Summary of Study ST002061
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 PR001305. The data can be accessed directly via it's Project DOI: 10.21228/M81H6N 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 | ST002061 |
| Study Title | Glutamine flux in macrophages treated with stable-isotope labeled analog 4 mM (U-13C5) glutamine |
| Study Type | Glutamine flux |
| Study Summary | 107 BMDMs per group (Sirt3 WT and Sirt3 K223R)were seeded in 10cm plates and incubated in RPMI-1640 cell culture medium with 10% FBS. Prior to isotopic labeling, the medium was replaced with RPMI-1640 without glutamine for 4 hrs. Then stable-isotope labeled analog 4 mM (U-13C5) glutamine (Cambridge Isotope) was added together with IL-4 (20ng/ml) for 4 hrs. |
| Institute | Shanghai Jiao Tong University affiliated Renji Hospital |
| Department | Department of Urology |
| Laboratory | Cheng Jinke's Lab |
| Last Name | Zhou |
| First Name | Wei |
| Address | N0.280 South Chongqing Road |
| joesphchou@alumni.sjtu.edu.cn | |
| Phone | +8615216716293 |
| Submit Date | 2022-01-19 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzXML |
| Analysis Type Detail | LC-MS |
| Release Date | 2022-02-07 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001305 |
| Project DOI: | doi: 10.21228/M81H6N |
| Project Title: | Glutamine Flux analysis in Macrophages |
| Project Type: | MS quantitive analysis |
| Project Summary: | To determine the effect of Sirt3 K223R on glutaminolysis, we traced glutamine metabolic influx in macrophages. The data demonstrated that Sirt3 K223R did not alter glutamine uptake and glutamate production in BMDMs after IL-4 treatment . However, 5C(M+5) labeled-αKG showed a higher ratio in IL-4-treated Sirt3 KR macrophages than that in IL-4-treated Sirt3 WT macrophages, suggesting that SENP1-Sirt3 axis mainly involves the conversion of glutamate to αKG of glutaminolysis in macrophage M2 polarization. |
| Institute: | Shanghai Jiao Tong University affiliated Renji Hospital |
| Department: | Department of Urology |
| Laboratory: | Cheng Jinke's Lab |
| Last Name: | Zhou |
| First Name: | Wei |
| Address: | N0.280 South Chongqing Road |
| Email: | joesphchou@alumni.sjtu.edu.cn |
| Phone: | +8615216716293 |
| Funding Source: | National Natural Science Foundation of China |
| Project Comments: | In summary, we reveal that SENP1-Sirt3 signaling plays a crucial role in promoting αKG production and M2 polarization. |
| Publications: | Cell Reports |
| Contributors: | Wei Zhou |
Subject:
| Subject ID: | SU002143 |
| Subject Type: | Cultured cells |
| Subject Species: | Mus musculus |
Factors:
Subject type: Cultured cells; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Genotype |
|---|---|---|
| SA194017 | KR3M2 | KR |
| SA194018 | KR2M2 | KR |
| SA194019 | KR1M2 | KR |
| SA194020 | KR4M2 | KR |
| SA194021 | KR2M0 | KR |
| SA194022 | KR1M0 | KR |
| SA194023 | KR3M0 | KR |
| SA194024 | KR5M2 | KR |
| SA194025 | WT5M2 | WT |
| SA194026 | WT3M2 | WT |
| SA194027 | WT3M0 | WT |
| SA194028 | WT2M0 | WT |
| SA194029 | WT1M2 | WT |
| SA194030 | WT2M2 | WT |
| SA194031 | WT1M0 | WT |
| SA194032 | WT4M2 | WT |
| Showing results 1 to 16 of 16 |
Collection:
| Collection ID: | CO002136 |
| Collection Summary: | The labelled cells were collected and washed with 2 mL of PBS, and then were quenched with 2 mL methanol (-20℃). To increase the polarity for the two phase extraction, 0.4 mL of 4℃ cold water was added. |
| Sample Type: | Macrophages |
Treatment:
| Treatment ID: | TR002155 |
| Treatment Summary: | Method was described in previous studies (Jha et al., 2015; Lauterbach et al., 2019). 107 BMDMs per group were seeded in 10cm plates and incubated in RPMI-1640 cell culture medium with 10% FBS. Prior to isotopic labeling, the medium was replaced with RPMI-1640 without glutamine for 4 hrs. Then stable-isotope labeled analog 4 mM (U-13C5) glutamine (Cambridge Isotope) was added together with IL-4 (20ng/ml) for 4 hrs. |
Sample Preparation:
| Sampleprep ID: | SP002149 |
| Sampleprep Summary: | The samples were processed by 5 cycles of 1 min ultra-sonication and 1 min interval in ice-water bath. After that samples were stood for 30 min at -40℃ and 10 min at 4 ℃. After centrifugation at 15000 g and 4℃ for 15 min. The supernatant was evaporated to dryness under mild nitrogen and reconstituted in 50 L of 50% acetonitrile (including 1g/mL phenylalanine-d5 internal standard) prior to perform further analysis. |
Combined analysis:
| Analysis ID | AN003360 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | ThermoFisher Ultimate 3000 UHPLC system |
| Column | Waters BEH Amide |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive HF hybrid Orbitrap |
| Ion Mode | NEGATIVE |
| Units | pmoles/I |
Chromatography:
| Chromatography ID: | CH002486 |
| Chromatography Summary: | Chromatographic separation was performed on a ThermoFisher Ultimate 3000 UHPLC system with a Waters BEH Amide column (2.1mm × 100 mm, 1.7 μm). The injection volume was 2μL and the flow rate was 0.25 mL/min. The column temperature was 15°C. The mobile phases consisted of water with 0.01% formic acid and 2 mM ammonium formate (phase A) and acetonitrile (phase B). A linear gradient elution was performed with the following program: 0 min, 90%B; 4 min, 85% B; 11 min, 75%B; 14 min, 70%B, 14.5min, 50%B and held to 17 min; 17.1 min, 90%B and held to 20.01 min. |
| Instrument Name: | ThermoFisher Ultimate 3000 UHPLC system |
| Column Name: | Waters BEH Amide |
| Column Temperature: | 15 |
| Flow Gradient: | 0 min, 90%B; 4 min, 85% B; 11 min, 75%B; 14 min, 70%B, 14.5min, 50%B and held to 17 min; 17.1 min, 90%B and held to 20.01 min. |
| Flow Rate: | 0.25ml/min |
| Solvent A: | 100% water; 0.01% formic acid; 2 mM ammonium formate |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS003129 |
| Analysis ID: | AN003360 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The eluents were analyzed on a ThermoFisher Q Exactive™ Hybrid Quadrupole-Orbitrap™ Mass Spectrometry (QE) in Heated Electrospray Ionization Negative (HESI-) mode, separately. Spray voltage was set to 4000 V. Capillary and Probe Heater Temperature were separately 320 °C and 320 °C. Sheath gas flow rate was 35 (Arb, arbitrary unit), and Aux gas flow rate was 10 (Arb). S-Lens RF Level was 50 (Arb). The full scan was operated at a high-resolution of 70000 FWHM (m/z=200) at a range of 70- 1050 m/z with AGC Target setting at 3×106. |
| Ion Mode: | NEGATIVE |
