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Muscle-specific and age-related changes in protein synthesis and protein degradation in response to hindlimb unloading in rats.

ABSTRACT: Disuse is a potent inducer of muscle atrophy, but the molecular mechanisms driving this loss of muscle mass are highly debated. In particular, the extent to which disuse triggers decreases in protein synthesis or increases in protein degradation, and whether these changes are uniform across muscles or influenced by age, is unclear. We aimed to determine the impact of disuse on protein synthesis and protein degradation in lower limb muscles of varied function and fiber type in adult and old rats. Alterations in protein synthesis and degradation were measured in the soleus, medial gastrocnemius, and tibialis anterior (TA) muscles of adult and old rats subjected to hindlimb unloading (HU) for 3, 7, or 14 days. Loss of muscle mass was progressive during the unloading period, but highly variable (-9 to -38%) across muscle types and between ages. Protein synthesis decreased significantly in all muscles, except for the old TA. Atrophy-associated gene expression was only loosely associated with protein degradation as muscle RING finger-1, muscle atrophy F-box (MAFbx), and Forkhead box O1 expression significantly increased in all muscles, but an increase in proteasome activity was only observed in the adult soleus. MAFbx protein levels were significantly higher in the old muscles compared with adult muscles, despite the old having higher expression of microRNA-23a. These results indicate that adult and old muscles respond similarly to HU, and the greatest loss in muscle mass occurs in predominantly slow-twitch extensor muscles due to a concomitant decrease in protein synthesis and increase in protein degradation.NEW & NOTEWORTHY In this study, we showed that age did not intensify the atrophy response to unloading in rats, but rather that the degree of atrophy was highly variable across muscles, indicating that changes in protein synthesis and protein degradation occur in a muscle-specific manner. Our data emphasize the importance of studying muscles of varying fiber-type and physiological function at multiple time points to fully understand the molecular mechanisms responsible for disuse atrophy.

SUBMITTER: Baehr LM

PROVIDER: S-EPMC5451534 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Muscle-specific and age-related changes in protein synthesis and protein degradation in response to hindlimb unloading in rats.

Baehr Leslie M LM West Daniel W D DWD Marshall Andrea G AG Marcotte George R GR Baar Keith K Bodine Sue C SC

Journal of applied physiology (Bethesda, Md. : 1985) 20170323 5

Disuse is a potent inducer of muscle atrophy, but the molecular mechanisms driving this loss of muscle mass are highly debated. In particular, the extent to which disuse triggers decreases in protein synthesis or increases in protein degradation, and whether these changes are uniform across muscles or influenced by age, is unclear. We aimed to determine the impact of disuse on protein synthesis and protein degradation in lower limb muscles of varied function and fiber type in adult and old rats. ...[more]

PMID: 28336537

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Project description:Skeletal muscle atrophy is associated with disease, aging, and disuse. Hindlimb unloading (HU) in animals provides an experimental model to study muscle atrophy. A comprehensive time course for how HU affects biomarkers of protein synthesis and degradation acutely and chronically and the associated resistance to an anabolic stimulus following disuse remain undocumented. Sixteen-week-old C57BL/6 mice underwent 0, 1, 12, 24, 72, 168, or 336 h of HU. Following 336 h of HU, mice were reloaded for 1, 24, or 72 h. Another group of mice underwent 120 h of HU, were fasted or refed, and were then compared with similar condition control animals (CTL). Protein content and phosphorylation of biomarkers of protein synthesis, degradation, and autophagy were assessed in the soleus muscle. Gastrocnemius, soleus, and plantaris muscles atrophied within 120 h of HU. Protein synthesis trended toward decrease following 24 h of HU. p70S6K phosphorylation and protein synthesis increased with reloading. Following HU, changes in MAFbx and DEPTOR expression and DEPTOR phosphorylation were consistent with development of a catabolic state. DEPTOR expression recovered following reloading. Animals that underwent 120 h of HU exhibited attenuation of refeeding-induced p70S6K phosphorylation compared with CTL counterparts. Following 120 h of HU, protein synthesis, eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) phosphorylation, and DEPTOR, MAFbx, and Sestrin1 expression indicated a catabolic state. Following 120 h of HU, autophagy markers, including p62 expression, REDD1 expression, LC3 ratio, and Unc-51-like autophagy-activating kinase 1 (ULK1) phosphorylation, indicated impaired autophagy. HU promotes a deleterious balance between protein synthesis and degradation. The time course herein provides scientists information about when the associated biomarkers become affected.NEW & NOTEWORTHY Hindlimb unloading causes significant skeletal muscle atrophy by adversely affecting the balance between protein synthesis and breakdown. This study demonstrates a more complete time course for changes in biomarkers associated with protein synthesis and breakdown and investigates the associated anabolic resistance to an anabolic stimulus following hindlimb unloading. These data in concert with information from other studies provide a basis for designing future experiments to optimally interrogate a desired cellular biomarker or pathway.

