Summary of Study ST002922

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 PR001814. The data can be accessed directly via it's Project DOI: 10.21228/M87D9M 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	ST002922
Study Title	Effects of DINCH and MINCH on adipocyte metabolism of human SGBS cells.
Study Summary	In the first part of the project, we investigated the effects of DINCH and MINCH on central carbon metabolism. For this purpose, the human SGBS preadipocyte cell line (Wabitsch et al., 2001) was exposed to DINCH and MINCH at concentrations ranging from 10 nM to 10 µM and compared with cells differentiated with rosiglitazone (adipogenic reference) and without rosiglitazone (undifferentiated control). Analysis of central carbon metabolism showed that MINCH, similar to rosiglitazone, induces lipid accumulation mainly through PPARG-mediated upregulation of the pyruvate cycle. In addition, increased lactate production suggests altered glucose homeostasis induced by MINCH-treatment. Our results suggest that MINCH could potentially lead to a weight-promoting effect, as observed with thiazolidinediones, because of the similarity of the observed changes to the effects of the thiazolidinedione rosiglitazone.
Institute	Helmholtz Centre for Environmental Research
Department	Molecular Systems Biology
Last Name	Engelmann
First Name	Beatrice
Address	Permoserstraße 15, Leipzipg, Saxony, 03418, Germany
Email	beatrice.engelmann@ufz.de
Phone	00493412351099
Submit Date	2023-10-04
Raw Data Available	Yes
Raw Data File Type(s)	wiff
Analysis Type Detail	LC-MS
Release Date	2023-11-03
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR001814
Project DOI:	doi: 10.21228/M87D9M
Project Title:	MINCH causes metabolic rewiring towards lipid accumulation and adipogenesis
Project Summary:	Humans are ubiquitously exposed to plastic additives, including plasticizers. There is growing evidence that exposure to certain plasticizers is associated with the development of obesity due to their metabolism-disrupting properties. Following the restriction of the use of the phthalate plasticizer di-(2-ethylhexyl) phthalate (DEHP) due to its adverse health effects, it has been replaced by new substitutes such as the plasticizer diisononylcyclohexane-1,2-dicarboxylate (DINCH). Despite recent studies suggesting that the primary metabolite monoisononylcyclohexane-1,2-dicarboxylic acid ester (MINCH) promotes human adipocyte differentiation, the adipogenic properties of MINCH remain controversial. Because the metabolome largely reflects the molecular phenotype and is sensitive to perturbation by external factors, we used targeted metabolomics to investigate the effects of DINCH and MINCH on key metabolic pathways of adipocytes. Analysis of central carbon metabolism is particularly relevant because it provides cellular energy through the degradation of organic compounds and metabolic precursors for anabolic functions that are critical for adipocyte function, such as de novo lipogenesis. The project consists of three main studies: analysis of the effects of DINCH and MINCH on central carbon metabolism of human SGSB cells, analysis of the insulin response of DINCH- and MINCH-treated SGSB cells, and analysis of the effects of DINCH and MINCH on central carbon metabolism of human SGSB cells in the presence of the PPARG inhibitor GW9662.
Institute:	Helmholtz Centre for Environmental Research
Last Name:	Engelmann
First Name:	Beatrice
Address:	Permoserstr. 15
Email:	beatrice.engelmann@ufz.de
Phone:	00493412351099

Subject:

Subject ID:	SU003035
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Gender:	Male

Factors:

Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id	local_sample_id	Treatment
SA317308	Ctrl_1	Control
SA317309	Ctrl_4	Control
SA317310	Ctrl_3	Control
SA317311	Ctrl_2	Control
SA317312	Ctrl_SN2	Control supernatant
SA317313	Ctrl_SN3	Control supernatant
SA317314	Ctrl_SN1	Control supernatant
SA317315	Ctrl_SN4	Control supernatant
SA317316	DINCH_100nM_2	DINCH 100nM
SA317317	DINCH_100nM_3	DINCH 100nM
SA317318	DINCH_100nM_1	DINCH 100nM
SA317319	DINCH_100nM_4	DINCH 100nM
SA317320	DINCH_100nM_SN3	DINCH 100nM supernatant
SA317321	DINCH_100nM_SN2	DINCH 100nM supernatant
SA317322	DINCH_100nM_SN1	DINCH 100nM supernatant
SA317323	DINCH_100nM_SN4	DINCH 100nM supernatant
SA317332	DINCH_10uM_2	DINCH 10µM
SA317333	DINCH_10uM_3	DINCH 10µM
SA317334	DINCH_10uM_4	DINCH 10µM
SA317335	DINCH_10uM_1	DINCH 10µM
SA317336	DINCH_10uM_SN1	DINCH 10µM supernatant
SA317337	DINCH_10uM_SN3	DINCH 10µM supernatant
SA317338	DINCH_10uM_SN2	DINCH 10µM supernatant
SA317339	DINCH_10uM_SN4	DINCH 10µM supernatant
SA317324	DINCH_10nM_2	DINCH 10nM
SA317325	DINCH_10nM_1	DINCH 10nM
SA317326	DINCH_10nM_3	DINCH 10nM
SA317327	DINCH_10nM_4	DINCH 10nM
SA317328	DINCH_10nM_SN3	DINCH 10nM supernatant
SA317329	DINCH_10nM_SN2	DINCH 10nM supernatant
SA317330	DINCH_10nM_SN4	DINCH 10nM supernatant
SA317331	DINCH_10nM_SN1	DINCH 10nM supernatant
SA317340	DINCH_1uM_2	DINCH 1µM
SA317341	DINCH_1uM_4	DINCH 1µM
SA317342	DINCH_1uM_3	DINCH 1µM
SA317343	DINCH_1uM_1	DINCH 1µM
SA317344	DINCH_1uM_SN3	DINCH 1µM supernatant
SA317345	DINCH_1uM_SN2	DINCH 1µM supernatant
SA317346	DINCH_1uM_SN4	DINCH 1µM supernatant
SA317347	DINCH_1uM_SN1	DINCH 1µM supernatant
SA317384	MINCH_10uM_blank3	Medium blank supernatant
SA317385	MINCH_10uM_blank2	Medium blank supernatant
SA317386	MINCH_10uM_blank1	Medium blank supernatant
SA317348	MINCH_100nM_4	MINCH 100nM
SA317349	MINCH_100nM_2	MINCH 100nM
SA317350	MINCH_100nM_1	MINCH 100nM
SA317351	MINCH_100nM_3	MINCH 100nM
SA317352	MINCH_100nM_SN4	MINCH 100nM supernatant
SA317353	MINCH_100nM_SN2	MINCH 100nM supernatant
SA317354	MINCH_100nM_SN3	MINCH 100nM supernatant
SA317355	MINCH_100nM_SN1	MINCH 100nM supernatant
SA317364	MINCH_10uM_2	MINCH 10µM
SA317365	MINCH_10uM_3	MINCH 10µM
SA317366	MINCH_10uM_4	MINCH 10µM
SA317367	MINCH_10uM_1	MINCH 10µM
SA317368	MINCH_10uM_SN2	MINCH 10µM supernatant
SA317369	MINCH_10uM_SN3	MINCH 10µM supernatant
SA317370	MINCH_10uM_SN4	MINCH 10µM supernatant
SA317371	MINCH_10uM_SN1	MINCH 10µM supernatant
SA317356	MINCH_10nM_3	MINCH 10nM
SA317357	MINCH_10nM_1	MINCH 10nM
SA317358	MINCH_10nM_4	MINCH 10nM
SA317359	MINCH_10nM_2	MINCH 10nM
SA317360	MINCH_10nM_SN3	MINCH 10nM supernatant
SA317361	MINCH_10nM_SN4	MINCH 10nM supernatant
SA317362	MINCH_10nM_SN2	MINCH 10nM supernatant
SA317363	MINCH_10nM_SN1	MINCH 10nM supernatant
SA317372	MINCH_1uM_4	MINCH 1µM
SA317373	MINCH_1uM_2	MINCH 1µM
SA317374	MINCH_1uM_1	MINCH 1µM
SA317375	MINCH_1uM_3	MINCH 1µM
SA317376	MINCH_1uM_SN4	MINCH 1µM supernatant
SA317377	MINCH_1uM_SN2	MINCH 1µM supernatant
SA317378	MINCH_1uM_SN3	MINCH 1µM supernatant
SA317379	MINCH_1uM_SN1	MINCH 1µM supernatant
SA317380	MINCH_GW_SN2	MINCH GW supernatant
SA317381	MINCH_GW_SN4	MINCH GW supernatant
SA317382	MINCH_GW_SN1	MINCH GW supernatant
SA317383	MINCH_GW_SN3	MINCH GW supernatant
SA317387	Rosi_4	Rosiglitazone
SA317388	Rosi_3	Rosiglitazone
SA317389	Rosi_2	Rosiglitazone
SA317390	Rosi_1	Rosiglitazone
SA317391	Rosi_SN1	Rosiglitazone supernatant
SA317392	Rosi_SN2	Rosiglitazone supernatant
SA317393	Rosi_SN3	Rosiglitazone supernatant
SA317394	Rosi_SN4	Rosiglitazone supernatant

