Summary of Study ST001311
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 PR000890. The data can be accessed directly via it's Project DOI: 10.21228/M8ND7K This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST001311 |
| Study Title | Lipid expression in liver after early lifer exposure to an endocrine disruptor at 70 days postnatal (part-II) |
| Study Type | Lipid expression after chemical exposure versus control |
| Study Summary | Our early-life environment has a profound influence on developing organs that impact metabolic function and determines disease susceptibility across the life-course. Using a rat model for exposure to an endocrine disrupting chemical (EDC), we show that early-life exposure causes metabolic dysfunction in adulthood and reprograms histone marks in the developing liver to accelerate acquisition of an adult epigenomic signature. This epigenomic reprogramming persists long after the initial exposure, but many reprogrammed genes remain transcriptionally silent with their impact on metabolism not revealed until a later life exposure to a Western-style diet. Diet-dependent metabolic disruption was largely driven by reprogramming of the Early Growth Response 1 (EGR1) transcriptome and production of metabolites in pathways linked to cholesterol, lipid and one-carbon metabolism. These findings demonstrate the importance of epigenome:environment interactions, which early in life accelerate epigenomic aging, and later in adulthood unlock metabolically restricted epigenetic reprogramming to drive metabolic dysfunction. |
| Institute | Baylor College of Medicine |
| Last Name | Walker |
| First Name | Cheryl |
| Address | 1 Baylor Plaza, Houston, TX, 77030, USA |
| Cheryl.walker@bcm.edu | |
| Phone | 713-798-8219 |
| Submit Date | 2020-01-24 |
| Num Groups | 2 |
| Total Subjects | 10 |
| Num Males | 10 |
| Analysis Type Detail | LC-MS |
| Release Date | 2020-03-11 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR000890 |
| Project DOI: | doi: 10.21228/M8ND7K |
| Project Title: | Metabolite and lipid profiling after early-life exposure to an endocrine disrupting chemical. |
| Project Type: | Targeted and Untargeted MS analysis |
| Project Summary: | Metabolic profiling in serum and liver tissue after early-life exposure to an endocrine disrupting chemical. |
| Institute: | Baylor College of Medicine |
| Last Name: | Walker |
| First Name: | Cheryl |
| Address: | 1 Baylor Plaza, Houston, TX, 77030, USA |
| Email: | Cheryl.walker@bcm.edu |
| Phone: | 713-798-8219 |
Subject:
| Subject ID: | SU001385 |
| Subject Type: | Mammal |
| Subject Species: | Rattus norvegicus |
| Taxonomy ID: | 10116 |
| Genotype Strain: | Sprague Dawley |
| Age Or Age Range: | 70 days old |
| Gender: | Male |
| Animal Animal Supplier: | Harlan |
| Animal Housing: | polycarbonate-free caging |
| Animal Light Cycle: | 14-hr light and 10-hr dark |
| Animal Feed: | Phytoestrogen Reduced II 18-5, Ziegler Bros, Inc |
Factors:
Subject type: Mammal; Subject species: Rattus norvegicus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Treatment |
|---|---|---|
| SA094659 | ncB3 | BPA |
| SA094660 | ncB4 | BPA |
| SA094661 | ncB5 | BPA |
| SA094662 | ncB2 | BPA |
| SA094663 | ncB1 | BPA |
| SA094664 | ncV2 | vehicle |
| SA094665 | ncV3 | vehicle |
| SA094666 | ncV4 | vehicle |
| SA094667 | ncV5 | vehicle |
| SA094668 | ncV1 | vehicle |
| Showing results 1 to 10 of 10 |
Collection:
| Collection ID: | CO001380 |
| Collection Summary: | Liver tissue was harvested on post-natal day 70 . Tissue was snap-frozen in liquid nitrogen. |
| Sample Type: | Liver |
Treatment:
| Treatment ID: | TR001400 |
| Treatment Summary: | Neonatal rats were treated with vehicle (sesame oil) or bisphenol A (BPA; 50 µg/kg dissolved in sesame oil) orally via pipette tip on post-natal days 1, 3, and 5. Littermates were randomly assigned to the treatment groups. BPA was obtained from the National Institute of Environmental Health Sciences (NIEHS). The dose and route of administration recapitulates human exposure to BPA. Tissue was harvested on postnatal day 70. |
Sample Preparation:
| Sampleprep ID: | SP001393 |
| Sampleprep Summary: | Lipids were extracted using a modified Bligh-Dyer method. Fifty µL of serum and 25 mg of crushed liver was used for the extraction. The extraction was carried out using 2:2:2 volume ratio of water/methanol/dichloromethane at room temperature after spiking internal standards 17:0 LPC, 17:0 PC, 17:0 PE, 17:0 PG, 17:0 ceramide, 17:0 SM, 17:0PS, 17:0PA, 17:0 TAG, 17:0 MAG, 16:0/18:1 DAG, 17:0 CE. The organic layer was collected and completely dried under nitrogen. Before MS analysis, the dried extract was resuspended in 100 μL of Buffer B (10:5:85 Acetonitrile/water/Isopropyl alcohol) containing 10 mM NH4OAc and subjected to LC/MS. The lipidome was separated using reverse-phase chromatography. |
| Sampleprep Protocol Filename: | unbiased.liver.MS.method.pdf |
Combined analysis:
