Summary of Study ST002190
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 PR001396. The data can be accessed directly via it's Project DOI: 10.21228/M88H8K This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST002190 |
| Study Title | Mass Spectrometry Imaging of Lipids In A Gut Epithelial Cell Model |
| Study Type | Mass spectromery imaging of cells. |
| Study Summary | Scope: The Caco2/HT29-MTX co-culture system is widely used as a cell model of the intestinal epithelium. Although the gut epithelium plays an important role in the uptake of free fatty acids and the resynthesis of triglycerides the lipid distribution profile of the co-culture system is not well understood. Desorption electrospray ionization (DESI) is a mass spectrometry (MS) technique which has been widely used to study the main classes of lipid molecules on different tissue surfaces. This has been used to map lipid species and their distribution in Caco2 and HT29-MTX co-culture system. Methods and results: Caco2 and HT29-MTX cells were seeded on coverslips either singly or as cocultures in ratios of 75:25 and 50:50. Cells were cultured for 21 days before MS imaging using a DESI source in both the positive and negative ionization modes. The identity of selected lipids was confirmed in negative and positive ionisation modes using tandem MS. Although many lipids were common to both cell lines, there were distinctive patterns in the lipidomes. Thus, the lipidome of Caco2 cells was more heterogeneous and rich in cholesterol esters and triglycerides whilst HT29-MTX cells has a distinctive lipidome relating to phosphatidylethanolamines, phosphatidylinositols and odd chain lipids, including C17 fatty acids. Conclusion: DESI-MSI has shown that Caco2 and HT29-MTX cells have distinctive lipidomes which are still evident when the cells are cocultured. It has potential to both allow further validation of these widely used cell models and provide insights into how dietary components may modify lipid metabolism in future. |
| Institute | Manchester Institute of Biotechnology, University of Manchester |
| Last Name | Mattar |
| First Name | Hadeer |
| Address | Manchester Institute of Biotechnology, Princess Street, Manchester, UK, M1 7DN |
| hmatar@bu.edu.sa | |
| Phone | 0161 306 6000 |
| Submit Date | 2022-05-11 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | MS(Dir. Inf.) |
| Release Date | 2022-07-06 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001396 |
| Project DOI: | doi: 10.21228/M88H8K |
| Project Title: | Mass Spectrometry Imaging of Lipids In A Gut Epithelial Cell Model |
| Project Type: | MS imaging of cells |
| Project Summary: | Mass spectrometry imaging of lipids in a gut epithelial cell model |
| Institute: | University of Manchester |
| Last Name: | Mattar |
| First Name: | Hadeer |
| Address: | Manchester Institute of Biotechnology, Princess Street, Manchester, UK, M1 7DN |
| Email: | hmatar@bu.edu.sa |
| Phone: | 0161 306 6000 |
Subject:
| Subject ID: | SU002276 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Caco2 | Ionisation_mode |
|---|---|---|---|
| SA210162 | 06DEC2018_Hadeer_neg_cell5_MS01 Analyte 3 | 0;100 | Negative |
| SA210163 | 06Decv18_manchesterUni_Hadeer_cell5_PosMS01 Analyte 1 | 0;100 | Positive |
| SA210164 | 06DEC2018_Hadeer_neg_cell1_MS01 Analyte 3 | 100;0 | Negative |
| SA210165 | 06Decv18_manchesterUni_Hadeer_cell1_PosMS01 Analyte 1 | 100;0 | Positive |
| SA210166 | 06DEC2018_Hadeer_neg_cell4_MS01 Analyte 3 | 25;75 | Negative |
| SA210167 | 06Decv18_manchesterUni_Hadeer_cell4_PosMS01 Analyte 1 | 25;75 | Positive |
| SA210168 | 06DEC2018_Hadeer_neg_cell3_MS01 Analyte 3 | 50;50 | Negative |
| SA210169 | 06Decv18_manchesterUni_Hadeer_cell3_PosMS01 Analyte 1 | 50;50 | Positive |
| SA210170 | 06DEC2018_Hadeer_neg_cell2_MS01 Analyte 3 | 75;25 | Negative |
| SA210171 | 06Decv18_manchesterUni_Hadeer_cell2_PosMS01 Analyte 1 | 75;25 | Positive |
| Showing results 1 to 10 of 10 |
Collection:
| Collection ID: | CO002269 |
