scholarly journals Leucine degradation in cell-free extracts of skeletal muscle

1979 ◽  
Vol 178 (2) ◽  
pp. 475-489 ◽  
Author(s):  
R Odessey ◽  
A L Goldberg
Keyword(s):  
Skeletal Muscle ◽  
Competitive Inhibition ◽  
Primary Source ◽  
Mitochondrial Fraction ◽  
The Body ◽  
Oxidative Decarboxylation ◽  
Transaminase Activity ◽  
Leucine Oxidation ◽  
Activity Measurements

Since skeletal muscle is the major site in the body for oxidation of leucine, isoleucine and valine, the pathway and control of leucine oxidation were investigated in cell-free preparations of rat muscle. Leucine was found to be transaminated to 4-methyl-2-oxopentanoate, which was then oxidatively decarboxylated. On differential centrifugation 70–80% of the transaminase activity was recovered in the soluble fraction of the cell, and the remaining amount in the mitochondrial fraction. The transaminase, from both fractions had similar pH optima and both were markedly inhibited by Ca2+. Thus changes in cellular Ca2+ concentration may regulate transaminase activity. Both transaminases had a much higher affinity for 2-oxoglutarate than for pyruvate. Therefore the utilization of amino groups from leucine for the biosynthesis of alanine in muscle [Odessey, Khairallah & Goldberg (1974) J. Biol. Chem. 249, 7623–7629] in vivo involves transamination with 2-oxoglutarate to produce glutamate, which is then transaminated with pyruvate to produce alanine. The dehydrogenase activity assayed by the decarboxylation of methyl-2-oxo[1-14C]pentanoate was localized exclusively in the fraction containing mitochondria and required NAD+, CoA and thiamin pyrophosphate for optimal activity. Measurements of competitive inhibition suggested that the oxo acids of leucine, isoleucine and valine are all decarboxylated by the same enzyme. The enzyme activity was decreased by 90% upon freezing or sonication and was stimulated severalfold by Mg2+, K+ and phosphate ions. In addition, it was markedly inhibited by ATP, but not by non-metabolizable analogues. This observation suggests that splitting of ATP is required for inhibition. The oxidative decarboxylation of 4-methyl-2-oxopentanoate by the dehydrogenase appears to be the rate-limiting step for leucine oxidation in muscle homogenates and also in intact tissues. In fact, rat muscles incubated with [1-14C]leucine release 1-14C-labelled oxo acid into the medium at rates comparable with the rate of decarboxylation. Intact muscles also released the oxo acids of [1-14C]valine or [1-14C]isoleucine, but not of other amino acids. These findings suggest that muscle is the primary source of the branched-chain oxo acids found in the blood.

scholarly journals Recent advances in understanding the role of FOXO3

F1000Research ◽  
2018 ◽  
Vol 7 ◽  
pp. 1372 ◽  
Author(s):  
Renae J. Stefanetti ◽  
Sarah Voisin ◽  
Aaron Russell ◽  
Séverine Lamon
Keyword(s):  
Cell Cycle ◽  
Skeletal Muscle ◽  
Protein Turnover ◽  
Genetic Basis ◽  
The Body ◽  
Biological Processes ◽  
Forkhead Box ◽  
Protein Pool

The forkhead box O3 (FOXO3, or FKHRL1) protein is a member of the FOXO subclass of transcription factors. FOXO proteins were originally identified as regulators of insulin-related genes; however, they are now established regulators of genes involved in vital biological processes, including substrate metabolism, protein turnover, cell survival, and cell death. FOXO3 is one of the rare genes that have been consistently linked to longevity in in vivo models. This review provides an update of the most recent research pertaining to the role of FOXO3 in (i) the regulation of protein turnover in skeletal muscle, the largest protein pool of the body, and (ii) the genetic basis of longevity. Finally, it examines (iii) the role of microRNAs in the regulation of FOXO3 and its impact on the regulation of the cell cycle.

scholarly journals Skeletal muscle releases extracellular vesicles with distinct protein and miRNA signatures that accumulate and function within the muscle microenvironment

2021 ◽  
Author(s):  
Sho Watanabe ◽  
Yuri Sudo ◽  
Satoshi Kimura ◽  
Kenji Tomita ◽  
Makoto Noguchi ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Extracellular Vesicles ◽  
Cell Types ◽  
The Body ◽  
C2c12 Cells ◽  
Ips Cell ◽  
Potential Marker ◽  
And Function ◽  
Intercellular Communications

