Effect of irradiation on Akt signaling in atrophying skeletal muscle

Muscle irradiation (IRR) exposure can accompany unloading during spaceflight or cancer treatment, and this has been shown to be sufficient by itself to induce skeletal muscle signaling associated with a remodeling response. Although protein kinase B/Akt has an established role in the regulation of muscle growth and metabolism, there is a limited understanding of how Akt signaling in unloaded skeletal muscle is affected by IRR. Therefore, we examined the combined effects of acute IRR and short-term unloading on muscle Akt signaling. Female C57BL/6 mice were subjected to load bearing or hindlimb suspension (HS) for 5 days ( n = 6/group). A single, unilateral hindlimb IRR dose (0.5 Gy X-ray) was administered on day 3. Gastrocnemius muscle protein expression was examined. HS resulted in decreased AktT308 phosphorylation, whereas HS+IRR resulted in increased AktT308 phosphorylation above baseline. HS resulted in reduced AktS473 phosphorylation, which was rescued by HS+IRR. Interestingly, IRR alone resulted in increased phosphorylation of AktS473, but not that of AktT308. HS resulted in decreased mTORC1 signaling, and this suppression was not altered by IRR. Both IRR and HS resulted in increased MuRF-1 expression, whereas atrogin-1 expression was not affected by either condition. These results demonstrate that either IRR alone or when combined with HS can differentially affect Akt phosphorylation, but IRR did not disrupt suppressed mTORC1 signaling by HS. Collectively, these findings highlight that a single IRR dose is sufficient to disrupt the regulation of Akt signaling in atrophying skeletal muscle.

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Ursolic acid stimulates mTORC1 signaling after resistance exercise in rat skeletal muscle

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00302.2013 ◽

2013 ◽

Vol 305 (6) ◽

pp. E760-E765 ◽

Cited By ~ 37

Author(s):

Riki Ogasawara ◽

Koji Sato ◽

Kazuhiko Higashida ◽

Koichi Nakazato ◽

Satoshi Fujita

Keyword(s):

Resistance Exercise ◽

Ursolic Acid ◽

Gastrocnemius Muscle ◽

Corn Oil ◽

Muscle Protein ◽

Akt Signaling ◽

Sprague Dawley ◽

Mtorc1 Signaling ◽

Exercise Induced ◽

Mtorc1 Activation

A recent study identified ursolic acid (UA) as a potent stimulator of muscle protein anabolism via PI3K/Akt signaling, thereby suggesting that UA can increase Akt-independent mTOR complex 1 (mTORC1) activation induced by resistance exercise via Akt signaling. The purpose of the present study was to investigate the effect of UA on resistance exercise-induced mTORC1 activation. The right gastrocnemius muscle of male Sprague-Dawley rats aged 11 wk was isometrically exercised via percutaneous electrical stimulation (stimulating ten 3-s contractions per set for 5 sets), while the left gastrocnemius muscle served as the control. UA or placebo (PLA; corn oil only) was injected intraperitoneally immediately after exercise. The rats were killed 1 or 6 h after the completion of exercise and the target tissues removed immediately. With placebo injection, the phosphorylation of p70S6K at Thr389 increased 1 h after resistance exercise but attenuated to the control levels 6 h after the exercise. On the other hand, the augmented phosphorylation of p70S6K was maintained even 6 h after exercise when UA was injected immediately after exercise. A similar trend of prolonged phosphorylation was observed in PRAS40 Thr246, whereas UA alone or resistance exercise alone did not alter its phosphorylation level at 6 h after intervention. These results indicate that UA is able to sustain resistance exercise-induced mTORC1 activity.

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Myostatin inhibits proliferation and insulin-stimulated glucose uptake in mouse liver cells

Biochemistry and Cell Biology ◽

10.1139/bcb-2014-0004 ◽

2014 ◽

Vol 92 (3) ◽

pp. 226-234 ◽

Cited By ~ 16

Author(s):

Rani Watts ◽

Mostafa Ghozlan ◽

Curtis C. Hughey ◽

Virginia L. Johnsen ◽

Jane Shearer ◽

...

