Summary of Study ST000393
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 PR000307. The data can be accessed directly via it's Project DOI: 10.21228/M8Z01Z This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
Study ID | ST000393 |
Study Title | Long-Chain Fatty Acid Combustion Rate Is Associated with Unique Metabolite Profiles in Skeletal Muscle Mitochondria |
Study Summary | Incomplete or limited long-chain fatty acid (LCFA) combustion in skeletal muscle has been associated with insulin resistance. Signals that are responsive to shifts in LCFA β-oxidation rate or degree of intramitochondrial catabolism are hypothesized to regulate second messenger systems downstream of the insulin receptor. Recent evidence supports a causal link between mitochondrial LCFA combustion in skeletal muscle and insulin resistance. We have used unbiased metabolite profiling of mouse muscle mitochondria with the aim of identifying candidate metabolites within or effluxed from mitochondria and that are shifted with LCFA combustion rate. This proof-of-principle study establishes that large-scale metabolomics methods can be applied to organelle-level models to discover metabolite patterns reflective of LCFA combustion, which may lead to identification of molecules linking muscle fat metabolism and insulin signaling. Our results suggest that future studies should focus on the fate of effluxed TCA cycle intermediates and on mechanisms ensuring their replenishment during LCFA metabolism in skeletal muscle. |
Institute | University of California, Davis |
Department | Genome and Biomedical Sciences Facility |
Laboratory | WCMC Metabolomics Core |
Last Name | Fiehn |
First Name | Oliver |
Address | 1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, CA 95616 |
ofiehn@ucdavis.edu | |
Phone | (530) 754-8258 |
Submit Date | 2016-05-06 |
Publications | http://dx.doi.org/10.1371/journal.pone.0009834 |
Raw Data Available | Yes |
Raw Data File Type(s) | peg |
Analysis Type Detail | GC-MS |
Release Date | 2016-06-18 |
Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
Project ID: | PR000307 |
Project DOI: | doi: 10.21228/M8Z01Z |
Project Title: | Long-Chain Fatty Acid Combustion Rate Is Associated with Unique Metabolite Profiles in Skeletal Muscle Mitochondria |
Project Summary: | Incomplete or limited long-chain fatty acid (LCFA) combustion in skeletal muscle has been associated with insulin resistance. Signals that are responsive to shifts in LCFA β-oxidation rate or degree of intramitochondrial catabolism are hypothesized to regulate second messenger systems downstream of the insulin receptor. Recent evidence supports a causal link between mitochondrial LCFA combustion in skeletal muscle and insulin resistance. We have used unbiased metabolite profiling of mouse muscle mitochondria with the aim of identifying candidate metabolites within or effluxed from mitochondria and that are shifted with LCFA combustion rate. This proof-of-principle study establishes that large-scale metabolomics methods can be applied to organelle-level models to discover metabolite patterns reflective of LCFA combustion, which may lead to identification of molecules linking muscle fat metabolism and insulin signaling. Our results suggest that future studies should focus on the fate of effluxed TCA cycle intermediates and on mechanisms ensuring their replenishment during LCFA metabolism in skeletal muscle. |
Institute: | University of California, Davis |
Department: | Genome and Biomedical Sciences Facility |
Laboratory: | WCMC Metabolomics Core |
Last Name: | Fiehn |
