Summary of Study ST002919

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	ST002919
Study Title	Short-term metabolic insulin response of DINCH- and MINCH-treated cells
Study Summary	In the second part of the project, we examined the short-term metabolic insulin response of DINAH- and MINCH-treated cells and compared them with rosiglitazone-differentiated cells. For this purpose, the human SGBS preadipocyte cell line was exposed to differentiation media conditioned with DINCH (10 nM or 10 µM), MINCH (10 nM or 10 µM), or rosiglitazone, and the insulin response was measured by analyzing the changes in glycolysis and PPP between insulin-stimulated and non-insulin-stimulated cells. In conclusion, the insulin response of glycolysis and PPP of cells treated with 10 µM MINCH, but not with 10 nM MINCH or DINCH, was similar to cells differentiated with rosiglitazone.
Institute	Helmholtz Centre for Environmental Research
Last Name	Engelmann
First Name	Beatrice
Address	Permoserstr. 15
Email	beatrice.engelmann@ufz.de
Phone	00493412351099
Submit Date	2023-10-08
Raw Data Available	Yes
Raw Data File Type(s)	wiff
Analysis Type Detail	LC-MS
Release Date	2023-11-01
Release Version	1

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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of the use of the phthalate plasticizer di-(2-ethylhexyl) phthalate (DEHP) due to its adverse health effects, it has been replaced by new substitutes such as the plasticizer diisononylcyclohexane-1,2-dicarboxylate (DINCH).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:	SU003032
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	Replicate
SA316808	MINCH_High_-_1	DINCH 10µM minus Insulin	1
SA316809	DINCH_High_-_1	DINCH 10µM minus Insulin	1
SA316810	DINCH_High_-_2	DINCH 10µM minus Insulin	2
SA316811	MINCH_High_-_2	DINCH 10µM minus Insulin	2
SA316812	MINCH_High_-_3	DINCH 10µM minus Insulin	3
SA316813	DINCH_High_-_3	DINCH 10µM minus Insulin	3
SA316814	MINCH_High_-_4	DINCH 10µM minus Insulin	4
SA316815	DINCH_High_-_4	DINCH 10µM minus Insulin	4
SA316816	MINCH_High_-_5	DINCH 10µM minus Insulin	5
SA316817	DINCH_High_-_5	DINCH 10µM minus Insulin	5
SA316798	MINCH_High_SN_-_1	DINCH 10µM minus Insulin supernatant	1
SA316799	DINCH_High_SN_-_1	DINCH 10µM minus Insulin supernatant	1
SA316800	DINCH_High_SN_-_2	DINCH 10µM minus Insulin supernatant	2
SA316801	MINCH_High_SN_-_2	DINCH 10µM minus Insulin supernatant	2
SA316802	DINCH_High_SN_-_3	DINCH 10µM minus Insulin supernatant	3
SA316803	MINCH_High_SN_-_3	DINCH 10µM minus Insulin supernatant	3
SA316804	DINCH_High_SN_-_4	DINCH 10µM minus Insulin supernatant	4
SA316805	MINCH_High_SN_-_4	DINCH 10µM minus Insulin supernatant	4
SA316806	DINCH_High_SN_-_5	DINCH 10µM minus Insulin supernatant	5
SA316807	MINCH_High_SN_-_5	DINCH 10µM minus Insulin supernatant	5
SA316828	DINCH_High_+_1	DINCH 10µM plus Insulin	1
SA316829	MINCH_High_+_1	DINCH 10µM plus Insulin	1
SA316830	DINCH_High_+_2	DINCH 10µM plus Insulin	2
SA316831	MINCH_High_+_2	DINCH 10µM plus Insulin	2
SA316832	MINCH_High_+_3	DINCH 10µM plus Insulin	3
SA316833	DINCH_High_+_3	DINCH 10µM plus Insulin	3
SA316834	MINCH_High_+_4	DINCH 10µM plus Insulin	4
SA316835	DINCH_High_+_4	DINCH 10µM plus Insulin	4
SA316836	DINCH_High_+_5	DINCH 10µM plus Insulin	5
SA316837	MINCH_High_+_5	DINCH 10µM plus Insulin	5
SA316818	MINCH_High_SN_+_1	DINCH 10µM plus Insulin supernatant	1
SA316819	DINCH_High_SN_+_1	DINCH 10µM plus Insulin supernatant	1
SA316820	MINCH_High_SN_+_2	DINCH 10µM plus Insulin supernatant	2
SA316821	DINCH_High_SN_+_2	DINCH 10µM plus Insulin supernatant	2
SA316822	MINCH_High_SN_+_3	DINCH 10µM plus Insulin supernatant	3