Keyword(s):

Skeletal Muscle ◽

Cell Proliferation ◽

Glucose Uptake ◽

Liver Cell ◽

Noncoding Rna ◽

Muscle Growth ◽

Liver Cells ◽

Negative Regulator ◽

Kinase Substrate ◽

Akt Phosphorylation

Although myostatin functions primarily as a negative regulator of skeletal muscle growth and development, accumulating biological and epidemiological evidence indicates an important contributing role in liver disease. In this study, we demonstrate that myostatin suppresses the proliferation of mouse Hepa-1c1c7 murine-derived liver cells (50%; p < 0.001) in part by reducing the expression of the cyclins and cyclin-dependent kinases that elicit G1-S phase transition of the cell cycle (p < 0.001). Furthermore, real-time PCR-based quantification of the long noncoding RNA metastasis associated lung adenocarcinoma transcript 1 (Malat1), recently identified as a myostatin-responsive transcript in skeletal muscle, revealed a significant downregulation (25% and 50%, respectively; p < 0.05) in the livers of myostatin-treated mice and liver cells. The importance of Malat1 in liver cell proliferation was confirmed via arrested liver cell proliferation (p < 0.05) in response to partial Malat1 siRNA-mediated knockdown. Myostatin also significantly blunted insulin-stimulated glucose uptake and Akt phosphorylation in liver cells while increasing the phosphorylation of myristoylated alanine-rich C-kinase substrate (MARCKS), a protein that is essential for cancer cell proliferation and insulin-stimulated glucose transport. Together, these findings reveal a plausible mechanism by which circulating myostatin contributes to the diminished regenerative capacity of the liver and diseases characterized by liver insulin resistance.

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Insulin Stimulates Human Skeletal Muscle Protein Synthesis via an Indirect Mechanism Involving Endothelial-Dependent Vasodilation and Mammalian Target of Rapamycin Complex 1 Signaling

Molecular Endocrinology ◽

10.1210/mend.24.6.9996 ◽

2010 ◽

Vol 24 (6) ◽

pp. 1306-1306

Author(s):

Kyle L. Timmerman ◽

Jessica L. Lee ◽

Hans C. Dreyer ◽

Shaheen Dhanani ◽

Erin L. Glynn ◽

...

Keyword(s):

Skeletal Muscle ◽

Protein Synthesis ◽

Muscle Protein ◽

Muscle Protein Synthesis ◽

Mammalian Target Of Rapamycin ◽

Skeletal Muscle Protein ◽

Capillary Recruitment ◽

Isotope Tracers ◽

Skeletal Muscle Protein Synthesis ◽

Mtorc1 Signaling

Abstract Objective: Our objective was to determine whether endothelial-dependent vasodilation is an essential mechanism by which insulin stimulates human skeletal muscle protein synthesis and anabolism. Subjects: Subjects were healthy young adults (n = 14) aged 31 ± 2 yr. Design: Subjects were studied at baseline and during local leg infusion of insulin alone (control, n = 7) or insulin plus the nitric oxide synthase inhibitor NG-monomethyl-l-arginine (L-NMMA, n = 7) to prevent insulin-induced vasodilation. Methods: We measured skeletal muscle protein metabolism with stable isotope tracers, blood flow with indocyanine green, capillary recruitment with contrast enhanced ultrasound, glucose metabolism with stable isotope tracers, and phosphorylation of proteins associated with insulin (Akt) and amino acid-induced mammalian target of rapamycin(mTOR) complex 1 (mTORC1) signaling (mTOR, S6 kinase 1, and eukaryotic initiation factor 4Ebinding protein 1) with Western blot analysis. Results: No basal differences between groups were detected. During insulin infusion, blood flow and capillary recruitment increased in the control (P < 0.05) group only; Akt phosphorylation and glucose uptake increased in both groups (P < 0.05), with no group differences; and mTORC1 signaling increased more in control (P < 0.05) than in l-NMMA. Phenylalanine net balance increased (P < 0.05) in both groups, but with opposite mechanisms: increased protein synthesis (basal, 0.051 ± 0.006%/h; insulin, 0.077 ± 0.008%/h; P < 0.05) with no change in proteolysis in control and decreased proteolysis (P < 0.05) with no change in synthesis (basal, 0.061 ± 0.004%/h; insulin, 0.050 ± 0.006%/h; P value not significant) in l-NMMA. Conclusions: Endothelial-dependent vasodilation and the consequent increase in nutritive flow and mTORC1 signaling, rather than Akt signaling, are fundamental mechanisms by which insulin stimulates muscle protein synthesis in humans. Additionally, these data underscore that insulin modulates skeletal muscle proteolysis according to its effects on nutritive flow.

