Summary of Study ST001975
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 PR001255. The data can be accessed directly via it's Project DOI: 10.21228/M8GT41 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 | ST001975 |
Study Title | Anti-oxidative metabolism measurement in mammalian cells and tissues by quantitative LC/MS method (II) |
Study Summary | Anti-oxidation metabolism measurement in mouse CSF by quantitative LC/MS method to establish MTX effects on mouse metabolism in mouse controls 0-48hrs in CSF (repeat of 20200124 ChP-MTX-Anti-oxidative-study-test) |
Institute | Boston Children's Hospital, Harvard Medical School |
Last Name | Petrova |
First Name | Boryana |
Address | 300 Longwood Ave |
boryana.petrova@childrens.harvard.edu | |
Phone | 6173557433 |
Submit Date | 2021-09-14 |
Raw Data Available | Yes |
Raw Data File Type(s) | raw(Thermo) |
Analysis Type Detail | LC-MS |
Release Date | 2022-08-29 |
Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
Project ID: | PR001255 |
Project DOI: | doi: 10.21228/M8GT41 |
Project Title: | Gene-therapy enhances CSF’s anti-oxidative activity to mitigate chemotherapy side effects |
Project Type: | Anti-oxidative Metabolism Measurement in CSF of MTX-treated mice under gene therapy by quantitative LC/MS method |
Project Summary: | In order to test the protective activity of SOD3 levels on the redox state of the CSF of MTX-treated mice, targeted metabolomics on CSF was employed at different timepoints after treatment, on mice of both sexes, at various levels of impacted hSOD3 expression. |
Institute: | Boston Childrens Hospital |
Department: | Pathology |
Laboratory: | Naama Kanarek |
Last Name: | Petrova |
First Name: | Boryana |
Address: | 300 Longwood Av, Boston, MA, 2115, USA |
Email: | boryana.petrova@childrens.harvard.edu |
Phone: | 6173557433 |
Subject:
Subject ID: | SU002055 |
Subject Type: | Mammal |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Gender: | Male and female |
Species Group: | Mammals |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
mb_sample_id | local_sample_id | Factor |
---|---|---|
SA185294 | pooled sample QCa | - |
SA185295 | pooled sample QCb | - |
SA185296 | pooled sample QC1c | - |
SA185297 | pooled sample QC 1/10 dilution b | - |
SA185298 | pooled sample QC1a | - |
SA185299 | pooled sample QC 1/3 dilution a | - |
SA185300 | pooled sample QC1b | - |
SA185301 | pooled sample QCd | - |
SA185302 | pooled sample QCc | - |
SA185303 | pooled sample QC 1/10 dilution 1a | - |
SA185304 | pooled sample QC 1/3 dilution | - |
SA185305 | pooled sample QCe | - |
SA185306 | pooled sample QC 1/10 dilution a | - |
SA185307 | blank start | - |
SA185308 | pooled sample QC 1/10 dilution 1b | - |
SA185309 | blank end | - |
SA185310 | blank mid | - |
SA185311 | mouse HC-R1 | female |
SA185312 | mouse HC-R3 | female |
SA185313 | mouse HC-L2 | female |
SA185314 | mouse HC-L1 | female |
SA185315 | mouse HC-R2 | female |
SA185316 | mouse CSF 19 | female 24h |
SA185317 | mouse CSF 22 | female 24h |
SA185318 | mouse CSF 20 | female 24h |
SA185319 | mouse CSF 21 | female 24h |
SA185320 | mouse CSF 13 | female 4h |
SA185321 | mouse CSF 12 | female 4h |
SA185322 | mouse CSF 14 | female 4h |
SA185323 | mouse CSF 11 | female 4h |
SA185324 | mouse CSF 5 | female ctl |
SA185325 | mouse CSF 4 | female ctl |
SA185326 | mouse CSF 6 | female ctl |
SA185327 | mouse CSF 15 | male 24h |
SA185328 | mouse CSF 18 | male 24h |
SA185329 | mouse CSF 17 | male 24h |
SA185330 | mouse CSF 16 | male 24h |
SA185331 | mouse CSF 25 | male 48h |
SA185332 | mouse CSF 26 | male 48h |
SA185333 | mouse CSF 23 | male 48h |
SA185334 | mouse CSF 24 | male 48h |
SA185335 | mouse CSF 7 | male 4h |
SA185336 | mouse CSF 8 | male 4h |
SA185337 | mouse CSF 10 | male 4h |
SA185338 | mouse CSF 9 | male 4h |
SA185339 | mouse CSF 2 | male ctl |
SA185340 | mouse CSF 1 | male ctl |
SA185341 | mouse CSF 3 | male ctl |
Showing results 1 to 48 of 48 |
Collection:
Collection ID: | CO002048 |
Collection Summary: | All animal care and experimental procedures were approved by the Institutional Animal Care and Use Committees of Boston Children’s Hospital. Mouse strain used was C57BL/6. All animal studies were performed under the protocol approved by the Institutional Animal Care and Use Committee (IACUC) of Boston Children’s Hospital (BCH). CD-1 mice (timed pregnant and 6-8 weeks) and Sprague Dawley rats (6-8 weeks) were purchased (Charles River Laboratories). Male and female animals were used equally for all studies. All animals were housed in a 12h light-dark cycle with ad libitum access to food and water. Hippocampus samples were collected and flash frozen.CSF was collected from the cisterna magna and centrifuged at 10,000g for 10 min. at 4 °C to remove any tissue debris (38). CSF samples were flash frozen and stored at −80°C until use. |
