InformationThe work was support by University of North Florida Transformational Learning Opportunity grants and a University of North Florida Foundation Board Grant to D.W.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Vitamin C deficiency causes muscle atrophy and a deterioration in physical performance

L-Ascorbic acid (AsA) is a water-soluble antioxidant. We examined the effect of AsA deficiency on skeletal muscle using senescence marker protein-30 (SMP30)-knockout (KO) mice that are defective in AsA biosynthesis, which makes this mouse model similar to humans, to clarify the function of AsA in skeletal muscle. Eight-week-old female SMP30-KO mice were divided into the following two groups: an AsA-sufficient group [AsA(+)] that was administered 1.5 g/L AsA and an AsA-deficient group [AsA(−)] that was administered tap (AsA-free) water. At 4 weeks, the AsA content in the gastrocnemius muscle of AsA(−) mice was 0.7% compared to that in the gastrocnemius muscle of AsA(+) mice. Significantly lower weights of all muscles were observed in AsA(−) mice than those in AsA(+) mice at 12 and 16 weeks. The cross-sectional area of the soleus was significantly smaller in AsA(−) mice at 16 weeks than that in AsA(+) mice. The physical performance of AsA(−) mice was significantly less than that of AsA(+) mice at 12 weeks. Following AsA deficiency for 12 weeks, the expression of ubiquitin ligases, such as atrogin1/muscle atrophy F-box (MAFbx) and muscle RING-finger protein 1 (MuRF1), was upregulated. Furthermore, all detected effects of AsA deficiency on muscles of the AsA(−) group at 12 weeks were restored following AsA supplementation for 12 weeks. Thus, longer-term AsA deficiency is associated with muscle wasting, that this can be reversed by restoring AsA levels.

Doxorubicin Exposure Causes Subacute Cardiac Atrophy Dependent on the Striated Muscle-Specific Ubiquitin Ligase MuRF1.

Background Anthracycline chemotherapeutics, such as doxorubicin, are used widely in the treatment of numerous malignancies. The primary dose-limiting adverse effect of anthracyclines is cardiotoxicity that often presents as heart failure due to dilated cardiomyopathy years after anthracycline exposure. Recent data from animal studies indicate that anthracyclines cause cardiac atrophy. The timing of onset and underlying mechanisms are not well defined, and the relevance of these findings to human disease is unclear. Methods and Results Wild-type mice were sacrificed 1 week after intraperitoneal administration of doxorubicin (1-25 mg/kg), revealing a dose-dependent decrease in cardiac mass ( R2=0.64; P<0.0001) and a significant decrease in cardiomyocyte cross-sectional area (336±29 versus 188±14 µm2; P<0.0001). Myocardial tissue analysis identified a dose-dependent upregulation of the ubiquitin ligase, MuRF1 (muscle ring finger-1; R2=0.91; P=0.003) and a molecular profile of muscle atrophy. To investigate the determinants of doxorubicin-induced cardiac atrophy, we administered doxorubicin 20 mg/kg to mice lacking MuRF1 (MuRF1-/-) and wild-type littermates. MuRF1-/- mice were protected from cardiac atrophy and exhibited no reduction in contractile function. To explore the clinical relevance of these findings, we analyzed cardiac magnetic resonance imaging data from 70 patients in the DETECT-1 cohort and found that anthracycline exposure was associated with decreased cardiac mass evident within 1 month and persisting to 6 months after initiation. Conclusions Doxorubicin causes a subacute decrease in cardiac mass in both mice and humans. In mice, doxorubicin-induced cardiac atrophy is dependent on MuRF1. These findings suggest that therapies directed at preventing or reversing cardiac atrophy might preserve the cardiac function of cancer patients receiving anthracyclines.

