A new in vitro screening bioassay for the ecotoxicological evaluation of the estrogenic responses of environmental chemicals using roach (Rutilus rutilus) liver explant culture

There is growing evidence that many chemicals released in the environment are able to disturb the normal endocrinology of organisms affecting the structure and function of their reproductive system. This has prompted the scientific community to develop appropriate testing methods to identify active compounds and elucidate mechanisms of action. Of particular interest are in vitro screening methods that can document the effects of these endocrine disrupting compounds on fish. In this study, an in vitro bioassay was developed in the roach (Rutilus rutilus) for evaluating the estrogenicity or antiestrogenicity potency of environmental pollutants by measuring vitellogenin (VTG) induction in cultured liver explants. The cell viability was assessed by the measurement of nonspecific esterase activity using a fluorescein diacetate hydrolysis assay. Results showed that explants could be cultured for 72 h without any significant loss of activity. Dose-dependent responses have been measured with estrogenic model compounds such as 17-β-estradiol (E2) and 17-α-ethynylestradiol (EE2) or antiestrogenic compounds such as tamoxifen. Lowest observable effective concentrations were 1 nM for E2, 1 nM for EE2, and 100 nM for tamoxifen, showing a good sensitivity of the test system. Estrogenicity of butyl 4-hydroxybenzoate, 4-nonylphenol, and bisphenol A was tested. bisphenol A (100 μM) or butylparaben induced a twofold increase in VTG production when compared with 100 nM E2, whereas this production was only 20% with 100 μM 4-nonylphenol. Overall, this study shows that the bioassay could provide valuable information on endocrine disrupting chemicals including metabolites and mixtures of compounds. © 2010 Wiley Periodicals, Inc

In vitro and in vivo studies of the endocrine disrupting potency of cadmium in roach (Rutilus rutilus) liver

International audienceCadmium has been reported to exert estrogenic, antiestrogenic or both effects in vertebrate species. To elucidate the endocrine disrupting action of CdCl2, ex vivo and in vivo experiments were performed in roach (Rutilus rutilus). Roach liver explants were exposed to a range of CdCl2 concentrations alone (0.1–50 μM) or with an effective concentration (100 nM) of 17β-estradiol (E2). In addition, juvenile roach were intraperitoneally injected with CdCl2 (0.1–2.5 mg/kg) with or without 1 mg E2/kg. Subsequent analysis evaluated the effect of CdCl2 on vitellogenin (VTG) synthesis both at the mRNA and protein level, on estrogen receptors (erα and erβ1) and on androgen receptor (ar) mRNA expression. Ex vivo and in vivo experiments indicated that CdCl2 is strongly anti-estrogenic as, when co-exposed to E2, CdCl2 significantly inhibited VTG production as well as vtg and erα mRNA expressions. Moreover, CdCl2 compromised the E2-mediated induction of the ar mRNA expression in vivo

Brain cytochrome P450 aromatase activity in roach (Rutilus rutilus) : Seasonal variations and impact of environmental contaminants

International audienceP450 aromatase catalyses the conversion of C19 androgens to C18 estrogens which is thought to be essential for the regulation of the reproductive function. In this study, brain aromatase activity (AA) was measured monthly over a reproductive cycle in wild roach (Rutilus rutilus) sampled in a reference site in Normandy. AA peaked during the breeding season, reaching 35 fmol mg-1 min-1 in both male and female fish, and was low during the rest of the year except for a significant rise in October. AA was correlated with ovary maturation (measured either as gonado-somatic index or by histological analysis of the gonads) and plasma sex-steroid levels (11-ketotestosterone in males and 17-beta-estradiol in females). Measurements of AA in polluted sites showed that activity was significantly upregulated in sites with fish showing high levels of plasma vitellogenin and large proportion of intersexuality (20-50%) thus suggesting the occurrence of estrogenic compounds and their involvement in AA modulation

