Corticosteroid production in H295R cells during exposure to 3 endocrine disrupters analyzed with LC-MS/MS

Christina S Winther; Frederik K Nielsen; Martin Hansen; Bjarne Styrishave

doi:10.1177/1091581813484366

Corticosteroid production in H295R cells during exposure to 3 endocrine disrupters analyzed with LC-MS/MS

Christina S Winther, Frederik K Nielsen, Martin Hansen, Bjarne Styrishave

14 Citations (Scopus)

Abstract

The adrenocortical human cell line H295R is a valuable tool for screening endocrine disrupting compounds. In general, previous research focus has been on the production of the 2 sex steroids, 17β-estradiol and testosterone, and less attention has been paid to other important steroid end points in the steroidogenesis with a wide range of physiological functions, such as the glucocorticoids (corticosterone and cortisol). A newly developed and validated solid phase extraction (SPE) liquid chromatography-mass spectroscopy (LC-MS/MS) method was used to measure the production of cortisol and corticosterone in the H295R cell line. The method was applied by studying the effects of 2 model endocrine disrupters, ketoconazole and prochloraz, the pharmaceutical budesonide, and the inducer forskolin on the steroid production in this cell line. Dose-response curves were obtained for the correlation between hormone concentrations and the concentration of the individual disruptors. Exposing cells to ketoconazole resulted in a decrease in cortisol and corticosterone concentrations in a dose-dependent manner with EC₅₀ values of 0.24 and 0.40 μmol/L, respectively. The same applied for cells exposed to prochloraz with EC₅₀ values of 0.06 and 0.09 μmol/L for cortisol and corticosterone, respectively. Budesonide also inhibited glucocorticoid secretion. The EC₅₀ value for cortisol was 19.50 μmol/L, whereas the EC₅₀ value for corticosterone was 71.42 μmol/L. Forskolin induced the secretion of both cortisol (EC₅₀ = 4.09 μmol/L) and corticosterone (EC₅₀ = 0.28 μmol/L). The results obtained demonstrated the validity of the method. Based on these findings, quality criteria for the production of these steroids in this cell line were suggested.

Original language	English
Journal	International Journal of Toxicology
Volume	32
Issue number	3
Pages (from-to)	219-27
Number of pages	9
ISSN	1091-5818
DOIs	https://doi.org/10.1177/1091581813484366
Publication status	Published - May 2013

Keywords

Adrenocortical Carcinoma
Budesonide
Cell Line, Tumor
Chromatography, Liquid
Colforsin
Endocrine Disruptors
Humans
Imidazoles
Ketoconazole
Tandem Mass Spectrometry

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https://journals.sagepub.com/doi/pdf/10.1177/1091581813484366

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title = "Corticosteroid production in H295R cells during exposure to 3 endocrine disrupters analyzed with LC-MS/MS",

abstract = "The adrenocortical human cell line H295R is a valuable tool for screening endocrine disrupting compounds. In general, previous research focus has been on the production of the 2 sex steroids, 17β-estradiol and testosterone, and less attention has been paid to other important steroid end points in the steroidogenesis with a wide range of physiological functions, such as the glucocorticoids (corticosterone and cortisol). A newly developed and validated solid phase extraction (SPE) liquid chromatography-mass spectroscopy (LC-MS/MS) method was used to measure the production of cortisol and corticosterone in the H295R cell line. The method was applied by studying the effects of 2 model endocrine disrupters, ketoconazole and prochloraz, the pharmaceutical budesonide, and the inducer forskolin on the steroid production in this cell line. Dose-response curves were obtained for the correlation between hormone concentrations and the concentration of the individual disruptors. Exposing cells to ketoconazole resulted in a decrease in cortisol and corticosterone concentrations in a dose-dependent manner with EC50 values of 0.24 and 0.40 μmol/L, respectively. The same applied for cells exposed to prochloraz with EC50 values of 0.06 and 0.09 μmol/L for cortisol and corticosterone, respectively. Budesonide also inhibited glucocorticoid secretion. The EC50 value for cortisol was 19.50 μmol/L, whereas the EC50 value for corticosterone was 71.42 μmol/L. Forskolin induced the secretion of both cortisol (EC50 = 4.09 μmol/L) and corticosterone (EC50 = 0.28 μmol/L). The results obtained demonstrated the validity of the method. Based on these findings, quality criteria for the production of these steroids in this cell line were suggested.",

keywords = "Adrenocortical Carcinoma, Budesonide, Cell Line, Tumor, Chromatography, Liquid, Colforsin, Endocrine Disruptors, Humans, Imidazoles, Ketoconazole, Tandem Mass Spectrometry",

author = "Winther, {Christina S} and Nielsen, {Frederik K} and Martin Hansen and Bjarne Styrishave",

year = "2013",

month = may,

doi = "10.1177/1091581813484366",

language = "English",

volume = "32",

pages = "219--27",

journal = "International Journal of Toxicology",

issn = "1091-5818",

publisher = "SAGE Publications",

number = "3",

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TY - JOUR

T1 - Corticosteroid production in H295R cells during exposure to 3 endocrine disrupters analyzed with LC-MS/MS

