Summary of Study ST000623

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 PR000455. The data can be accessed directly via it's Project DOI: 10.21228/M8Q415 This work is supported by NIH grant, U2C- DK119886.

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Study IDST000623
Study TitleNon-targeted Metabolomics Analysis of Golden Retriever Muscular Dystrophy-Affected Muscles Reveals Alterations in Arginine and Proline Metabolism, and Elevations in Glutamic and Oleic Acid In Vivo
Study SummaryLike Duchenne muscular dystrophy (DMD), the Golden Retriever Muscular Dystrophy (GRMD) dog model of DMD exhibits is characterized by muscle necrosis, progressive paralysis, and pseudohypertrophy in specific skeletal muscles. This severe GRMD phenotype included atrophy of the biceps femoris (BF) and compared to unaffected normal dogs, while the long digital extensor (LDE) of the pelvic limb, serving as a hip flex and stifle extensor, is unaffected. A recent microarray analysis of GRMD identified alterations in genes associated with lipid metabolism and energy production. We, therefore, undertook a non-targeted metabolomics analysis of the GRMD BF (affected) and LDE (unaffected) using GC-MS to identify underlying metabolic defects specific for affected GRMD skeletal muscle. Of the 134 metabolites identified in BF, eight were significantly altered in GRMD BF compared to control BF (Glutamic Acid (2.48 fold vs. controls); Oleic Acid (1.76 fold vs. controls); Proline (1.73 fold vs. controls); Myoinositol-2- Phosphate (0.44 fold vs. controls); Fumaric Acid (0.40 fold vs. controls); Carnosine (0.40 fold vs. controls); Lactamide (0.33 fold vs. controls); and Stearamide (0.23 fold vs. controls). Pathway analysis of the T-test significant metabolites identified BF muscle metabolites significantly enriched for Arginine and proline metabolism (p=5.8E-4, FDR=0.04) and Alanine, aspartate, and glutamate metabolism (p=1.3E-3, FDR=0.05). The GRMD LDE previously reported to be unaffected, in contrast, had only one significantly altered metabolite (3-Phosphoglyceric Acid (0.35 Fold vs. controls)).The identification of elevated BF Oleic acid (a long-chain fatty acid) is consistent with recent microarray studies identifying altered lipid metabolism genes, while alterations in Arginine and Proline metabolism are consistent with recent studies identifying elevated L-arginine in DMD patient sera as a biomarker of disease (alterations in DMD or GRMD muscle itself have not previously been reported).Together, these studies demonstrate muscle-specific alterations in GRMD-affected muscle, which illustrate previously unidentified metabolic changes.
Institute
University of North Carolina at Chapel Hill
DepartmentMcAllister heart Institute, Department of Internal medicine, Department of Pathology & Laboratory, Department of Pharmacology
LaboratoryMultiple Centers
Last NameWillis
First NameMonte
Address111 Mason Farm Road, MBRB 2336, Chapel Hill, North Carolina, 27599-7126, USA
Emailmonte_willis@med.unc.edu
Phone(984) 999-5431
Submit Date2017-04-27
Study CommentsSkeletal Muscle (Biceps femoris (BF), long digital extensor(LDE))
Raw Data AvailableYes
Raw Data File Type(s)d
Analysis Type DetailGC-MS
Release Date2017-11-20
Release Version1
Monte Willis Monte Willis
https://dx.doi.org/10.21228/M8Q415
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR000455
Project DOI:doi: 10.21228/M8Q415
Project Title:Non-targeted Metabolomics Analysis of Golden Retriever Muscular Dystrophy-Affected Muscles Dog
Project Type:GC-MS non targeted analysis
Project Summary:Non-targeted Metabolomics Analysis of Golden Retriever Muscular Dystrophy-Affected Muscles Dog
Institute:University of North Carolina at Chapel Hill
Department:McAllister heart Institute, Department of Internal medicine
Laboratory:Multiple Centers
Last Name:Willis
First Name:Monte
Address:111 Mason Farm road, Chapel Hill, North Carolina, 27599-7126, USA
Email:monte_willis@med.unc.edu
Phone:(984) 999-5431
Funding Source:NIH, Fondation Leducq

