High glucuronidation activity of environmental estrogens in the carp (Cyprinus carpino) intestine

Many adverse effects on carp reproductive organs have been reported to be caused by exposure to environmental estrogens, such as nonylphenol and bisphenol A, which contaminate the aquatic environment. The glucuronidation activities of xenoestrogens (bisphenol A and diethylstilbestrol) and phytoestrogens (coumestrol, genistein and biochanin A), but not nonylphenol and octylphenol, were observed in microsomes prepared from carp organs. The highest levels of glucuronidation of environmental estrogens, for which the optimum temperature was 25-30 degrees C, were observed in the intestinal microsomes of 2-year-old carp. These activities in carp intestine increased developmentally, and the maximum levels corresponded to 5-10 % of that in rat liver microsomes. However, the glucuronidation of phytoestrogen by carp intestinal microsomes corresponded to that of rat liver microsomes. Only bisphenol A-glucuronide was excreted from the everted intestine, indicating that bisphenol A is metabolized in the carp intestine mainly as glucuronide.These results suggest that glucuronidation by carp intestine plays an important role for the detoxification of xenoestrogens and phytoestrogens, except for nonylphenol and octylphenol.     

Distinct effects of 4-nonylphenol and estrogen-17 beta on expression of estrogen receptor alpha gene in smolting sockeye salmon

Xenoestrogens such as 4-nonylphenol (4-NP) have been shown to affect the parr-smolt transformation, but their mechanisms of action are not known. We therefore examined effects of 4-NP and estradiol-17beta (E2) on expression of estrogen receptor (ER) alpha gene in the liver, gill, pituitary and brain of sockeye salmon to elucidate molecular mechanisms of 4-NP and E2 and developmental differences in response during smolting. Fish were treated twice within a week with 4-NP (15 and 150 mg/kg BW), E2 (2 mg/kg BW) or only vehicle at three stages of smolting, pre-smolting in March, early smolting in April and late smolting in May. The absolute amounts of ERalpha mRNA were determined by real-time PCR. The basal amounts of ERalpha mRNA peaked in April in the liver, gill and pituitary. In March, E2 extensively increased the amounts in the liver, while 4-NP had no effects at this stage. In contrast, 4-NP (but not E2) decreased liver ERalpha mRNA in April. 4-NP also decreased the amount of ERalpha mRNA in the gill in April. In the pituitary, 4-NP increased ERalpha mRNA in March but decreased it in May. There were no significant effects in the brain. Changes in basal ERalpha mRNA observed in this study indicate that estrogen responsiveness of tissues may change during salmon smolting. Furthermore, 4-NP and E2 have different effects on expression of ERalpha gene in the liver and gill during smolting, and the response is dependent on smolt stage.     

WISP-2 is a secreted protein and can be a marker of estrogen exposure in MCF-7 cells

As many structurally diverse chemicals have been reported to function as estrogens, evaluations for estrogenicity of compounds are of widespread concern. Recently, we identified WISP-2 (Wnt-1 inducible signaling pathway protein 2) as a novel estrogen-inducible gene in human breast cancer cells. In this study, we examined whether WISP-2 could be utilized as a marker for screening environmentally relevant compounds for estrogenicity. In MCF-7 cells, progesterone, dexamethasone, tri-iodothyronine, and 2,3,7,8-tetrachlorodibenzo-p-dioxin did not regulate the expression of WISP-2, indicating that its induction is highly specific for hormones that interact with the estrogen receptor. Western blot analysis detected WISP-2 protein induced by 17-beta-estradiol (E2), not only in the cell lysates but also in the culture supernatant of exposed cells, indicating that WISP-2 was a secreted protein. The induction of WISP-2 protein by E2 in the culture supernatant was dose-dependent with estimated EC(50) levels between 10 and 100 pM. Our results demonstrated the capacity to screen environmental compounds for estrogenicity via WISP-2 induction.     

Developmental exposure to xenoestrogen enhances spatial learning in male rats

Steroid hormones have profound effects on the development and function of the nervous system. Environmental estrogens or xenoestrogens are manmade or are natural compounds, which mimics the action of estrogen hormones. The experimental evidence for impairment of cognitive functions in humans and mammals following exposure to xenoestrogens has been fiercely debated. The strongest arguments against such studies have been that the route, time course, and intensity of exposure did not simulate environmental exposure, and that the chemicals tested have additional, non-estrogenic toxic effects, hindering a generalization of actual "xenoestrogenic" effects. Here we show that an environmental-like exposure to the pure estrogen, 17alpha-ethynylestradiol (EE2) during development enhances spatial learning abilities in adult male Sprague-Dawley rats. To simulate an environmental exposure, we used a very low dose (4 ng/kg/day) of EE2 equivalent to concentrations measured in European and US streams which was given orally with a non-invasive method, and we extended the treatment for the entire course of development, from conception to puberty. The animals were tested in a Morris water maze protocol at 6 months of age. Male rats treated with EE2 during development showed a faster learning during the training phase, and remembered better the position of the hidden platform in the short term. Our study demonstrates that actual levels of exposure to xenoestrogens can permanently alter cognitive abilities of a mammalian species.     

Synthesis of phytoestrogenic isoflavonoid disulfates

Di-O-sulfates of six phytoestrogenic isoflavonoids, daidzein (1), genistein (2), glycitein (3), and the reduced metabolites dihydrodaidzein (4), dihydrogenistein (5) and equol (6) were synthesized. These compounds are known or potential inhibitors of steroid sulfatase enzymes. The new compounds were characterized by NMR and mass spectrometry.