Summary of Study ST004300
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 PR002716. The data can be accessed directly via it's Project DOI: 10.21228/M8KK1D This work is supported by NIH grant, U2C- DK119886. See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST004300 |
| Study Title | Two-dimensional liquid chromatography-mass spectrometry (2DLC-MS) platform to conduct an in-depth lipidomic analysis of liver and brain tissues from fat-1 transgenic mice exposed to ethanol. |
| Study Summary | To gain deeper insights into how alcohol and fat-1 gene influence lipid metabolism, we employed a two-dimensional liquid chromatography-mass spectrometry (2DLC-MS) platform to conduct an in-depth lipidomic analysis of liver and brain tissues from fat-1 transgenic mice exposed to ethanol. The 2DLC consisted of a hydrophilic interaction liquid chromatography (HILIC) column and a reversed phase liquid chromatography (RPC) column configured in a comprehensive mode. We aimed to delineate the lipid regulation changes in both the mouse liver and brain at a systems level and provide insights for understanding the ALD pathogenesis and the protective role of fat-1. |
| Institute | University of Louisville |
| Department | Chemistry |
| Laboratory | Dr. Xiang Zhang lab |
| Last Name | Feng |
| First Name | Jing |
| Address | 2310 S Brook St. Louisville, KY 40208 |
| jing.feng@louisville.edu | |
| Phone | 5026187846 |
| Submit Date | 2025-10-06 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-12-01 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002716 |
| Project DOI: | doi: 10.21228/M8KK1D |
| Project Title: | Integrated liver and brain lipidomics reveal tissue-specific alcohol effects and systemic intervention of fat-1 gene in mice with alcohol-associated liver disease |
| Project Type: | Original Research |
| Project Summary: | Alcohol-associated liver disease (ALD) involves profound systemic lipid disruption. Using comprehensive 2DLC-MS-based untargeted lipidomics, we characterized the hepatic and cerebral lipidomes in wild-type and fat-1 transgenic (endogenous omega-3 PUFA-producing) mice subjected to a chronic-plus-binge ethanol model. Compared to fat-1 gene, ethanol intake was the dominant factor that induced 228 lipids in the liver and 316 in the brain with significant abundance changes. . Those significantly changed lipids were consistent across wild-type and fat-1 genotypes but highly tissue-specific. Furthermore, fat-1 genotype significantly modulated ethanol's impact and even reversed the regulation of some lipids, including sphingolipids and monosaturated and saturated fatty acyl-containing lipids. Four lipids in the liver and seven lipids in the brain were co-regulated by ethanol, genotype, and their interaction, with excellent discriminatory power in linear discriminant analysis. The odd-chain lipids of those 11 lipids suggest gut microbiome contributions. While ethanol effects were tissue-specific, fat-1 induced consistent responses in the two tissues, indicating conserved protective pathways. Our findings revealed complex lipid network remodeling in ALD through a multi-factorial affecting the liver-brain axis, highlighting the potential of fat-1 gene to mitigate tissue-specific metabolic dysfunction and informing future therapeutic strategies. |
| Institute: | University of Louisville |
| Department: | Chemistry |
| Laboratory: | Dr. Xiang Zhang lab |
| Last Name: | Feng |
| First Name: | Jing |
| Address: | 2310 S Brooks St. Louisville, KY 40208 |
| Email: | jing.feng@louisville.edu |
| Phone: | 5026187846 |
Subject:
| Subject ID: | SU004453 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | fat-1 knock-in mice, fat-1 gene is from Caenorhabditis elegans |
| Age Or Age Range: | 8 weeks |
| Gender: | Female |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Sample source | Treatment |
|---|---|---|---|
| SA505133 | brain_354 | brain | Alcohol |
| SA505134 | brain_359 | brain | Alcohol |
| SA505135 | brain_358 | brain | Alcohol |
| SA505136 | brain_357 | brain | Alcohol |
| SA505137 | brain_355 | brain | Alcohol |
| SA505138 | brain_352 | brain | Alcohol |
| SA505139 | brain_353 | brain | Alcohol |
| SA505140 | brain_318 | brain | Control |
| SA505141 | brain_317 | brain | Control |
| SA505142 | brain_351 | brain | Control |
| SA505143 | brain_349 | brain | Control |
| SA505144 | brain_326 | brain | Control |
| SA505145 | brain_325 | brain | Control |
| SA505146 | brain_350 | brain | Control |
| SA505147 | liver_353 | liver | Alcohol |
| SA505148 | liver_359 | liver | Alcohol |
| SA505149 | liver_358 | liver | Alcohol |
| SA505150 | liver_357 | liver | Alcohol |
| SA505151 | liver_355 | liver | Alcohol |
| SA505152 | liver_354 | liver | Alcohol |
| SA505153 | liver_352 | liver | Alcohol |