| Analysis ID | AN002182 | AN002183 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Shimadzu Nexera-x2 | Shimadzu Nexera-x2 |
| Column | 1.8 μm particle 50 × 2.1 mm | 1.8 μm particle 50 × 2.1 mm |
| MS Type | ESI | ESI |
| MS instrument type | Triple TOF | Triple TOF |
| MS instrument name | ABI Sciex 5600 TripleTOF | ABI Sciex 5600 TripleTOF |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | peak intensity | peak intensity |
Chromatography:
| Chromatography ID: | CH001597 |
| Instrument Name: | Shimadzu Nexera-x2 |
| Column Name: | 1.8 μm particle 50 × 2.1 mm |
| Column Temperature: | 55 |
| Flow Gradient: | linear gradient over a 20 min total run time, with 60% solvent A and 40% solvent B gradient in the first 10 minutes, then the gradient was ramped in a linear fashion to 100% solvent B which was maintained for 7 minutes. After that the system was switched back to 60% solvent B and 40% solvent A for 3 minutes. |
| Flow Rate: | 0.4 mL/min |
| Solvent A: | 40% acetonitrile/60% water; 10 mM ammonium acetate |
| Solvent B: | 10% acetonitrile/5% water/85% isopropanol; 10 mM ammonium acetate |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH001598 |
| Instrument Name: | Shimadzu Nexera-x2 |
| Column Name: | 1.8 μm particle 50 × 2.1 mm |
| Column Temperature: | 55 |
| Flow Gradient: | linear gradient over a 20 min total run time, with 60% solvent A and 40% solvent B gradient in the first 10 minutes, then the gradient was ramped in a linear fashion to 100% solvent B which was maintained for 7 minutes. After that the system was switched back to 60% solvent B and 40% solvent A for 3 minutes. |
| Flow Rate: | 0.4 mL/min |
| Solvent A: | 40% acetonitrile/60% water; 10 mM ammonium acetate |
| Solvent B: | 10% acetonitrile/5% water/85% isopropanol; 10 mM ammonium acetate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS002029 |
| Analysis ID: | AN002182 |
| Instrument Name: | ABI Sciex 5600 TripleTOF |
| Instrument Type: | Triple TOF |
| MS Type: | ESI |
| MS Comments: | For data acquisition through LC/MS analysis, we used a Shimadzu CTO-20A Nexera X2 UHPLC system equipped with a degasser, binary pump, thermostatted auto sampler, and a column oven for chromatographic separation. The column heater temperature was set at 55°C. For lipid separation, the 5 uL of the lipid extract was injected into a 1.8 μm particle 50 × 2.1 mm Acquity HSS UPLC T3 column (Waters, Milford, MA) which heats to 55°C. Acetonitrile/water (40:60, v/v) with 10 mM ammonium acetate was solvent A and acetonitrile/water/isopropanol (10:5:85 v/v) with 10 mM ammonium acetate was solvent B. For chromatographic elution we used a linear gradient over a 20 min total run time, with 60% solvent A and 40% solvent B gradient in the first 10 minutes, then the gradient was ramped in a linear fashion to 100% solvent B which was maintained for 7 minutes. After that the system was switched back to 60% solvent B and 40% solvent A for 3 minutes. The flow rate used for these experiments was 0.4 mL/min and the injection volume was 5μL. The column was equilibrated for 3 min before the next injection and run at a flow rate of 0.4 mL/min for a total run time of 20 min. The data acquisition of each sample was performed in both positive and negative ionization modes using a TripleTOF 5600 equipped with a Turbo VTM ion source (AB Sciex, Concord, Canada). The column effluent was directed to the electrospray ionization source. The voltage of source was set to 5500 V for positive ionization and 4500 V for negative ionization mode, the declustering potential was set to 60 V, and the source temperature to 450oC for both modes. The curtain gas flow, nebulizer, and heater gas were set to 30, 40, and 45 units, respectively. The instrument performed one TOF MS survey scan (150 ms) and 15 MS/MS scans with a total duty cycle time of 2.4 s. The mass range in both modes was 50-1200 m/z. We controlled the acquisition of MS/MS spectra by data-dependent acquisition (DDA) function of the Analyst TF software (AB Sciex, Concord, Canada) with the following parameters: dynamic background subtraction, charge monitoring to exclude multiply charged ions and isotopes, and dynamic exclusion of former target ions for 9 s. Rolling collision energy spread was set whereby the software calculated the collision energy value to be applied as a function of m/z. Mass accuracy was maintained by the use of an automated calibrant delivery system interfaced to the second inlet of the DuoSpray source. Lipidomics Data Analysis. Raw data was converted to the mgf data format using proteoWizard software (2).The NIST MS PepSearch Program was used to search the converted files against LipidBlast libraries. The m/z width was determined by the mass accuracy of internal standards and were set to 0.001 for positive mode and to 0.005 for negative mode with an overall mass error of less than 2 parts per million. The minimum match factor used was set to 400. All raw data files were searched against the library of identified lipids based on mass and retention time using Multiquant 