Role of SIRT2 in regulating the dexamethasone-activated autophagy pathway in skeletal muscle atrophy

The proteolytic autophagy system is involved in a major regulatory pathway in dexamethasone (Dex)-induced muscle atrophy. Sirtuin 2 (SIRT2) is known to participate in modulating autophagy signaling, exerting effects in skeletal muscle atrophy. We aimed to determine the effects of SIRT2 on autophagy in Dex-induced myoatrophy. Mice were randomly divided into the normal, Dex, and sirtinol groups. C2C12 cells were differentiated into myotubes and transfected with short hairpin (sh)-Sirt2-green fluorescent protein (GFP) or Sirt2-GFP lentivirus. To evaluate the mass and function of skeletal muscles, we measured the myofiber cross-sectional area, myotube size, gastrocnemius muscle wet weight/body weight ratio (%), and time-to-exhaustion. The SIRT2, myosin heavy chain (MyHC), LC3, and Beclin-1 expression levels were detected by western blotting and quantitative reverse transcription-polymerase chain reaction. Inhibition of SIRT2 markedly attenuated the muscle mass and endurance capacity. The same phenotype was observed in Sirt2-shRNA-treated myotubes, as evidenced by their decreased size. Conversely, SIRT2 overexpression alleviated Dex-induced myoatrophy in vitro. Moreover, SIRT2 negatively regulated the expression of the LC3b and Beclin-1 in skeletal muscles. These findings suggested that SIRT2 activation protects myotubes against Dex-induced atrophy through the inhibition of the autophagy system; this phenomenon may potentially serve as a target for treating glucocorticoid-induced myopathy.

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AMP-activated protein kinase agonists increase mRNA content of the muscle-specific ubiquitin ligases MAFbx and MuRF1 in C2C12 cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00622.2006 ◽

2007 ◽

Vol 292 (6) ◽

pp. E1555-E1567 ◽

Cited By ~ 93

Author(s):

Brian J. Krawiec ◽

Gerald J. Nystrom ◽

Robert A. Frost ◽

Leonard S. Jefferson ◽

Charles H. Lang

Keyword(s):

Skeletal Muscle ◽

Protein Kinase ◽

Mrna Expression ◽

Muscle Atrophy ◽

Ubiquitin Ligases ◽

C2c12 Cells ◽

Skeletal Muscle Atrophy ◽

Compound C ◽

Mrna Content ◽

Amp Activated Protein Kinase

The hypothesis of the present study was that exposure of differentiated muscle cells to agonists of the AMP-activated protein kinase (AMPK) would increase the mRNA content of the muscle-specific ubiquitin ligases muscle atrophy F-box (MAFbx) and muscle RING finger 1 (MuRF1). C2C12 cells were incubated with incremental doses of 5-aminoimidazol-4-carboximide ribonucleoside (AICAR) or metformin for 24 h. Both MAFbx and MuRF1 mRNA increased dose dependently in response to these AMPK activators. AICAR, metformin, and 2-deoxy-d-glucose produced time-dependent alterations in ubiquitin ligase expression, typified by a biphasic pattern of expression marked by an acute repression followed by a sustained induction. AMPK-activating treatments in conjunction with dexamethasone produced a pronounced synergistic effect on ligase mRNA expression at later time points. This cooperative response occurred in the absence of a dexamethasone-dependent increase in AMPK expression or activity, as determined by immunoblotting for phosphorylation and expression of AMPKα and its downstream target acetyl-CoA carboxylase (ACC). These responses elicited by AMPK activation singly or in combination with dexamethasone did not extend to the mRNA expression of the UBR box family E3s UBR1/E3αI and UBR2/E3αII. Treatment with the AMPK inhibitor compound C prevented increases in MAFbx and MuRF1 mRNA in response to serum deprivation, as well as AICAR and dexamethasone treatment individually or jointly. Stimulation of AMPK activity in vivo via AICAR injection increased both MAFbx and MuRF1 mRNA in murine skeletal muscle. These data suggest that activation of AMPK in skeletal muscle results in a specific upregulation of MAFbx and MuRF1, responses that are reminiscent of the proposed atrophic transcriptional program executed under various conditions of skeletal muscle wasting. Therefore, AMPK may be a critical component of the intercalated network of signaling pathways governing skeletal muscle atrophy, where its input acts to modify anti- and proatrophic signals to influence gene expression in reaction to catabolic perturbations.