First Name: | Oliver |
Address: | 1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, CA 95616 |
Email: | ofiehn@ucdavis.edu |
Phone: | (530) 754-8258 |
Funding Source: | NIH U24DK097154 |
Publications: | http://dx.doi.org/10.1371/journal.pone.0009834 |
Subject:
Subject ID: | SU000414 |
Subject Type: | Animal |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Genotype Strain: | C57Bl/6J |
Age Or Age Range: | 4 months |
Gender: | Female |
Human Smoking Status: | Former/Current |
Animal Animal Supplier: | Jackson Laboratories |
Animal Housing: | housed in a facility at 23°C |
Animal Light Cycle: | 12∶12 hr light cycle, lights on 0700 |
Animal Feed: | with free access to chow (4.5% fat/weight), for at least 1 week before being studied |
Species Group: | Mammals |
Factors:
Subject type: Animal; Subject species: Mus musculus (Factor headings shown in green)
mb_sample_id | local_sample_id | Organ | Time | Dose |
---|---|---|---|---|
SA018676 | 061215byusa1091_1 | cytosol | 0 min | palmitic acid 2.4 uM |
SA018677 | 061215byusa1127_1 | cytosol | 0 min | palmitic acid 2.4 uM |
SA018678 | 061215byusa1092_1 | cytosol | 0 min | palmitic acid 2.4 uM |
SA018679 | 061215byusa1119_1 | cytosol | 0 min | palmitic acid 2.4 uM |
SA018680 | 061215byusa1096_1 | cytosol | 0 min | palmitic acid 2.4 uM |
SA018681 | 061215byusa1088_1 | cytosol | 0 min | palmitic acid 2.4 uM |
SA018682 | 061215byusa1134_1 | cytosol | 0 min | palmitic acid 9.6 uM |
SA018683 | 061215byusa1090_1 | cytosol | 0 min | palmitic acid 9.6 uM |
SA018684 | 061215byusa1086_1 | cytosol | 0 min | palmitic acid 9.6 uM |
SA018685 | 061215byusa1103_1 | cytosol | 0 min | palmitic acid 9.6 uM |
SA018686 | 061215byusa1079_1 | cytosol | 0 min | palmitic acid 9.6 uM |
SA018687 | 061215byusa1065_1 | cytosol | 0 min | palmitic acid 9.6 uM |
SA018688 | 061215byusa1112_1 | cytosol | 0 min | palmitoyl carnitine 9.6 uM |
SA018689 | 061215byusa1075_1 | cytosol | 0 min | palmitoyl carnitine 9.6 uM |
SA018690 | 061215byusa1124_1 | cytosol | 0 min | palmitoyl carnitine 9.6 uM |
SA018691 | 061215byusa1087_1 | cytosol | 0 min | palmitoyl carnitine 9.6 uM |
SA018692 | 061215byusa1077_1 | cytosol | 0 min | palmitoyl carnitine 9.6 uM |
SA018693 | 061215byusa1114_1 | cytosol | 0 min | palmitoyl carnitine 9.6 uM |
SA018694 | 061215byusa1128_1 | cytosol | 20 min | palmitic acid 2.4 uM |
SA018695 | 061215byusa1074_1 | cytosol | 20 min | palmitic acid 2.4 uM |
SA018696 | 061215byusa1105_1 | cytosol | 20 min | palmitic acid 2.4 uM |
SA018697 | 061215byusa1129_1 | cytosol | 20 min | palmitic acid 2.4 uM |
SA018698 | 061215byusa1116_1 | cytosol | 20 min | palmitic acid 2.4 uM |
SA018699 | 061215byusa1123_1 | cytosol | 20 min | palmitic acid 2.4 uM |
SA018700 | 061215byusa1100_1 | cytosol | 20 min | palmitic acid 9.6 uM |
SA018701 | 061215byusa1080_1 | cytosol | 20 min | palmitic acid 9.6 uM |
SA018702 | 061215byusa1085_1 | cytosol | 20 min | palmitic acid 9.6 uM |
SA018703 | 061215byusa1076_1 | cytosol | 20 min | palmitic acid 9.6 uM |
SA018704 | 061215byusa1078_1 | cytosol | 20 min | palmitic acid 9.6 uM |
SA018705 | 061215byusa1093_1 | cytosol | 20 min | palmitic acid 9.6 uM |
SA018706 | 061215byusa1133_1 | cytosol | 20 min | palmitoyl carnitine 9.6 uM |
SA018707 | 061215byusa1109_1 | cytosol | 20 min | palmitoyl carnitine 9.6 uM |
SA018708 | 061215byusa1084_1 | cytosol | 20 min | palmitoyl carnitine 9.6 uM |
SA018709 | 061215byusa1068_1 | cytosol | 20 min | palmitoyl carnitine 9.6 uM |
SA018710 | 061215byusa1106_1 | cytosol | 20 min | palmitoyl carnitine 9.6 uM |
SA018711 | 061215byusa1125_1 | cytosol | 20 min | palmitoyl carnitine 9.6 uM |