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Integrin signaling: linking mechanical stimulation to skeletal muscle hypertrophy

AJP Cell Physiology ◽

10.1152/ajpcell.00009.2019 ◽

2019 ◽

Vol 317 (4) ◽

pp. C629-C641 ◽

Cited By ~ 13

Author(s):

Marni D. Boppart ◽

Ziad S. Mahmassani

Keyword(s):

Skeletal Muscle ◽

Mechanical Load ◽

Muscle Protein ◽

Muscle Protein Synthesis ◽

Muscle Growth ◽

Mammalian Target Of Rapamycin ◽

Integrin Subunit ◽

Resistance Exercise Training ◽

Adhesion Protein ◽

Subunit Mrna

The α7β1-integrin is a transmembrane adhesion protein that connects laminin in the extracellular matrix (ECM) with actin in skeletal muscle fibers. The α7β1-integrin is highly expressed in skeletal muscle and is concentrated at costameres and myotendious junctions, providing the opportunity to transmit longitudinal and lateral forces across the membrane. Studies have demonstrated that α7-integrin subunit mRNA and protein are upregulated following eccentric contractions as a mechanism to reinforce load-bearing structures and resist injury with repeated bouts of exercise. It has been hypothesized for many years that the integrin can also promote protein turnover in a manner that can promote beneficial adaptations with resistance exercise training, including hypertrophy. This review provides basic information about integrin structure and activation and then explores its potential to serve as a critical mechanosensor and activator of muscle protein synthesis and growth. Overall, the hypothesis is proposed that the α7β1-integrin can contribute to mechanical-load induced skeletal muscle growth via an mammalian target of rapamycin complex 1-independent mechanism.

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Analysis of the Molecular Signaling Signatures of Muscle Protein Wasting Between the Intercostal Muscles and the Gastrocnemius Muscles in db/db Mice

International Journal of Molecular Sciences ◽

10.3390/ijms20236062 ◽

2019 ◽

Vol 20 (23) ◽

pp. 6062 ◽

Cited By ~ 1

Author(s):

Kun Woo Kim ◽

Mi-Ock Baek ◽

Ji-Young Choi ◽

Kuk Hui Son ◽

Mee-Sup Yoon

Keyword(s):

Gastrocnemius Muscle ◽

Muscle Wasting ◽

Muscle Protein ◽

Respiratory Muscles ◽

Diabetic Mouse ◽

Molecular Signaling ◽

Akt Phosphorylation ◽

Intercostal Muscles ◽

The Difference ◽

Gastrocnemius Muscles

Type 2 diabetes (T2D) patients suffer from dyspnea, which contributes to disease-related morbidity. Although T2D has been reported to induce a catabolic state in skeletal muscle, whether T2D induces muscle wasting in respiratory muscles has not yet been investigated. In this study, we examine the difference in the molecular signaling signature of muscle wasting between the intercostal and gastrocnemius muscles using db/db mice, a well-known diabetic mouse model. Akt phosphorylation was significantly decreased in both the intercostal and gastrocnemius muscles of db/db mice and was accompanied by a decrease in mTORC1 activity. In addition, FoxO phosphorylation was suppressed, and ubiquitin-proteasome degradation, characterized by the level of Atrogin-1 and MuRF1, was subsequently enhanced in both muscle types of db/db mice. An increase in LC3BII levels and a decrease in p62 levels marked the occurrence of substantial autophagy in the gastrocnemius muscle but not in the intercostal muscles of db/db mice. Therefore, we suggest that the signaling events of muscle wasting in the intercostal muscles of db/db mice are different from those in the gastrocnemius muscle of db/db mice.

