An approach to detecting estrogenic endocrine disruption via choriogenin expression in an estuarine model fish species

Abstract

A large body of work has established a link between endocrine disrupting compounds (EDCs) and a number of abnormalities in fishes. However, most EDC studies use several standard laboratory denizens to assess impacts, so assumptions about sensitivity are primarily based on these few species. Additionally, existing methods rely on obtaining sufficient plasma to measure EDC biomarkers. Our objectives were (a) to establish a new model species for estuarine fishes, (b) to evaluate endocrine impacts with a highly sensitive and specific biomarker, and (c) to develop a method for the analysis of this biomarker in small fish that do not possess sufficient blood plasma for protein measurement. As such, we created a polyclonal antibody (Ab) to the estrogen-responsive proteins chorion (Ch) and choriogenin (Chg) in Menidia beryllina, found throughout coastal North America and already utilized in EPA Whole Effluent Testing. We then validated the Ab by using it to measure the response to aqueous ethinylestradiol (EE2) through the development an ELISA using Menidia whole body homogenate (WBH). Sensitivity of the Ab to Menidia WBH is greater than that of the commercially available option. ELISA sensitivity, with a detection limit of 5 ng/ml and a working range of 22.6-1370.9 ng/ml, is comparable to ELISAs developed to measure plasma Chg. To our knowledge this is the first ELISA method developed for the detection of Chg using WBH. Including additional model species and methods allowing the evaluation of alternative sample matrices will contribute to an enhanced understanding of inter-species differences in EDC response.

The in vivo estrogenic and in vitro anti‐estrogenic activity of permethrin and bifenthrin

“…Equivalents were then divided by the protein concentration per well to normalize the data as described in detail previously [21]. Then two types of statistical analyses were performed on Chg data: a traditional ANOVA followed by a Tukey HSD test to detect whether there were differences between Chg expression induced by different compounds (i.e.…”

“…Our second objective was to utilize a biomarker of estrogenic endocrine disruption that is potentially more sensitive than vitellogenin [19], the estrogen-dependent egg yolk protein typically measured following exposure to suspected endocrine disrupting compounds (EDCs) [20]. Although vitellogenin is a reliable marker of estrogenic endocrine disruption, we instead measured expression of choriogenin (Chg), an estrogen-dependent egg coat protein, using a Menidia specific antibody [21]. Chg, the precursor to the chorion or zona radiata, has been demonstrated to be more sensitive than vitellogenin (Vtg) [19, 22], and was shown recently to have increased mRNA expression in larval fish following bifenthrin exposure [23].…”

Gene Structure and Estrogen-Responsive mRNA Expression of a Novel Choriogenin H Isoform from a Marine Medaka Oryzias dancena

“…Among these pollutants, xenoestrogens are potent endocrine-disrupting chemicals (EDCs) that deregulate gene expression and enzymatic functions associated with the reproduction of the exposed organisms (Rotchell and Ostrander, 2003;Tokumoto et al, 2004Tokumoto et al, , 2011. Previous studies have proposed that exposure to EDCs results in adverse effects on other physiologic attributes, including the innate immunity and host defense mechanisms, of the exposed animals (Lam et al, 2011;Brander et al, 2012). For this reason, understanding the gene expression pattern following EDC exposure is important for developing effective risk management strategies in relation to EDC-caused pollution in marine and coastal waters.…”

From ‘Omics to Otoliths: Responses of an Estuarine Fish to Endocrine Disrupting Compounds across Biological Scales

“…The adaptive significance of this pattern is that females have more time to grow larger and hence have the ability to carry more eggs [27], [28]. Few EDC studies have been performed with fish that have temperature sensitive sex determination (TSD), but recent findings indicate that Menidia species are highly sensitive to EDCs [29], [30], that exposure may disrupt the adaptive benefits of TSD [31], and that the potential for Menidia species to be widely-utilized North American estuarine bioindicators is unparalleled [29], [32].…”