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

TitleQuantitative RT-PCR methods for evaluating toxicant-induced effects on steroidogenesis using the H295R cell line
Authors
KeywordsMolecular Sequence Numbers
Issue Date2005
PublisherAmerican Chemical Society. The Journal's web site is located at http://pubs.acs.org/est
Citation
Environmental Science And Technology, 2005, v. 39 n. 8, p. 2777-2785 How to Cite?
Abstract
Gene 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.
Persistent Identifierhttp://hdl.handle.net/10722/92789
ISSN
2013 Impact Factor: 5.481
ISI Accession Number ID
References

 

Author Affiliations
  1. Michigan State University
  2. ENTRIX Inc.
  3. City University of Hong Kong
  4. Utrecht University
DC FieldValueLanguage
dc.contributor.authorZhang, Xen_HK
dc.contributor.authorYu, RMKen_HK
dc.contributor.authorJones, PDen_HK
dc.contributor.authorLam, GKWen_HK
dc.contributor.authorNewsted, JLen_HK
dc.contributor.authorGracia, Ten_HK
dc.contributor.authorHecker, Men_HK
dc.contributor.authorHilscherova, Ken_HK
dc.contributor.authorSanderson, JTen_HK
dc.contributor.authorWu, RSSen_HK
dc.contributor.authorGiesy, JPen_HK
dc.date.accessioned2010-09-17T10:57:12Z-
dc.date.available2010-09-17T10:57:12Z-
dc.date.issued2005en_HK
dc.identifier.citationEnvironmental Science And Technology, 2005, v. 39 n. 8, p. 2777-2785en_HK
dc.identifier.issn0013-936Xen_HK
dc.identifier.urihttp://hdl.handle.net/10722/92789-
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.en_HK
dc.languageengen_HK
dc.publisherAmerican Chemical Society. The Journal's web site is located at http://pubs.acs.org/esten_HK
dc.relation.ispartofEnvironmental Science and Technologyen_HK
dc.subjectMolecular Sequence Numbersen_HK
dc.titleQuantitative RT-PCR methods for evaluating toxicant-induced effects on steroidogenesis using the H295R cell lineen_HK
dc.typeArticleen_HK
dc.identifier.emailWu, RSS: rudolfwu@hku.hken_HK
dc.identifier.authorityWu, RSS=rp01398en_HK
dc.description.naturelink_to_subscribed_fulltext-
dc.identifier.doi10.1021/es048679ken_HK
dc.identifier.pmid15884376en_HK
dc.identifier.scopuseid_2-s2.0-20244384266en_HK
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-20244384266&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume39en_HK
dc.identifier.issue8en_HK
dc.identifier.spage2777en_HK
dc.identifier.epage2785en_HK
dc.identifier.isiWOS:000228428900053-
dc.publisher.placeUnited Statesen_HK
dc.identifier.scopusauthoridZhang, X=8606600100en_HK
dc.identifier.scopusauthoridYu, RMK=9278574900en_HK
dc.identifier.scopusauthoridJones, PD=34771015600en_HK
dc.identifier.scopusauthoridLam, GKW=8606600400en_HK
dc.identifier.scopusauthoridNewsted, JL=6603677236en_HK
dc.identifier.scopusauthoridGracia, T=6506058618en_HK
dc.identifier.scopusauthoridHecker, M=35247848500en_HK
dc.identifier.scopusauthoridHilscherova, K=6603438103en_HK
dc.identifier.scopusauthoridSanderson, JT=7004489858en_HK
dc.identifier.scopusauthoridWu, RSS=7402945079en_HK
dc.identifier.scopusauthoridGiesy, JP=35459135300en_HK

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