AU - Winther, Christina S

AU - Nielsen, Frederik K

AU - Hansen, Martin

AU - Styrishave, Bjarne

PY - 2013/5

Y1 - 2013/5

N2 - The adrenocortical human cell line H295R is a valuable tool for screening endocrine disrupting compounds. In general, previous research focus has been on the production of the 2 sex steroids, 17β-estradiol and testosterone, and less attention has been paid to other important steroid end points in the steroidogenesis with a wide range of physiological functions, such as the glucocorticoids (corticosterone and cortisol). A newly developed and validated solid phase extraction (SPE) liquid chromatography-mass spectroscopy (LC-MS/MS) method was used to measure the production of cortisol and corticosterone in the H295R cell line. The method was applied by studying the effects of 2 model endocrine disrupters, ketoconazole and prochloraz, the pharmaceutical budesonide, and the inducer forskolin on the steroid production in this cell line. Dose-response curves were obtained for the correlation between hormone concentrations and the concentration of the individual disruptors. Exposing cells to ketoconazole resulted in a decrease in cortisol and corticosterone concentrations in a dose-dependent manner with EC50 values of 0.24 and 0.40 μmol/L, respectively. The same applied for cells exposed to prochloraz with EC50 values of 0.06 and 0.09 μmol/L for cortisol and corticosterone, respectively. Budesonide also inhibited glucocorticoid secretion. The EC50 value for cortisol was 19.50 μmol/L, whereas the EC50 value for corticosterone was 71.42 μmol/L. Forskolin induced the secretion of both cortisol (EC50 = 4.09 μmol/L) and corticosterone (EC50 = 0.28 μmol/L). The results obtained demonstrated the validity of the method. Based on these findings, quality criteria for the production of these steroids in this cell line were suggested.

AB - The adrenocortical human cell line H295R is a valuable tool for screening endocrine disrupting compounds. In general, previous research focus has been on the production of the 2 sex steroids, 17β-estradiol and testosterone, and less attention has been paid to other important steroid end points in the steroidogenesis with a wide range of physiological functions, such as the glucocorticoids (corticosterone and cortisol). A newly developed and validated solid phase extraction (SPE) liquid chromatography-mass spectroscopy (LC-MS/MS) method was used to measure the production of cortisol and corticosterone in the H295R cell line. The method was applied by studying the effects of 2 model endocrine disrupters, ketoconazole and prochloraz, the pharmaceutical budesonide, and the inducer forskolin on the steroid production in this cell line. Dose-response curves were obtained for the correlation between hormone concentrations and the concentration of the individual disruptors. Exposing cells to ketoconazole resulted in a decrease in cortisol and corticosterone concentrations in a dose-dependent manner with EC50 values of 0.24 and 0.40 μmol/L, respectively. The same applied for cells exposed to prochloraz with EC50 values of 0.06 and 0.09 μmol/L for cortisol and corticosterone, respectively. Budesonide also inhibited glucocorticoid secretion. The EC50 value for cortisol was 19.50 μmol/L, whereas the EC50 value for corticosterone was 71.42 μmol/L. Forskolin induced the secretion of both cortisol (EC50 = 4.09 μmol/L) and corticosterone (EC50 = 0.28 μmol/L). The results obtained demonstrated the validity of the method. Based on these findings, quality criteria for the production of these steroids in this cell line were suggested.

KW - Adrenocortical Carcinoma

KW - Budesonide

KW - Cell Line, Tumor

KW - Chromatography, Liquid

KW - Colforsin

KW - Endocrine Disruptors

KW - Humans

KW - Imidazoles

KW - Ketoconazole

KW - Tandem Mass Spectrometry

U2 - 10.1177/1091581813484366

DO - 10.1177/1091581813484366

M3 - Journal article

C2 - 23616146

SN - 1091-5818

VL - 32

SP - 219

EP - 227

JO - International Journal of Toxicology

JF - International Journal of Toxicology

IS - 3

ER -

Corticosteroid production in H295R cells during exposure to 3 endocrine disrupters analyzed with LC-MS/MS

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