| Collection Summary: | Caco2 and HT29-MTX cells were cultured separately in bulk in 25 cm2 flasks at 37 ºC, 5% CO2 for 2-3 days. DMEM complete media supplemented with 20% (v/v) foetal serum bovine, 1% (w/v) non-essential amino acids, 1% (w/v) L-glutamine and 1% (w/v) penicillin-streptomycin was used as culture medium. On reaching 80–90% confluency cells were trypsinised and then seeded on a coverslip at a density of 1x105 cells/ml either as separate cultures or in a co-culture system seeded at ratios of 75:25, 50:50 and 25:75 (Caco2: HT29-MTXcells). After seeding, each coverslip was placed in the well of a 6-well cell culture plate, and incubated at 37 ºC, 5% CO2. Media was changed every 48 h for 21 days until cells were confluent. Cells were prepared for DESI MSI analysis by rinsing coverslips in 150 mM ammonium acetate, pH 7.1 for 30 s before being allowed to dry in the air stream of a biological safety cabinet for 15 min. Coverslips were then thoroughly dried using a vacuum desiccator for 15 min before storage at -80 °C in petri dishes until required. |
| Sample Type: | Cultured cells |
Treatment:
| Treatment ID: | TR002288 |
| Treatment Summary: | Caco2 and HT29-MTX cells were cultured separately in bulk in 25 cm2 flasks at 37 ºC, 5% CO2 for 2-3 days. DMEM complete media supplemented with 20% (v/v) foetal serum bovine, 1% (w/v) non-essential amino acids, 1% (w/v) L-glutamine and 1% (w/v) penicillin-streptomycin was used as culture medium. On reaching 80–90% confluency cells were trypsinised and then seeded on a coverslip at a density of 1x105 cells/ml either as separate cultures or in a co-culture system seeded at ratios of 75:25, 50:50 and 25:75 (Caco2: HT29-MTXcells). After seeding, each coverslip was placed in the well of a 6-well cell culture plate, and incubated at 37 ºC, 5% CO2. Media was changed every 48 h for 21 days until cells were confluent. Cells were prepared for DESI MSI analysis by rinsing coverslips in 150 mM ammonium acetate, pH 7.1 for 30 s before being allowed to dry in the air stream of a biological safety cabinet for 15 min. Coverslips were then thoroughly dried using a vacuum desiccator for 15 min before storage at -80 °C in petri dishes until required. |
Sample Preparation:
| Sampleprep ID: | SP002282 |
| Sampleprep Summary: | Cells were prepared for DESI MSI analysis by rinsing coverslips in 150 mM ammonium acetate, pH 7.1 for 30 s before being allowed to dry in the air stream of a biological safety cabinet for 15 min. Coverslips were then thoroughly dried using a vacuum desiccator for 15 min before storage at -80 °C in petri dishes until required. |
Combined analysis:
| Analysis ID | AN003584 | AN003585 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | None (Direct infusion) | None (Direct infusion) |
| Chromatography system | none | none |
| Column | none | none |
| MS Type | ESI | ESI |
| MS instrument type | QTOF | QTOF |
| MS instrument name | Waters Xevo-G2-XS | Waters Xevo-G2-XS |
| Ion Mode | NEGATIVE | POSITIVE |
| Units | Peak area | Peak area |
Chromatography:
| Chromatography ID: | CH002649 |
| Instrument Name: | none |
| Column Name: | none |
| Chromatography Type: | None (Direct infusion) |
MS:
| MS ID: | MS003339 |
| Analysis ID: | AN003584 |
| Instrument Name: | Waters Xevo-G2-XS |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | Mass spectrometry imaging using 2D DESI source (Prosolia) was conducted at Waters Corporation (Wilmslow, UK). Dried cell cultures gown on coverslips were mounted on microscope glass slides using double sided tape. Slides were scanned using Epson perfection V600 photo scanner. Scanned images were imported and the area where cells were confluent was selected using the co-registered photographic image of the samples in High-Definition Imaging (HDI) 1.5. Imaging experiments were carried out on a DESI (Prosolia, USA) mounted on a Xevo-G2-XS quadrupole-time of flight (QTOF) mass spectrometer (Waters Corporation, Wilmslow, UK). The DESI spray was composed of a solvent mixture of 98:2% MeOH: water (v/v) delivered at a flow rate of 2 μl/min with nebulizing gas pressure of 5 bar. The sprayer geometric positions were set that the sprayer was 1.5 mm above sample surface and the distance between sprayer to capillary was 6mm. The source temperature was 100 °C. For both positive and negative ionization modes, the acquisition mass range was 50-1200 m/z. DESI MSI experiments were performed using