Is regulation of proteolysis associated with redox-state changes in rat skeletal muscle?
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ABSTRACT:
SUBMITTER: Tischler ME
PROVIDER: S-EPMC1162426 | biostudies-other | 1980 Dec
REPOSITORIES: biostudies-other
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Project description:BackgroundCalpain-3 deficiency causes oxidative and nitrosative stress-induced damage in skeletal muscle of LGMD2A patients, but mitochondrial respiratory chain function and anti-oxidant levels have not been systematically assessed in this clinical population previously.MethodsWe identified 14 patients with phenotypes consistent with LGMD2A and performed CAPN3 gene sequencing, CAPN3 expression/autolysis measurements, and in silico predictions of pathogenicity. Oxidative damage, anti-oxidant capacity, and mitochondrial enzyme activities were determined in a subset of muscle biopsies.ResultsTwenty-one disease-causing variants were detected along the entire CAPN3 gene, five of which were novel (c.338 T>C, c.500 T>C, c.1525-1 G>T, c.2115+4 T>G, c.2366 T>A). Protein- and mRNA-based tests confirmed in silico predictions and the clinical diagnosis in 75% of patients. Reductions in antioxidant defense mechanisms (SOD-1 and NRF-2, but not SOD-2), coupled with increased lipid peroxidation and protein ubiquitination, were observed in calpain-3 deficient muscle, indicating a redox imbalance primarily affecting non-mitochondrial compartments. Although ATP synthase levels were significantly lower in LGMD2A patients, citrate synthase, cytochrome c oxidase, and complex I+III activities were not different from controls.ConclusionsDespite significant oxidative damage and redox imbalance in cytosolic/myofibrillar compartments, mitochondrial respiratory chain function is largely maintained in skeletal muscle of LGMD2A patients.
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Project description:In intact animals, time of drug administration may be an important factor influencing drug response. Our general goal seeks to incorporate circadian time into the study of corticosteroid regulated gene expression. This study is designed to examine fluctuations in gene expression in skeletal muscle within the 24 hour circadian cycle in normal animals. Circadian time is relevant to designing optimal corticosteroid dosing regimens. Since levels of endogenous steroid exhibit circadian fluctuations, it is our hypothesis that the expression of genes controlled by corticosteroids either directly or indirectly will also exhibit a circadian pattern in normal animals. Keywords: time series design
2007-09-11 | GSE8989 | GEO
Project description:The relationship between the redox state and lactate accumulation in contracting human skeletal muscle was investigated. Ten men performed bicycle exercise for 10 min at 40 and 75% of maximal oxygen uptake [VO2(max.)], and to fatigue (4.8 +/- 0.6 min; mean +/- S.E.M.) at 100% VO2(max.). Biopsies from the quadriceps femoris muscle were analysed for NADH, high-energy phosphates and glycolytic intermediates. Muscle NADH was 0.20 +/- 0.02 mmol/kg dry wt. of muscle at rest, and decreased to 0.12 +/- 0.01 (P less than 0.01) after exercise at 40% VO2(max.), but no change occurred in the [lactate]/[pyruvate] ratio. These data, together with previous results on isolated cyanide-poisoned soleus muscle, where NADH increased while [lactate]/[pyruvate] ratio was unchanged [Sahlin & Katz (1986) Biochem. J. 239, 245-248], suggest that the observed changes in muscle NADH occurred within the mitochondria. After exercise at 75 and 100% VO2(max.), muscle NADH increased above the value at rest to 0.27 +/- 0.03 (P less than 0.05) and 0.32 +/- 0.04 (P less than 0.001) mmol/kg respectively. Muscle lactate was unchanged after exercise at 40% VO2(max.), but increased substantially at the higher work loads. At 40% VO2(max.), phosphocreatine decreased by 11% compared with the values at rest, and decreased further at the higher work loads. The decrease in phosphocreatine reflects increased ADP and Pi. It is concluded that muscle NADH decreases during low-intensity exercise, but increases above the value at rest during high-intensity exercise. The increase in muscle NADH is consistent with the hypothesis that the accelerated lactate production during submaximal exercise is due to a limited availability of O2 in the contracting muscle. It is suggested that the increases in NADH, ADP and Pi are metabolic adaptations, which primarily serve to activate the aerobic ATP production, and that the increased anaerobic energy production (phosphocreatine breakdown and lactate formation) is a consequence of these changes.