Effects of leucine supplementation on muscle protein synthesis and degradation pathways in broilers at constant dietary concentrations of isoleucine and valine

ABSTRACTThe present study investigated the hypothesis that dietary concentrations of leucine (Leu) in excess of the breeder´s recommendations activates protein synthesis and decreases protein degradation in muscle of broilers. Day-old male Ross 308 broilers (n = 450) were phase-fed corn-soybean meal-based diets during starter (d 1–10), grower (d 11–22), and finisher (d 23–34) period. The basal diets fed to the control group (L0) met the broilers’ requirements for nutrients and amino acids, and contained Leu, Leu:isoleucine (Ile) and Leu:valine (Val) ratios, close to those recommended by the breeder (Leu:Ile: 100:54, 100:52, 100:51; Leu:Val 100:64, 100:61, 100:58; in starter, grower and finisher diet, resp.). Basal diets were supplemented with Leu to exceed the breeder’s recommendations by 35% (group L35) and 60% (group L60). Growth performance during 34 d, and carcass weights, and breast and thigh muscle weights on d 34 were similar among groups. Hepatic and muscle mRNA levels of genes involved in the somatotropic axis [growth hormone receptor, insulin-like growth factor (IGF)-1, IGF binding protein 2, IGF receptor] on d 34 were not influenced by Leu. In the breast muscle, relative mRNA abundances of genes involved in the mammalian target of rapamycin (mTOR) pathway of protein synthesis (mTOR, ribosomal p70 S6 kinase) and the ubiquitin-proteasome pathway of protein degradation (F-box only protein 32, Forkhead box protein O1, Muscle RING-finger protein-1) on d 34 were largely similar among groups. Likewise, relative phosphorylation and thus activation of mTOR and ribosomal protein S6 involved in the mTOR pathway, and of eukaryotic translation initiation factor 2A (eIF2a) involved in the general control nonderepressible 2 (GCN2)/eIF2a pathway of protein synthesis inhibition, were not influenced. These data indicate that dietary Leu concentrations exceeding the broiler´s requirements up to 60% neither influence protein synthesis nor degradation pathways nor muscle growth in growing broilers.

Myricanol rescues dexamethasone-induced muscle dysfunction via a sirtuin 1-dependent mechanism.

BackgroundMuscle atrophy and weakness are adverse effects of high dose or the sustained usage of glucocorticoids. Loss of mitochondria and degradation of protein are highly correlated with muscle dysfunction. The deacetylase sirtuin 1 (SIRT1) plays a vital role in muscle remodelling. The current study was designed to identify myricanol as a SIRT1 activator, which could protect skeletal muscle against dexamethasone‐induced wasting.MethodsThe dexamethasone‐induced atrophy in C2C12 myotubes was evaluated by expression of myosin heavy chain, muscle atrophy F‐box (atrogin‐1), and muscle ring finger 1 (MuRF1), using western blots. The mitochondrial content and oxygen consumption were assessed by MitoTracker staining and extracellular flux analysis, respectively. Muscle dysfunction was established in male C57BL/6 mice (8–10 weeks old, n = 6) treated with a relatively high dose of dexamethasone (25 mg/kg body weight, i.p., 10 days). Body weight, grip strength, forced swimming capacity, muscle weight, and muscle histology were assessed. The expression of proteolysis‐related, autophagy‐related, apoptosis‐related, and mitochondria‐related proteins was analysed by western blots or immunoprecipitation.ResultsMyricanol (10 μM) was found to rescue dexamethasone‐induced muscle atrophy and dysfunction in C2C12 myotubes, indicated by increased expression of myosin heavy chain (0.33 ± 0.14 vs. 0.89 ± 0.21, *P < 0.05), decreased expression of atrogin‐1 (2.31 ± 0.67 vs. 1.53 ± 0.25, *P < 0.05) and MuRF1 (1.55 ± 0.08 vs. 0.99 ± 0.12, **P < 0.01), and elevated ATP production (3.83 ± 0.46 vs. 5.84 ± 0.79 nM/mg protein, **P < 0.01), mitochondrial content (68.12 ± 10.07% vs. 116.38 ± 5.12%, *P < 0.05), and mitochondrial oxygen consumption (166.59 ± 22.89 vs. 223.77 ± 22.59 pmol/min, **P < 0.01). Myricanol directly binds and activates SIRT1, with binding energy of −5.87 kcal/mol. Through activating SIRT1 deacetylation, myricanol inhibits forkhead box O 3a transcriptional activity to reduce protein degradation, induces autophagy to enhance degraded protein clearance, and increases peroxisome proliferator‐activated receptor γ coactivator‐1α activity to promote mitochondrial biogenesis. In dexamethasone‐induced muscle wasting C57BL/6 mice, 5 mg/kg myricanol treatment reduces the loss of muscle mass; the percentages of quadriceps and gastrocnemius muscle in myricanol‐treated mice are 1.36 ± 0.02% and 0.87 ± 0.08%, respectively (cf. 1.18 ± 0.06% and 0.78 ± 0.05% in dexamethasone‐treated mice, respectively). Myricanol also rescues dexamethasone‐induced muscle weakness, indicated by improved grip strength (70.90 ± 4.59 vs. 120.58 ± 7.93 g, **P < 0.01) and prolonged swimming exhaustive time (48.80 ± 11.43 vs. 83.75 ± 15.19 s, **P < 0.01). Myricanol prevents dexamethasone‐induced muscle atrophy and weakness by activating SIRT1, to reduce muscle protein degradation, enhance autophagy, and promote mitochondrial biogenesis and function in mice.ConclusionsMyricanol ameliorates dexamethasone‐induced skeletal muscle wasting by activating SIRT1, which might be developed as a therapeutic agent for treatment of muscle atrophy and weakness.