| SA505154 | liver_350 | liver | Control |
| SA505155 | liver_351 | liver | Control |
| SA505156 | liver_349 | liver | Control |
| SA505157 | liver_326 | liver | Control |
| SA505158 | liver_325 | liver | Control |
| SA505159 | liver_318 | liver | Control |
| SA505160 | liver_317 | liver | Control |
| Showing results 1 to 28 of 28 |
Collection:
| Collection ID: | CO004446 |
| Collection Summary: | Collection summary: At the end of the experiment, mice were gavaged with vehicle (saline) or 31.5% (v/v) ethanol (in saline) at 5 g/kg. Nine hours later, mice were deeply anesthetized with 100 mg/kg ketamine and 16 mg/kg xylazine. Following collection of blood from the inferior vena cava, organs were collected, snap-frozen in liquid nitrogen, and stored at -80⁰C. |
| Collection Protocol ID: | IACUC 15423 |
| Sample Type: | Liver cells, brain cells |
| Collection Location: | Clinical Translational Research Building, University of Louisville Health Sciences Campus, Louisville, Kentucky. |
| Collection Frequency: | Once (endpoint) |
| Collection Duration: | 5 minutes |
| Storage Conditions: | -80℃ |
| Collection Vials: | 1.5 ml self-standing microcentrifuge tubes with O-ring and screw top |
| Storage Vials: | 1.5 ml self-standing microcentrifuge tubes with O-ring and screw top |
| Collection Tube Temp: | room temperature |
| Additives: | 10 U/ml sodium heparin to blood samples |
Treatment:
| Treatment ID: | TR004462 |
| Treatment Summary: | The study design involved wild type (WT) C57BL6/J and fat-1+/- transgenic female mice (8 week-old, bred in house), which were fed with high-fat (60%) diets for 12 weeks. Diets were purchased from the Research Diets (D12492i, high fat). On the final day of the dietary intervention, mice received a single dose of EtOH (5 g/kg) by oral gavage. The control mice were received with the same amount of saline by oral gavage. In this study, only fat-1+/- transgenic mice were used. Control mice were assayed in study ST004281. |
| Treatment Protocol ID: | IACUC 15423 |
| Treatment: | Ethanol binge |
| Treatment Compound: | Ethanol |
| Treatment Route: | Oral |
| Treatment Dose: | 5 g ethanol/kg body weight |
| Treatment Dosevolume: | 20 µL of 31.5% (v/v) ethanol solution/g body weight |
| Treatment Vehicle: | Pharmaceutical (USP)-grade saline |
| Animal Anesthesia: | Endpoint only |
| Animal Acclimation Duration: | 12 weeks (bred in-house) |
| Animal Endp Euthanasia: | 100 mg/kg ketamine+16mg/kg xylazine |
| Animal Endp Tissue Coll List: | Liver, brain |
| Animal Endp Clinical Signs: | >10% body weight loss, body condition score, facial grimace scale for pain and distress. |
Sample Preparation:
| Sampleprep ID: | SP004459 |
| Sampleprep Summary: | The procedure began by homogenizing the tissues in methanol at a ratio of 3 mg tissue per 20 μL. Subsequently, 200 μL of the resulting homogenate was mixed with 400 μL of chloroform, vortexed for 2 min, and then incubated for 1 hour. Following incubation, 120 μL of deionized water was added to the mixture. The mixture was vortexed for 2 min and then centrifuged at 5000 rpm for 5 min at room temperature. The bottom chloroform phase (300 μL) was collected and dried under a stream of nitrogen gas. The dried lipid extract was reconstituted in 150 μL of a chloroform/methanol solution (2:1 v/v) containing 1 mM butylated hydroxytoluene (BHT) as an antioxidant. The sample was then transferred to a glass vial for 2DLC-MS analysis. |
| Processing Storage Conditions: | On ice |
| Extraction Method: | Folch Method |
| Extract Storage: | -80℃ |
Combined analysis:
| Analysis ID | AN007152 | AN007153 | AN007154 | AN007155 |
|---|---|---|---|---|
| Chromatography ID | CH005434 | CH005434 | CH005435 | CH005435 |
| MS ID | MS006847 | MS006848 | MS006849 | MS006850 |
| Analysis type | MS | MS | MS | MS |
| Chromatography type | HILIC | HILIC | Reversed phase | Reversed phase |
| Chromatography system | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 |
| Column | Waters ACQUITY UPLC BEH HILIC (150 x 2.1 mm, 1.7 μm) | Waters ACQUITY UPLC BEH HILIC (150 x 2.1 mm, 1.7 μm) | Waters ACQUITY UPLC BEH C18 (50 x 2.1 mm, 1.7 μm) | Waters ACQUITY UPLC BEH C18 (50 x 2.1 mm, 1.7 μm) |
| MS Type | ESI | ESI | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive HF hybrid Orbitrap | Thermo Q Exactive HF hybrid Orbitrap | Thermo Q Exactive HF hybrid Orbitrap | Thermo Q Exactive HF hybrid Orbitrap |
| Ion Mode | POSITIVE | NEGATIVE | POSITIVE | NEGATIVE |
| Units | Relative Abundance | Relative Abundance | Relative Abundance | Relative Abundance |
Chromatography:
| Chromatography ID: | CH005434 |
