Summary of Study ST004339

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 PR002751. The data can be accessed directly via it's Project DOI: 10.21228/M82G23 This work is supported by NIH grant, U2C- DK119886. See: https://www.metabolomicsworkbench.org/about/howtocite.php

Study ID	ST004339
Study Title	Glutathione-deficiency promotes basal hyperinsulinemia in the insulin secreting cell line INS-1 (832/13)
Study Summary	This study sought to determine the impact of Glutathione (GSH) synthesis on β-cell function. Glutamate-cysteine ligase catalytic subunit (Gclc) is essential for GSH synthesis. To induce GSH deficiency, GCLC was inhibited in INS-1 (832/13) cells using buthionine sulfoximine (BSO). The 24-hour treatment of INS-1 cells with BSO (250 µM) reduced GSH levels to 33% of untreated, however, glucose-stimulated insulin secretion was unchanged. The 24-hour treatment of INS-1 cells with BME-free medium containing BSO further reduced GSH levels to 9% of untreated and elicited significantly higher basal insulin secretion. We sought to determine if higher basal insulin secretion was related to metabolic alteration therefore, we performed a metabolomics screen. This data is presented here. Out of 53 identified metabolites, we found 6 with significantly lower abundance in -BME+BSO as compared to -BME. Those include, adenosine monophosphate, deoxycytidine monophosphate, glucose 1-phosphate, ADP-glucose, thymidine monophostphase, and deoxycytidine diphosphate.
Institute	Yale University
Last Name	Davidson
First Name	Emily
Address	300 Cedar St, S110, New Haven, CT 06510
Email	emily.davidson@yale.edu
Phone	8104797570
Submit Date	2025-11-06
Num Groups	2
Raw Data Available	Yes
Raw Data File Type(s)	mzML
Analysis Type Detail	LC-MS
Release Date	2026-01-02
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR002751
Project DOI:	doi: 10.21228/M82G23
Project Title:	Glutathione-deficiency promotes basal hyperinsulinemia in the insulin secreting cell line INS-1 (832/13)
Project Summary:	This study sought to determine the impact of Glutathione (GSH) synthesis on β-cell function. Glutamate-cysteine ligase catalytic subunit (Gclc) is essential for GSH synthesis. To induce GSH deficiency, GCLC was inhibited in INS-1 (832/13) cells using buthionine sulfoximine (BSO). The 24-hour treatment of INS-1 cells with BSO (250 µM) reduced GSH levels to 33% of untreated, however, glucose-stimulated insulin secretion was unchanged. The 24-hour treatment of INS-1 cells with BME-free medium containing BSO further reduced GSH levels to 9% of untreated and elicited significantly higher basal insulin secretion. We sought to determine if higher basal insulin secretion was related to metabolic alteration therefore, we performed a metabolomics screen. This data is presented here. Out of 53 identified metabolites, we found 6 with significantly lower abundance in -BME+BSO as compared to -BME. Those include, adenosine monophosphate, deoxycytidine monophosphate, glucose 1-phosphate, ADP-glucose, thymidine monophostphase, and deoxycytidine diphosphate.
Institute:	Yale University
Last Name:	Davidson
First Name:	Emily
Address:	300 Cedar St, S110, New Haven, CT 06510
Email:	emily.davidson@yale.edu
Phone:	8104797570

Subject:

Subject ID:	SU004498
Subject Type:	Cultured cells
Subject Species:	Rattus norvegicus
Taxonomy ID:	10116
Cell Biosource Or Supplier:	Millipore Sigma
Cell Strain Details:	INS-1 832/13

Factors:

Subject type: Cultured cells; Subject species: Rattus norvegicus (Factor headings shown in green)

mb_sample_id	local_sample_id	Treatment	Sample source	Injection order
SA509351	bb_7	BME-free (BSO)	INS-1 cells	10
SA509352	bb_6	BME-free (BSO)	INS-1 cells	11
SA509353	bb_10	BME-free (BSO)	INS-1 cells	3
SA509354	bb_8	BME-free (BSO)	INS-1 cells	4
SA509355	bb_9	BME-free (BSO)	INS-1 cells	7
SA509356	b_1	BME-free (control)	INS-1 cells	12
SA509357	b_3	BME-free (control)	INS-1 cells	13
SA509358	b_4	BME-free (control)	INS-1 cells	14
SA509359	b_2	BME-free (control)	INS-1 cells	5
SA509360	b_5	BME-free (control)	INS-1 cells	6
SA509364	eb_1	extraction blank	media	1
SA509365	eb_3	extraction blank	media	16
SA509366	eb_2	extraction blank	media	9
SA509361	QC_10_20ev	MSe	INS-1 cells	17
SA509362	QC_10_30ev	MSe	INS-1 cells	18
SA509363	QC_20_40ev	MSe	INS-1 cells	19
SA509367	QC_3	quality control	INS-1 cells	15
SA509368	QC_1	quality control	INS-1 cells	2
SA509369	QC_2	quality control	INS-1 cells	8

Showing results 1 to 19 of 19

Showing results 1 to 19 of 19

Collection:

Collection ID:	CO004491
Collection Summary:	INS-1 (832/13) cells (#SCC207, Sigma-Aldrich) were cultured as monolayers in RPMI 1640 medium (#11875, Gibco™) supplemented with 10% (v/v) fetal bovine serum (#16140-071, Gibco™), antibiotics (10,000 units/mL penicillin and 10 mg/mL streptomycin) (#15140122, Gibco™), 10mM HEPES (#15630-080, Gibco™), 2 mM L-glutamine (#35050-061, Gibco™), 1 mM sodium pyruvate (#11360-070, Gibco™), and 50 µM β-mercaptoethanol (BME) (#M6250, Sigma-Aldrich).
Sample Type:	INS-1 cells

Treatment:

Treatment ID:	TR004507
Treatment Summary:	INS-1 cells (~2.5 million cells per dish) were seeded into 100 mm tissue culture-treated dishes (#0877222, Corning). After overnight incubation, +BME medium was aspirated and replaced with 10 mL of experimental medium (-BME or -BME+BSO medium) for 24 hours. Prior to metabolite extraction, treated cells underwent a glucose-stimulated insulin secretion assay (2.5mM glucose).