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Project description:Nitric oxide (NO), produced by NO-synthases via L-arginine oxidation, is an essential trigger for signaling processes involved in structural and metabolic changes in muscle fibers. Recently, it was shown that L-arginine administration prevented the decrease in levels of the muscle cytoskeletal proteins, desmin and dystrophin, in rat soleus muscle after 14 days of hindlimb unloading. Therefore, in this study, we investigated the effect of L-arginine administration on the degree of atrophy changes in the rat soleus muscles under unloading conditions, and on the content, gene expression, and phosphorylation level of titin, the giant protein of striated muscles, able to form a third type of myofilaments-elastic filaments. A 7-day gravitational unloading [hindlimb suspension (HS) group] resulted in a decrease in the soleus weight:body weight ratio (by 31.8%, p < 0.05), indicating muscle atrophy development. The content of intact titin (T1) decreased (by 22.4%, p < 0.05) and the content of proteolytic fragments of titin (T2) increased (by 66.7%, p < 0.05) in the soleus muscle of HS rats, compared to control rats. The titin gene expression and phosphorylation level of titin between these two groups were not significantly different. L-Arginine administration under 7-day gravitational unloading decreased the degree of atrophy changes and also prevented the decrease in levels of T1 in the soleus muscle as compared to HS group. Furthermore, L-arginine administration under unloading resulted in increased titin mRNA level (by 76%, p < 0.05) and decreased phosphorylation level of T2 (by 28%, p < 0.05), compared to those in the HS group. These results suggest that administration of L-arginine, the NO precursor, under unloading decreased the degree of atrophy changes, increased gene expression of titin and prevented the decrease in levels of T1 in the rat soleus muscle. The results can be used to search for approaches to reduce the development of negative changes caused by gravitational unloading in the muscle.

Project description:BackgroundSpaceflights-induced microgravity can alter various physiological processes in human's body including the functional status of the reproductive system. Rodent model of tail-suspension hindlimb unloading is extensively used to stimulate the organs responses to the microgravity condition. This study explores the potential effects of hindlimb unloading on testicular functions and spermatogenesis in adult male rats and the underlying mechanism/s.MethodsTwenty Sprague-Dawley rats were allotted into two groups: normally loaded group (control; all arms were in touch with the grid floor) and hindlimb unloaded group (HU; only the forearms were in contact with the grid floor).ResultsFollowing 30 days of exposure, the HU group saw a decline in body weight, testicular and epidydimal weights, and all semen parameters. The circulating concentrations of gonadotropin-releasing hormone (GnRH), follicle stimulating hormone (FSH), luteinizing hormone (LH) and testosterone significantly decreased, while levels of kisspeptin, corticosterone, inhibin, prolactin and estradiol (E2) increased in the HU group. Intratesticular levels of 5α-reductase enzyme and dihydrotestosterone (DHT) were suppressed, while the levels of aromatase and kisspeptin were significantly elevated in the HU group. Hypothalamic kisspeptin (Kiss1) mRNA expression levels were downregulated while its receptors (Kiss1R) were upregulated in the HU group. On the contrary, the mRNA expression levels of testicular Kiss1 were upregulated while Kiss1R were downregulated. The pituitary mRNA expression levels of FSHβ and LHβ decreased in the HU group. The levels of the antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and nitric oxide (NO) concentrations, and total antioxidant capacity (TAC) were elevated while malondialdehyde (MDA) concentrations declined in the testes of HU group. The testes of the HU rats showed positive immunostaining of caspase-3, heat shock protein 70 (HSP70) and Bcl2.ConclusionsAltogether, these results revealed an inhibitory effect of hindlimb unloading on kisspeptin signaling in the hypothalamic-pituitary-testicular axis with impaired spermatogenesis and steroidogenesis.

Dataset Information

Muscle-specific and age-related changes in protein synthesis and protein degradation in response to hindlimb unloading in rats.

Publications

Muscle-specific and age-related changes in protein synthesis and protein degradation in response to hindlimb unloading in rats.

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