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Project description:Skeletal muscle accounts for the largest proportion of human body mass, on average, and is a key tissue in complex diseases and mobility. It is composed of several different cell and muscle fiber types. Here, we optimize single-nucleus ATAC-seq (snATAC-seq) to map skeletal muscle cell-specific chromatin accessibility landscapes in frozen human and rat samples, and single-nucleus RNA-seq (snRNA-seq) to map cell-specific transcriptomes in human. We additionally perform multi-omics profiling (gene expression and chromatin accessibility) on human and rat muscle samples.
| EGAS00001005730 | EGA
Project description:We investigated the relationship between markers of mitochondrial biogenesis, cell signaling, and antioxidant enzymes by depleting skeletal muscle glutathione with diethyl maleate (DEM) which resulted in a demonstrable increase in oxidative stress during exercise. Animals were divided into six groups: (1) sedentary control rats; (2) sedentary rats + DEM; (3) exercise control rats euthanized immediately after exercise; (4) exercise rats + DEM; (5) exercise control rats euthanized 4 h after exercise; and (6) exercise rats + DEM euthanized 4 h after exercise. Exercising animals ran on the treadmill at a 10% gradient at 20 m/min for the first 30 min. The speed was then increased every 10 min by 1.6 m/min until exhaustion. There was a reduction in total glutathione in the skeletal muscle of DEM treated animals compared to the control animals (P < 0.05). Within the control group, total glutathione was higher in the sedentary group compared to after exercise (P < 0.05). DEM treatment also significantly increased oxidative stress, as measured by increased plasma F2-isoprostanes (P < 0.05). Exercising animals given DEM showed a significantly greater increase in peroxisome proliferator activated receptor γ coactivator-1α (PGC-1α) mRNA compared to the control animals that were exercised (P < 0.05). This study provides novel evidence that by lowering the endogenous antioxidant glutathione in skeletal muscle and inducing oxidative stress through exercise, PGC-1α gene expression was augmented. These findings further highlight the important role of exercise induced oxidative stress in the regulation of mitochondrial biogenesis.
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Project description:Insulin resistance is a major cause of muscle wasting in patients with ESRD. Uremic metabolic acidosis impairs insulin signaling, which normally suppresses proteolysis. The low pH may inhibit the SNAT2 l-Glutamine (L-Gln) transporter, which controls protein synthesis via amino acid-dependent insulin signaling through mammalian target of rapamycin (mTOR). Whether SNAT2 also regulates signaling to pathways that control proteolysis is unknown. In this study, inhibition of SNAT2 with the selective competitive substrate methylaminoisobutyrate or metabolic acidosis (pH 7.1) depleted intracellular L-Gln and stimulated proteolysis in cultured L6 myotubes. At pH 7.1, inhibition of the proteasome led to greater depletion of L-Gln, indicating that amino acids liberated by proteolysis sustain L-Gln levels when SNAT2 is inhibited by acidosis. Acidosis shifted the dose-response curve for suppression of proteolysis by insulin to the right, confirming that acid increases proteolysis by inducing insulin resistance. Blocking mTOR or phosphatidylinositol-3-kinase (PI3K) increased proteolysis, indicating that both signaling pathways are involved in its regulation. When both mTOR and PI3K were inhibited, methylaminoisobutyrate or acidosis did not stimulate proteolysis further. Moreover, partial silencing of SNAT2 expression in myotubes and myoblasts with small interfering RNA stimulated proteolysis and impaired insulin signaling through PI3K. In conclusion, SNAT2 not only regulates mTOR but also regulates proteolysis through PI3K and provides a link among acidosis, insulin resistance, and protein wasting in skeletal muscle cells.
| S-EPMC2573006 | biostudies-literature
Project description:Nitrogen balances were measured in isolated perfused rat livers in the presence and absence of nitrogen donors. In all instances the balance apparently was incomplete. The expression [alanine][alpha-oxoglutarate]/[pyruvate][glutamate] remained fairly constant under the metabolic conditions studied, indicating that it may be at near-equilibrium. The source of the extra nitrogen seems to be derived from increased hepatic proteolysis. The addition of a nitrogen donor to the perfusate arrested proteolysis, as did the addition of pyruvate. The free mitochondrial [NAD(+)]/[NADH] ratio, calculated from the glutamate dehydrogenase and beta-hydroxybutyrate dehydrogenase reactants, showed similar values and exhibited parallel changes under most metabolic situations studied. These results suggest that, under the reported experimental conditions, both dehydrogenases share a common mitochondrial NAD pool. Glutamate dehydrogenase plays an important role in hepatic nitrogen metabolism in vivo.
