RESUMO
Oestrogens participate in various biological processes such as oogenesis, vitellogenesis and testicular development, but studies regarding the distribution and protein levels of oestrogen receptors (ERα and ERß) and aromatase (Cyp19) in testis are rarely investigated in fish species. The aim of the present study was to analyse the expression pattern of ERα, ERß and Cyp19 in testis of Astyanax rivularis and, in addition, to verify if oestrogenic contamination interferes in the expression levels of these proteins. Quarterly, field samplings were carried out during a reproductive cycle in a stream of the Upper Velhas River with a good conservation status (site S1). In the gonadal maturation peak (June), when ripe stage was most abundant, fish collection was made in three streams: S1, reference site, and S2 and S3, sites contaminated by untreated sewage. The results of immunohistochemistry demonstrated labelling of Cyp19 in Leydig cells and acidophilic granulocytes, but spermatogonia, Sertoli cells, spermatids and spermatozoa were also labelled. ERα was more widely distributed than ERß being found in all developmental germ cell phases. On the other hand, ERß was found only in spermatogonia and spermatocytes. During testicular maturation, ELISA levels for Cyp19, ERα and ERß followed the gonadosomatic index (GSI) with significant higher values in the ripe stage. Regarding to endocrine disruption, the males exposed to domestic sewage presented significant higher expression of Cyp19 and ERα when compared to the non-exposed fish. Together, our results demonstrate expression patterns of Cyp19, ERα and ERß in the testis of A. rivularis. In addition, we indicate ERα and Cyp19 as sensitive biomarkers for monitoring of oestrogenic contamination in freshwater environments.
Assuntos
Characidae/metabolismo , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Testículo/metabolismo , Poluentes Químicos da Água/análise , Animais , Aromatase/metabolismo , Brasil , Ecotoxicologia/métodos , Disruptores Endócrinos/análise , Disruptores Endócrinos/toxicidade , Proteínas de Peixes/metabolismo , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/metabolismo , Masculino , Rios , Células de Sertoli/efeitos dos fármacos , Células de Sertoli/metabolismo , Espermatozoides/efeitos dos fármacos , Espermatozoides/metabolismo , Testículo/citologia , Testículo/efeitos dos fármacos , Poluentes Químicos da Água/toxicidadeRESUMO
The Velhas River is the most polluted river in the state of Minas Gerais, south-eastern Brazil. Due to its historical and environmental relevance, the aim of this study was to evaluate the effects of oestrogenic endocrine disruptors on the reproduction of the lambari Astyanax rivularis, a small-sized species found in headwaters of the São Francisco River basin. Quarterly field samplings were carried out during a reproductive cycle in three streams of the upper Velhas River: S1 (reference site) and S2 and S3 (sites contaminated by untreated sewage). The main oestrogenic compounds were evaluated in water using HPLC/MS. Molecular, histological and reproductive biomarkers were assessed in liver and gonad. The results showed higher average concentrations of oestradiol (>200ng/l) in S2 and S3, oestrone (>250ng/l) in S2 as well as oestriol (>200ng/l), bisphenol A (>190ng/l), and nonylphenol (>600ng/l) in S3 compared to S1 (<70ng/l for all compounds). In S2 and S3, there was an increase in the proportion of females, higher ELISA levels of vitellogenin (Vtg) and proteins of the zona radiata (Zrp) in liver males. Insulin-like growth factor (IGF-I) levels were lower in S2 males, which also had a smaller body size, a smaller seminiferous tubule diameter, a higher proportion of spermatogonia, and lower proportion of spermatozoa in relation to S1. Histopathological analyses detected an increase in yolk deficient oocytes and over-ripening in the contaminated sites, and these alterations were associated to a reduction of hepatic Vtg levels and a delay in spawning, respectively. Intersex specimens with perinucleolar follicles in a multifocal distribution in the testis were detected in S2 and S3. These results indicate that chronic exposure to oestrogenic compounds induced endocrine disruption that may affect wild populations of A. rivularis in the Velhas River.