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Skeletal-muscle growth and protein turnover

Biochemical Journal ◽

10.1042/bj1500235 ◽

1975 ◽

Vol 150 (2) ◽

pp. 235-243 ◽

Cited By ~ 222

Author(s):

D J Millward ◽

P J Garlick ◽

R J C Stewart ◽

D O Nnanyelugo ◽

J C Waterlow

Keyword(s):

Skeletal Muscle ◽

Growth Rate ◽

Protein Synthesis ◽

Muscle Protein ◽

Muscle Growth ◽

Muscle Size ◽

Synthesis Rate ◽

Protein Breakdown ◽

Breakdown Rate ◽

Breakdown Rates

Because of turnover, protein synthesis and breakdown can each be involved in the regulation of the growth of tissue protein. To investigate the regulation of skeletal-muscle-protein growth we measured rates of protein synthesis and breakdown in growing rats during development on a good diet, during development on a marginally low-protein diet and during rehabilitation on a good diet after a period of severe protein deficiency. Rates of protein synthesis were measured in vivo with a constant intravenous infusion of [14C]tyrosine. The growth rate of muscle protein was measured and the rate of breakdown calculated as breakdown rate=synthesis rate-growth rate. These measurements showed that during development on a good diet there was a fall with age in the rate of protein synthesis resulting from a fall in capacity (RNA concentration) and activity (synthesis rate per unit of RNA). There was a fall with age in the breakdown rate so that the rate was highest in the weaning rats, with a half-life of 3 days. There was a direct correlation between the fractional growth and breakdown rates. During rehabilitation on the good diet, rapid growth was also accompanied by high rates of protein breakdown. During growth on the inadequate diet protein synthesis rates were lesss than in controls, but growth occurred because of decreased rates of protein breakdown. This compression was not complete, however, since ultimate muscle size was only one-half that of controls. It is suggested that increased rates of protein breakdown are a necessary accompaniment to muscle growth and may result from the way in which myofibrils proliferate.

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Muscle mTORC1 suppression by IL-6 during cancer cachexia: a role for AMPK

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00410.2012 ◽

2013 ◽

Vol 304 (10) ◽

pp. E1042-E1052 ◽

Cited By ~ 64

Author(s):

James P. White ◽

Melissa J. Puppa ◽

Song Gao ◽

Shuichi Sato ◽

Stephen L. Welle ◽

...

Keyword(s):

Skeletal Muscle ◽

Cancer Cachexia ◽

Muscle Protein ◽

Muscle Protein Synthesis ◽

Mammalian Target Of Rapamycin ◽

Mtor Signaling ◽

Glucose Administration ◽

Mtorc1 Signaling ◽

Dose Dependent ◽

Mtor Activity

Although catabolic signaling has a well-established role in muscle wasting during cancer cachexia, the suppression of anabolic signaling also warrants further investigation. In cachectic tumor-bearing mice, circulating IL-6 levels are associated with suppressed muscle protein synthesis and mTORC1 signaling. We have found AMPK and IGF-I/insulin signaling, two well-known regulators of the mammalian target of rapamycin (mTOR), are altered with the progression of cachexia. How IL-6 can induce suppression of mTORC1 signaling remains to be established. The purpose of this study was to examine mTOR complex 1 (mTORC1) activation and regulation by IL-6 during cancer cachexia. IL-6 effects on mTOR activation were examined in Apc Min/+ mouse skeletal muscle and C2C12 myotubes. Systemic IL-6 overexpression in Apc Min/+ mice produced a dose-dependent suppression of mTOR signaling that corresponded to induction of STAT3 and AMPK phosphorylation. This result was also evident in IL-6-treated myotubes. Basal mTOR activation and mTOR responsiveness to glucose administration were suppressed in cachectic skeletal muscle. However, insulin induction of mTOR activity was maintained in IL-6-treated myotubes. Whereas IL-6 suppression of myotube mTOR activity was rescued by AMPK inhibition, inhibition of STAT3 signaling was not sufficient to rescue IL-6 suppression of mTOR activity. Last, treadmill exercise training was able to prevent IL-6-induced inhibition of mTOR signaling in Apc Min/+ mice independently of activated STAT. In conclusion, we report dose-dependent suppression of mTOR activity by IL-6 and suppressed mTOR responsiveness to glucose administration in Apc Min/+ mice. IL-6 suppression of mTOR activity was dependent on AMPK activation and independent of STAT signaling in myotubes.