| S-EPMC1166219 | biostudies-other
Project description:BMP9, a member of the TGF? superfamily, is a homodimer that forms a signaling complex with two type I and two type II receptors. Signaling through high-affinity activin receptor-like kinase 1 (ALK1) in endothelial cells, circulating BMP9 acts as a vascular quiescence factor, maintaining endothelial homeostasis. BMP9 is also the most potent BMP for inducing osteogenic signaling in mesenchymal stem cells in vitro and promoting bone formation in vivo. This activity requires ALK1, the lower affinity type I receptor ALK2, and higher concentrations of BMP9. In adults, BMP9 is constitutively expressed in hepatocytes and secreted into the circulation. Optimum concentrations of BMP9 are essential to maintain the highly specific endothelial-protective function. Factors regulating BMP9 stability and activity remain unknown. Here, we showed by chromatography and a 1.9 Å crystal structure that stable BMP9 dimers could form either with (D-form) or without (M-form) an intermolecular disulfide bond. Although both forms of BMP9 were capable of binding to the prodomain and ALK1, the M-form demonstrated less sustained induction of Smad1/5/8 phosphorylation. The two forms could be converted into each other by changing the redox potential, and this redox switch caused a major alteration in BMP9 stability. The M-form displayed greater susceptibility to redox-dependent cleavage by proteases present in serum. This study provides a mechanism for the regulation of circulating BMP9 concentrations and may provide new rationales for approaches to modify BMP9 levels for therapeutic purposes.
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Project description:It is not clear whether the muscle wasting commonly observed in hyperthyroidism is due to alteration in the rate of protein synthesis or degradation. The effect of experimental hyperthyroidism on skeletal-muscle proteolysis in the rat was studied by measuring alanine and tyrosine release from isolated skeletal muscles in vitro and 3-methyl-histidine excretion in vivo. Alanine release from the isolated epitrochlaris-muscle preparation was increased as soon as 24h after a 25 microgram dose of L-tri-iodothyronine in vivo. Conversely, alanine release from muscles of hypothyroid rats was decreased, but restored by L-tri-iodothyronine supplementation before death. Furthermore, 3-methylhistidine excretion was increased in hyperthyroid rats throughout an 18-day treatment period. The increased amino acid release from isolated muscles and the increased 3-methylhistidine excretion in vivo strongly suggests that hyperthyroidism increases skeletal-muscle proteolysis. Furthermore, the thyroid-hormone concentration may be an important factor in regulating muscle proteolysis.
| S-EPMC1162802 | biostudies-other
Project description:Thioredoxins (Trx's) regulate redox signaling and are localized to various cellular compartments. Specific redox-regulated pathways for adaptation of skeletal muscle to contractions are attenuated during aging, but little is known about the roles of Trx's in regulating these pathways. This study investigated the susceptibility of Trx1 and Trx2 in skeletal muscle to oxidation and reduction in vitro and the effects of aging and contractions on Trx1, Trx2, and thioredoxin reductase (TrxR) 1 and 2 contents and nuclear and cytosolic Trx1 and mitochondrial Trx2 redox potentials in vivo. The proportions of cytosolic and nuclear Trx1 and mitochondrial Trx2 in the oxidized or reduced forms were analyzed using redox Western blotting. In myotubes, the mean redox potentials were nuclear Trx1, -251 mV; cytosolic Trx1, -242mV; mitochondrial Trx2, -346mV, data supporting the occurrence of differing redox potentials between cell compartments. Exogenous treatment of myoblasts and myotubes with hydrogen peroxide or dithiothreitol modified glutathione redox status and nuclear and cytosolic Trx1, but mitochondrial Trx2 was unchanged. Tibialis anterior muscles from young and old mice were exposed to isometric muscle contractions in vivo. Aging increased muscle contents of Trx1, Trx2, and TrxR2, but neither aging nor endogenous ROS generated during contractions modified Trx redox potentials, although oxidation of glutathione and other thiols occurred. We conclude that glutathione redox couples in skeletal muscle are more susceptible to oxidation than Trx and that Trx proteins are upregulated during aging, but do not appear to modulate redox-regulated adaptations to contractions that fail during aging.
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