Extracellular vesicles (EVs) contain various regulatory molecules and mediate intercellular communications. Although EVs are secreted from various cell types, including skeletal muscle cells, and present in the blood, their identity is poorly characterized in vivo, limiting the identification of their origin in the blood. Since the skeletal muscle is the largest organ in the body, it could substantially contribute to circulating EVs as their source. However, due to the lack of defined markers that distinguish SkM-EVs from others, whether the skeletal muscle releases EVs in vivo and how much the skeletal muscle-derived EVs (SkM-EVs) account for plasma EVs remain poorly understood. In this work, we perform quantitative proteomic analyses on EVs released from C2C12 cells and human iPS cell-derived myocytes and identify potential marker proteins that mark SkM-EVs. These markers we identified apply to in vivo tracking of SkM-EVs. The results show that skeletal muscle makes only a subtle contribution to plasma EVs as their source in both control and exercise conditions in mice. On the other hand, we demonstrate that SkM-EVs are concentrated in the skeletal muscle interstitium. Furthermore, we show that interstitium EVs are highly enriched with the muscle-specific miRNAs and repress the expression of the paired box transcription factor Pax7, a master regulator for myogenesis. Taken together, our findings reveal that the skeletal muscle releases exosome-like small EVs with distinct protein and miRNA profiles in vivo and that SkM-EVs mainly play a role within the muscle microenvironment where they accumulate.

scholarly journals Characterization of proprioceptive system dynamics in behaving Drosophila larvae using high-speed volumetric microscopy

2018 ◽  
Author(s):  
Rebecca Vaadia ◽  
Wenze Li ◽  
Venkatakaushik Voleti ◽  
Aditi Singhania ◽  
Elizabeth M.C. Hillman ◽  
...  
Keyword(s):  
High Speed ◽  
Body Position ◽  
Activity Patterns ◽  
The Body ◽  
Larval Body ◽  
Activity Measurements ◽  
Natural Movement ◽  
Rigid Joints

SummaryProprioceptors provide feedback about body position that is essential for coordinated movement. Proprioceptive sensing of the position of rigid joints has been described in detail in several systems, however it is not known how animals with an elastic skeleton encode their body positions. Understanding how diverse larval body positions are dynamically encoded requires knowledge of proprioceptor activity patterns in vivo during natural movement. Here we applied high-speed volumetric SCAPE microscopy to simultaneously track the position, physical deformation, and temporal patterns of intracellular calcium activity of multidendritic proprioceptors in crawling Drosophila larvae. During the periodic segment contraction and relaxation that occurs during crawling, proprioceptors with diverse morphologies showed sequential onset of activity throughout each periodic episode. A majority of these proprioceptors showed activity during segment contraction with one neuron type activated by segment extension. Different timing of activity of contraction-sensing proprioceptors was related to distinct dendrite terminal targeting, providing a continuum of position encoding during all phases of crawling. These dynamics could endow different proprioceptors with specific roles in monitoring the progression of contraction waves, as well as body shape during other behaviors. We provide activity measurements during exploration as one example. Our results provide powerful new insights into the body-wide neuronal dynamics of the proprioceptive system in crawling Drosophila, and demonstrate the utility of our approach for characterization of neural encoding throughout the nervous system of a freely behaving animal.

Factors Controlling Movement of Skeletal Muscles

Leonardo ◽  
2015 ◽  
Vol 48 (3) ◽  
pp. 270-271
Author(s):  
Miranda D. Grounds
Keyword(s):  
Skeletal Muscle ◽  
Extracellular Matrix ◽  
Cell Membrane ◽  
Skeletal Muscles ◽  
Muscle Cells ◽  
The Body ◽  
Skeletal Muscle Cells ◽  
3 Dimensional

The contraction of specialized skeletal muscle cells results in classic movements of bones and other parts of the body that are vital for life. There is exquisite control over the movement of diverse types of muscles. This paper indicates the way in which skeletal muscles (myofibres) are formed; then factors that contribute to generating the movement are outlined: these include the nerve, sarcomeres, cytoskeleton, cell membrane and the extracellular matrix. The factors controlling the movement of mature myofibres in 3-dimensional tissues in vivo are vastly more complex than for tissue cultured immature muscle cells in an artificial in vitro environment.

scholarly journals The contribution of muscle, kidney, and splanchnic tissues to leucine transamination in humans

2018 ◽  
Vol 96 (4) ◽  
pp. 382-387 ◽  
Author(s):  
Giacomo Garibotto ◽  
Daniela Verzola ◽  
Monica Vettore ◽  
Paolo Tessari
Keyword(s):  
Skeletal Muscle ◽  
Regulatory Role ◽  
Whole Body ◽  
Leucine Oxidation ◽  
Irreversible Oxidation ◽  
Isotope Kinetics

