Determining the contributions of protein synthesis and breakdown to muscle atrophy requires non‐steady‐state equations

Kobak, Kamil; Lawrence, Marcus M.; Pharaoh, Gavin; Borowik, Agnieszka; Peelor, Frederick F.; Shipman, Patrick D.; Griffin, Timothy M.; Remmen, Holly Van; Miller, Benjamin F.

doi:10.1002/jcsm.12772

Cited by 15 publications

(11 citation statements)

References 41 publications

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“…For body water enrichment, 100 μL of plasma was placed in the inner well of an o‐ring cap of inverted screw‐capped tubes and placed in a heat block for overnight distillation at 80°C. Distilled samples were diluted 1:300 in ddH2O and analyzed on a liquid water isotope analyzer (Los Gatos Research, Los Gatos, CA, USA) against a standard curve prepared with samples containing different concentrations of D 2 O (Abbott et al., 2021; Borowik et al., 2023; Brown et al., 2021; Kobak et al., 2021).…”

Section: Methodsmentioning

confidence: 99%

High‐fat diet increases electron transfer flavoprotein synthesis and lipid respiration in skeletal muscle during exercise training in female mice

Batterson,

McGowan,

Borowik

et al. 2023

Physiological Reports

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High‐fat diet (HFD) and exercise remodel skeletal muscle mitochondria. The electron transfer flavoproteins (ETF) transfer reducing equivalents from β‐oxidation into the electron transfer system. Exercise may stimulate the synthesis of ETF proteins to increase lipid respiration. We determined mitochondrial remodeling for lipid respiration through ETF in the context of higher mitochondrial abundance/capacity seen in female mice. We hypothesized HFD would be a greater stimulus than exercise to remodel ETF and lipid pathways through increased protein synthesis alongside increased lipid respiration. Female C57BL/6J mice (n = 15 per group) consumed HFD or low‐fat diet (LFD) for 4 weeks then remained sedentary (SED) or completed 8 weeks of treadmill training (EX). We determined mitochondrial lipid respiration, RNA abundance, individual protein synthesis, and abundance for ETFα, ETFβ, and ETF dehydrogenase (ETFDH). HFD increased absolute and relative lipid respiration (p = 0.018 and p = 0.034) and RNA abundance for ETFα (p = 0.026), ETFβ (p = 0.003), and ETFDH (p = 0.0003). HFD increased synthesis for ETFα and ETFDH (p = 0.0007 and p = 0.002). EX increased synthesis of ETFβ and ETFDH (p = 0.008 and p = 0.006). Higher synthesis rates of ETF were not always reflected in greater protein abundance. Greater synthesis of ETF during HFD indicates mitochondrial remodeling which may contribute higher mitochondrial lipid respiration through enhanced ETF function.

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Section: Methodsmentioning

confidence: 99%

High‐fat diet increases electron transfer flavoprotein synthesis and lipid respiration in skeletal muscle during exercise training in female mice

Batterson,

McGowan,

Borowik

et al. 2023

Physiological Reports

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“…Heart tissue was fractionated to cytoplasmic and mitochondrial proteins by using differential centrifugation as previously described [ 39 , 40 ]. The pentafluorobenzyl-N,Ndi (pentafluorobenzyl) derivative of alanine was analyzed on an Agilent 7890A GC (Agilent, Santa Clara, USA) coupled to an Agilent 5975C MS (Agilent, Santa Clara, USA) as previously described [ 39 , 40 ]. To assay body water enrichment, serum was analyzed on LWIA-45-EP (Los Gatos Research).…”

Section: Methodsmentioning

confidence: 99%

The loss of cardiac SIRT3 decreases metabolic flexibility and proteostasis in an age-dependent manner

Newhardt

Matsuzaki

et al. 2022

GeroScience

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SIRT3 is a longevity factor that acts as the primary deacetylase in mitochondria. Although ubiquitously expressed, previous global SIRT3 knockout studies have shown primarily a cardiac-specific phenotype. Here, we sought to determine how specifically knocking out SIRT3 in cardiomyocytes (SIRTcKO mice) temporally affects cardiac function and metabolism. Mice displayed an age-dependent increase in cardiac pathology, with 10-month-old mice exhibiting significant loss of systolic function, hypertrophy, and fibrosis. While mitochondrial function was maintained at 10 months, proteomics and metabolic phenotyping indicated SIRT3 hearts had increased reliance on glucose as an energy substrate. Additionally, there was a significant increase in branched-chain amino acids in SIRT3cKO hearts without concurrent increases in mTOR activity. Heavy water labeling experiments demonstrated that, by 3 months of age, there was an increase in protein synthesis that promoted hypertrophic growth with a potential loss of proteostasis in SIRT3cKO hearts. Cumulatively, these data show that the cardiomyocyte-specific loss of SIRT3 results in severe pathology with an accelerated aging phenotype.