Sample Preparation:

Sampleprep ID:	SP004504
Sampleprep Summary:	For extraction, cells were washed 2x with 1 mL cold DBPS to remove culture medium and scrapped from the dish in 1 mL ice-cold acetonitrile:methanol:water (2:2:1 %v/v/v) and transferred to 1.5 mL centrifuge tubes. Cells were lysed through three snap-freeze thaw cycles, following centrifugation at 15,000 RPM for 10 minutes at 4ºC. The supernatants containing soluble metabolites were transferred to clean 1.5 mL centrifuge tubes and evaporated in a vacuum concentrator (SPD111V#, Savant) The remaining pellets were processed for total protein quantification using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). Dry extracts were reconstituted in 100 μL acetonitrile:water (1:1, %v/v) containing 10 μL of IROA Internal Standard U-13C, 95% (IROA® TrueQuant IQQ Kit, IROA Technologies™) and centrifuged (10,000 RPM) for 10 minutes at 4°C (to remove insoluble debris). One hundred μL of the supernatant was transferred into liquid chromatography-mass spectrometry vials (TrueView LC–MS Certified, Waters Corporation) for liquid chromatography-mass spectrometry analysis. Quality Control (QC) samples: 80 μL of the sample supernatant was removed before the evaporation step. The aliquots were all pooled, sample volume matched, and aliquots were evaporated to dryness, stored, and reconstituted as described above.

Sampleprep ID:

SP004504

Sampleprep Summary:

For extraction, cells were washed 2x with 1 mL cold DBPS to remove culture medium and scrapped from the dish in 1 mL ice-cold acetonitrile:methanol:water (2:2:1 %v/v/v) and transferred to 1.5 mL centrifuge tubes. Cells were lysed through three snap-freeze thaw cycles, following centrifugation at 15,000 RPM for 10 minutes at 4ºC. The supernatants containing soluble metabolites were transferred to clean 1.5 mL centrifuge tubes and evaporated in a vacuum concentrator (SPD111V#, Savant) The remaining pellets were processed for total protein quantification using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). Dry extracts were reconstituted in 100 μL acetonitrile:water (1:1, %v/v) containing 10 μL of IROA Internal Standard U-13C, 95% (IROA® TrueQuant IQQ Kit, IROA Technologies™) and centrifuged (10,000 RPM) for 10 minutes at 4°C (to remove insoluble debris). One hundred μL of the supernatant was transferred into liquid chromatography-mass spectrometry vials (TrueView LC–MS Certified, Waters Corporation) for liquid chromatography-mass spectrometry analysis. Quality Control (QC) samples: 80 μL of the sample supernatant was removed before the evaporation step. The aliquots were all pooled, sample volume matched, and aliquots were evaporated to dryness, stored, and reconstituted as described above.

Combined analysis:

Analysis ID	AN007246
Chromatography ID	CH005500
MS ID	MS006940
Analysis type	MS
Chromatography type	HILIC
Chromatography system	Waters Acquity I-Class
Column	Waters ACQUITY UPLC BEH Amide (100 x 2.1mm,1.7um)
MS Type	ESI
MS instrument type	QTOF
MS instrument name	Waters Xevo-G2-XS
Ion Mode	NEGATIVE
Units	abundance

Chromatography:

Chromatography ID:	CH005500
Instrument Name:	Waters Acquity I-Class
Column Name:	Waters ACQUITY UPLC BEH Amide (100 x 2.1mm,1.7um)
Column Temperature:	30
Flow Gradient:	95% B (0–0.5 min), 95%-65% B (0.5–7 min), 65–40% B (7–8 min), 40% B (8–9 min), 40–95% B (9–9.1 min) and continuing at 95% B (9.1–12.0 min)
Flow Rate:	0.5mL/min
Solvent A:	100% water; 25 mM ammonium hydroxide; 25 mM ammonium acetate
Solvent B:	100% acetonitrile
Chromatography Type:	HILIC

MS:

MS ID:	MS006940
Analysis ID:	AN007246
Instrument Name:	Waters Xevo-G2-XS
Instrument Type:	QTOF
MS Type:	ESI
MS Comments:	Q-ToF–MS scan data (300 ms/scan; mass scan range 50–1200 Da) were first acquired for each sample. Thereafter, MSe fragmentation data were acquired for metabolite identification (low energy scan: 200 ms/scan, collision energy 6 eV; high energy scan: 100 ms/scan, collision energy 10, 20, 30 and 40 eV, mass scan range 25–1000 Da). ESI source parameters were as follows: 1.8 kV capillary voltage, 40 V sampling cone, 50°C source temperature, 420°C desolvation temperature, 80 L/hr cone gas flow, 850 L/hr desolvation gas flow
Ion Mode:	NEGATIVE