Transcriptional changes in African clawed frogs (Xenopus laevis) exposed to 17α-ethynylestradiol during early development.

Although the past two decades have witnessed a significant increase in the number of studies investigating effects of estrogenic chemicals on amphibians, to date little is known about specific molecular interactions of estrogens with the hypothalamus-pituitary-gonadal-hepatic axis in developing amphibians. Here, tissue-specific functional sets of genes, derived previously from studies of fishes exposed to endocrine active chemicals, were evaluated in Xenopus laevis exposed to 17α-ethynylestradiol (EE2) throughout their early development. Specifically, transcriptional responses of X. laevis exposed to 0.09, 0.84, or 8.81 µg EE2/L were characterized during sexual differentiation [31 day post hatch (dph)] and after completion of metamorphosis during the juvenile stage (89 dph). While at 31 dph there were no consistent effects of EE2 on abundances of transcripts,at 89 dph X. laevis exhibited significant alterations in expression of genes involved in steroid signaling and metabolism, synthesis of cholesterol, and vitellogenesis. Specifically, expression of androgen receptor, farnesyl diphosphate synthase, estrogen receptor α, and vitellogenin A2 was significantly greater (>2-fold) than in controls while expression of farnesoid x-activated receptors α and ß was significantly less (>2-fold reduction) than in controls. These results support the hypothesis that sets of genes derived from studies in teleost fish can be extrapolated for use in amphibians during the juvenile stage but not in sexually undifferentiated individuals. Furthermore, changes in abundances of transcripts of the here utilized sets of genes in animals sampled post sexual differentiation were in accordance with developmental effects and alterations of gonadal histology reported in a parallel study. This set of genes might be useful for predicting potential adverse outcomes at later life-stages.

Effects of 17α-ethynylestradiol on sexual differentiation and development of the African clawed frog (Xenopus laevis).

Several studies have shown that exposure of amphibians, including the African clawed frog (Xenopus laevis), to potent estrogens at critical times during development results in feminization and/or demasculinization. However, genotyping of X. laevis has only recently become possible, so studies performed in the past were rarely able to make explicit linkages between genetic and phenotypic sex. Therefore, to further characterize this relationship, X. laevis tadpoles were exposed during development to 0.09, 0.84, or 8.81 µg/L 17α-ethynylestradiol (EE2), which is the estrogen analog commonly used in oral contraceptives. Exposure to all concentrations of EE2 tested resulted in significant delays in time to metamorphosis. Genotyping showed that genetic sex ratios were similar among treatments. However, morphological evaluation revealed that a significant number of individuals with a male genotype displayed mixed sex and abnormal phenotypes. Additionally, both genetic males and females exposed to EE2 exhibited greater presence of vitellogenin protein relative to the respective controls. Since estrogens function downstream of the initial molecular signals of sexual differentiation, it is likely that genetic male animals received mixed endogenous male and exogenous female signals that caused disordered sexual development. The production of vitellogenin was probably temporally separated and independent from primary effects on sexual differentiation, and might have contributed to delays in metamorphosis observed in individuals exposed to EE2.