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Hydrogen sulfide alleviates hyperhomocysteinemia-mediated skeletal muscle atrophy via mitigation of oxidative and endoplasmic reticulum stress injury

AJP Cell Physiology ◽

10.1152/ajpcell.00147.2018 ◽

2018 ◽

Vol 315 (5) ◽

pp. C609-C622 ◽

Cited By ~ 17

Author(s):

Avisek Majumder ◽

Mahavir Singh ◽

Jyotirmaya Behera ◽

Nicholas T. Theilen ◽

Akash K. George ◽

...

Keyword(s):

Oxidative Stress ◽

Skeletal Muscle ◽

Endoplasmic Reticulum ◽

Hydrogen Sulfide ◽

Er Stress ◽

Muscle Atrophy ◽

C2c12 Cells ◽

Skeletal Muscle Atrophy ◽

Stress Injury ◽

Western Blots

Although hyperhomocysteinemia (HHcy) occurs because of the deficiency in cystathionine-β-synthase (CBS) causing skeletal muscle dysfunction, it is still unclear whether this effect is mediated through oxidative stress, endoplasmic reticulum (ER) stress, or both. Nevertheless, there is no treatment option available to improve HHcy-mediated muscle injury. Hydrogen sulfide (H2S) is an antioxidant compound, and patients with CBS mutation do not produce H2S. In this study, we hypothesized that H2S mitigates HHcy-induced redox imbalance/ER stress during skeletal muscle atrophy via JNK phosphorylation. We used CBS+/−mice to study HHcy-mediated muscle atrophy, and treated them with sodium hydrogen sulfide (NaHS; an H2S donor). Proteins and mRNAs were examined by Western blots and quantitative PCR. Proinflammatory cytokines were also measured. Muscle mass and strength were studied via fatigue susceptibility test. Our data revealed that HHcy was detrimental to skeletal mass, particularly gastrocnemius and quadriceps muscle weight. We noticed that oxidative stress was reversed by NaHS in homocysteine (Hcy)-treated C2C12 cells. Interestingly, ER stress markers (GRP78, ATF6, pIRE1α, and pJNK) were elevated in vivo and in vitro, and NaHS mitigated these effects. Additionally, we observed that JNK phosphorylation was upregulated in C2C12 after Hcy treatment, but NaHS could not reduce this effect. Furthermore, inflammatory cytokines IL-6 and TNF-α were higher in plasma from CBS as compared with wild-type mice. FOXO1-mediated Atrogin-1 and MuRF-1 upregulation were attenuated by NaHS. Functional studies revealed that NaHS administration improved muscle fatigability in CBS+/−mice. In conclusion, our work provides evidence that NaHS is beneficial in mitigating HHcy-mediated skeletal injury incited by oxidative/ER stress responses.

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Two Weeks of Smoking Cessation Reverse Cigarette Smoke-Induced Skeletal Muscle Atrophy and Mitochondrial Dysfunction in Mice

Nicotine & Tobacco Research ◽

10.1093/ntr/ntaa016 ◽

2020 ◽

Cited By ~ 2

Author(s):

Tom Tanjeko Ajime ◽

Jef Serré ◽

Rob C I Wüst ◽

Guy Anselme Mpaka Messa ◽

Chiel Poffé ◽

...

Keyword(s):

