| pubmed-article:11751013 | pubmed:abstractText | Several predictive test methods for endocrine disrupters have been evaluated by international organizations. In this study, we performed a series of predictive tests for endocrine disrupters, i.e. the receptor binding assay, reporter gene assay, and immature rat uterotrophic assay, on all-trans retinoic acid (tRA), which may cause antiestrogenic activity via their receptors, interfere with estrogenic action at estrogen responsive element level, and we examine the efficacy of endocrine disruptor screening tests in detecting anti-estrogenic effects downstream of receptor-ligand interactions. Despite showing complete lack of binding affinity to ER in the receptor binding assay, tRA exhibited clear antagonist activity without any agonist activity in the reporter gene assay. In the in vivo test, tRA was subcutaneously administered to immature Crj:CD (SD) IGS rats at doses of 5 and 25 mg/kg per day for 3 days, beginning at 20 days of age. Additional groups of rats given tRA at the above doses were also subcutaneously injected with ethinyl estradiol (EE) at a dose of 0.6 microg per rat per day. A vehicle control group given olive oil alone and a positive control group given EE alone were also established. Although no uterotrophic activity was detected in any of the rats given only tRA, co-treatment with 5 and 25 mg/kg tRA and EE reduced the EE-induced increases in uterine weight. We confirmed that the ER antagonist activity of tRA may be mediated by transcriptional interference after ER-ligand complex binding to an estrogen responsive element of the gene by the gel mobility shift analysis. These findings suggest the reporter gene assay and uterotrophic assay can detect anti-estrogenic effects downstream of receptor-ligand interactions, but the receptor binding assay can not detect this type of interference. In any case, a screening strategy for endocrine disrupters, especially the primary screening battery for prioritizing the chemicals to be tested in the higher screening stages, should be designed to detect various kinds of chemicals possessing endocrine modulating activity including a retinoid-like endocrine modulator. Accordingly, reporter gene assay or uterotrophic assay should be conducted in the early stage of screening process for endocrine disrupting chemicals, because they can detect antagonist activity caused by both inhibition of receptor-ligand interaction and transcriptional interference. Particularly, the reporter gene assay may be a promising prescreening procedure, because it can be adopted in the high throughput screening process for thousands of chemicals and it requires no use of experimental animals. | lld:pubmed |