SA018712 | 061215byusa1107_1 | mitochondria | 0 min | palmitic acid 2.4 uM |
SA018713 | 061215byusa1072_1 | mitochondria | 0 min | palmitic acid 2.4 uM |
SA018714 | 061215byusa1066_1 | mitochondria | 0 min | palmitic acid 2.4 uM |
SA018715 | 061215byusa1089_1 | mitochondria | 0 min | palmitic acid 2.4 uM |
SA018716 | 061215byusa1083_1 | mitochondria | 0 min | palmitic acid 2.4 uM |
SA018717 | 061215byusa1081_1 | mitochondria | 0 min | palmitic acid 2.4 uM |
SA018718 | 061215byusa1113_1 | mitochondria | 0 min | palmitic acid 9.6 uM |
SA018719 | 061215byusa1095_1 | mitochondria | 0 min | palmitic acid 9.6 uM |
SA018720 | 061215byusa1115_1 | mitochondria | 0 min | palmitic acid 9.6 uM |
SA018721 | 061215byusa1099_1 | mitochondria | 0 min | palmitic acid 9.6 uM |
SA018722 | 061215byusa1094_1 | mitochondria | 0 min | palmitic acid 9.6 uM |
SA018723 | 061215byusa1122_1 | mitochondria | 0 min | palmitoyl carnitine 9.6 uM |
SA018724 | 061215byusa1136_1 | mitochondria | 0 min | palmitoyl carnitine 9.6 uM |
SA018725 | 061215byusa1098_1 | mitochondria | 0 min | palmitoyl carnitine 9.6 uM |
SA018726 | 061215byusa1111_1 | mitochondria | 0 min | palmitoyl carnitine 9.6 uM |
SA018727 | 061215byusa1110_1 | mitochondria | 0 min | palmitoyl carnitine 9.6 uM |
SA018728 | 061215byusa1101_1 | mitochondria | 0 min | palmitoyl carnitine 9.6 uM |
SA018729 | 061215byusa1097_1 | mitochondria | 20 min | palmitic acid 2.4 uM |
SA018730 | 061215byusa1104_1 | mitochondria | 20 min | palmitic acid 2.4 uM |
SA018731 | 061215byusa1067_1 | mitochondria | 20 min | palmitic acid 2.4 uM |
SA018732 | 061215byusa1130_1 | mitochondria | 20 min | palmitic acid 2.4 uM |
SA018733 | 061215byusa1120_1 | mitochondria | 20 min | palmitic acid 2.4 uM |
SA018734 | 061215byusa1131_1 | mitochondria | 20 min | palmitic acid 9.6 uM |
SA018735 | 061215byusa1071_1 | mitochondria | 20 min | palmitic acid 9.6 uM |
SA018736 | 061215byusa1082_1 | mitochondria | 20 min | palmitic acid 9.6 uM |
SA018737 | 061215byusa1102_1 | mitochondria | 20 min | palmitic acid 9.6 uM |
SA018738 | 061215byusa1135_1 | mitochondria | 20 min | palmitic acid 9.6 uM |
SA018739 | 061215byusa1118_1 | mitochondria | 20 min | palmitoyl carnitine 9.6 uM |
SA018740 | 061215byusa1069_1 | mitochondria | 20 min | palmitoyl carnitine 9.6 uM |
SA018741 | 061215byusa1132_1 | mitochondria | 20 min | palmitoyl carnitine 9.6 uM |
SA018742 | 061215byusa1070_1 | mitochondria | 20 min | palmitoyl carnitine 9.6 uM |
SA018743 | 061215byusa1121_1 | mitochondria | 20 min | palmitoyl carnitine 9.6 uM |
Showing results 1 to 68 of 68 |
Collection:
Collection ID: | CO000408 |
Collection Summary: | Skeletal muscle mitochondria were isolated essentially according to Chappell and Perry [Chappell JB, Perry SV (1954) Biochemical and osmotic properties of skeletal muscle mitochondria. Nature 173: 1094–1095.]. All media were ice-cold, and procedures done on ice or at 4°C. Briefly, pectoral, forelimb and hindlimb muscles were rapidly dissected and placed in basic medium [BM (mM): KCl (140), HEPES (20), MgCl2 (5), EGTA (2); pH 7.0]. Together, these muscle groups comprise a mixed population of mainly type II oxidative and glycolytic fibers. Muscle was cleaned of connective tissue and fat, minced and placed in 15 vol of homogenizing medium (HM: BM with 1 mM ATP and 1% BSA (w/v)) containing one unit of protease (Subtilisin A) per g muscle wet weight. |
Collection Protocol Filename: | Long-Chain_Fatty_Acid_Metabolite_Profiles_in_Skeletal_Muscle_Mitochondria.PDF |
Sample Type: | Mitochondria |
Treatment:
Treatment ID: | TR000428 |