The alleviative effects and related mechanisms of taurine supplementation on growth performance and carcass characteristics in broilers exposed to chronic heat stress.

To investigate the alleviative effects and molecular mechanisms of taurine supplementation on growth performance and carcass characteristics in broilers exposed to chronic heat stress, 144 male Arbor Acres broilers (28 d old) were randomly distributed to positive control (PC, 22°C, basal diet), heat stress (HS, consistent 32°C, basal diet), or heat stress + taurine (HS + T, consistent 32°C, basal diet + 5.00g/kg taurine) groups, with 6 cages per group and 8 birds per cage. Chronic heat stress significantly decreased body weight, average daily gain, and average daily feed intake, and increased cloacal temperature and feed conversion ratio (FCR, P < 0.05). Though taurine supplementation tended to decrease the FCR in the HS + T group compared with the HS group after 14 d of heat exposure (P = 0.071), there were no significant alleviative effects of taurine supplementation on the increased cloacal temperature and decreased growth performance in chronic heat-stressed broilers (P > 0.05). After 7 and 14 d of heat exposure, taurine supplementation significantly increased the proportion of breast muscle and hormone-sensitive lipase activity in the abdominal fat (P < 0.05), and decreased the mRNA expressions of muscle atrophy F-box protein (MAFbx) and muscle ring-finger protein-1 (MuRF1) in breast muscle compared with the HS group (P < 0.05). After 7 d of heat exposure, taurine supplementation significantly increased serum non-esterified fatty acid concentration (P < 0.05), and decreased the mRNA expressions of acetyl-CoA carboxylase 1c (ACC) and muscular isoform of carnitine palmitoyl transferase 1 (M-CPT1) compared with the HS group (P < 0.05). In addition, the mRNA expressions of M-CPT1 and ribosomal protein S6 kinase, 70kDa (p70S6K) in the HS + T group were significantly higher than those of the other two groups after 14 d of heat exposure (P < 0.05). In conclusion, taurine supplementation can improve carcass characteristics of chronic heat-stressed broilers by facilitating lipolysis for energy, enhancing protein synthesis, and suppressing protein degradation of the breast muscles.

Dietary intake of probiotic kimchi ameliorated IL-6-driven cancer cachexia.

Cancer cachexia is a syndrome accompanying weight loss, skeletal muscle atrophy, and loss of adipose tissue in patients with advanced cancer. Since interleukin-6 (IL-6) is one of core mediators causing cancer cachexia and kimchi can modulate IL-6 response, we hypothesized dietary intake of kimchi can ameliorate cancer cachexia. In this study, we studied preemptive administration of kimchi can ameliorate mouse colon carcinoma cells colon (C26) adenocarcinoma-induced cancer cachexia and explored anti-cachexic mechanisms of kimchi focused on the changes of muscle atrophy, cachexic inflammation, and catabolic catastrophe. As results, dietary intake of kimchi significantly attenuated the development of cancer cachexia, presented with lesser weight loss, higher muscle preservation as well as higher survival from cancer cachexia in mice. Starting from significant inhibition of IL-6 and its signaling, kimchi afforded significant inhibition of muscle specific ubiquitin-proteasome system including inhibition of atrogin-1 and muscle ring finger protein-1 (MuRF-1) with other muscle related genes including mitofusin-2 (Mfn-2) and PGC-1α. Significant inhibition of lipolysis gene such as adipose triglyceride lipase (ATGL) and hormone-sensitive ligase (HSL) accompanied with significant induction of fatty acid synthase (FAS) and sterol response element binding protein 1 (SREBP1) was achieved with kimchi. As gene regulation, IL-6 and their receptor as well as Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) were significantly attenuated with kimchi. In conclusion, dietary intake of cancer preventive kimchi can be an anticipating option to ameliorate cancer cachexia via suppressive action of IL-6 accompanied with decreased muscle atrophy and lipolysis.

