Summary of study ST000379

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 PR000298. The data can be accessed directly via it's Project DOI: 10.21228/M8HW28 This work is supported by NIH grant, U2C- DK119886.

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Study IDST000379
Study TitleTemporal metabolomic responses of cultured HepG2 liver cells to high fructose and high glucose exposures
Study SummaryHigh fructose consumption has been implicated with deleterious effects on human health, including hyperlipidemia elicited through de novo lipogenesis. However, more global effects of fructose on cellular metabolism have not been elucidated. In order to explore the metabolic impact of fructose-containing nutrients, we applied both GC-TOF and HILIC-QTOF mass spectrometry metabolomic strategies using extracts from cultured HepG2 cells exposed to fructose, glucose, or fructose + glucose. Cellular responses were analyzed in a time-dependent manner, incubated in media containing 5.5 mM glucose + 5.0 mM fructose in comparison to controls incubated in media containing either 5.5 mM glucose or 10.5 mM glucose. Mass spectrometry identified 156 unique known metabolites and a large number of unknown compounds, which revealed metabolite changes due to both utilization of fructose and high-carbohydrate loads independent of hexose structure. Fructose was shown to be partially converted to sorbitol, and generated higher levels of fructose-1-phosphate as a precursor for glycolytic intermediates. Differentially regulated ratios of 3-phosphoglycerate to serine pathway intermediates in high fructose media indicated a diversion of carbon backbones away from energy metabolism. Additionally, high fructoseconditions changed levels of complex lipids toward phosphatidylethanolamines. Patterns of acylcarnitines in response to high hexose exposure (10.5 mM glucose or glucose/fructose combination) suggested a reduction in mitochondrial beta-oxidation.
Institute
University of California, Davis
DepartmentGenome and Biomedical Sciences Facility
LaboratoryWCMC Metabolomics Core
Last NameFiehn
First NameOliver
Address1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, CA 95616
Emailofiehn@ucdavis.edu
Phone(530) 754-8258
Submit Date2016-04-14
Raw Data AvailableYes
Raw Data File Type(s).peg
Analysis Type DetailGC-MS
Release Date2016-04-25
Release Version1
Oliver Fiehn Oliver Fiehn
https://dx.doi.org/10.21228/M8HW28
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000298
Project DOI:doi: 10.21228/M8HW28
Project Title:Temporal metabolomic responses of cultured HepG2 liver cells to high fructose and high glucose exposures
Project Type:Time-course Study
Project Summary:High fructose consumption has been implicated with deleterious effects on human health, including hyperlipidemia elicited through de novo lipogenesis. However, more global effects of fructose on cellular metabolism have not been elucidated. In order to explore the metabolic impact of fructose-containing nutrients, we applied both GC-TOF and HILIC-QTOF mass spectrometry metabolomic strategies using extracts from cultured HepG2 cells exposed to fructose, glucose, or fructose + glucose. Cellular responses were analyzed in a time-dependent manner, incubated in media containing 5.5 mM glucose + 5.0 mM fructose in comparison to controls incubated in media containing either 5.5 mM glucose or 10.5 mM glucose. Mass spectrometry identified 156 unique known metabolites and a large number of unknown compounds, which revealed metabolite changes due to both utilization of fructose and high-carbohydrate loads independent of hexose structure. Fructose was shown to be partially converted to sorbitol, and generated higher levels of fructose-1-phosphate as a precursor for glycolytic intermediates. Differentially regulated ratios of 3-phosphoglycerate to serine pathway intermediates in high fructose media indicated a diversion of carbon backbones away from energy metabolism. Additionally, high fructoseconditions changed levels of complex lipids toward phosphatidylethanolamines. Patterns of acylcarnitines in response to high hexose exposure (10.5 mM glucose or glucose/fructose combination) suggested a reduction in mitochondrial beta-oxidation.
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

Subject:

Subject ID:SU000400
Subject Type:Cells
Subject Species:Homo sapiens
Species Group:Human

Factors:

Subject type: Cells; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Treatment Time
SA017079110625baasa27_110.5mM Glu 10min
SA017080110625baasa04_110.5mM Glu 10min
SA017081110624baasa30_110.5mM Glu 10min
SA017082110624baasa23_110.5mM Glu 10min
SA017083110625baasa14_110.5mM Glu 10min
SA017084110625baasa11_110.5mM Glu 10min
SA017085110624baasa12_110.5mM Glu 1hr
SA017086110625baasa03_110.5mM Glu 1hr
SA017087110625baasa19_110.5mM Glu 1hr
SA017088110624baasa46_110.5mM Glu 1hr
SA017089110624baasa15_110.5mM Glu 1hr
SA017090110625baasa22_110.5mM Glu 1hr
SA017091110624baasa33_110.5mM Glu 24hr
SA017092110624baasa35_110.5mM Glu 24hr
SA017093110624baasa24_110.5mM Glu 24hr
SA017094110625baasa18_110.5mM Glu 24hr
SA017095110624baasa47_110.5mM Glu 24hr
SA017096110624baasa03_110.5mM Glu 24hr
SA017097110624baasa20_110.5mM Glu 6hr
SA017098110624baasa05_110.5mM Glu 6hr
SA017099110624baasa42_110.5mM Glu 6hr
SA017100110624baasa31_110.5mM Glu 6hr
SA017101110625baasa21_110.5mM Glu 6hr
SA017102110625baasa09_110.5mM Glu 6hr
SA017103110624baasa44_15.5mM Glu 5mM Fru 10min
SA017104110624baasa16_15.5mM Glu 5mM Fru 10min
SA017105110624baasa27_15.5mM Glu 5mM Fru 10min
SA017106110624baasa43_15.5mM Glu 5mM Fru 10min
SA017107110625baasa01_15.5mM Glu 5mM Fru 10min
SA017108110624baasa28_15.5mM Glu 5mM Fru 10min
SA017109110625baasa23_15.5mM Glu 5mM Fru 1hr
SA017110110624baasa01_15.5mM Glu 5mM Fru 1hr
SA017111110625baasa08_15.5mM Glu 5mM Fru 1hr
SA017112110624baasa08_15.5mM Glu 5mM Fru 1hr
SA017113110625baasa15_15.5mM Glu 5mM Fru 1hr
SA017114110624baasa10_15.5mM Glu 5mM Fru 1hr
SA017115110624baasa40_15.5mM Glu 5mM Fru 24hr
SA017116110624baasa06_15.5mM Glu 5mM Fru 24hr
SA017117110624baasa37_15.5mM Glu 5mM Fru 24hr
SA017118110625baasa28_15.5mM Glu 5mM Fru 24hr
SA017119110625baasa07_15.5mM Glu 5mM Fru 24hr
SA017120110624baasa45_15.5mM Glu 5mM Fru 24hr
SA017121110625baasa05_15.5mM Glu 5mM Fru 6hr
SA017122110624baasa19_15.5mM Glu 5mM Fru 6hr
SA017123110624baasa39_15.5mM Glu 5mM Fru 6hr
SA017124110624baasa17_15.5mM Glu 5mM Fru 6hr
SA017125110624baasa13_15.5mM Glu 5mM Fru 6hr
SA017126110624baasa49_15.5mM Glu 5mM Fru 6hr
SA017127110624baasa34_15.5mM Glu 10min
SA017128110624baasa09_15.5mM Glu 10min
SA017129110625baasa25_15.5mM Glu 10min
SA017130110624baasa14_15.5mM Glu 10min
SA017131110625baasa16_15.5mM Glu 10min
SA017132110624baasa50_15.5mM Glu 10min
SA017133110624baasa02_15.5mM Glu 1hr
SA017134110625baasa17_15.5mM Glu 1hr
SA017135110624baasa41_15.5mM Glu 1hr
SA017136110624baasa11_15.5mM Glu 1hr
SA017137110624baasa26_15.5mM Glu 1hr
SA017138110624baasa22_15.5mM Glu 1hr
SA017139110624baasa38_15.5mM Glu 24hr
SA017140110625baasa20_15.5mM Glu 24hr
SA017141110625baasa24_15.5mM Glu 24hr
SA017142110625baasa02_15.5mM Glu 24hr
SA017143110625baasa10_15.5mM Glu 24hr
SA017144110624baasa48_15.5mM Glu 24hr
SA017145110624baasa18_15.5mM Glu 6hr
SA017146110624baasa21_15.5mM Glu 6hr
SA017147110624baasa04_15.5mM Glu 6hr
SA017148110624baasa25_15.5mM Glu 6hr
SA017149110625baasa06_15.5mM Glu 6hr
SA017150110625baasa13_15.5mM Glu 6hr
SA017151110624baasa36_1Glucosamine 10min
SA017152110624baasa32_1Glucosamine 10min
SA017153110624baasa29_1Glucosamine 10min
SA017154110625baasa26_1Glucosamine 10min
SA017155110624baasa07_1Glucosamine 10min
SA017156110625baasa12_1Glucosamine 10min
Showing results 1 to 78 of 78