| Chromatography Summary: | The first dimension (1D) lipid separation was achieved on a BEH hydrophilic interaction chromatography (HILIC) column (150 × 2.1 mm, 1.7 µm, Waters Corporation, Milford, MA, USA). Lipids eluted from the HILIC column were collected into five fractions based on their retention time (Rt) using a 10-port divert valve and two Acquity BEH C18 trap columns (Waters Corporation). |
| Methods Filename: | Chromatography_method.txt |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | Waters ACQUITY UPLC BEH HILIC (150 x 2.1 mm, 1.7 μm) |
| Column Temperature: | 55⁰C |
| Flow Gradient: | 0-5 min, 0%; 5-90 min, 0% to 90%; 90-95 min, 90% to 0% |
| Flow Rate: | 0.15 mL/min |
| Solvent A: | 97.5% acetonitrile; 2.5% water; 10 mM ammonium formate |
| Solvent B: | 50% methanol; 50% water; 10 mM ammonium formate |
| Chromatography Type: | HILIC |
| Chromatography ID: | CH005435 |
| Chromatography Summary: | Each HILIC fraction was then subjected to an RPC column, Acquity UPLC BEH C18 column (50 × 2.1 mm, 1.8 μm, Waters Corporation) for further separation. |
| Methods Filename: | Chromatography_method.txt |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | Waters ACQUITY UPLC BEH C18 (50 x 2.1 mm, 1.7 μm) |
| Column Temperature: | 55⁰C |
| Flow Gradient: | 0-5 min, 0%; 5-53 min, 8%; 53-73 min, 8% to 30%; 73-77 min 30% to 60%; 77.1-95 min, 60% to 0% |
| Flow Rate: | 0.5 mL/min |
| Solvent A: | 60% acetonitrile; 40% water; 10 mM ammonium formate |
| Solvent B: | 90% isopropanol; 7.5% acetonitrile; 2.5% water; 10 mM ammonium formate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS006847 |
| Analysis ID: | AN007152 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The mass spectrometer was operated using electrospray ionization (ESI) in both positive and negative ionization modes. Full MS scans were acquired over a mass-to-charge (m/z) range of 200-1500 with a resolution of 30,000 at m/z 200. The ESI source parameters were configured as follows: sheath gas flow rate 50; auxiliary gas flow rate 13; spray voltage 3.5 kV; capillary temperature 263 °C; S-lens RF level 55; and heater temperature 425°C. For tandem mass spectrometry (MS/MS), the data-dependent acquisition (DDA) settings included: 2 micro scans, resolution of 15,000 at m/z 200, maximum injection time 100 ms, and collision energies 20 and 30 eV. |
| Ion Mode: | POSITIVE |
| Analysis Protocol File: | MS_Analysis_Protocol.txt |
| MS ID: | MS006848 |
| Analysis ID: | AN007153 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The mass spectrometer was operated using electrospray ionization (ESI) in both positive and negative ionization modes. Full MS scans were acquired over a mass-to-charge (m/z) range of 200-1500 with a resolution of 30,000 at m/z 200. The ESI source parameters were configured as follows: sheath gas flow rate 50; auxiliary gas flow rate 13; spray voltage 3.5 kV; capillary temperature 263°C; S-lens RF level 55; and heater temperature 425°C. For tandem mass spectrometry (MS/MS), the data-dependent acquisition (DDA) settings included: 2 micro scans, resolution of 15,000 at m/z 200, maximum injection time 100 ms, and collision energies 20 and 30 eV. |
| Ion Mode: | NEGATIVE |
| Analysis Protocol File: | MS_Analysis_Protocol.txt |
| MS ID: | MS006849 |
| Analysis ID: | AN007154 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The mass spectrometer was operated using electrospray ionization (ESI) in both positive and negative ionization modes. Full MS scans were acquired over a mass-to-charge (m/z) range of 200-1500 with a resolution of 30,000 at m/z 200. The ESI source parameters were configured as follows: sheath gas flow rate 50; auxiliary gas flow rate 13; spray voltage 3.5 kV; capillary temperature 263°C; S-lens RF level 55; and heater temperature 425°C. For tandem mass spectrometry (MS/MS), the data-dependent acquisition (DDA) settings included: 2 micro scans, resolution of 15,000 at m/z 200, maximum injection time 100 ms, and collision energies 20 and 30 eV. |
| Ion Mode: | POSITIVE |
| Analysis Protocol File: | MS_Analysis_Protocol.txt |
| MS ID: | MS006850 |
| Analysis ID: | AN007155 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The mass spectrometer was operated using electrospray ionization (ESI) in both positive and negative ionization modes. Full MS scans were acquired over a mass-to-charge (m/z) range of 200-1500 with a resolution of 30,000 at m/z 200. The ESI source parameters were configured as follows: sheath gas flow rate 50; auxiliary gas flow rate 13; spray voltage 3.5 kV; capillary temperature 263°C; S-lens RF level 55; and heater temperature 425°C. For tandem mass spectrometry (MS/MS), the data-dependent acquisition (DDA) settings included: 2 micro scans, resolution of 15,000 at m/z 200, maximum injection time 100 ms, and collision energies 20 and 30 eV. |
| Ion Mode: | NEGATIVE |
| Analysis Protocol File: | MS_Analysis_Protocol.txt |