Constant light exposure aggravates POMC-mediated muscle wasting associated with hypothalamic alteration of circadian clock and SIRT1 in endotoxemia rats

Constant light exposure is widespread in the intensive care unit (ICU) and could increase the rate of brain dysfunction as delirium and sleep disorders in critical patients. And the activation of hypothalamic neuropeptides is proved to play a crucial role in regulating hypercatabolism, especially skeletal muscle wasting in critical patients, which could lead to serious complications and poor prognosis. Here we investigated the hypothesis that constant light exposure could aggravate skeletal muscle wasting in endotoxemia rats and whether it was associated with alterations of circadian clock and hypothalamic proopiomelanocortin(POMC) expression. Fifty-four adult male Sprague-Dawley rats were intraperitoneally injected with lipopolysaccharide(LPS) or saline, subjected to constant light or a 12:12 h light-dark cycle for 7 days. On day 8, rats were sacrificed across six time points in 24 h and hypothalamus tissues and skeletal muscle were obtained. Rates of muscle wasting were measured by 3-methylhistidine(3-MH) and tyrosine release as well as expression of two muscle atrophic genes, muscle ring finger 1(MuRF-1) and muscle atrophy F-box(MAFbx). The expression of circadian clock genes, silent information regulator 1(SIRT1), POMC and hypothalamic inflammatory cytokines were also detected. Results showed that LPS administration significantly increased hypothalamic POMC expression, inflammatory cytokine levels and muscle wasting rates. Meanwhile constant light exposure disrupted the circadian rhythm, declined the expression of SIRT1 as well as aggravated hypothalamic POMC overexpression and skeletal muscle wasting in rats with endotoxemia. Taken together, the results demonstrated that constant light exposure could aggravate POMC-mediated skeletal muscle wasting in endotoxemia rats, which is associated with alteration of circadian clocks and SIRT1 in the hypothalamus.

Isobavachalcone attenuates myotube atrophy induced by TNF-α through muscle atrophy F-box signaling and the nuclear factor erythroid 2-related factor 2 cascade.

Skeletal muscle atrophy is a condition characterized by damaged muscle fibers and reduced numbers of muscle cells due to various causes. Muscle atrophy is associated with chronic diseases, such as heart failure, diabetes, and aging-related diseases. Isobavachalcone (IBC) is a flavonoid found in various foods and natural products, and studies have investigated its diverse effects, including its neuroprotective and anticancer effects. However, no studies have evaluated the effects of IBC on muscle atrophy. Thus, in this study, we assessed the effects of IBC on prevention of muscle atrophy. To evaluate the preventive effects of IBC on muscle atrophy, we used C2C12 myoblasts and induced muscle atrophy by tumor necrosis factor (TNF)-α. IBC regulated the expression levels of muscle atrophy F-box and muscle RING finger-1 in response to damaged muscle cells, thereby restoring the expression of myosin heavy chain and myogenin. Moreover, IBC regulated the phosphorylation of the nuclear factor-κB and p38 and upregulated the expression of nuclear factor erythroid 2-related factor 2 and heme oxygenase-1, which are involved in regulating oxidative stress. Our results indicated that IBC acted to relieve TNF-α-induced skeletal muscle atrophy by regulating the factors related to inflammation and oxidative stress.

Beneficial effects of metformin on muscle atrophy induced by obesity in rats.

