{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002061","ANALYSIS_ID":"AN003360","VERSION":"1","CREATED_ON":"01-20-2022"},

"PROJECT":{"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","DOI":"http://dx.doi.org/10.21228/M81H6N","CONTRIBUTORS":"Wei Zhou"},

"STUDY":{"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","EMAIL":"joesphchou@alumni.sjtu.edu.cn","PHONE":"+8615216716293","SUBMIT_DATE":"2022-01-19"},

"SUBJECT":{"SUBJECT_TYPE":"Cultured cells","SUBJECT_SPECIES":"Mus musculus"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"KR1M0",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR1M0.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR1M2",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR1M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR2M0",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR2M0.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR2M2",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR2M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR3M0",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR3M0.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR3M2",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR3M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR4M2",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR4M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"KR5M2",
"Factors":{"Genotype":"KR"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"KR5M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT1M0",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT1M0.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT1M2",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT1M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT2M0",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT2M0.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT2M2",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT2M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT3M0",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT3M0.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT3M2",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT3M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT4M2",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT4M2.mzXML"}
},
{
"Subject ID":"-",
"Sample ID":"WT5M2",
"Factors":{"Genotype":"WT"},
"Additional sample data":{"time":"4hrs","treatment":"(U-13C5) glutamine(4mM)","RAW_FILE_NAME":"WT5M2.mzXML"}
}
],
"COLLECTION":{"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_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."},

"SAMPLEPREP":{"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 1g/mL phenylalanine-d5 internal standard) prior to perform further analysis."},

"CHROMATOGRAPHY":{"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","CHROMATOGRAPHY_TYPE":"HILIC"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"ThermoFisher Q Exactive™ Hybrid Quadrupole-Orbitrap™ Mass Spectrometry (QE)","INSTRUMENT_TYPE":"Hybrid Quadrupole-Orbitrap™","MS_TYPE":"Other","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"},

"MS_METABOLITE_DATA":{
"Units":"pmoles/I",

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