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PO-184 Effects of resistance training and aerobic training on Fibronectin of Skeletal Muscle Extracellular Matrix and Satellite Cell in Aging Mice

Exercise Biochemistry Review ◽

10.14428/ebr.v1i4.12773 ◽

2018 ◽

Vol 1 (4) ◽

Author(s):

Jian Lu ◽

Shanshan Li ◽

Caizhen Chen

Keyword(s):

Skeletal Muscle ◽

Extracellular Matrix ◽

Resistance Training ◽

Protein Content ◽

Grip Strength ◽

Satellite Cell ◽

Gastrocnemius Muscle ◽

Aerobic Training ◽

Significant Difference ◽

Group A

Objective To investigate the effects of resistance and aerobic training on the content of fibronectin in skeletal muscle extracellular matrix and satellite cell in 9-month-old BALB/c mice, and then explore the effect of fibronectin content change on satellite cell and its intrinsic mechanism, which will provide a new research perspective and theoretical evidence for delaying sarcopenia. Methods 27 one-month-old BALB / c mice of SPF grade were purchased and fed for natural aging. At 9 months of age, the mice were divided into three groups randomly. Group R was resistance exercise training group (n=9), group A was aerobic exercise training group (n=9), and group C was control group (n=9). The load ladder model for resistance training in group R, the pyramid training program for 9 week training, 5 sets of load ladder training every time. 2 min for rest between sets, 1 min for rest between repetitions, 3 times a week. The treadmill training for aerobic training in group A for 9 weeks, the speed of 0.8km/h, 40min every time, 3 times a week. There is no training in group C. During the training, mouse grip strength was tested by the BIOSEB grip instrument once a week. After exercise intervention, the blood of mice was taken from the eyeball, and the gastrocnemius muscles were removed and placed in -80℃ temperature refrigerator to be freezed for tested . Immunofluorescence was used to detect FN and Pax7; The Real-time PCR was used to detect mRNA of FN, Sdc4, Fzd7, Wnt7a, c-Jun, Pax7; Western Blotting was used to detect the FN, sdc4, Fzd7, Wnt7a, c-Jun, p-c-Jun, Pax7 protein content. Results (1) Body weight, grip strength and skeletal muscle mass of mice: The body weight of group C and group R were significantly decreased after 9 weeks to compared with group A (P <0.05; P <0.05). The grip strength of group R and A was significantly increased in the ninth week to compared with group C, (P <0.01; P <0.05); The grip strength of group R and A in the ninth week were significantly higher than that in the first week (P <0.01; P <0.01)； (2) FN in skeletal muscle extracellular matrix and it’s receptor Sdc4: The integrated optical density (IOD) of FN in group R was higher than that in group C and group A, but there was no significant difference among the three groups. The FN mRNA in group R was significantly increased to compared with group A (P<0.05). The FN protein content in group R was significantly increased to compared with group C and group A (P <0.01; P <0.01). There was no significant difference in the expression of Sdc4 mRNA in gastrocnemius muscle among the three groups. Compared with group C, the Sdc4 protein content was significantly down-regulated in both group R and group A (P<0.01; P<0.05). (3) Wnt7a/PCP signaling pathway: The Wnt7a mRNA and Wnt7a protein content in the gastrocnemius muscle of group R were significantly increased to compared with group C (P<0.05; P<0.01)). Compared with group A, Wnt7a protein content in group R was also significantly increased (P<0.05). There was no significant difference in Fzd7 mRNA in gastrocnemius muscle among the three groups; but the Fzd7 protein content in group R was significantly increased to compared with group A (P<0.05). The c-Jun mRNA in group R was significantly increased to compared with group C and A (P<0.05; P< 0.05). The content of c-Jun protein in group R was significantly increased to compared with group C and group A (P<0.01; P<0.01). The content of p-c-Jun protein in group R was significantly increased to compared with group C (P<0.05). (4) Pax7 : The number of Pax7 positive cells in group R was higher than that in group C and group A, but there was no significant difference among the three groups. Pax7 mRNA in group R and group A were significantly lower than that in group C (P<0.05; P<0.01), but Pax7 protein content in group R was significantly higher than group C and group A (P <0.05; P <0.05) Conclusions (1) Exercise can improve muscle strength of aging mice. (2) Resistance training can promote FN in skeletal muscle extracellular matrix and improve skeletal muscle extracellular matrix components of aging mice. (3) Resistance exercise training can promote Pax7 expression through the increase of FN and up-regulation of Wnt7a / PCP signaling pathway, thus make it possible for satellite cell proliferation.