SA316823	DINCH_High_SN_+_3	DINCH 10µM plus Insulin supernatant	3
SA316824	MINCH_High_SN_+_4	DINCH 10µM plus Insulin supernatant	4
SA316825	DINCH_High_SN_+_4	DINCH 10µM plus Insulin supernatant	4
SA316826	DINCH_High_SN_+_5	DINCH 10µM plus Insulin supernatant	5
SA316827	MINCH_High_SN_+_5	DINCH 10µM plus Insulin supernatant	5
SA316768	DINCH_Low_-_1	DINCH 10nM minus Insulin	1
SA316769	MINCH_Low_-_1	DINCH 10nM minus Insulin	1
SA316770	MINCH_Low_-_2	DINCH 10nM minus Insulin	2
SA316771	DINCH_Low_-_2	DINCH 10nM minus Insulin	2
SA316772	MINCH_Low_-_3	DINCH 10nM minus Insulin	3
SA316773	DINCH_Low_-_3	DINCH 10nM minus Insulin	3
SA316774	MINCH_Low_-_4	DINCH 10nM minus Insulin	4
SA316775	DINCH_Low_-_4	DINCH 10nM minus Insulin	4
SA316776	MINCH_Low_-_5	DINCH 10nM minus Insulin	5
SA316777	DINCH_Low_-_5	DINCH 10nM minus Insulin	5
SA316758	DINCH_Low_SN_-_1	DINCH 10nM minus Insulin supernatant	1
SA316759	MINCH_Low_SN_-_1	DINCH 10nM minus Insulin supernatant	1
SA316760	DINCH_Low_SN_-_2	DINCH 10nM minus Insulin supernatant	2
SA316761	MINCH_Low_SN_-_2	DINCH 10nM minus Insulin supernatant	2
SA316762	DINCH_Low_SN_-_3	DINCH 10nM minus Insulin supernatant	3
SA316763	MINCH_Low_SN_-_3	DINCH 10nM minus Insulin supernatant	3
SA316764	DINCH_Low_SN_-_4	DINCH 10nM minus Insulin supernatant	4
SA316765	MINCH_Low_SN_-_4	DINCH 10nM minus Insulin supernatant	4
SA316766	MINCH_Low_SN_-_5	DINCH 10nM minus Insulin supernatant	5
SA316767	DINCH_Low_SN_-_5	DINCH 10nM minus Insulin supernatant	5
SA316788	MINCH_Low_+_1	DINCH 10nM plus Insulin	1
SA316789	DINCH_Low_+_1	DINCH 10nM plus Insulin	1
SA316790	DINCH_Low_+_2	DINCH 10nM plus Insulin	2
SA316791	MINCH_Low_+_2	DINCH 10nM plus Insulin	2
SA316792	DINCH_Low_+_3	DINCH 10nM plus Insulin	3
SA316793	MINCH_Low_+_3	DINCH 10nM plus Insulin	3
SA316794	DINCH_Low_+_4	DINCH 10nM plus Insulin	4
SA316795	MINCH_Low_+_4	DINCH 10nM plus Insulin	4
SA316796	MINCH_Low_+_5	DINCH 10nM plus Insulin	5
SA316797	DINCH_Low_+_5	DINCH 10nM plus Insulin	5
SA316778	DINCH_Low_SN_+_1	DINCH 10nM plus Insulin supernatant	1
SA316779	MINCH_Low_SN_+_1	DINCH 10nM plus Insulin supernatant	1
SA316780	DINCH_Low_SN_+_2	DINCH 10nM plus Insulin supernatant	2
SA316781	MINCH_Low_SN_+_2	DINCH 10nM plus Insulin supernatant	2
SA316782	MINCH_Low_SN_+_3	DINCH 10nM plus Insulin supernatant	3
SA316783	DINCH_Low_SN_+_3	DINCH 10nM plus Insulin supernatant	3
SA316784	DINCH_Low_SN_+_4	DINCH 10nM plus Insulin supernatant	4
SA316785	MINCH_Low_SN_+_4	DINCH 10nM plus Insulin supernatant	4
SA316786	MINCH_Low_SN_+_5	DINCH 10nM plus Insulin supernatant	5
SA316787	DINCH_Low_SN_+_5	DINCH 10nM plus Insulin supernatant	5
SA316843	Rosi_-_1	Rosiglitazone minus insulin	1
SA316844	Rosi_-_2	Rosiglitazone minus insulin	2
SA316845	Rosi_-_3	Rosiglitazone minus insulin	3
SA316846	Rosi_-_4	Rosiglitazone minus insulin	4
SA316847	Rosi_-_5	Rosiglitazone minus insulin	5
SA316838	Rosi_SN_-_1	Rosiglitazone minus insulin supernatant	1
SA316839	Rosi_SN_-_2	Rosiglitazone minus insulin supernatant	2
SA316840	Rosi_SN_-_3	Rosiglitazone minus insulin supernatant	3
SA316841	Rosi_SN_-_4	Rosiglitazone minus insulin supernatant	4
SA316842	Rosi_SN_-_5	Rosiglitazone minus insulin supernatant	5
SA316853	Rosi_+_1	Rosiglitazone plus insulin	1
SA316854	Rosi_+_2	Rosiglitazone plus insulin	2
SA316855	Rosi_+_3	Rosiglitazone plus insulin	3
SA316856	Rosi_+_4	Rosiglitazone plus insulin	4
SA316857	Rosi_+_5	Rosiglitazone plus insulin	5
SA316848	Rosi_SN_+_1	Rosiglitazone plus insulin supernatant	1
SA316849	Rosi_SN_+_2	Rosiglitazone plus insulin supernatant	2
SA316850	Rosi_SN_+_3	Rosiglitazone plus insulin supernatant	3
SA316851	Rosi_SN_+_4	Rosiglitazone plus insulin supernatant	4
SA316852	Rosi_SN_+_5	Rosiglitazone plus insulin supernatant	5

