title: An in vitro/in vivo screening assay as a sensitive tool to assess endocrine disruptive activity in surface water.
abstract: Adult male fathead minnow were exposed for 14 or 28-days under flow-through conditions to undiluted filtered water samples from the rivers Meuse and Rhine in the Netherlands. The experiment included two vessels per treatment each containing 10 fish and samples of five fish were taken after 14 and 28 days. Additional groups were exposed to 17alpha-ethinylestradiol (EE2) as a reference and untreated drinking water as a negative control. Major endpoints examined included induction of vitellogenin (VTG) synthesis, VTG mRNA activity, hepato- and gonadosomatic indices (HSI and GSI) and gonadal histology. No significant difference was recorded in body weight or mean GSI values between the various treatments. Only exposure to Meuse water resulted in significantly higher HSI means after 14 days. Histological examination showed no apparent effects on gonadal tissue except for eosinophilic blood plasma in fish exposed to Meuse water or EE2. After 14 and 28 days, elevated VTG and VTG mRNA levels were measured in most livers of the fish exposed to Meuse water, but not in the fish exposed to Rhine water. This was confirmed by measuring estrogenic responses in the in vitro ER CALUX assay. Induction of VTG synthesis proved to be the most sensitive endpoint in the Non Spawning Male Fish Assay for in vivo detection of bio-available estrogenic activity supplementary to a sensitive in vitro assay. The other endpoints examined varied too much and required a higher number of fish or replicates to achieve sufficient power for statistical testing making them less animal friendly.
authors.affiliation: NOTOX B.V., Hambakenwetering 7, 5231 DD's-Hertogenbosch, The Netherlands. rinus.bogers@notox.nl
authors.forename: Rinus
authors.initials: R
authors.lastname: Bogers
authors.forename: Selinda
authors.initials: S
authors.lastname: De Vries-Buitenweg
authors.forename: Ine
authors.initials: I
authors.lastname: Geuijen
authors.forename: Beppy
authors.initials: B
authors.lastname: van de Waart
authors.forename: Raoul
authors.initials: R
authors.lastname: Kuiper
authors.forename: Sander
authors.initials: S
authors.lastname: Van Der Linden
authors.forename: Leo
authors.initials: L
authors.lastname: Puijker
authors.forename: Albertinka
authors.initials: A
authors.lastname: Murk
authors.forename: Bart
authors.initials: B
authors.lastname: Van Der Burg
authors.forename: Juliette
authors.initials: J
authors.lastname: Legler
mesh_terms: D000818:Animals; D001681:Biological Assay; D045744:Cell Line, Tumor; D003530:Cyprinidae; D052244:Endocrine Disruptors / Q000633:toxicity*; D006066:Gonads / Q000033:anatomy & histology / Q000187:drug effects / Q000254:growth & development; D006801:Humans; D008099:Liver / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism; D008297:Male; D009426:Netherlands; D012333:RNA, Messenger / Q000096:biosynthesis; D011960:Receptors, Estrogen / Q000378:metabolism; D045483:Rivers; D014819:Vitellogenins / Q000096:biosynthesis* / Q000235:genetics; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: D052244:Endocrine Disruptors; D012333:RNA, Messenger; D011960:Receptors, Estrogen; D014819:Vitellogenins; D014874:Water Pollutants, Chemical