Effects of Ethinylestradiol, EE2, exposure on Poecilia reticulta male specific genes, DMRT1 and CYP11b 

Abstract: Endocrine disrupting chemicals (EDCs) found in every day product, can either be natural or man-made and usually exists in a concerning concentration in our aquatic environment which in turn could affect teleost reproduction. EDCs exposure in fishes may result in changes in their reproductive physiology and morphology resulting in for example induction in gonopodia in female fish when exposed to androgens and in males exposed to female estrogen a reduced sperm count and skewed sex ration. Exposures during development have effects on organisms that are permanent and irreversible. Exogenous estrogens are known to direct gonadal in male fish into differentiation of functional ovary while androgens in the other hand can masculine female fish. The contraceptive pills, ethinylestradiol (EE2), secreted by women, is one of those chemicals that affects organisms by demasculizing or feminizing it. One of the most useful biomarkers in distinguishing male fish that are feminized or desmaskuliniseras is the detection of vitellogenin, VTG a female yolk protein in samples. For this study Poecilia reticulata, guppy, is chosen since they are live bearers, reproduce readily and one can easily distinguish the male from the female and they grow quickly. The most significant hormone in fish, were outstanding secondary sexual characterization, is 11-ketotestosterone whose production starts in Leydig cells. The biosynthesis of these potent 11-oxygenated androgens includes a first enzyme, 11-β hydroxylase cytochrome P45011β(CYP11b), involved from precursors such as testosterone (T) or androtestedione (AT). Another important protein involved in the role of sex determination is Double sex and map-3 related transcription factor 1, a gene that is essential for the testicular differentiation. Main objective of this study is to study the effects of administration of estrogen in male fish that has been exposed during the two week period at different levels of estrogen; 0 ng / L, 3 ng / L and 10ng / L, where the expected results should show that these fishes becomes feminized or demaskuliniserade, because the masculine genes such as CYP11b and DMRT-1 will decreases in gene expression or result in no expression at all. A molecular approach in the manner of RNA isolation by TRIZOL protocols, obtaining cDNA, PCR with RNA/ cDNA and semi-quantitive PCR with cDNA will be performed. Results in the study does not correlate with the expected results, which meant that of the fishes that are exposed to maximum dose of estrogen still gave transcription of CYP11b and DMRT1. Although this was a declining pattern discern in some cases. Despite that results were not as expected these genes are definitely good biomarkers for indentifying males and for the use of identifying desmasculinization in male. Considering the ecological disturbance these EDCs are having, they are for certain affecting aquatic organisms due to the huge amount of pollution that are being released in their environment. It is therefore important to continue already proceed studies.