Showing results 1 to 87 of 87

Showing results 1 to 87 of 87

Collection:

Collection ID:	CO003028
Collection Summary:	The SGBS cells were obtained from Prof. Martin Wabitsch laboratory at the University Clinic Ulm. SGBS preadipocytes were differentiated according to the standard protocol described previously (Wabitsch et al., 2001).
Sample Type:	Adipose tissue
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR003044
Treatment Summary:	SGSB preadipocytes were maintained at 37°C and 5% CO2 in 95% humidity. To investigate the effects of the plasticizer DINCH and its primary metabolite MINCH on adipocyte differentiation, SGBS preadipocytes were treated for 12 days with differentiation media without the PPARG agonist rosiglitazone supplemented with DINCH or MINCH (10 nM, 100 nM, 1 µM, and 10 µM). To obtain an adipogenesis reference, SGBS cells were differentiated in the presence of rosiglitazone; to obtain an untreated control, they were differentiated in the absence of rosiglitazone. A final concentration of 0.01% (v/v) MeOH and 0.02% (v/v) DMSO was added to all conditioned differentiation media. Continuous exposure was mimicked by replacing the cell culture medium every second day. Each treatment was performed in four biological replicates (n=4).

Treatment ID:

TR003044

Treatment Summary:

SGSB preadipocytes were maintained at 37°C and 5% CO2 in 95% humidity. To investigate the effects of the plasticizer DINCH and its primary metabolite MINCH on adipocyte differentiation, SGBS preadipocytes were treated for 12 days with differentiation media without the PPARG agonist rosiglitazone supplemented with DINCH or MINCH (10 nM, 100 nM, 1 µM, and 10 µM). To obtain an adipogenesis reference, SGBS cells were differentiated in the presence of rosiglitazone; to obtain an untreated control, they were differentiated in the absence of rosiglitazone. A final concentration of 0.01% (v/v) MeOH and 0.02% (v/v) DMSO was added to all conditioned differentiation media. Continuous exposure was mimicked by replacing the cell culture medium every second day. Each treatment was performed in four biological replicates (n=4).

Sample Preparation:

Sampleprep ID:	SP003041
Sampleprep Summary:	Extraction of intracellular and extracellular metabolites was performed by a 1:1:1 methanol:water:chloroform extraction protocol. For the extraction of intracellular metabolites, the culture medium was removed and the cells were rinsed twice with 1 ml of 0.9% ice-cold NaCl. The rinsing solution was removed, and the metabolism of the cells was stopped by adding 400 µL of MeOH (-20 °C) followed by 400 µL of ice-cold H2O containing 10 µM d6-glutarate. Cells were collected using a cell lifter and 400 µL of chloroform was added. After shaking at 1,400 rpm and 4 °C for 20 min, the extraction mixture was centrifuged at 18,000 g and 4 °C for 5 min. Subsequently, 300 µL volume of the polar upper phase was collected and evaporated to complete dryness. For the extraction of extracellular metabolites, 300 µL of the supernatant was extracted by adding 400 µL MeOH (-20 °C) containing 100 nM MEHP, 100 µL ice-cold H2O containing 40 µM d6-glutarate, and 400 µL chloroform (-20 °C). Subsequent sample preparation was identical to the extraction of intracellular metabolites. Note: After measurement of the samples by LC-MS, the raw AUC values uploaded here were normalized to the internal standard (d6-glutarate, if applicable) and DNA content per well (measured by DAPI fluorescence). After normalization, log2 fold changes were calculated by dividing the normalized peak area from each replicate of each treatment by the normalized peak area from each control. Insulin data were not normalized to DAPI because fold changes were calculated by dividing the intensities of the insulin-stimulated cells by the noninsulin-stimulated cells from each treatment.