The first steps of leucine utilization are reversible deamination to α-ketoisocaproic acid (α-KIC) and irreversible oxidation. Recently, the regulatory role of leucine deamination over oxidation was underlined in rodents. Our aim was to measure leucine deamination and reamination in the whole body, in respect to previously determined rates across individual organs, in humans. By leucine and KIC isotope kinetics, we determined whole-body leucine deamination and reamination, and we compared these rates with those already reported across the sampled organs. As an in vivo counterpart of the “metabolon” concept, we analysed ratios between oxidation and either deamination or reamination. Leucine deamination to KIC was greater than KIC reamination to leucine in the whole body (p = 0.005), muscles (p = 0.005), and the splanchnic area (p = 0.025). These rates were not significantly different in the kidneys. Muscle accounted for ≈60% and ≈78%, the splanchnic bed for ≈15% and ≈15%, and the kidney for ≈12% and ≈18%, of whole-body leucine deamination and reamination rates, respectively. In the kidney, percent leucine oxidation over either deamination or reamination was >3-fold greater than muscle and the splanchnic bed. Skeletal muscle contributes by the largest fraction of leucine deamination, reamination, and oxidation. However, in relative terms, the kidney plays a key role in leucine oxidation.

scholarly journals Defective endoplasmic reticulum-mitochondria contacts and bioenergetics in SEPN1-related myopathy

2020 ◽  
Vol 28 (1) ◽  
pp. 123-138 ◽  
Author(s):  
Anne Filipe ◽  
Alexander Chernorudskiy ◽  
Sandrine Arbogast ◽  
Ersilia Varone ◽  
Rocío-Nur Villar-Quiles ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Body Composition ◽  
Muscle Weakness ◽  
Super Resolution ◽  
In Vitro Models ◽  
The Body ◽  
Mitochondrial Bioenergetics

AbstractSEPN1-related myopathy (SEPN1-RM) is a muscle disorder due to mutations of the SEPN1 gene, which is characterized by muscle weakness and fatigue leading to scoliosis and life-threatening respiratory failure. Core lesions, focal areas of mitochondria depletion in skeletal muscle fibers, are the most common histopathological lesion. SEPN1-RM underlying mechanisms and the precise role of SEPN1 in muscle remained incompletely understood, hindering the development of biomarkers and therapies for this untreatable disease. To investigate the pathophysiological pathways in SEPN1-RM, we performed metabolic studies, calcium and ATP measurements, super-resolution and electron microscopy on in vivo and in vitro models of SEPN1 deficiency as well as muscle biopsies from SEPN1-RM patients. Mouse models of SEPN1 deficiency showed marked alterations in mitochondrial physiology and energy metabolism, suggesting that SEPN1 controls mitochondrial bioenergetics. Moreover, we found that SEPN1 was enriched at the mitochondria-associated membranes (MAM), and was needed for calcium transients between ER and mitochondria, as well as for the integrity of ER-mitochondria contacts. Consistently, loss of SEPN1 in patients was associated with alterations in body composition which correlated with the severity of muscle weakness, and with impaired ER-mitochondria contacts and low ATP levels. Our results indicate a role of SEPN1 as a novel MAM protein involved in mitochondrial bioenergetics. They also identify a systemic bioenergetic component in SEPN1-RM and establish mitochondria as a novel therapeutic target. This role of SEPN1 contributes to explain the fatigue and core lesions in skeletal muscle as well as the body composition abnormalities identified as part of the SEPN1-RM phenotype. Finally, these results point out to an unrecognized interplay between mitochondrial bioenergetics and ER homeostasis in skeletal muscle. They could therefore pave the way to the identification of biomarkers and therapeutic drugs for SEPN1-RM and for other disorders in which muscle ER-mitochondria cross-talk are impaired.

Towards bioengineered skeletal muscle: recent developments in vitro and in vivo

2021 ◽  
Author(s):  
Anita Quigley ◽  
Catherine Ngan ◽  
Kate Firipis ◽  
Cathal D. O’Connell ◽  
Elena Pirogova ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Ex Vivo ◽  
Neuromuscular Disorders ◽  
Structural Features ◽  
The Body ◽  
Muscle Loss ◽  
Volumetric Muscle Loss ◽  
Free Flap Transfer

