Summary of Study ST000165

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 PR000143. The data can be accessed directly via it's Project DOI: 10.21228/M8GP4N This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

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Download additional data:  the amino acids were measured under negative ion chemical ionisation conditions using isobutane as reactant gas
Study IDST000165
Study TitleSparing of muscle mass and function by passive loading in an experimental intensive care unit model
Study Typetime course + intervention
Study SummaryA unique experimental rat ICU model has been used allowing long-term (weeks) time-resolved analyses of the effects of standardized unilateral passive mechanical loading on skeletal muscle size and function and underlying mechanisms. Results show that passive mechanical loading alleviated the muscle wasting and the loss of force-generation associated with the ICU intervention, resulting in a doubling of the functional capacity of the loaded versus the unloaded muscles after a 2-week ICU intervention.
Institute
Uppsala University
DepartmentDepartment of Neuroscience
Last NameLarsson
First NameLars
EmailLars.larsson@neuro.uu.se
Submit Date2015-05-14
Num Groups2
Total Subjects13
Raw Data AvailableNo
Analysis Type DetailIR-MS
Release Date2015-05-10
Release Version1
Lars Larsson Lars Larsson
https://dx.doi.org/10.21228/M8GP4N
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000143
Project DOI:doi: 10.21228/M8GP4N
Project Title:Sparing of muscle mass and function by passive loading in an experimental intensive care unit model
Project Summary:The response to mechanical stimuli, i.e. tensegrity, plays an important role in regulating cell physiological and pathophysiological function, and the mechanical silencing observed in intensive care unit (ICU) patients leads to a severe and specific muscle wasting condition. This study aims to unravel the underlying mechanisms and the effects of passive mechanical loading on skeletal muscle mass and function at the gene, protein and cellular levels. A unique experimental rat ICU model has been used allowing long-term (weeks) time-resolved analyses of the effects of standardized unilateral passive mechanical loading on skeletal muscle size and function and underlying mechanisms. Results show that passive mechanical loading alleviated the muscle wasting and the loss of force-generation associated with the ICU intervention, resulting in a doubling of the functional capacity of the loaded versus the unloaded muscles after a 2-week ICU intervention. We demonstrate that the improved maintenance of muscle mass and function is probably a consequence of a reduced oxidative stress revealed by lower levels of carbonylated proteins, and a reduced loss of the molecular motor protein myosin. A complex temporal gene expression pattern, delineated by microarray analysis, was observed with loading-induced changes in transcript levels of sarcomeric proteins, muscle developmental processes, stress response, extracellular matrix/cell adhesion proteins and metabolism. Thus, the results from this study show that passive mechanical loading alleviates the severe negative consequences on muscle size and function associated with the mechanical silencing in ICU patients, strongly supporting early and intense physical therapy in immobilized ICU patients.
Institute:Uppsala University
Department:Department of Neuroscience
Last Name:Larsson
First Name:Lars
Email:Lars.larsson@neuro.uu.se

Subject:

Subject ID:SU000184
Subject Type:Animal
Subject Species:Rattus norvegicus
Taxonomy ID:10116
Genotype Strain:Sprague–Dawley
Gender:female
Species Group:Mammal

Factors:

Subject type: Animal; Subject species: Rattus norvegicus (Factor headings shown in green)