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PL - 033 A translational model of muscle protein synthetic bioactivity in vitro, ex vivo and in vivo

Exercise Biochemistry Review ◽

10.14428/ebr.v1i1.8513 ◽

2018 ◽

Vol 1 (1) ◽

Cited By ~ 1

Author(s):

Brian Carson ◽

Robert Davies ◽

Joseph Bass ◽

Catherine Norton ◽

Bijal Patel ◽

...

Keyword(s):

Skeletal Muscle ◽

Stable Isotope ◽

Resistance Exercise ◽

Ex Vivo ◽

Muscle Protein ◽

Muscle Growth ◽

Translational Model ◽

Nutrient Feeding

Objective The aim of this research was the development and validation of a translational model for the evaluation of exercise and nutrient stimulated muscle protein synthesis (MPS). To achieve this overall aim, three primary objectives had to be realised: (i) Development of an in vitro skeletal muscle cell bioassay to measure muscle growth and MPS; (ii) Development of an ex vivo model to evaluate the humoral effect on MPS in response to nutrient feeding and exercise; (iii) Use of a stable isotope technique to evaluate MPS in response to nutrient feeding and exercise in vivo. Methods To develop a novel in vitro skeletal muscle cell bioassay to measure muscle growth and MPS, C2C12 myoblasts were proliferated and subsequently differentiated to myotubes over 8 days in DMEM (2% HS). Changes in cell behavior and adhesion properties were monitored by measuring impedance via interdigitated microelectrodes using the xCELLigence system. MPS was measured by puromycin incorporation using the SUnSET technique, intracellular signalling measured by western blot, and myotube thickness by microscopy. To demonstrate the capability to monitor nutrient regulation of muscle growth, media was conditioned with a known potent regulator of MPS (leucine) in a dose response experiment (0.20 - 2.0 mM). To establish the ability of the bioassay to measure the humoral effect of MPS in response to feeding and exercise, media was conditioned by ex vivo human serum from fasted, rested, fed (protein and isonitrogenous non-essential amino acid (NEAA) control) and post-exercise conditions. To evaluate MPS in response to nutrient feeding and exercise in vivo, acute MPS (5 h) was assessed by measuring stable isotope deuterium oxide (D2O) incorporation into m. vastus lateralis skeletal muscle following consumption of either a Whey Protein (WP) or an isonitrogenous NEAA control combined with resistance exercise in resistance trained males. Results In vitro experiments observed a dose-response effect with a 32 % increase in cell index and a 27 % increase in cell thickness after 2 h in the presence of 2.0 mM leucine when compared with control myotubes. Ex vivo serum following ingestion of NEAA had no effect on protein signalling or MPS whereas WP fed serum significantly increased mTOR, P70S6K and 4E-BP1 phosphorylation (p<0.01, p<0.05) compared to fasted serum. Furthermore, the effect of WP fed serum on protein signalling and MPS was significantly increased (p<0.01, p<0.05) compared to NEAA fed serum. Ex vivo human serum following resistance exercise was also increased MPS (29 %) and phosphorylation of mTOR (6 %), p70S6K (12 %) and 4EBP1 (7 %), compared with resting serum. These ex vivo/in vitro findings translated to the in vivo model as myofibrillar fractional synthetic rates (myoFSR) (Basal 0.068±0.002%h-1 vs. WP 0.084±0.006 %h-1, p=0.033) and absolute synthetic rates (ASR) (Basal 10.34±1.01 vs. WP 13.18±0.71 g.day-1, p=0.026) were increased from basal levels only when resistance exercise was combined with WP ingestion and not the NEAA control (NEAA MPS 0.072±0.004%h-1, NEAA ASR 10.23±0.80 g.day-1). Thus, ingestion of WP in combination with resistance training augments acute MPS responses in resistance trained young men. Conclusions We have developed a translational model of muscle protein synthetic bioactivity using in vitro, ex vivo and in vivo methodologies. We have shown that we can impact MPS in vitro using ex vivo human serum to condition media, that MPS in vitro is differentially regulated by ex vivo serum containing bioactive WP compared to a non-bioactive NEAA control, and that this tranlates for resistance exercise combined with WP in humans when MyoFSR is measured using stable isotope technology. These experiments demonstrate that ex vivo/in vitro experiments translate to the in vivo model and these methods can be used to inform both exercise and nutrient human interventions.