A growing interest to understand the signaling pathways mediating obesity-induced muscle atrophy is given. Metformin (Met) was reported to possess positive effects on preventing muscle damage and promoting muscle mass maintenance. The aim of the present study to investigate pathways involved in Met effect on obesity induced muscle atrophy. Thirty adult male albino rats were assigned into two groups: normal chew diet fed group as control group (C; n = 10) and high-fat-diet (HFD) fed group ( n = 20). After 16 weeks, the HFD-fed animals were subdivided into two groups; HFD group ( n = 10) and HFD fed treated with oral Met (320 mg/day) treatment (Met, n = 10) for 4 weeks. At the end of the experiment; final body weight, visceral fat weight, fasting blood glucose, insulin, lactate, total cholesterol, triglycerides were measured and calculated homeostatic model assessment insulin resistant (HOMA-IR) for all groups. Soleus muscle weight, histopathlogical examination and expression of peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), forkhead box O3 (FoxO3), atrogin-1/MAFbx, and muscle RING finger 1 (MuRF-1) were performed. HFD-fed animals showed significant increase in final body weight, visceral fat mass, fasting blood glucose, insulin, calculated HOMA-IR, lactate, total cholesterol and triglycerides with significant decrease in soleus muscle weight, PGC-1α and significant increase in FoxO3, atrogin-1/MAFbx, and MuRF-1 expression. Also, there was significant decrease in fiber diameter, myosin heavy chain (MHC) I content while collagen content and myosin heavy chain IIa were increased compared with control group. Met-treated group showed a significant decrease in the measured parameters compared with the HFD group. It also restored the gene expression, morphometric measures and MHC composition toward normal. The current study is the first to provide evidence that Met could ameliorate muscle atrophy in high-fat diet induced obesity and this effect may be in part due to regulation of PGC-1α-FoxO3 pathway.

Combination of Soy Protein, Amylopectin, and Chromium Stimulates Muscle Protein Synthesis by Regulation of Ubiquitin-Proteasome Proteolysis Pathway after Exercise.

The present study was undertaken to investigate the effect of the combination of soy protein, amylopectin, and chromium (SAC) on muscle protein synthesis and signal transduction pathways involved in protein synthesis (mTOR pathways, IGF-1, and AktSer473) and proteolysis (FOXO1Ser256; MURF1, MAFbx) after exercise. Thirty-five Wistar rats were randomly divided into five groups: (1) control (C); (2) exercise (E); (3) exercise + soy protein (3.1g/kg/day) (E + S); (4) exercise + soy protein + chromium (E + S + Cr); (5) exercise + soy protein + amylopectin + chromium (E + S + A + Cr). Post-exercise ingestion of SAC significantly increased the fractional rate of protein synthesis (FSR), insulin, glycogen, and amino acid levels with the highest effect observed in E + S + A + Cr group (P˂0.05). However, SAC supplementation decreased the lactic acid concentration (P˂0.05). A reduction in forkhead box protein O1 (FOXO1) and forkhead box protein O3 (FOXO3) (regulators of ubiquitin-related proteolysis) and muscle atrophy F-box (MAFbx) levels was noted after treatment with SAC (P< 0.05). Insulin-like growth factor 1(IGF-1) level was increased in the E + S, E + S + Cr, and E + S + A + Cr groups (P< 0.05). While the phosphorylation of 4E-BP1Thr37/46, AktSer473, mTORSer2448, and S6K1Thr389 levels increased after SAC supplementation, phosphorylated muscle ring finger 1 (MuRF-1, an E3-ubiquitin ligase gene) was found to be significantly lower compared with the E group (P˂0.05). These results indicate that SAC supplementation improves FSR, insulin, and glycogen levels after exercise. SAC improves protein synthesis by inhibiting the ubiquitin-proteasome pathway and inducing anabolic metabolism.