Skeletal Muscle ◽

Smoking Cessation ◽

Mitochondrial Dysfunction ◽

Muscle Mass ◽

Muscle Atrophy ◽

Skeletal Muscles ◽

Skeletal Muscle Atrophy ◽

Limb Muscle ◽

Male Mice ◽

And Function

Abstract Introduction Apart from its adverse effects on the respiratory system, cigarette smoking also induces skeletal muscle atrophy and dysfunction. Whether short-term smoking cessation can restore muscle mass and function is unknown. We, therefore, studied the impact of 1- and 2-week smoking cessation on skeletal muscles in a mouse model. Methods Male mice were divided into four groups: Air-exposed (14 weeks); cigarette smoke (CS)-exposed (14 weeks); CS-exposed (13 weeks) followed by 1-week cessation; CS-exposed (12 weeks) followed by 2 weeks cessation to examine exercise capacity, physical activity levels, body composition, muscle function, capillarization, mitochondrial function and protein expression in the soleus, plantaris, and diaphragm muscles. Results CS-induced loss of body and muscle mass was significantly improved within 1 week of cessation due to increased lean and fat mass. Mitochondrial respiration and protein levels of the respiratory complexes in the soleus were lower in CS-exposed mice, but similar to control values after 2 weeks of cessation. Exposing isolated soleus muscles to CS extracts reduced mitochondrial respiration that was reversed after removing the extract. While physical activity was reduced in all groups, exercise capacity, limb muscle force, fatigue resistance, fiber size and capillarization, and diaphragm cytoplasmic HIF-1α were unaltered by CS-exposure. However, CS-induced diaphragm atrophy and increased capillary density were not seen after 2 weeks of smoking cessation. Conclusion In male mice, 2 weeks of smoking cessation reversed smoking-induced mitochondrial dysfunction, limb muscle mass loss, and diaphragm muscle atrophy, highlighting immediate benefits of cessation on skeletal muscles. Implications Our study demonstrates that CS-induced skeletal muscle mitochondrial dysfunction and atrophy are significantly improved by 2 weeks of cessation in male mice. We show for the first time that smoking cessation as short as 1 to 2 weeks is associated with immediate beneficial effects on skeletal muscle structure and function with the diaphragm being particularly sensitive to CS-exposure and cessation. This could help motivate smokers to quit smoking as early as possible. The knowledge that smoking cessation has potential positive extrapulmonary effects is particularly relevant for patients referred to rehabilitation programs and those admitted to hospitals suffering from acute or chronic muscle deterioration yet struggling with smoking cessation.

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Exercise preconditioning diminishes skeletal muscle atrophy after hindlimb suspension in mice

Journal of Applied Physiology ◽

10.1152/japplphysiol.00137.2018 ◽

2018 ◽

Vol 125 (4) ◽

pp. 999-1010 ◽

Cited By ~ 7

Author(s):

Nicholas T. Theilen ◽

Nevena Jeremic ◽

Gregory J. Weber ◽

Suresh C. Tyagi

Keyword(s):

Skeletal Muscle ◽

Exercise Training ◽

Muscle Atrophy ◽

Mitochondrial Biogenesis ◽

Weight Ratio ◽

Hindlimb Suspension ◽

Skeletal Muscle Atrophy ◽

Short Term ◽

Concurrent Exercise ◽

And Function

The aim of the present study was to investigate whether short-term, concurrent exercise training before hindlimb suspension (HLS) prevents or diminishes both soleus and gastrocnemius atrophy and to analyze whether changes in mitochondrial molecular markers were associated. Male C57BL/6 mice were assigned to control at 13 ± 1 wk of age, 7-day HLS at 12 ± 1 wk of age (HLS), 2 wk of exercise training before 7-day HLS at 10 ± 1 wk of age (Ex+HLS), and 2 wk of exercise training at 11 ± 1 wk of age (Ex) groups. HLS resulted in a 27.1% and 21.5% decrease in soleus and gastrocnemius muscle weight-to-body weight ratio, respectively. Exercise training before HLS resulted in a 5.6% and 8.1% decrease in soleus and gastrocnemius weight-to-body weight ratio, respectively. Exercise increased mitochondrial biogenesis- and function-associated markers and slow myosin heavy chain (SMHC) expression, and reduced fiber-type transitioning marker myosin heavy chain 4 (Myh4). Ex+HLS revealed decreased reactive oxygen species (ROS) and oxidative stress compared with HLS. Our data indicated the time before an atrophic setting, particularly caused by muscle unloading, may be a useful period to intervene short-term, progressive exercise training to prevent skeletal muscle atrophy and is associated with mitochondrial biogenesis, function, and redox balance. NEW & NOTEWORTHY Mitochondrial dysfunction is associated with disuse-induced skeletal muscle atrophy, whereas exercise is known to increase mitochondrial biogenesis and function. Here we provide evidence of short-term concurrent exercise training before an atrophic event protecting skeletal muscle from atrophy in two separate muscles with different, dominant fiber-types, and we reveal an association with the adaptive changes of mitochondrial molecular markers to exercise.