Sample Type: | CSF |
Treatment:
Treatment ID: | TR002067 |
Treatment Summary: | Control CSF + hippocampus samples + buffers from Study Design |
Sample Preparation:
Sampleprep ID: | SP002061 |
Sampleprep Summary: | CSF was collected by puncture of the cisterna magna with a glass capillary [39] and flash-frozen for further analysis. Per condition, 3 µl of precleared CSF were extracted by brief sonication in 200 µl of the indicated extraction buffers. After centrifugation for 10 min at maximum speed on a benchtop centrifuge (Eppendorf) the cleared supernatant was dried using a nitrogen dryer and reconstituted in 30 µl water by brief sonication. Extracted metabolites were spun again and cleared supernatant was transferred to LC-MS micro vials. A small amount of each sample was pooled and serially diluted 3- and 10-fold to be used as quality controls throughout the run of each batch. For hippocampus tissues – chunks were crushed using a hand-held homogenizer (VWR 47747-370) with several pulses while keeping the samples on ice. 300 µl of prechilled extraction buffer was used per 2 mg of tissue. For characterization by mass spectrometry, CSF was acquired 4h, 24h, and 48h following a single 75 mg/kg MTX injection from 6-8 mice and flash frozen for further analysis. Per condition, 3 μl of CSF were extracted by brief sonication in 240 μl 100% methanol, supplemented with isotopically labeled internal standards (17 amino acids and reduced glutathione, Cambridge Isotope Laboratories, MSK-A2-1.2 and CNLM-6245-10) and 60 μl 20 mM Ellman’s reagent in water (Sigma-Aldrich, D8130). After centrifugation for 10min. at maximum speed on a benchtop centrifuge (Eppendorf) the cleared supernatant was dried using a nitrogen dryer and reconstituted in 30 µl water by brief sonication. Extracted metabolites were spun again and cleared supernatant was transferred in LC-MS microvials. A small amount of each sample was pooled and serially diluted 3 and 10-fold to be used as quality controls throughout the batch run. Two microliters (equivalent to 0.2 ul of CSF) of reconstituted sample were injected into a ZIC-pHILIC 150 × 2.1 mm (5 µm particle size) column (EMD Millipore) operated on a Dionex UltiMate 3000 UPLC system (Thermo Fisher Scientific). Chromatographic separation was achieved using the following conditions: buffer A was acetonitrile; buffer B was 20 mM ammonium carbonate, 0.1% ammonium hydroxide. Gradient conditions were: linear gradient from 20% to 80% B; 20–20.5 min: from 80% to 20% B; 20.5–28 min: hold at 20% B. The column oven and autosampler tray were held at 25 °C and 4 °C, respectively. The MS data acquisition was on a QExactive benchtop orbitrap mass spectrometer equipped with an Ion Max source and a HESI II probe and was performed in a range of m/z= 70–1000, with the resolution set at 70,000, the AGC target at 1x106, and the maximum injection time (Max IT) at 20 msec. For tSIM scans, the resolution was set at 70,000, the AGC target was 1x105, and the max IT was 100 msec. Relative quantitation of polar metabolites was performed with TraceFinder 4.1 (Thermo Fisher Scientific) using a 5 ppm mass tolerance and referencing an in-house library of chemical standards. Pooled samples and fractional dilutions were prepared as quality controls and only those metabolites were taken for further analysis, for which the correlation between the dilution factor and the peak area was >0.95 (high confidence metabolites). Normalization for biological material amounts was based on the total integrated peak area values of high-confidence metabolites within an experimental batch after normalizing to the averaged factor from all mean-centered chromatographic peak areas of isotopically labeled amino acids internal standards (Cambridge Isotope Laboratories). The data were Log transformed and Pareto scaled for MetaboAnalyst-based statistical or pathway analysis (41). We profiled 200 metabolites, 85 of which were detected in CSF and passed our quality control protocol. |
Combined analysis:
Analysis ID | AN003223 |
---|---|
Analysis type | MS |
Chromatography type | HILIC |
Chromatography system | Thermo Q Exactive Orbitrap |
Column | EMD Millipore ZIC-HILIC (100 x 2.1mm,3.5um) |
MS Type | ESI |
MS instrument type | Orbitrap |
MS instrument name | Thermo Exactive Plus Orbitrap |
Ion Mode | UNSPECIFIED |
Units | ppm |
Chromatography:
Chromatography ID: | CH002377 |
Instrument Name: | Thermo Q Exactive Orbitrap |
Column Name: | EMD Millipore ZIC-HILIC (100 x 2.1mm,3.5um) |
Chromatography Type: | HILIC |
MS:
MS ID: | MS002997 |
Analysis ID: | AN003223 |
Instrument Name: | Thermo Exactive Plus Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | Xcalibur + Tracefinder |
Ion Mode: | UNSPECIFIED |