mb_sample_id local_sample_id Mechanical loading hours/day Group
SA0089384412 Passive Mechanical Loading
SA0089394612 Passive Mechanical Loading
SA0089404212 Passive Mechanical Loading
SA0089414012 Passive Mechanical Loading
SA0089423612 Passive Mechanical Loading
SA0089433812 Passive Mechanical Loading
SA0089444812 Passive Mechanical Loading
SA0089455012 Passive Mechanical Loading
SA0089467012 Passive Mechanical Loading
SA0089477212 Passive Mechanical Loading
SA0089486812 Passive Mechanical Loading
SA0089496612 Passive Mechanical Loading
SA0089505412 Passive Mechanical Loading
SA0089515612 Passive Mechanical Loading
SA0089523412 Passive Mechanical Loading
SA0089535212 Passive Mechanical Loading
SA00895439None Passive Mechanical Loading
SA00895549None Passive Mechanical Loading
SA00895641None Passive Mechanical Loading
SA00895751None Passive Mechanical Loading
SA00895855None Passive Mechanical Loading
SA00895953None Passive Mechanical Loading
SA00896037None Passive Mechanical Loading
SA00896135None Passive Mechanical Loading
SA00896243None Passive Mechanical Loading
SA00896369None Passive Mechanical Loading
SA00896467None Passive Mechanical Loading
SA00896565None Passive Mechanical Loading
SA00896645None Passive Mechanical Loading
SA00896747None Passive Mechanical Loading
SA00896871None Passive Mechanical Loading
SA00896933None Passive Mechanical Loading
SA00897082None Sham
SA00897181None Sham
SA00897279None Sham
SA00897374None Sham
SA00897473None Sham
SA0089752None Sham
SA0089763None Sham
SA00897775None Sham
SA00897876None Sham
SA00897983None Sham
SA00898078None Sham
SA00898177None Sham
SA00898280None Sham
SA00898385None Sham
SA00898499None Sham
SA00898598None Sham
SA00898697None Sham
SA00898796None Sham
SA008988100None Sham
SA008989101None Sham
SA008990104None Sham
SA008991103None Sham
SA008992102None Sham
SA00899395None Sham
SA00899494None Sham
SA00899588None Sham
SA00899687None Sham
SA00899786None Sham
SA0089984None Sham
SA00899989None Sham
SA00900090None Sham
SA00900193None Sham
SA00900292None Sham
SA00900391None Sham
SA00900484None Sham
SA00900560None Sham
SA00900629None Sham
SA00900728None Sham
SA00900827None Sham
SA00900930None Sham
SA00901031None Sham
SA00901115None Sham
SA00901216None Sham
SA00901332None Sham
SA00901426None Sham
SA00901525None Sham
SA00901620None Sham
SA00901719None Sham
SA00901818None Sham
SA00901921None Sham
SA00902022None Sham
SA00902124None Sham
SA00902223None Sham
SA00902314None Sham
SA00902413None Sham
SA00902517None Sham
SA00902659None Sham
SA00902758None Sham
SA00902861None Sham
SA00902962None Sham
SA00903064None Sham
SA00903163None Sham
SA00903257None Sham
SA0090336None Sham
SA00903410None Sham
SA00903511None Sham
SA00903612None Sham
SA0090379None Sham
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Collection:

Collection ID:CO000170
Collection Summary:The tibialis anterior (TA), extensor digitorum longus (EDL), plantaris, gastrocnemius and soleus muscles were dissected from the loaded left leg and the unloaded right leg immediately after death. One half of the soleus and EDL muscles together with TA and gastrocnemius were quickly frozen in liquid propane cooled by liquid nitrogen, and stored at ?160°C for further analyses. In the other halves of the soleus and EDL muscles, bundles of approximately 50 fibres were dissected from the muscles in relaxing solution at 4°C and tied to glass capillaries, stretched to about 110% of their resting slack length. The bundles were chemically skinned for 24 h in relaxing solution containing 50% (v/v) glycerol for 24 h at 4°C and were subsequently stored at ?20°C (Larsson & Moss, 1993). All the bundles were cryo-protected within 1 week after skinning by transferring the bundles every 30 min to relax solution containing increasing concentrations of sucrose, i.e. 0, 0.5, 1.0, 1.5 and 2.0 m, and subsequently frozen in liquid propane chilled with liquid nitrogen (Frontera & Larsson, 1997). The frozen bundles were stored at ?160°C pending use. One day before the experiments, a bundle was transferred to a 2.0 m sucrose solution for 30 min, subsequently incubated in solutions of decreasing sucrose concentration (1.5–0.5 m) and finally kept in a skinning solution at ?20°C.
Sample Type:Muscle

