Article: Quantitative RT-PCR methods for evaluating toxicant-induced effects on steroidogenesis using the H295R cell line

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TitleQuantitative RT-PCR methods for evaluating toxicant-induced effects on steroidogenesis using the H295R cell line
AuthorsZhang, X1 3
Yu, RMK3
Jones, PD1
Lam, GKW3
Newsted, JL2
Gracia, T1
Hecker, M1
Hilscherova, K1
Sanderson, JT4
Wu, RSS3
Giesy, JP1 3
KeywordsMolecular Sequence Numbers
Issue Date2005
PublisherAmerican Chemical Society. The Journal's web site is located at http://pubs.acs.org/est
CitationEnvironmental Science And Technology, 2005, v. 39 n. 8, p. 2777-2785 [How to Cite?]
DOI: http://dx.doi.org/10.1021/es048679k
AbstractGene expression profiles show considerable promise for the evaluation of the toxic potential of environmental contaminants. For example, any alterations in the pathways of steroid synthesis or breakdown have the potential to cause endocrine disruption. Therefore monitoring these pathways can provide information relative to a chemical's ability to impact endocrine function. One approach to monitoring these pathways has been to use a human adrenocortical carcinoma cell line (H295R) that expresses all the key enzymes necessary for steroidogenesis. In this study we have further developed these methods using accurate and specific quantification methods utilizing molecular beacon-based quantitative RT-PCR (Q-RT-PCR). The assay system was used to analyze the expression patterns of 11 steroidogenic genes in H295R cells. The expression of gene transcripts was measured using a real-time PCR system and quantified based on both a standard curve method using a dilution series of RNA standards and a comparative C t method. To validate the optimized method, cells were exposed to specific and nonspecific model compounds (inducers and inhibitors of various steroidogenic enzymes) for gene expression profiling. Similar gene expression profiles were exhibited by cells treated with chemicals acting through common mechanisms of action. Overall, our findings demonstrated that the present assay can facilitate the development of compound-specific response profiles, and will provide a sensitive and integrative screen for the effects of chemicals on steroidogenesis. © 2005 American Chemical Society.
ISSN0013-936X
2011 Impact Factor: 5.228
2011 SCImago Journal Rankings: 0.305
DOIhttp://dx.doi.org/10.1021/es048679k
ReferencesReferences in Scopus
DC Field
Value
dc.contributor.authorZhang, X
dc.contributor.authorYu, RMK
dc.contributor.authorJones, PD
dc.contributor.authorLam, GKW
dc.contributor.authorNewsted, JL
dc.contributor.authorGracia, T
dc.contributor.authorHecker, M
dc.contributor.authorHilscherova, K
dc.contributor.authorSanderson, JT
dc.contributor.authorWu, RSS
dc.contributor.authorGiesy, JP
dc.date.accessioned2010-09-17T10:57:12Z
dc.date.available2010-09-17T10:57:12Z
dc.date.issued2005
dc.description.abstractGene expression profiles show considerable promise for the evaluation of the toxic potential of environmental contaminants. For example, any alterations in the pathways of steroid synthesis or breakdown have the potential to cause endocrine disruption. Therefore monitoring these pathways can provide information relative to a chemical's ability to impact endocrine function. One approach to monitoring these pathways has been to use a human adrenocortical carcinoma cell line (H295R) that expresses all the key enzymes necessary for steroidogenesis. In this study we have further developed these methods using accurate and specific quantification methods utilizing molecular beacon-based quantitative RT-PCR (Q-RT-PCR). The assay system was used to analyze the expression patterns of 11 steroidogenic genes in H295R cells. The expression of gene transcripts was measured using a real-time PCR system and quantified based on both a standard curve method using a dilution series of RNA standards and a comparative C t method. To validate the optimized method, cells were exposed to specific and nonspecific model compounds (inducers and inhibitors of various steroidogenic enzymes) for gene expression profiling. Similar gene expression profiles were exhibited by cells treated with chemicals acting through common mechanisms of action. Overall, our findings demonstrated that the present assay can facilitate the development of compound-specific response profiles, and will provide a sensitive and integrative screen for the effects of chemicals on steroidogenesis. © 2005 American Chemical Society.
dc.description.natureLink_to_subscribed_fulltext
dc.identifier.citationEnvironmental Science And Technology, 2005, v. 39 n. 8, p. 2777-2785 [How to Cite?]
DOI: http://dx.doi.org/10.1021/es048679k
dc.identifier.doihttp://dx.doi.org/10.1021/es048679k
dc.identifier.epage2785
dc.identifier.isiWOS:000228428900053
dc.identifier.issn0013-936X
2011 Impact Factor: 5.228
2011 SCImago Journal Rankings: 0.305
dc.identifier.issue8
dc.identifier.pmid15884376
dc.identifier.scopuseid_2-s2.0-20244384266
dc.identifier.spage2777
dc.identifier.urihttp://hdl.handle.net/10722/92789
dc.identifier.volume39
dc.languageeng
dc.publisherAmerican Chemical Society. The Journal's web site is located at http://pubs.acs.org/est
dc.publisher.placeUnited States
dc.relation.ispartofEnvironmental Science and Technology
dc.relation.referencesReferences in Scopus
dc.subjectMolecular Sequence Numbers
dc.titleQuantitative RT-PCR methods for evaluating toxicant-induced effects on steroidogenesis using the H295R cell line
dc.typeArticle
Author Affiliations
  1. Michigan State University
  2. ENTRIX Inc.
  3. City University of Hong Kong
  4. Utrecht University