Subject:

Subject ID:SU001168
Subject Type:Animal
Subject Species:Canis lupus familiaris
Taxonomy ID:9615

Factors:

Subject type: Animal; Subject species: Canis lupus familiaris (Factor headings shown in green)

mb_sample_id local_sample_id Gentotype
SA076743K 8Control BF
SA076744K 9Control BF
SA076745K 7Control BF
SA076746K 10Control BF
SA076747K 19Control LDE
SA076748K 20Control LDE
SA076749K 18Control LDE
SA076750K17Control LDE
SA076751K 1GRMD BF
SA076752K 2GRMD BF
SA076753K 3GRMD BF
SA076754K 4GRMD BF
SA076755K 6GRMD BF
SA076756K 5GRMD BF
SA076757K 15GRMD LDE
SA076758K 11GRMD LDE
SA076759K 12GRMD LDE
SA076760K 13GRMD LDE
SA076761K 14GRMD LDE
SA076762K 16GRMD LDE
Showing results 1 to 20 of 20

Collection:

Collection ID:CO001162
Collection Summary:Skeletal muscle (BF, LDE) was surgically biosied and flash frozen in liquid nitrogen.
Sample Type:Muscle

Treatment:

Treatment ID:TR001182
Treatment Summary:Fraction of cardiac tissue weighed (25-50 mg wet weight), then the finely cut up tissue quickly added to fresh pre-made buffer (50 % acetyl-nitrile, 50% water, 0.3% formic acid) at a standard concentration of 25 mg/475 mcl buffer then fully homogenized on ice for 20-25 s and placed on dry ice/stored at - 80C

Sample Preparation:

Sampleprep ID:SP001175
Sampleprep Summary:The samples were crash deprotonized by methanol precipitation and spiked with D27-deuterated myristic acid (D27-C14:0) as an internal standard for retention-time locking and dried. The trimethylsilyl-D27-C14:0 standard retention time (RT) was set at 16.727 min. Reactive carbonyls were stabilized at 50C with methoxyamine hydrochloride in dry pyridine. Metabolites were made volatile with TMS groups using N-methyl-N-(trimethylsilyl) trifluoroacetamide or MSTFA with catalytic trimethylchlorosilane at 50C.

Combined analysis:

Analysis ID AN001820
Analysis type MS
Chromatography type GC
Chromatography system Agilent 6890N
Column Agilent DB5-MS (15m x 0.25mm,0.25um)
MS Type EI
MS instrument type Single quadrupole
MS instrument name Agilent 5975
Ion Mode POSITIVE
Units Peak values (Log transformed)

Chromatography:

Chromatography ID:CH001289
Chromatography Summary:GC/MS methods follow previous studies using a 6890 N GC connected to a 5975 Inert single quadrupole MS (Agilent Technologies, Santa Clara, CA) (Bonikos et al. 1975; Fiehn 2008; Kind et al. 2009). The two wall-coated, open-tubular GC columns connected in series are both from J&W/Agilent (part 122-5512), DB5-MS, 15 meters in length, 0.25 mm in diameter, with an 0.25-l m luminal film. Positive ions generated with conventional electron-ionization at 70 eV are scanned broadly from 600 to 50 m/z in the detector throughout the 45 min cycle time.
Instrument Name:Agilent 6890N
Column Name:Agilent DB5-MS (15m x 0.25mm,0.25um)
Chromatography Type:GC

MS:

MS ID:MS001683
Analysis ID:AN001820
Instrument Name:Agilent 5975
Instrument Type:Single quadrupole
MS Type:EI
MS Comments:none
Ion Mode:POSITIVE
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