Endocrine effects of the tapeworm Ligula Intestinalis in its teleost host, the roach (Rutilis rutilis)

International audienceThe effects of parasite infection by the cestode Ligula intestinalis on the reproductive function and endocrine system of wild roach Rutilus rutilus were evaluated. Gonad maturation, plasma vitellogenin, plasma steroid concentrations (i.e. progesterone, 11-keto-testosterone and 17 beta-estradiol) and brain aromatase activity were investigated in relation with parasitization. A low prevalence (8%) of ligulosed roach and a moderate impact of parasitization (mean parasitization index of 8.8%) were found in the studied population. Inhibition of gonad maturation generally resulted from infestation but 55 of the ligulosed roach nevertheless reached maturity. Main sex steroid plasma content was depleted in both genders. Male 11-keto-testosterone, female 17-beta-estradiol and progesterone plasma concentrations of both genders were, respectively, 27, 5 and 3 times lower m ligulosed fish when compared to their non-infected counterparts. Progesterone levels were negatively correlated with the parasitization index in females. Brain aromatase activity of infected roach was reduced to 50% of that of the non-infected fish. These results demonstrate significant negative effects on the reproductive function of wild roach infected by the tapeworm L. intestinalis collected from a site with low contamination

CYP19A and B gene expression and activity in roach (Rutilus rutilus)

Estrogens are steroid hormones involved in the control of important reproduction-related processes including sexual differentiation and maturation. Careful spatial and temporal balance of estrogens in the body is crucial for proper functioning. At the final step of estrogen biosynthesis, cytochrome P450 aromatase, encoded by the cyp19 gene, converts androgens into estrogens. Modulation of aromatase CYP19 expression and function can dramatically alter the rate of estrogen production, disturbing the local and systemic levels of estrogens. This activity has been shown to be sensitive to many endocrine disruptive compounds (EDC) released in the environment and thus could be used as a biomarker to screen for EDC effects. Teleosts are becoming increasingly important indicators of environmental health, and roach, Rutilus rutilus, was suggested as a sensitive model organism to assess the impact of xenobiotic in freshwater. A study has been undertaken to characterise both gene expression and activity of aromatase in the brain and gonad. Subsequently, seasonal variations, incidence of physical factors, biological factors (sex, age), presence of chemicals and association with pathological features (parasites, intersex) have been studied

Mnémonique et poétique : La figure et son lieu dans la peinture des Tre-Quattrocento

International audienc

Molecular isolation and characterization of the kisspeptin system, KISS and GPR54 genes in roach Rutilus rutilus.

European program Interreg III and IV

Roach (Rutilus rutilus) reproductive cycle: a study of biochemical and histological parameters in a low contaminated site

Fish reproduction is subjected to worrying trends in many aquatic environments. In this study, we report the absence of histological and biochemical alterations in fish sampled in a low contaminated site (characterised by the absence of detectable oestrogenic activity and mutagenicity in sediment extracts). A total of 474 roach (Rutilus rutilus) were monthly sampled during 18 months, and no intersex fish were recorded after careful histological examination, thus indicating that the incidence of this phenomenon may be very low under natural conditions. Furthermore, mean male plasma vitellogenin concentration was 24 ng ml(-1) and was only slightly elevated during the spawning period (up to 120 ng ml(-1)) indicating that these low values may be characteristic of a low contaminated site. Of the male roach, 45.3% were sampled, a sex-ratio that did not significantly deviated from the expected 1:1 ratio between male and female. Results also showed that natural conditions can greatly affect the reproductive cycle of roach. Gametogenesis showed a biphasic pattern with first gonad maturation between September and December and a final maturation occurring at the end of winter/early spring. Under decreasing temperatures, particularly below 6A degrees C, gametogenesis was stopped or even regressed with secondary oocytes becoming rare under histological observation. Conversely, elevated temperatures during the winter lead to an earlier gonad maturation