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Low Levels of Serum Tryptophan Underlie Skeletal Muscle Atrophy

Nutrients ◽

10.3390/nu12040978 ◽

2020 ◽

Vol 12 (4) ◽

pp. 978 ◽

Cited By ~ 3

Author(s):

Soranobu Ninomiya ◽

Nobuhiko Nakamura ◽

Hiroshi Nakamura ◽

Taku Mizutani ◽

Yuto Kaneda ◽

...

Keyword(s):

Skeletal Muscle ◽

Muscle Atrophy ◽

Skeletal Muscles ◽

B Cell Lymphoma ◽

Tryptophan Metabolism ◽

Skeletal Muscle Atrophy ◽

C2c12 Myoblast ◽

Standard Diet ◽

Deficient Diet ◽

Low Levels

Sarcopenia is a poor prognosis factor in some cancer patients, but little is known about the mechanisms by which malignant tumors cause skeletal muscle atrophy. Tryptophan metabolism mediated by indoleamine 2,3-dioxygenase is one of the most important amino acid changes associated with cancer progression. Herein, we demonstrate the relationship between skeletal muscles and low levels of tryptophan. A positive correlation was observed between the volume of skeletal muscles and serum tryptophan levels in patients with diffuse large B-cell lymphoma. Low levels of tryptophan reduced C2C12 myoblast cell proliferation and differentiation. Fiber diameters in the tibialis anterior of C57BL/6 mice fed a tryptophan-deficient diet were smaller than those in mice fed a standard diet. Metabolomics analysis revealed that tryptophan-deficient diet downregulated glycolysis in the gastrocnemius and upregulated the concentrations of amino acids associated with the tricarboxylic acid cycle. The weights and muscle fiber diameters of mice fed the tryptophan-deficient diet recovered after switching to the standard diet. Our data showed a critical role for tryptophan in regulating skeletal muscle mass. Thus, the tryptophan metabolism pathway may be a promising target for preventing or treating skeletal muscle atrophies.

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Effect of Quercetin on Dexamethasone-Induced C2C12 Skeletal Muscle Cell Injury

Molecules ◽

10.3390/molecules25143267 ◽

2020 ◽

Vol 25 (14) ◽

pp. 3267 ◽

Cited By ~ 1

Author(s):

Chun Chen ◽

Jai-Sing Yang ◽

Chi-Cheng Lu ◽

Yu-Jen Chiu ◽

Hung-Che Chen ◽

...

Keyword(s):

Skeletal Muscle ◽

Muscle Atrophy ◽

Fruits And Vegetables ◽

Cell Injury ◽

C2c12 Cells ◽

Skeletal Muscle Atrophy ◽

Protective Effects ◽

Cross Sectional ◽

Hydroxyl Free Radical ◽

C2c12 Skeletal Muscle Cells

Glucocorticoids are widely used anti-inflammatory drugs in clinical settings. However, they can induce skeletal muscle atrophy by reducing fiber cross-sectional area and myofibrillar protein content. Studies have proven that antioxidants can improve glucocorticoid-induced skeletal muscle atrophy. Quercetin is a potent antioxidant flavonoid widely distributed in fruits and vegetables and has shown protective effects against dexamethasone-induced skeletal muscle atrophy. In this study, we demonstrated that dexamethasone significantly inhibited cell growth and induced cell apoptosis by stimulating hydroxyl free radical production in C2C12 skeletal muscle cells. Our results evidenced that quercetin increased C2C12 skeletal cell viability and exerted antiapoptotic effects on dexamethasone-treated C2C12 cells by regulating mitochondrial membrane potential (ΔΨm) and reducing oxidative species. Quercetin can protect against dexamethasone-induced muscle atrophy by regulating the Bax/Bcl-2 ratio at the protein level and abnormal ΔΨm, which leads to the suppression of apoptosis.

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Butyrate ameliorate skeletal muscle atrophy in Diabetic Nephropathy via enhancing gut barrier function and GPR43 mediated PI3K/AKT/mTOR signals

10.22541/au.160688559.90642598/v1 ◽

2020 ◽

Author(s):

Gang Tang ◽

Yi Du ◽

Haochen Guan ◽

Jieshuang Jia ◽

Nan Zhu ◽

...