Showing results 1 to 100 of 100

Showing results 1 to 100 of 100

Collection:

Collection ID:	CO003025
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:	TR003041
Treatment Summary:	SGSB preadipocytes were maintained at 37°C and 5% CO2 in 95% humidity. To examine the insulin response of SGBS treated with DINCH and MINCH, SGBS preadipocytes were treated for 18 days with differentiation media without the PPARG agonist rosiglitazone supplemented with DINCH or MINCH (10 nM and 10 µM). To obtain an adipogenesis reference, SGBS cells were differentiated with rosiglitazone for 18 days according to the standard differentiation protocol. After replacing the medium on day 17 with differentiation medium without insulin and overnight incubation (insulin starvation), cells were starved for 1 hour with differentiation medium without glucose and insulin (glucose + insulin starvation). Subsequently, cells were incubated with either differentiation medium with insulin (insulin-stimulated cells correspond to plus-insulin samples) or without insulin (unstimulated cells correspond to minus-insulin samples). During differentiation, a final concentration of 0.01% (v/v) MeOH and 0.02% (v/v) DMSO was added to all differentiation media. Continuous exposure was mimicked by replacing the cell culture medium every other day. Each treatment was performed in five biological replicates (n=5).

Treatment ID:

TR003041

Treatment Summary:

SGSB preadipocytes were maintained at 37°C and 5% CO2 in 95% humidity. To examine the insulin response of SGBS treated with DINCH and MINCH, SGBS preadipocytes were treated for 18 days with differentiation media without the PPARG agonist rosiglitazone supplemented with DINCH or MINCH (10 nM and 10 µM). To obtain an adipogenesis reference, SGBS cells were differentiated with rosiglitazone for 18 days according to the standard differentiation protocol. After replacing the medium on day 17 with differentiation medium without insulin and overnight incubation (insulin starvation), cells were starved for 1 hour with differentiation medium without glucose and insulin (glucose + insulin starvation). Subsequently, cells were incubated with either differentiation medium with insulin (insulin-stimulated cells correspond to plus-insulin samples) or without insulin (unstimulated cells correspond to minus-insulin samples). During differentiation, a final concentration of 0.01% (v/v) MeOH and 0.02% (v/v) DMSO was added to all differentiation media. Continuous exposure was mimicked by replacing the cell culture medium every other day. Each treatment was performed in five biological replicates (n=5).

Sample Preparation:

Sampleprep ID:	SP003038
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:

SP003038

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	AN004789
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Agilent 1290 Infinity II
Column	Waters Xselect XP HSS T3 (150 x 2.1mm, 2.5µm)
MS Type	ESI
MS instrument type	QTRAP
MS instrument name	ABI Sciex 6500+
Ion Mode	NEGATIVE
Units	Peak AUC

Chromatography:

Chromatography ID:	CH003620
Instrument Name:	Agilent 1290 Infinity II
Column Name:	Waters Xselect XP HSS T3 (150 x 2.1mm, 2.5µm)
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:	MS004535
Analysis ID:	AN004789
Instrument Name:	ABI Sciex 6500+
Instrument Type:	QTRAP
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