Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.

Xenoestrogens are either natural or synthetic compounds that mimic the effects of endogenous estrogen. These compounds, such as bisphenol-A (BPA), and phthalates, are commonly found in plastic wares. Exposure to these compounds poses major risk to human health because of the potential to cause endoc...

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Autores principales:	Philip Wing-Lok Ho, Zero Ho-Man Tse, Hui-Fang Liu, Song Lu, Jessica Wing-Man Ho, Michelle Hiu-Wai Kung, David Boyer Ramsden, Shu-Leong Ho
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/c8c5755a6a68475fb068dd562e0f1055
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spelling	oai:doaj.org-article:c8c5755a6a68475fb068dd562e0f10552021-11-18T08:56:29ZAssessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.1932-620310.1371/journal.pone.0074065https://doaj.org/article/c8c5755a6a68475fb068dd562e0f10552013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24040167/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203Xenoestrogens are either natural or synthetic compounds that mimic the effects of endogenous estrogen. These compounds, such as bisphenol-A (BPA), and phthalates, are commonly found in plastic wares. Exposure to these compounds poses major risk to human health because of the potential to cause endocrine disruption. There is huge demand for a wide range of chemicals to be assessed for such potential for the sake of public health. Classical in vivo assays for endocrine disruption are comprehensive but time-consuming and require sacrifice of experimental animals. Simple preliminary in vitro screening assays can reduce the time and expense involved. We previously demonstrated that catechol-O-methyltransferase (COMT) is transcriptionally regulated by estrogen via estrogen receptor (ER). Therefore, detecting corresponding changes of COMT expression in estrogen-responsive cells may be a useful method to estimate estrogenic effects of various compounds. We developed a novel cell-based ELISA to evaluate cellular response to estrogenicity by reduction of soluble-COMT expression in ER-positive MCF-7 cells exposed to estrogenic compounds. In contrast to various existing methods that only detect bioactivity, this method elucidates direct physiological effect in a living cell in response to a compound. We validated our assay using three well-characterized estrogenic plasticizers - BPA, benzyl butyl phthalate (BBP), and di-n-butyl phthalate (DBP). Cells were exposed to either these plasticizers or 17β-estradiol (E2) in estrogen-depleted medium with or without an ER-antagonist, ICI 182,780, and COMT expression assayed. Exposure to each of these plasticizers (10(-9)-10(-7)M) dose-dependently reduced COMT expression (p<0.05), which was blocked by ICI 182,780. Reduction of COMT expression was readily detectable in cells exposed to picomolar level of E2, comparable to other in vitro assays of similar sensitivity. To satisfy the demand for in vitro assays targeting different cellular components, a cell-based COMT assay provides useful initial screening to supplement the current assessments of xenoestrogens for potential estrogenic activity.Philip Wing-Lok HoZero Ho-Man TseHui-Fang LiuSong LuJessica Wing-Man HoMichelle Hiu-Wai KungDavid Boyer RamsdenShu-Leong HoPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 9, p e74065 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Philip Wing-Lok Ho Zero Ho-Man Tse Hui-Fang Liu Song Lu Jessica Wing-Man Ho Michelle Hiu-Wai Kung David Boyer Ramsden Shu-Leong Ho Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.
description	Xenoestrogens are either natural or synthetic compounds that mimic the effects of endogenous estrogen. These compounds, such as bisphenol-A (BPA), and phthalates, are commonly found in plastic wares. Exposure to these compounds poses major risk to human health because of the potential to cause endocrine disruption. There is huge demand for a wide range of chemicals to be assessed for such potential for the sake of public health. Classical in vivo assays for endocrine disruption are comprehensive but time-consuming and require sacrifice of experimental animals. Simple preliminary in vitro screening assays can reduce the time and expense involved. We previously demonstrated that catechol-O-methyltransferase (COMT) is transcriptionally regulated by estrogen via estrogen receptor (ER). Therefore, detecting corresponding changes of COMT expression in estrogen-responsive cells may be a useful method to estimate estrogenic effects of various compounds. We developed a novel cell-based ELISA to evaluate cellular response to estrogenicity by reduction of soluble-COMT expression in ER-positive MCF-7 cells exposed to estrogenic compounds. In contrast to various existing methods that only detect bioactivity, this method elucidates direct physiological effect in a living cell in response to a compound. We validated our assay using three well-characterized estrogenic plasticizers - BPA, benzyl butyl phthalate (BBP), and di-n-butyl phthalate (DBP). Cells were exposed to either these plasticizers or 17β-estradiol (E2) in estrogen-depleted medium with or without an ER-antagonist, ICI 182,780, and COMT expression assayed. Exposure to each of these plasticizers (10(-9)-10(-7)M) dose-dependently reduced COMT expression (p<0.05), which was blocked by ICI 182,780. Reduction of COMT expression was readily detectable in cells exposed to picomolar level of E2, comparable to other in vitro assays of similar sensitivity. To satisfy the demand for in vitro assays targeting different cellular components, a cell-based COMT assay provides useful initial screening to supplement the current assessments of xenoestrogens for potential estrogenic activity.
format	article
author	Philip Wing-Lok Ho Zero Ho-Man Tse Hui-Fang Liu Song Lu Jessica Wing-Man Ho Michelle Hiu-Wai Kung David Boyer Ramsden Shu-Leong Ho
author_facet	Philip Wing-Lok Ho Zero Ho-Man Tse Hui-Fang Liu Song Lu Jessica Wing-Man Ho Michelle Hiu-Wai Kung David Boyer Ramsden Shu-Leong Ho
author_sort	Philip Wing-Lok Ho
title	Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.
title_short	Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.
title_full	Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.
title_fullStr	Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.
title_full_unstemmed	Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.
title_sort	assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-o-methyltransferase (comt) expression in an elisa-based system.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/c8c5755a6a68475fb068dd562e0f1055
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Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system.

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