Keyword(s):

Oxidative Stress ◽

Skeletal Muscle ◽

Diabetic Nephropathy ◽

Muscle Atrophy ◽

Barrier Function ◽

Muscle Protein ◽

C2c12 Cells ◽

Skeletal Muscle Atrophy ◽

Intestinal Barrier Function ◽

Gut Barrier Function

Muscle protein catabolism in patients with diabetic nephropathy (DN) results in striking losses of muscle proteins, which increases morbidity and mortality risks. Emerging evidence shows that short-chain fatty acids (SCFAs) play an important role in the maintenance of health and disease development. Recently, the connection between butyrate (a SCFA) and DN has been revealed, although the relationship between butyrate and muscle atrophy is still not clear. In our study, we found a significant decrease in butyrate in DN using metabolomics analyses. The addition of butyrate remarkably intestinal barrier function. Concurrently, butyrate could alleviate muscle atrophy and promote PI3K/AKT/mTOR signals, and suppress oxidative stress and autophagy in the skeletal muscle of db/db mice as well as high glucose/lipopolysaccharide (HG/LPS)-induced C2C12 cells. To further explore the mechanism, we found that GPR43, the key SCFAs signaling molecule, was significantly decreased in the skeletal muscle of db/db mice and HG/LPS-induced C2C12 cells. Overexpression of GPR43 could activate PI3K/AKT/mTOR signals and inhibit oxidative stress and autophagy in HG/LPS-induced C2C12 cells. Silencing of GPR43 blocked PI3K/AKT/mTOR signals improved by butyrate, as well as suppression of oxidative stress and reduction of autophagy. Ultimately, butyrate alleviated muscle atrophy in DN via GPR43-mediated PI3K/AKT/mTOR pathway

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Angiotensin-(1-7) Prevents Lipopolysaccharide-Induced Autophagy via the Mas Receptor in Skeletal Muscle

International Journal of Molecular Sciences ◽

10.3390/ijms21249344 ◽

2020 ◽

Vol 21 (24) ◽

pp. 9344

Author(s):

Juan Carlos Rivera ◽

Johanna Abrigo ◽

Franco Tacchi ◽

Felipe Simon ◽

Enrique Brandan ◽

...

Keyword(s):

Skeletal Muscle ◽

Muscle Atrophy ◽

B Cell Lymphoma ◽

Mapk Signaling ◽

C2c12 Cells ◽

Skeletal Muscle Atrophy ◽

Autophagic Flux ◽

Mrna Levels ◽

Mas Receptor ◽

The Impact

Skeletal muscle atrophy, which occurs in lipopolysaccharide (LPS)-induced sepsis, causes a severe muscle function reduction. The increased autophagy contributes to sepsis-induced skeletal muscle atrophy in a model of LPS injection, increasing LC3II/LC3I ratio, autophagy flux, and autophagosomes. Angiotensin-(1-7) (Ang-(1-7)) has anti-atrophic effects via the Mas receptor in skeletal muscle. However, the impact of Ang-(1-7) on LPS-induced autophagy is unknown. In this study, we determined the effect of Ang-(1-7) on sepsis-induced muscle autophagy. C57BL6 wild-type (WT) mice and mice lacking the Mas receptor (KO Mas) were injected with LPS together with the systemic administration of Ang-(1-7) to determine autophagy in skeletal muscle. We also evaluated autophagy and p38 and c-Jun N-terminal kinase (JNK)activation. Our results show that Ang-(1-7) prevents LPS-induced autophagy in the diaphragm, tibialis anterior, and gastrocnemius of WT mice, which is demonstrated by a decrease in the LC3II/LC3I ratio and mRNA levels of lc3b and ctsl. This effect was lost in KO Mas mice, suggesting the role of the Mas receptor. The results in C2C12 cells show that Ang-(1-7) reduces several LPS-dependent effects, such as autophagy (LC3II/LC3I ratio, autophagic flux, and autophagosomes), activation of p38 and JNK, B-cell lymphoma-2 (BCL2) phosphorylation, and disassembly of the Beclin1/BCL2 complex. In conclusion, Ang-(1-7)/Mas receptor reduces LPS-induced autophagy in skeletal muscle. In vitro assays indicate that Ang-(1-7) prevents LPS-induced autophagy and modifies the MAPK signaling and the disassembly of a complex involved at the beginning of autophagy.

