title: Time-dependent effects of the flame retardant tris(1,3-dichloro-2-propyl) phosphate (TDCPP) on mRNA expression, in vitro and in ovo, reveal optimal sampling times for rapidly metabolized compounds.
abstract: The flame retardant, tris(1,3-dichloro-2-propyl) phosphate (TDCPP), was previously shown to affect chicken embryo growth, gallbladder size, and lipid homeostasis. A microarray study, however, revealed only modest transcriptional alterations in liver tissue of pipping embryos (days 20-21), which was attributed to the rapid metabolism of TDCPP throughout incubation. To identify the most appropriate sampling time for rapidly metabolized compounds, the present study assessed the time-dependent effects of TDCPP on 27 genes, in ovo (50 µg [116 nmol] TDCPP/g egg) and in vitro (10 µM), using a chicken ToxChip polymerase chain reaction array. The greatest magnitude in dysregulation (up to 362-fold) occurred on day 8 of incubation (in ovo) with alterations of genes involved in phase I, II, and III metabolism, among others. Gallbladder hypotrophy was observed by embryonic day 12, corroborating the finding in pipping embryos from our previous study. From days 12 to 19, genes involved in lipid homeostasis, steroid hormone metabolism, and oxidative stress were affected. In chicken embryonic hepatoctyes (CEHs), TDCPP was completely metabolized to bis(1,3-dichloro-2-propyl) phosphate (BDCPP) within 36 h, but transcriptional changes remained significant up to 36 h. These changes were not attributed to BDCPP exposure as it only altered 1 gene (CYP1A4). An 18-h exposure in CEHs altered the greatest number of genes, making it an appropriate time point for high-throughput chemical screening; however, depending on the biological pathways of interest, shorter or longer incubation times may be more informative. Overall, TDCPP elicits the transcriptional and phenotypic alterations observed in vitro and in ovo, whereas its major metabolite, BDCPP, is far less biologically active.
authors.affiliation: National Wildlife Research Centre, Environment Canada, Ottawa, Ontario, Canada.
authors.forename: Amani
authors.initials: A
authors.lastname: Farhat
authors.forename: Doug
authors.initials: D
authors.lastname: Crump
authors.forename: Emily
authors.initials: E
authors.lastname: Porter
authors.forename: Suzanne
authors.initials: S
authors.lastname: Chiu
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Letcher
authors.forename: Guanyong
authors.initials: G
authors.lastname: Su
authors.forename: Sean W
authors.initials: SW
authors.lastname: Kennedy
mesh_terms: D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism; D030161:Avian Proteins / Q000235:genetics / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002642:Chick Embryo; D002645:Chickens / Q000254:growth & development; D002851:Chromatography, High Pressure Liquid; D005411:Flame Retardants / Q000032:analysis / Q000378:metabolism / Q000633:toxicity*; D005786:Gene Expression Regulation / Q000187:drug effects*; D050356:Lipid Metabolism / Q000187:drug effects / Q000235:genetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D010755:Organophosphates / Q000032:analysis / Q000378:metabolism / Q000633:toxicity*; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics*; D053719:Tandem Mass Spectrometry; D013997:Time Factors; D059467:Transcriptome
chemical_list: D030161:Avian Proteins; D005411:Flame Retardants; D010755:Organophosphates; D012333:RNA, Messenger; C012085:tris(2,3-dibromopropyl)phosphate; D001189:Aryl Hydrocarbon Hydroxylases; C416342:CYP1A4 protein, Gallus gallus
keywords: Chicken embryo; Hepatocytes; Organophosphate flame retardant; Polymerase chain reaction array; Time course; mRNA expression