Treatment:

Treatment ID:TR000190
Treatment Summary:Mechanical stimulation|Sham operation
Treatment Protocol Comments:Sham operated controls and 46 anaesthetized and mechanically ventilated female Sprague–Dawley rats treated with ?-cobratoxin for durations varying from 6 h to 14 days were included in this study. The experimental model has previously been described in detail (Dworkin & Dworkin, 1990, 2004). For mechanical stimulation animals, left leg was mechanically stimulated and right leg was not. |Sham operated controls and 46 anaesthetized and mechanically ventilated female Sprague–Dawley rats treated with ?-cobratoxin for durations varying from 6 h to 14 days were included in this study. The experimental model has previously been described in detail (Dworkin & Dworkin, 1990, 2004). For sham operated controls, no stimulation was performed on either legs

Sample Preparation:

Sampleprep ID:SP000184
Sampleprep Summary:An i.v. bolus dose of [ring-13C6]phenylalanine (15 ?g g?1 body weight) was given 15 min prior to the animals being killed. Immediately after death, the gastrocnemius muscle was removed from the left and right hindlimb and split into a medial and lateral deep red and superficial white portion and frozen in liquid propane chilled by liquid nitrogen. Tissue fluid and mixed gastrocnemius muscle proteins were isolated according to the method of Ljungqvist et al. (1997). The mixed muscle protein precipitate from the isolation was hydrolysed by heating with 6 m HCl overnight at 110°C. Both tissue fluid and hydrolysed mixed gastrocnemius muscle amino acids were purified using a BioRad AG-50 × 8 ion exchange resin prior to mass spectrometry analysis.
Tissue fluid
The level of enrichment of [ring-13C6]phenylalanine in tissue fluid was analysed using ThermoFisher Quantum gas chromatography tandem mass spectrometry (GC/MS/MS) (San Jose, CA, USA). The heptafluorobutyryl isobutyl ester derivative was prepared as described by Ford et al. (1985) and the amino acids were measured under negative ion chemical ionisation conditions using isobutane as reactant gas. The [M-HF]? fragments reflecting the m0 and m+6 species were monitored (m/z transitions 397?377 and 403?383, respectively) and the enrichment of the label was measured against a calibration curve prepared from known amounts of labelled and unlabelled phenylalanine (range 0–30%).

Combined analysis:

Analysis ID AN000258
Analysis type MS
Chromatography type GC
Chromatography system
Column
MS Type IR MS
MS instrument type IR MS
MS instrument name Thermo DeltaPlus Isotope Ratio MS
Ion Mode NEGATIVE
Units

Chromatography:

Chromatography ID:CH000182
Chromatography Summary:The level of enrichment of [ring-13C6]phenylalanine derived from hydrolysed mixed muscle proteins were analysed using a ThermoFisher DeltaPlus Isotope Ratio mass spectrometer (IR/MS) (Bremen, Germany) fitted with an on-line gas chromatograph with oxidation and reduction furnaces as previously described (Balagopal et al. 1996). The amino acids were derivatized to their trimethyl acetyl, methyl esters according to the method of Metges et al. (1996). Any amino acid eluting from the gas chromatograph is converted to CO2 and N2 prior to entry into the IR/MS. The amino acids were derivatized to their trimethyl acetyl, methyl esters according to the method of Metges et al. (1996). Enrichment of the tracer was measured by monitoring the ratio of 13C to 12CO2 in the IR/MS and again referenced to a calibration curve (00.1%).
Chromatography Type:GC

MS:

MS ID:MS000208
Analysis ID:AN000258
Instrument Name:Thermo DeltaPlus Isotope Ratio MS
Instrument Type:IR MS
MS Type:IR MS
MS Comments:the amino acids were measured under negative ion chemical ionisation conditions using isobutane as reactant gas
Ion Mode:NEGATIVE
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