Sampleprep ID:

SP003041

Sampleprep Summary:

Extraction of intracellular and extracellular metabolites was performed by a 1:1:1 methanol:water:chloroform extraction protocol. For the extraction of intracellular metabolites, the culture medium was removed and the cells were rinsed twice with 1 ml of 0.9% ice-cold NaCl. The rinsing solution was removed, and the metabolism of the cells was stopped by adding 400 µL of MeOH (-20 °C) followed by 400 µL of ice-cold H2O containing 10 µM d6-glutarate. Cells were collected using a cell lifter and 400 µL of chloroform was added. After shaking at 1,400 rpm and 4 °C for 20 min, the extraction mixture was centrifuged at 18,000 g and 4 °C for 5 min. Subsequently, 300 µL volume of the polar upper phase was collected and evaporated to complete dryness. For the extraction of extracellular metabolites, 300 µL of the supernatant was extracted by adding 400 µL MeOH (-20 °C) containing 100 nM MEHP, 100 µL ice-cold H2O containing 40 µM d6-glutarate, and 400 µL chloroform (-20 °C). Subsequent sample preparation was identical to the extraction of intracellular metabolites. Note: After measurement of the samples by LC-MS, the raw AUC values uploaded here were normalized to the internal standard (d6-glutarate, if applicable) and DNA content per well (measured by DAPI fluorescence). After normalization, log2 fold changes were calculated by dividing the normalized peak area from each replicate of each treatment by the normalized peak area from each control. Insulin data were not normalized to DAPI because fold changes were calculated by dividing the intensities of the insulin-stimulated cells by the noninsulin-stimulated cells from each treatment.

Combined analysis:

Analysis ID	AN004792
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Agilent 1290 Infinity II
Column	Waters XSelect XP HSS T3 (150 x 2.1mm, 2.5um)
MS Type	ESI
MS instrument type	Triple quadrupole
MS instrument name	ABI Sciex 6500+
Ion Mode	NEGATIVE
Units	Peak AUC

Chromatography:

Chromatography ID:	CH003623
Instrument Name:	Agilent 1290 Infinity II
Column Name:	Waters XSelect XP HSS T3 (150 x 2.1mm, 2.5um)
Column Temperature:	40
Flow Gradient:	0-5 min 0% B, 5-9 min 0%- 2% B, 9-9.5 min 2-6% B, 9.5-11.5 min 6% B, 11.5-12 min 6-11% B, 12-13.5 min 11% B, 13.5-15.5 min 11-28% B, 15.5-16.5 min 28-53% B, 16.5-22.5 53% B, 22.5-23 min 53-0% B, 23-33 min 0% B
Flow Rate:	0-15.5 min 0.4 mL/min, 15.5-16.5 min 0.4-0.15 mL/min, 16.5-23 min 0.15 mL/min, 23-27 min 0.15-0.4 mL/min, 27-33 min 0.4 mL/min
Solvent A:	10mM TBA, 10mM acetic acid, 5% MeOH, 2% IPA in water
Solvent B:	100% IPA
Chromatography Type:	Reversed phase

MS:

MS ID:	MS004538
Analysis ID:	AN004792
Instrument Name:	ABI Sciex 6500+
Instrument Type:	Triple quadrupole
MS Type:	ESI
MS Comments:	For identification and quantitation, a scheduled MRM method was used, with specific transitions for every metabolite. Data acquisition and peak integration were performed in Analyst® software (Version 1.7.1).
Ion Mode:	NEGATIVE