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“…11,23,24 To determine ribosomal biogenesis, we assessed RNA synthesis using isolated RNA as previously described. 8,25 Derivatized samples of alanine and RNA were run on an Agilent 7890A GC coupled to an Agilent 5975C MS. 23,24 Body water enrichment was determined on serum using a Liquid Water Isotope Analyzer (LWIA-45-EP, Los Gatos Research, San Jose, CA). 26 For protein and RNA turnover, we calculated the rise to plateau of fraction new over the time-course according to our previously published study.…”

Section: Protein and Rna Turnover Proteomics And Bioinformaticsmentioning

confidence: 99%

Impaired proteostatic mechanisms other than decreased protein synthesis limit old skeletal muscle recovery after disuse atrophy

Fuqua,

Lawrence,

Hettinger

et al. 2023

J cachexia sarcopenia muscle

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BackgroundSkeletal muscle mass and strength diminish during periods of disuse but recover upon return to weight bearing in healthy adults but are incomplete in old muscle. Efforts to improve muscle recovery in older individuals commonly aim at increasing myofibrillar protein synthesis via mammalian target of rapamycin (mTOR) stimulation despite evidence demonstrating that old muscle has chronically elevated levels of mammalian target of rapamycin complex 1 (mTORC1) activity. We hypothesized that protein synthesis is higher in old muscle than adult muscle, which contributes to a proteostatic stress that impairs recovery.MethodsWe unloaded hindlimbs of adult (10‐month) and old (28‐month) F344BN rats for 14 days to induce atrophy, followed by reloading up to 60 days with deuterium oxide (D2O) labelling to study muscle regrowth and proteostasis.ResultsWe found that old muscle has limited recovery of muscle mass during reloading despite having higher translational capacity and myofibrillar protein synthesis (0.029 k/day ± 0.002 vs. 0.039 k/day ± 0.002, P < 0.0001) than adult muscle. We showed that collagen protein synthesis was not different (0.005 k (1/day) ± 0.0005 vs. 0.004 k (1/day) ± 0.0005, P = 0.15) in old compared to adult, but old muscle had higher collagen concentration (4.5 μg/mg ± 1.2 vs. 9.8 μg/mg ± 0.96, P < 0.01), implying that collagen breakdown was slower in old muscle than adult muscle. This finding was supported by old muscle having more insoluble collagen (4.0 ± 1.1 vs. 9.2 ± 0.9, P < 0.01) and an accumulation of advanced glycation end products (1.0 ± 0.06 vs. 1.5 ± 0.08, P < 0.001) than adult muscle during reloading. Limited recovery of muscle mass during reloading is in part due to higher protein degradation (0.017 1/t ± 0.002 vs. 0.028 1/t ± 0.004, P < 0.05) and/or compromised proteostasis as evidenced by accumulation of ubiquitinated insoluble proteins (1.02 ± 0.06 vs. 1.22 ± 0.06, P < 0.05). Last, we showed that synthesis of individual proteins related to protein folding/refolding, protein degradation and neural‐related biological processes was higher in old muscle during reloading than adult muscle.ConclusionsOur data suggest that the failure of old muscle to recover after disuse is not due to limitations in the ability to synthesize myofibrillar proteins but because of other impaired proteostatic mechanisms (e.g., protein folding and degradation). These data provide novel information on individual proteins that accumulate in protein aggregates after disuse and certain biological processes such as protein folding and degradation that likely play a role in impaired recovery. Therefore, interventions to enhance regrowth of old muscle after disuse should be directed towards the identified impaired proteostatic mechanisms and not aimed at increasing protein synthesis.

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Determining the contributions of protein synthesis and breakdown to muscle atrophy requires non‐steady‐state equations

Cited by 15 publications

References 41 publications

High‐fat diet increases electron transfer flavoprotein synthesis and lipid respiration in skeletal muscle during exercise training in female mice

High‐fat diet increases electron transfer flavoprotein synthesis and lipid respiration in skeletal muscle during exercise training in female mice

The loss of cardiac SIRT3 decreases metabolic flexibility and proteostasis in an age-dependent manner

Impaired proteostatic mechanisms other than decreased protein synthesis limit old skeletal muscle recovery after disuse atrophy

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