Treatment Summary: | Mitochondria (0.6 mg/ml) were supplied with three concentrations of palmitate corresponding to rates of β-oxidation: 1. low (2 µM) 2. medium (9 µM) 3. high (19 µM) Three ml aliquots of incubation medium [IM, (mM): KCl (120), HEPES (5), KH2PO4 (5), MgCl2 (5) and EGTA (1); pH 7.4] were supplemented with (mM) ATP (1), malate (0.05), coenzyme A (0.025), and carnitine (0.5) and added to 20-ml glass reaction vials. Solutions of low, medium and high palmitate concentrations were added to vials in a 6∶1 FA:BSA complex. Two additional incubations were performed as controls: 1. 0 µM palmitate 2. 9 µM palmitate + inhibitors |
Treatment Protocol Filename: | Long-Chain_Fatty_Acid_Metabolite_Profiles_in_Skeletal_Muscle_Mitochondria.PDF |
Treatment Protocol Comments: | The first control condition evaluated the metabolic profile of mitochondria oxidizing only malate, and included ATP, carnitine and CoA and ethanol (0.5%). The second control condition assessed effects of FA in the absence of complete oxidative catabolism, and consisted of malate, 9 µM palmitate, ATP, carnitine and CoA, and supplemented with the TCA cycle inhibitor malonate (10 mM) and the electron transport chain complex I inhibitor rotenone (5 µM) |
Sample Preparation:
Sampleprep ID: | SP000421 |
Sampleprep Summary: | Tissue was homogenized using a glass/Teflon Potter-Elvehjem tissue grinder (240 rpm) and fractionated by centrifugation at 800 g (10 min), and the supernatant collected and spun at 12000 g (9 min). The pellet was resuspended in 20 ml BM and incubated on ice for 5 min (myofibrillar repolymerization). Samples were spun at 800 g (8 min) to pellet actin-myosin polymers. The supernatant was then spun at 12000 g (9 min). The final pellet was resuspended in 220 µl of BM. |
Sampleprep Protocol Filename: | Long-Chain_Fatty_Acid_Metabolite_Profiles_in_Skeletal_Muscle_Mitochondria.PDF |
Sampleprep Protocol Comments: | This isolation procedure yields mitochondria with high respiratory control ratios (state 3/state 4; ∼8–10 when supplied with 10 mM pyruvate/5 mM malate), and which are capable of activating palmitate [29], a process dependent on the integrity of enzymes on the mitochondrial outer membrane. Protein concentration was determined by a modified Lowry method with BSA as standard. |
Processing Method: | Homogenized |
Combined analysis:
Analysis ID | AN000629 |
---|---|
Analysis type | MS |
Chromatography type | GC |
Chromatography system | Leco Pegasus III GC |
Column | Restek Corporation Rtx-5Sil MS |
MS Type | EI |
MS instrument type | GC-TOF |
MS instrument name | Leco Pegasus III GC TOF |
Ion Mode | POSITIVE |
Units | counts |
Chromatography:
Chromatography ID: | CH000454 |
Instrument Name: | Leco Pegasus III GC |
Column Name: | Restek Corporation Rtx-5Sil MS |
Column Pressure: | 7.7 PSI |
Column Temperature: | 50-330C |
Flow Rate: | 1 ml/min |
Injection Temperature: | 50 C ramped to 250 C by 12 C/s |
Sample Injection: | 0.5 uL |
Transferline Temperature: | 230C |
Washing Buffer: | Ethyl Acetate |
Sample Loop Size: | 30 m length x 0.25 mm internal diameter |
Randomization Order: | Excel generated |
Chromatography Type: | GC |
MS:
MS ID: | MS000562 |
Analysis ID: | AN000629 |
Instrument Name: | Leco Pegasus III GC TOF |
Instrument Type: | GC-TOF |
MS Type: | EI |
Ion Mode: | POSITIVE |
Ion Source Temperature: | 250 C |
Ionization: | Pos |
Ionization Energy: | 70 eV |
Mass Accuracy: | Nominal |
Source Temperature: | 250 C |
Scan Range Moverz: | 85-500 Da |
Scanning Cycle: | 17 Hz |
Scanning Range: | 85-500 Da |
Skimmer Voltage: | 1850 V |