Summary of study ST000645

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000460. The data can be accessed directly via it's Project DOI: 10.21228/M8M027 This work is supported by NIH grant, U2C- DK119886.

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Study IDST000645
Study TitleEffects of Exercise on Dystrophic Mouse Muscle Amino Acids (part I)
Study SummaryWe will use male C57BL10, mdx, and nNOS-/- mice (n = 10 each group) to characterize the skeletal and cardiac muscle metabolomes. Half of the mice in each group will remain sedentary while the other half will be subjected to a single bout of treadmill exercise to exhaustion. Mice will be euthanized immediately postexercise and blood, hearts, and hindlimb muscles will be harvested and frozen as detailed in the General Methods. The heart and gastrocnemius, soleus, and quadriceps muscles of one hindlimb will be sent to the Mayo Clinic Metabolomics Resource Core. The heart and quadriceps muscle will be used for untargeted metabolomics profiling (LC/MS) while the gastrocnemius and soleus muscles will be used for targeted analyses of amino acids plus amino metabolites, non-esterified fatty acids, and citric acid cycle intermediates.
Institute
Mayo Clinic
Last NameThomas
First NameGail
AddressPenn State Hershey Heart and Vascular Institute Penn State College of Medicine 500 University Drive, MC H047 Hershey, PA 17033
Emailgthomas4@hmc.psu.edu
Phone717-531-0003, ext. 287087
Submit Date2017-06-23
Analysis Type DetailLC-MS
Release Date2019-07-17
Release Version1
Gail Thomas Gail Thomas
https://dx.doi.org/10.21228/M8M027
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000460
Project DOI:doi: 10.21228/M8M027
Project Title:The dystrophic muscle metabolome: effects of exercise and NO donor therapy
Project Summary:"In Duchenne and Becker muscular dystrophy (DMD, BMD), loss of the cytoskeletal protein dystrophin weakens the sarcolemma and disrupts cellular signaling, rendering the diseased muscles susceptible to contractioninduced damage. We and others have shown that loss of neuronal nitric oxide synthase (nNOSμ) from the sarcolemma of dystrophin-deficient muscle causes functional muscle ischemia during exercise due to unopposed sympathetic vasoconstriction, thereby exacerbating fatigue and injury of the diseased muscles. Genetic and pharmacologic strategies targeting nNOSμ-NO signaling ameliorate functional muscle ischemia, as well as many other features of the dystrophic phenotype in the mdx mouse model of DMD/BMD. These findings suggest that the therapeutic benefit of NO likely extends beyond its vascular effects. A growing body of evidence indicates that NO directly influences muscle metabolism through effects on glucose transport as well as mitochondrial biogenesis and function. Both nNOS-/- mice and mdx mice exhibit muscle mitochondrial dysfunction, decreased resistance to fatigue, and exercise-induced muscle injury, suggesting a causal role of nNOSμ-NO deficiency. However, the specific metabolic changes resulting from reduced NO signaling that might render dystrophic muscle susceptible to fatigue and use-dependent injury remain poorly defined. Therefore, the goal of this pilot metabolomics study is to identify the unique biochemical profiles of skeletal and cardiac muscles of mdx mice to gain further mechanistic insight into the pathophysiological role of NO deficiency in muscular dystrophy. In Aim 1, we will characterize the skeletal and cardiac muscle metabolomes of mdx and nNOS-/- mice at rest and following a single bout of treadmill exercise with the goal of discovering common metabolic signatures caused by loss of NO signaling. In Aim 2, we will evaluate the potential of a NO donor drug that is under development as a therapeutic for DMD/BMD to improve the skeletal and cardiac muscle metabolomes in mdx mice. As a result of this pilot study, we hope to gain new understanding of the metabolic derangements in dystrophin-deficient muscle, insight into the therapeutic effects of NO replacement, and to identify new pathogenic mechanisms and putative therapeutic targets that will form the basis of future grant applications."
Institute:Mayo Clinic
Last Name:Thomas
First Name:Gail
Address:Penn State Hershey Heart and Vascular Institute Penn State College of Medicine 500 University Drive, MC H047 Hershey, PA 17033
Email:gthomas4@hmc.psu.edu
Phone:717-531-0003, ext. 287087

Subject:

Subject ID:SU000668
Subject Type:Mouse
Subject Species:Mus musculus
Taxonomy ID:10090
Species Group:Mammal