OR-017 Supplementation of Ala-Gln inhibits protein breakdown of skeletal muscle in rats with altitude training through TNF-α/NF-κB/MuRF1 pathway

Objective Objective: To explore the effects of alanyl-glutamine（Ala-Gln）or glutamine（Gln） supplementation on protein metabolism in rat skeletal muscle during simulated altitude training，and compare the intervention of Gln or Ala-Gln to provide the necessary experimental basis for finding nutritional interventions to inhibit skeletal muscle protein degradation during altitude training.&#x0D; Methods Methods: Forty SD rats aged 6 weeks were randomly divided into normoxic control group（NC group，n=10），hypoxic exercise group（HE group， n=10），hypoxic exercise + glutamine + alanine group（HEG group，n=10）， hypoxic exercise + alanyl glutamine group（HEAG group，n=10）. Rats were subjected to 6 weeks of 13.6% hypoxic exposure and 90% lactic acid threshold intensity weight-bearing swimming（load weight of 2.1% of body weight）exercise training，30 minutes after the end of each training，the mixed solution of Ala and Gln was administered according to the dose of 0.75g/Kg body weight in HEG group，and the solution of Ala-Gln was administered in the HEAG group at a dose of 1.5 g/kg body weight. After 6 weeks，the contents of rat skeletal muscle total protein（Pro），myosin heavy chain（Myo），tumor necrosis factor-α（TNF-α），nuclear transcription factor-κB （NF-κB），NF-κB inhibitory protein α（IkBα），and mRNA expression of muscle atrophy box F gene（MAFbx），muscle ring finger gene 1（MuRF1），and inhibitor of kappa B kinase complex-beta（IKKβ）were measured.&#x0D; Results Results:（1）Compared with NC group，the content of Pro and Myo in skeletal muscle in HE group was significantly decreased（P&lt;0.05，P&lt;0.01），and the mRNA expression of MAFbx and MuRF1 in skeletal muscle was significantly increased（P&lt;0.05，P&lt; 0.01），the levels of TNF-α and NF-κB were significantly increased（P&lt;0.05），the content of IkBα was significantly decreased（P&lt;0.05），and the expression of IKKβ mRNA was significantly increased（P&lt;0.01）. （2）Compared with HE group，the content of Pro and Myo in skeletal muscle in HEG group increased，but there was no significant difference（P&gt;0.05）. The expression of MuRF1 mRNA and the content of TNF-α and NF-κB in skeletal muscle decreased，IkBα content increased，there were no significant difference，but mRNA expression of MAFbx and IKKβ was significantly decreased（P&lt;0.05， P&lt;0.01）. （3）Compared with HE group，the content of Pro and Myo in skeletal muscle in HEAG group increased significantly（P&lt;0.05），mRNA expression of IKKβ，MuRF1 and MAFb（P&lt;0.01）and TNF-α，NF-κB content（P&lt;0.05）in skeletal muscle was significantly decreased，and the IkBα content was significantly increased（P&lt;0.05）.&#x0D; Conclusions Conclusion:（1）Simulated altitude training can activate TNF-α/NF-κB/MuRF1 pathway and enhance the catabolism of skeletal muscle protein，which is one of the important mechanisms for the reduction of skeletal muscle protein content caused by altitude training. （2）Supplementation of Ala-Gln during altitude training can significantly reduce the activation of TNF-α/NF-κB/MuRF1 pathway in skeletal muscle，and reduce the catabolism of skeletal muscle protein during altitude training，which plays a very important role in preventing the loss of skeletal muscle protein caused by altitude training. supplementation of Gln monomer during altitude training has little inhibitory effect on the activation of TNF-α/NF-κB/MuRF1 pathway in skeletal muscle.

Muscle-specific regulation of right ventricular transcriptional responses to chronic hypoxia-induced hypertrophy by the muscle ring finger-1 (MuRF1) ubiquitin ligase in mice

BackgroundWe recently identified a role for the muscle-specific ubiquitin ligase MuRF1 in right-sided heart failure secondary to pulmonary hypertension induced by chronic hypoxia (CH). MuRF1−/− mice exposed to CH are resistant to right ventricular (RV) dysfunction whereas MuRF1 Tg + mice exhibit impaired function indicative of heart failure. The present study was undertaken to understand the underlying transcriptional alterations in the RV of MuRF1−/− and MuRF1 Tg + mice.MethodsMicroarray analysis was performed on RNA isolated from the RV of MuRF1−/−, MuRF1 Tg+, and wild-type control mice exposed to CH.ResultsMuRF1−/− RV differentially expressed 590 genes in response to CH. Analysis of the top 66 genes (> 2-fold or < − 2-fold) revealed significant associations with oxidoreductase, transcription regulation, and transmembrane component annotations. The significant genes had promoters enriched for HOXD12, HOXC13, and RREB-1 protein transcription factor binding sites. MuRF1 Tg + RV differentially expressed 150 genes in response to CH. Analysis of the top 45 genes (> 3-fold or < − 3-fold) revealed significant associations with oxidoreductase-metabolic, glycoprotein-transmembrane-integral proteins, and alternative splicing/splice variant annotations. The significant genes were enriched for promoters with ZIC1 protein transcription factor binding sites.ConclusionsThe differentially expressed genes in MuRF1−/− and MuRF1 Tg + RV after CH have common functional annotations related to oxidoreductase (including antioxidant) and transmembrane component functions. Moreover, the functionally-enhanced MuRF1−/− hearts regulate genes related to transcription, homeobox proteins, and kinases/phosphorylation. These studies also reveal potential indirect effects of MuRF1 through regulating Rreb-1, and they reveal mechanisms by which MuRF1 may transcriptionally regulate anti-oxidant systems in the face of right heart failure.