the scan rate of 4 scan/ second in negative mode. The X and Y pixel sizes were set at 20 μm. Selected precursor ions in both positive and negative ionisation modes (including m/z 810.55 and m/z 773.53) were further analysed using MS/MS. The experiments were carried out on a DESI (Prosolia, place, USA) mounted on a Xevo-G2-XS Q-TOF mass spectrometer (Waters Corporation, Wilmslow, UK). Spray conditions were the same as DESI imaging. Spectra were visualised using MassLynx (Waters Corporation, Wilmslow, UK). Raw data from each biological condition were processed using HDI software version 1.5 (Waters Corporation, Wilmslow, UK) to provide ion images from a consolidated list of m/z common to the different datasets as well as the unique m/z. Ion images were normalised to total ion current (TIC). Regions of interest (ROIs) were drawn directly from the DESI images that produced a .csv file containing average TIC normalised intensities which was used for statistical analysis using MetaboAnalyst1 (https://www.metaboanalyst.ca/MetaboAnalyst/faces/home.xhtml) to compare between selected lipids that are presented in the two cell lines. Similarly, MSI analysis of Caco2/HT29-MTX co-culture was performed using the same criteria of DESI images with the same precursor ions mentioned above. For pixel classification of DESI imaging datasets, a Waters Corporation (WRC, Budapest, Hungary) prototype AMX MS imaging software was used in combination with HDI software. |
| Ion Mode: | NEGATIVE |
| MS ID: | MS003340 |
| Analysis ID: | AN003585 |
| Instrument Name: | Waters Xevo-G2-XS |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | Mass spectrometry imaging using 2D DESI source (Prosolia) was conducted at Waters Corporation (Wilmslow, UK). Dried cell cultures gown on coverslips were mounted on microscope glass slides using double sided tape. Slides were scanned using Epson perfection V600 photo scanner. Scanned images were imported and the area where cells were confluent was selected using the co-registered photographic image of the samples in High-Definition Imaging (HDI) 1.5. Imaging experiments were carried out on a DESI (Prosolia, USA) mounted on a Xevo-G2-XS quadrupole-time of flight (QTOF) mass spectrometer (Waters Corporation, Wilmslow, UK). The DESI spray was composed of a solvent mixture of 98:2% MeOH: water (v/v) delivered at a flow rate of 2 μl/min with nebulizing gas pressure of 5 bar. The sprayer geometric positions were set that the sprayer was 1.5 mm above sample surface and the distance between sprayer to capillary was 6mm. The source temperature was 100 °C. For both positive and negative ionization modes, the acquisition mass range was 50-1200 m/z. DESI MSI experiments were performed using the scan rate of 2 scan/ second in positive mode. The X and Y pixel sizes were set at 20 μm. Selected precursor ions in both positive and negative ionisation modes (including m/z 810.55 and m/z 773.53) were further analysed using MS/MS. The experiments were carried out on a DESI (Prosolia, place, USA) mounted on a Xevo-G2-XS Q-TOF mass spectrometer (Waters Corporation, Wilmslow, UK). Spray conditions were the same as DESI imaging. Spectra were visualised using MassLynx (Waters Corporation, Wilmslow, UK). Raw data from each biological condition were processed using HDI software version 1.5 (Waters Corporation, Wilmslow, UK) to provide ion images from a consolidated list of m/z common to the different datasets as well as the unique m/z. Ion images were normalised to total ion current (TIC). Regions of interest (ROIs) were drawn directly from the DESI images that produced a .csv file containing average TIC normalised intensities which was used for statistical analysis using MetaboAnalyst1 (https://www.metaboanalyst.ca/MetaboAnalyst/faces/home.xhtml) to compare between selected lipids that are presented in the two cell lines. Similarly, MSI analysis of Caco2/HT29-MTX co-culture was performed using the same criteria of DESI images with the same precursor ions mentioned above. For pixel classification of DESI imaging datasets, a Waters Corporation (WRC, Budapest, Hungary) prototype AMX MS imaging software was used in combination with HDI software. |
| Ion Mode: | POSITIVE |