1.1.0.26 (ABsciex, Concord, Canada). Relative abundance of peak spectra was used for the analyses. Lipids that were identified in both positive and negative ion modes were initially analyzed separately for their relationship with outcome to ensure persistent results. As the relationship with outcome was not different in such lipids by ion modes, the values from positive mode were used in the final analysis. For lipid features with multiple adducts, the sum of spectral peaks from different adducts was used for the corresponding lipid. Identified lipids were quantified by normalizing against their respective internal standard. Quality control samples were used to monitor the overall quality of the lipid extraction and mass spectrometry analyses. The distributions of detected lipid species across the quality control samples indicated high reproducibility. |
| Ion Mode: | POSITIVE |
| MS ID: | MS002030 |
| Analysis ID: | AN002183 |
| Instrument Name: | ABI Sciex 5600 TripleTOF |
| Instrument Type: | Triple TOF |
| MS Type: | ESI |
| MS Comments: | For data acquisition through LC/MS analysis, we used a Shimadzu CTO-20A Nexera X2 UHPLC system equipped with a degasser, binary pump, thermostatted auto sampler, and a column oven for chromatographic separation. The column heater temperature was set at 55°C. For lipid separation, the 5 uL of the lipid extract was injected into a 1.8 μm particle 50 × 2.1 mm Acquity HSS UPLC T3 column (Waters, Milford, MA) which heats to 55°C. Acetonitrile/water (40:60, v/v) with 10 mM ammonium acetate was solvent A and acetonitrile/water/isopropanol (10:5:85 v/v) with 10 mM ammonium acetate was solvent B. For chromatographic elution we used a linear gradient over a 20 min total run time, with 60% solvent A and 40% solvent B gradient in the first 10 minutes, then the gradient was ramped in a linear fashion to 100% solvent B which was maintained for 7 minutes. After that the system was switched back to 60% solvent B and 40% solvent A for 3 minutes. The flow rate used for these experiments was 0.4 mL/min and the injection volume was 5μL. The column was equilibrated for 3 min before the next injection and run at a flow rate of 0.4 mL/min for a total run time of 20 min. The data acquisition of each sample was performed in both positive and negative ionization modes using a TripleTOF 5600 equipped with a Turbo VTM ion source (AB Sciex, Concord, Canada). The column effluent was directed to the electrospray ionization source. The voltage of source was set to 5500 V for positive ionization and 4500 V for negative ionization mode, the declustering potential was set to 60 V, and the source temperature to 450oC for both modes. The curtain gas flow, nebulizer, and heater gas were set to 30, 40, and 45 units, respectively. The instrument performed one TOF MS survey scan (150 ms) and 15 MS/MS scans with a total duty cycle time of 2.4 s. The mass range in both modes was 50-1200 m/z. We controlled the acquisition of MS/MS spectra by data-dependent acquisition (DDA) function of the Analyst TF software (AB Sciex, Concord, Canada) with the following parameters: dynamic background subtraction, charge monitoring to exclude multiply charged ions and isotopes, and dynamic exclusion of former target ions for 9 s. Rolling collision energy spread was set whereby the software calculated the collision energy value to be applied as a function of m/z. Mass accuracy was maintained by the use of an automated calibrant delivery system interfaced to the second inlet of the DuoSpray source. Lipidomics Data Analysis. Raw data was converted to the mgf data format using proteoWizard software (2).The NIST MS PepSearch Program was used to search the converted files against LipidBlast libraries. The m/z width was determined by the mass accuracy of internal standards and were set to 0.001 for positive mode and to 0.005 for negative mode with an overall mass error of less than 2 parts per million. The minimum match factor used was set to 400. All raw data files were searched against the library of identified lipids based on mass and retention time using Multiquant 1.1.0.26 (ABsciex, Concord, Canada). Relative abundance of peak spectra was used for the analyses. Lipids that were identified in both positive and negative ion modes were initially analyzed separately for their relationship with outcome to ensure persistent results. As the relationship with outcome was not different in such lipids by ion modes, the values from positive mode were used in the final analysis. For lipid features with multiple adducts, the sum of spectral peaks from different adducts was used for the corresponding lipid. Identified lipids were quantified by normalizing against their respective internal standard. Quality control samples were used to monitor the overall quality of the lipid extraction and mass spectrometry analyses. The distributions of detected lipid species across the quality control samples indicated high reproducibility. |
| Ion Mode: | NEGATIVE |