Role of altered proteostasis network in chronic hypobaric hypoxia induced skeletal muscle atrophy.

BackgroundHigh altitude associated hypobaric hypoxia is one of the cellular and environmental perturbation that alters proteostasis network and push the healthy cell towards loss of muscle mass. The present study has elucidated the robust proteostasis network and signaling mechanism for skeletal muscle atrophy under chronic hypobaric hypoxia (CHH).MethodsMale Sprague Dawley rats were exposed to simulated hypoxia equivalent to a pressure of 282 torr for different durations (1, 3, 7 and 14 days). After CHH exposure, skeletal muscle tissue was excised from the hind limb of rats for biochemical analysis.ResultsChronic hypobaric hypoxia caused a substantial increase in protein oxidation and exhibited a greater activation of ER chaperones, glucose-regulated protein-78 (GRP-78) and protein disulphide isomerase (PDI) till 14d of CHH. Presence of oxidized proteins triggered the proteolytic systems, 20S proteasome and calpain pathway which were accompanied by a marked increase in [Ca2+]. Upregulated Akt pathway was observed upto 07d of CHH which was also linked with enhanced glycogen synthase kinase-3β (GSk-3β) expression, a negative regulator of Akt. Muscle-derived cytokines, tumor necrosis factor-α (TNF-α), interferon-ϒ (IFN-©) and interleukin-1β (IL-1β) levels significantly increased from 07d onwards. CHH exposure also upregulated the expression of nuclear factor kappa-B (NF-κB) and E3 ligase, muscle atrophy F-box-1 (Mafbx-1/Atrogin-1) and MuRF-1 (muscle ring finger-1) on 07d and 14d. Further, severe hypoxia also lead to increase expression of ER-associated degradation (ERAD) CHOP/ GADD153, Ub-proteasome and apoptosis pathway.ConclusionsThe disrupted proteostasis network was tightly coupled to degradative pathways, altered anabolic signaling, inflammation, and apoptosis under chronic hypoxia. Severe and prolonged hypoxia exposure affected the protein homeostasis which overwhelms the muscular system and tends towards skeletal muscle atrophy.

Effects of mesenchymal stromal cells on type 1 diabetes mellitus rat muscles.

Type 1 diabetes mellitus (DM) causes marked skeletal muscle atrophy. Mesenchymal stromal cells (MSC) are an attractive therapy to avoid diabetic complications because of their ability to modify the microenvironment at sites of tissue injury. The objective of this study was to evaluate the effects of MSC transplantation on muscle adaptation caused by diabetes. DM was induced by streptozotocin (STZ), and the diabetic animals received systemic MSC transplantation. The von Frey test and footprint analysis were used to assess sensation and sensory motor performance, respectively. Tibialis anterior muscles were investigated by morphology; molecular markers atrogin-1/muscle RING-finger protein-1, nuclear factor κB/p38 mitogen-activated protein kinase, tumor necrosis-like weak inducer of apoptosis/fibroblast growth factor-inducible 14, myostatin, myogenic differentiation 1, and insulin-like growth factor 1 were also assessed. MSC transplantation improved sensation and walking performance and also decreased muscle fibrosis in DM rats by modulating atrogenes but did not prevent muscle atrophy. MSCs can reduce muscle and functional complications that result from type 1 DM in rats. Muscle Nerve 58: 583-591, 2018.