Collection:

Collection ID:CO000394
Collection Summary:HepG2 cells (ATCC HB-8065) were cultured in MEM containing 10 % (v:v) FBS, 100 U/mL penicillin and 100 mg/mL streptomycin (Invitrogen, Carlsbad, CA), 1 x MEM non-essential amino acids, and 5.5 mM glucose at 37 °C in a 5 % CO2 environment. Cells were grown for four to six passages in 10 cm tissue culture dishes with 14 mL MEM, and transferred to MULTIWELL™ 12 well culture dishes for incubation in 2 mL of the treatment medium. Upon reaching 80 % confluency, the cell culture medium was changed to media representative of experimental condition: 5.5 mM glucose MEM, 5.5 mM glucose MEM + 5 mM glucose, or 5.5 mM glucose MEM + 5 mM fructose. Media was replenished after 24 and 48 h. Cell material was collected at 10 min, 1, 6, and 24 h time points following the 48 h media replacement. Culture dishes were placed on ice and each well was washed twice with 1 mL ice-cold PBS. 4 mL ice-cold 3:1 methanol/H2O extraction solvent was added to each well. Cell material was manually scraped from each well, and the extraction solvent cell material suspension was transferred to collection tubes and frozen at −80 °C prior to further processing.
Collection Protocol Filename:metabolomic_responses_of_cultured_HepG2_liver_cells.pdf
Sample Type:Cell
Collection Location:Invitrogen, Carlsbad, CA
Collection Frequency:Cell material was collected at 10 min, 1, 6, and 24 h time points following the 48 h media replacement.
Tissue Cell Identification:HepG2

Treatment:

Treatment ID:TR000414
Treatment Summary:3 Treatments: A) Control group of HepG2 cells incubated in media with 5.5 mM glucose (Glc5) B) HepG2 cells incubated in media containing 5.5 mM glucose + 5.0 mM fructose (Glc5Fru5) C) HepG2 cells incubated in media containing 10.5 mM glucose (Glc10)
Treatment Protocol Filename:metabolomic_responses_of_cultured_HepG2_liver_cells.pdf
Treatment Protocol Comments:HepG2 cells incubated in media containing 10.5 mM glucose (Glc10) was included to enable differentiation of metabolic effects caused by fructose from metabolic effects caused by increased hexose resources
Cell Media:Cells were acclimated to their respective media condition for 48 h prior to collection of sample material to obtain a metabolite profile representative of continued exposure instead of response to a sudden change in carbohydrate resources. Media was replenished after 24 and 48 h

Sample Preparation:

Sampleprep ID:SP000407
Sampleprep Summary:Sample material was thawed on ice, vortexed for 20 s, sonicated for 5 min with a VWR 50HT Ultrasonic Bath (VWR International Inc., Bridgeport, NJ), and separated into 500 μL aliquots. Each aliquot was centrifuged for 5 min @ 14,000 rcf, and supernatant was collected and lyophilized to dryness. Samples was kept on ice and removed only for sonication, centrifugation, and lyophilization steps. Lyophilized material was used for HILIC-QTOF metabolite profiling without additional clean-up steps. Lyophilized material for GC-TOF analysis was redissolved in 1:1 acetonitrile/H2O, vortexed for 10 s, and centrifuged for 5 min @ 14,000 rcf. Supernatant was collected and lyophilized to dryness.
Sampleprep Protocol Filename:metabolomic_responses_of_cultured_HepG2_liver_cells.pdf
Processing Method:Lysophilization

Combined analysis:

Analysis ID AN000613
Analysis type MS
Chromatography type GC
Chromatography system Agilent 6890N
Column Restek Corporation Rtx-5Sil MS
MS Type EI
MS instrument type GC-TOF
MS instrument name Leco Pegasus IV TOF
Ion Mode POSITIVE
Units counts

Chromatography:

Chromatography ID:CH000439
Methods Filename:Data_Dictionary_Fiehn_laboratory_GCTOF_MS_primary_metabolism_10-15-2013_general.pdf
Instrument Name:Agilent 6890N
Column Name:Restek Corporation Rtx-5Sil MS
Column Temperature:50-330C
Flow Rate:1 ml/min
Oven Temperature:50°C for 1 min, then ramped at 20°C/min to 330°C, held constant for 5 min
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:MS000546
Analysis ID:AN000613
Instrument Name:Leco Pegasus IV TOF
Instrument Type:GC-TOF
MS Type:EI
Ion Mode:POSITIVE
Ion Source Temperature:250 C
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
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