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Immobilization-induced activation of key proteolytic systems in skeletal muscles is prevented by a mitochondria-targeted antioxidant

Journal of Applied Physiology ◽

10.1152/japplphysiol.00471.2013 ◽

2013 ◽

Vol 115 (4) ◽

pp. 529-538 ◽

Cited By ~ 79

Author(s):

Erin E. Talbert ◽

Ashley J. Smuder ◽

Kisuk Min ◽

Oh Sung Kwon ◽

Hazel H. Szeto ◽

...

Keyword(s):

Skeletal Muscle ◽

Muscle Atrophy ◽

Skeletal Muscles ◽

Mammalian Target Of Rapamycin ◽

Skeletal Muscle Atrophy ◽

Muscle Disuse ◽

Mitochondrial Ros ◽

Disuse Muscle Atrophy ◽

Muscle Fiber Atrophy ◽

First Time

Long periods of skeletal muscle disuse result in muscle fiber atrophy, and mitochondrial production of reactive oxygen species (ROS) appears to be a required signal for the increase in protein degradation that occurs during disuse muscle atrophy. The experiments detailed here demonstrate for the first time in limb muscle that the inactivity-induced increases in E3 ligase expression and autophagy biomarkers result from increases in mitochondrial ROS emission. Treatment of animals with a mitochondrial-targeted antioxidant also prevented the disuse-induced decrease in anabolic signaling (Akt/mammalian target of rapamycin signaling) that is normally associated with prolonged inactivity in skeletal muscles. Additionally, our results confirm previous findings that treatment with a mitochondrial-targeted antioxidant is sufficient to prevent casting-induced skeletal muscle atrophy, mitochondrial dysfunction, and activation of the proteases calpain and caspase-3. Collectively, these data reveal that inactivity-induced increases in mitochondrial ROS emission play a required role in activation of key proteolytic systems and the downregulation of important anabolic signaling molecules in muscle fibers exposed to prolonged inactivity.

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Daily Oral Administration of Protease-Treated Royal Jelly Protects Against Denervation-Induced Skeletal Muscle Atrophy

Nutrients ◽

10.3390/nu12103089 ◽

2020 ◽

Vol 12 (10) ◽

pp. 3089

Author(s):

Tomohiko Shirakawa ◽

Aki Miyawaki ◽

Takuma Matsubara ◽

Nobuaki Okumura ◽

Hideto Okamoto ◽

...

Keyword(s):

Skeletal Muscle ◽

Muscle Atrophy ◽

Royal Jelly ◽

Muscle Metabolism ◽

Sebacic Acid ◽

C2c12 Cells ◽

Skeletal Muscle Atrophy ◽

Muscle Weight ◽

Age Related ◽

Myoblast Cells

Honeybees produce royal jelly (RJ) from their cephalic glands. Royal jelly is a source of nutrition for the queen honey bee throughout its lifespan and is also involved in fertility and longevity. Royal jelly has long been considered beneficial to human health. We recently observed that RJ delayed impairment of motor function during aging, affecting muscle fiber size. However, how RJ affects skeletal muscle metabolism and the functional component of RJ is as of yet unidentified. We demonstrate that feeding mice with RJ daily prevents a decrease in myofiber size following denervation without affecting total muscle weight. RJ did not affect atrophy-related genes but stimulated the expression of myogenesis-related genes, including IGF-1 and IGF receptor. Trans-10-hydroxy-2-decenoic acid (10H2DA) and 10-hydroxydecanoic acid (10HDAA), two major fatty acids contained in RJ. After ingestion, 10H2DA and 10HDAA are metabolized into 2-decenedioic acid (2DA) and sebacic acid (SA) respectively. We found that 10H2DA, 10HDAA, 2DA, and SA all regulated myogenesis of C2C12 cells, murine myoblast cells. These novel findings may be useful for potential preventative and therapeutic applications for muscle atrophy disease included in Sarcopenia, an age-related decline in skeletal muscle mass and strength.

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