Abstract Skeletal muscle is a functional tissue that accounts for approximately 40% of the human body mass. It has remarkable regenerative potential, however, trauma and volumetric muscle loss, progressive disease and aging can lead to significant muscle loss that the body cannot recover from. Clinical approaches to address this range from free-flap transfer for traumatic events involving volumetric muscle loss, to myoblast transplantation and gene therapy to replace muscle loss due to sarcopenia and hereditary neuromuscular disorders, however, these interventions are often inadequate. The adoption of engineering paradigms, in particular materials engineering and materials/tissue interfacing in biology and medicine, has given rise to the rapidly growing, multidisciplinary field of bioengineering. These methods have facilitated the development of new biomaterials that sustain cell growth and differentiation based on bionic biomimicry in naturally occurring and synthetic hydrogels and polymers, as well as additive fabrication methods to generate scaffolds that go some way to replicate the structural features of skeletal muscle. Recent advances in biofabrication techniques have resulted in significant improvements to some of these techniques and have also offered promising alternatives for the engineering of living muscle constructs ex vivo to address the loss of significant areas of muscle. This review highlights current research in this area and discusses the next steps required towards making muscle biofabrication a clinical reality.

scholarly journals In vivo measurement of synthesis rate of individual skeletal muscle mitochondrial proteins

2008 ◽  
Vol 295 (5) ◽  
pp. E1255-E1268 ◽  
Author(s):  
Abdul Jaleel ◽  
Kevin R. Short ◽  
Yan W. Asmann ◽  
Katherine A. Klaus ◽  
Dawn M. Morse ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Dysfunction ◽  
Translational Regulation ◽  
Protein Identification ◽  
Mitochondrial Protein ◽  
Mitochondrial Fraction ◽  
Mitochondrial Proteins ◽  
Synthesis Rate ◽  
Gene Transcripts

Skeletal muscle mitochondrial dysfunction occurs in many conditions including aging and insulin resistance, but the molecular pathways of the mitochondrial dysfunction remain unclear. Presently, no methodologies are available to measure synthesis rates of individual mitochondrial proteins, which limits our ability to fully understand the translational regulation of gene transcripts. Here, we report a methodology to measure synthesis rates of multiple muscle mitochondrial proteins, which, along with large-scale measurements of mitochondrial gene transcripts and protein concentrations, will enable us to determine whether mitochondrial alteration is due to transcriptional or translational changes. The methodology involves in vivo labeling of muscle proteins with l-[ ring-13C6]phenylalanine, protein purification by two-dimensional gel electrophoresis of muscle mitochondrial fraction, and protein identification and stable isotope abundance measurements by tandem mass spectrometry. Synthesis rates of 68 mitochondrial and 23 nonmitochondrial proteins from skeletal muscle mitochondrial fraction showed a 10-fold range, with the lowest rate for a structural protein such as myosin heavy chain (0.16 ± 0.04%/h) and the highest for a mitochondrial protein such as dihydrolipoamide branched chain transacylase E2 (1.5 ± 0.42%/h). This method offers an opportunity to better define the translational regulation of proteins in skeletal muscle or other tissues.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

Effects of Unfractionated and Low Molecular Weight Heparins on Platelet Thromboxane Biosynthesis “In Vivo”

1994 ◽  
Vol 72 (06) ◽  
pp. 942-946 ◽  
Author(s):  
Raffaele Landolfi ◽  
Erica De Candia ◽  
Bianca Rocca ◽  
Giovanni Ciabattoni ◽  
Armando Antinori ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Unfractionated Heparin ◽  
Low Molecular Weight Heparin ◽  
Primary Source ◽  
In Vivo Studies ◽  
Low Molecular Weight ◽  
Heparin Administration ◽  
To Receive

SummarySeveral “in vitro” and “in vivo” studies indicate that heparin administration may affect platelet function. In this study we investigated the effects of prophylactic heparin on thromboxane (Tx)A2 biosynthesis “in vivo”, as assessed by the urinary excretion of major enzymatic metabolites 11-dehydro-TxB2 and 2,3-dinor-TxB2. Twenty-four patients who were candidates for cholecystectomy because of uncomplicated lithiasis were randomly assigned to receive placebo, unfractionated heparin, low molecular weight heparin or unfractionaed heparin plus 100 mg aspirin. Measurements of daily excretion of Tx metabolites were performed before and during the treatment. In the groups assigned to placebo and to low molecular weight heparin there was no statistically significant modification of Tx metabolite excretion while patients receiving unfractionated heparin had a significant increase of both metabolites (11-dehydro-TxB2: 3844 ± 1388 vs 2092 ±777, p <0.05; 2,3-dinor-TxB2: 2737 ± 808 vs 1535 ± 771 pg/mg creatinine, p <0.05). In patients randomized to receive low-dose aspirin plus unfractionated heparin the excretion of the two metabolites was largely suppressed thus suggesting that platelets are the primary source of enhanced thromboxane biosynthesis associated with heparin administration. These data indicate that unfractionated heparin causes platelet activation “in vivo” and suggest that the use of low molecular weight heparin may avoid this complication.

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