Factors:

Subject type: Mouse; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Time point Grouping
SA036439ms5942-7Post Run RUN-BL10 Control
SA036440ms5942-9Post Run RUN-BL10 Control
SA036441ms5942-10Post Run RUN-BL10 Control
SA036442ms5942-6Post Run RUN-BL10 Control
SA036443ms5942-8Post Run RUN-BL10 Control
SA036444ms5942-28Post Run RUN-MDX
SA036445ms5942-26Post Run RUN-MDX
SA036446ms5942-30Post Run RUN-MDX
SA036447ms5942-27Post Run RUN-MDX
SA036448ms5942-29Post Run RUN-MDX
SA036449ms5942-18Post Run RUN-nNOS-/-
SA036450ms5942-19Post Run RUN-nNOS-/-
SA036451ms5942-20Post Run RUN-nNOS-/-
SA036452ms5942-17Post Run RUN-nNOS-/-
SA036453ms5942-16Post Run RUN-nNOS-/-
SA036454ms5942-5Sedentary SED-BL10 Control
SA036455ms5942-4Sedentary SED-BL10 Control
SA036456ms5942-3Sedentary SED-BL10 Control
SA036457ms5942-1Sedentary SED-BL10 Control
SA036458ms5942-2Sedentary SED-BL10 Control
SA036459ms5942-25Sedentary SED-MDX
SA036460ms5942-24Sedentary SED-MDX
SA036461ms5942-21Sedentary SED-MDX
SA036462ms5942-22Sedentary SED-MDX
SA036463ms5942-23Sedentary SED-MDX
SA036464ms5942-15Sedentary SED-nNOS-/-
SA036465ms5942-14Sedentary SED-nNOS-/-
SA036466ms5942-11Sedentary SED-nNOS-/-
SA036467ms5942-12Sedentary SED-nNOS-/-
SA036468ms5942-13Sedentary SED-nNOS-/-
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Collection:

Collection ID:CO000662
Collection Summary:Blood and Tissue Harvesting: Mice will be euthanized and blood will be taken by cardiac puncture, centrifuged, and the plasma will be stored at -80ºC. The heart, diaphragm, and muscles of both hindlimbs (gastrocnemius, soleus, quadriceps) will be dissected and snap frozen in liquid nitrogen. Tibialis anterior and plantaris muscles from both hindlimbs also will be dissected and one set will be snap frozen in liquid nitrogen while the other set will be mounted in OCT and frozen in isopentane cooled by liquid nitrogen. Cryosections will be used to evaluate disease activity and exercise-induced muscle injury by staining with: (a) hematoxylin and eosin to assess gross morphology, cellular infiltration, and necrosis, (b) anti-F4/80 to label macrophages, and (c) anti-IgG or IgM to label damaged muscle fibers.
Sample Type:Heart tissue

Treatment:

Treatment ID:TR000682
Treatment Summary:"Studies will be performed in 12-16 week old male mdx mice, C57BL10 control mice, and nNOS-/- mice obtained from Jackson Laboratory. All protocols will be approved by the Penn State College of Medicine Institutional Animal Care and Use Committee. Treadmill Exercise: Mice will run on a horizontal treadmill (Columbus Instruments) to assess fatigue by a single bout of exercise beginning at 5 m/min for 5 min followed by 1 m/min increases every minute until exhaustion. Electric shocks will not be used to stimulate running due to adverse effects in mdx mice."

Sample Preparation:

Sampleprep ID:SP000675
Sampleprep Summary:mouse gastrocnemius amino acids

Combined analysis:

Analysis ID AN000977
Analysis type MS
Chromatography type Reversed phase
Chromatography system Waters Acquity
Column Waters Acquity BEH C18 (150 x 2.1mm, 1.7um)
MS Type ESI
MS instrument type Triple quadrupole
MS instrument name Thermo Quantum Ultra
Ion Mode POSITIVE
Units picomol/mg

Chromatography:

Chromatography ID:CH000702
Instrument Name:Waters Acquity
Column Name:Waters Acquity BEH C18 (150 x 2.1mm, 1.7um)
Chromatography Type:Reversed phase

MS:

MS ID:MS000872
Analysis ID:AN000977
Instrument Name:Thermo Quantum Ultra
Instrument Type:Triple quadrupole
MS Type:ESI
Ion Mode:POSITIVE
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