Summary of Study ST000431
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 PR000334. The data can be accessed directly via it's Project DOI: 10.21228/M8D02C This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST000431 |
| Study Title | C2C12 stretch cessation models muscle atrophy and anaplerotic changes in metabolism |
| Study Type | GC-MS non-targeted metabolomic profiling |
| Study Summary | Studies of skeletal muscle disuse either in patients on bed rest or experimentally in animals(immobilization) have demonstrated that decreased protein synthesis is common, with transient parallel increases in protein degradation. Muscle disuse atrophy involves a process of transition from slow to fast myosin fiber types 6 . A shift toward glycolysis, decreased capacity for fat oxidation, and substrate accumulation in atrophied muscles have been reported as has accommodation of the liver with an increased gluconeogenic capacity. Recent studies have modeled skeletal muscle disuse by using cyclic stretch of differentiated myotubes (C2C12), which mimics the loading pattern of mature skeletal muscle, followed by cessation of stretch.We utilized this model to determine the metabolic changes using non-targeted metabolomics analysis of the media. We identified increases in amino acids resulting from protein degradation (largely sarcomere) that occurs with muscle atrophy that are involved in feeding the Kreb’s cycle through anaplerosis. Specifically, we identified increased alanine/proline metabolism (significantly elevated proline, alanine, glutamine, and asparagine) and increased -ketoglutaric acid, the proposed Kreb’s cycle intermediate being fed by the alanine/proline metabolic anaplerotic mechanism. Additionally, several unique pathways not clearly delineated in previous studies of muscle unloading were seen, including: 1) elevated ethanolamine and elevated keto-acids (e.g. 2-ketoleucine and 2-keovaline) represent intermediates in the Ehlrich amino acid degradation pathway, which feeds into a metabolic pathway supplying acetyl-CoA and 2-hydroxybutyrate (also significantly increased); and 2) elevated guanine, an intermediate of purine metabolism, was seen at 12 hours unloading. Given the interest in targeting different aspects of the ubiquitin proteasome system to inhibit protein degradation, this C2C12 system may allow the identification of direct and indirect alterations in metabolism due to anaplerosis or through other yet to be identified mechanisms using a non-targeted metabolomics approach. |
| Institute | University of North Carolina at Chapel Hill |
| Department | McAllister Heart Institute |
| Laboratory | Mutliple Centers |
| Last Name | Ilaiwy;WIllis |
| First Name | Amro;Monte |
| Address | 111 Mason Farm road, Chapel Hill, North Carolina, 27599-7126, USA |
| amroilaiwy@gmail.com, monte_willis@med.unc.edu | |
| Phone | 919-3607599 |
| Submit Date | 2016-05-28 |
| Analysis Type Detail | GC-MS |
| Release Date | 2016-09-23 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR000334 |
| Project DOI: | doi: 10.21228/M8D02C |
| Project Title: | C2C12 stretch cessation models muscle atrophy and anaplerotic changes in metabolism |
| Project Type: | GC-MS non targeted qualitative analysis |
| Project Summary: | Studies of skeletal muscle disuse either in patients on bed rest or experimentally in animals(immobilization) have demonstrated that decreased protein synthesis is common, with transient parallel increases in protein degradation. Muscle disuse atrophy involves a process of transition from slow to fast myosin fiber types 6 . A shift toward glycolysis, decreased capacity for fat oxidation, and substrate accumulation in atrophied muscles have been reported as has accommodation of the liver with an increased gluconeogenic capacity. Recent studies have modeled skeletal muscle disuse by using cyclic stretch of