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Multiomics-Identified Intervention to Restore Ethanol-Induced Dysregulated Proteostasis and Secondary Sarcopenia in Alcoholic Liver Disease

Cellular Physiology and Biochemistry ◽

10.33594/000000327 ◽

2021 ◽

Vol 55 (1) ◽

pp. 91-116

Keyword(s):

Skeletal Muscle ◽

Protein Synthesis ◽

Liver Disease ◽

Mouse Model ◽

Muscle Protein ◽

Protein Homeostasis ◽

Intermediary Metabolites ◽

Intermediary Metabolite ◽

Mtorc1 Signaling ◽

Oxidative Function

BACKGROUND/AIMS: Signaling and metabolic perturbations contribute to dysregulated skeletal muscle protein homeostasis and secondary sarcopenia in response to a number of cellular stressors including ethanol exposure. Using an innovative multiomics-based curating of unbiased data, we identified molecular and metabolic therapeutic targets and experimentally validated restoration of protein homeostasis in an ethanol-fed mouse model of liver disease. METHODS: Studies were performed in ethanol-treated differentiated C2C12 myotubes and physiological relevance established in an ethanol-fed mouse model of alcohol-related liver disease (mALD) or pair-fed control C57BL/6 mice. Transcriptome and proteome from ethanol treated-myotubes and gastrocnemius muscle from mALD and pair-fed mice were analyzed to identify target pathways and molecules. Readouts including signaling responses and autophagy markers by immunoblots, mitochondrial oxidative function and free radical generation, and metabolic studies by gas chromatography-mass spectrometry and sarcopenic phenotype by imaging. RESULTS: Multiomics analyses showed that ethanol impaired skeletal muscle mTORC1 signaling, mitochondrial oxidative pathways, including intermediary metabolite regulatory genes, interleukin-6, and amino acid degradation pathways are β-hydroxymethyl-butyrate targets. Ethanol decreased mTORC1 signaling, increased autophagy flux, impaired mitochondrial oxidative function with decreased tricarboxylic acid cycle intermediary metabolites, ATP synthesis, protein synthesis and myotube diameter that were reversed by HMB. Consistently, skeletal muscle from mALD had decreased mTORC1 signaling, reduced fractional and total muscle protein synthesis rates, increased autophagy markers, lower intermediary metabolite concentrations, and lower muscle mass and fiber diameter that were reversed by β-hydroxymethyl-butyrate treatment. CONCLUSION: An innovative multiomics approach followed by experimental validation showed that β-hydroxymethyl-butyrate restores muscle protein homeostasis in liver disease.

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