differentiated myotubes (C2C12), which mimics the loading pattern of mature skeletal muscle, followed by cessation of stretch.We utilized this model to determine the metabolic changes using non-targeted metabolomics analysis of the media. We identified increases in amino acids resulting from protein degradation (largely sarcomere) that occurs with muscle atrophy that are involved in feeding the Kreb’s cycle through anaplerosis. Specifically, we identified increased alanine/proline metabolism (significantly elevated proline, alanine, glutamine, and asparagine) and increased -ketoglutaric acid, the proposed Kreb’s cycle intermediate being fed by the alanine/proline metabolic anaplerotic mechanism. Additionally, several unique pathways not clearly delineated in previous studies of muscle unloading were seen, including: 1) elevated ethanolamine and elevated keto-acids (e.g. 2-ketoleucine and 2-keovaline) represent intermediates in the Ehlrich amino acid degradation pathway, which feeds into a metabolic pathway supplying acetyl-CoA and 2-hydroxybutyrate (also significantly increased); and 2) elevated guanine, an intermediate of purine metabolism, was seen at 12 hours unloading. Given the interest in targeting different aspects of the ubiquitin proteasome system to inhibit protein degradation, this C2C12 system may allow the identification of direct and indirect alterations in metabolism due to anaplerosis or through other yet to be identified mechanisms using a non-targeted metabolomics approach. |
| Institute: | University of North Carolina at Chapel Hill |
| Department: | McAllister heart Institute, Department of Internal medicine |
| Laboratory: | Multiple Centers |
| Last Name: | Ilaiwy; Willis |
| First Name: | Amro; Monte |
| Address: | 111 Mason Farm road, Chapel Hill, North Carolina, 27599-7126, USA |
| Email: | amroilaiwy@gmail.com, monte_willis@med.unc.edu |
| Phone: | 210-596-0171 |
| Funding Source: | NIH, Fondation Leducq |
Subject:
| Subject ID: | SU000452 |
| Subject Type: | Animal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Species Group: | Mammal |
Factors:
Subject type: Animal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Group |
|---|---|---|
| SA021671 | p12 #2 | 12 hour reversal |
| SA021672 | p12 #1 | 12 hour reversal |
| SA021673 | p12 #3 | 12 hour reversal |
| SA021668 | p1 #3 | 1 hour reversal |
| SA021669 | p1 #1 | 1 hour reversal |
| SA021670 | p1 #2 | 1 hour reversal |
| SA021674 | p3 #3 | 3 hour reversal |
| SA021675 | p3 #1 | 3 hour reversal |
| SA021676 | p3 #2 | 3 hour reversal |
| SA021677 | p6 #3 | 6 hour reversal |
| SA021678 | p6 #2 | 6 hour reversal |
| SA021679 | p6 #1 | 6 hour reversal |
| SA021680 | p0 #1 | 6 hour stretch |
| SA021681 | p0 #2 | 6 hour stretch |
| SA021682 | p0 #3 | 6 hour stretch |
| SA021683 | p6 no stretch #1 | no stretch-serum free |
| SA021684 | p1 no stretch #1 | no stretch-serum free |
| SA021685 | p3 no stretch #1 | no stretch-serum free |
| SA021686 | p0 no stretch #1 | no stretch-serum free |
| SA021687 | serum 6hr no stretch #3 | serum-no stretch |
| SA021688 | serum 6hr no stretch #2 | serum-no stretch |
| SA021689 | serum 6hr no stretch #1 | serum-no stretch |
| Showing results 1 to 22 of 22 |
Collection:
| Collection ID: | CO000446 |
| Collection Summary: | Media was collected from all different 6 wells, and stored in -80 C |
| Sample Type: | Muscle |
Treatment:
| Treatment ID: | TR000466 |
| Treatment Summary: | N/A |
Sample Preparation:
| Sampleprep ID: | SP000459 |
| Sampleprep Summary: | N/A |
Combined analysis:
| Analysis ID | AN000681 |
|---|---|
| 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: | CH000493 |
| 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: | MS000607 |
| Analysis ID: | AN000681 |
| Instrument Name: | Agilent 5975 |
| Instrument Type: | Single quadrupole |
| MS Type: | EI |
| Ion Mode: | POSITIVE |
