In vitro studies on sex steroid-binding protein in rainbow trout (Oncorhynchus mykiss) hepatocytes : influence of 17â-estradiol and environmental estrogens

Abstract

Environmental estrogens may modulate the endocrine system through interactions with sex steroid-binding protein (SBP), and these processes may be a novel mechanism for endocrine disruption. The endogenous hormone 17â-estradiol (E2) together with weakly estrogen compounds as di-(n-butyl) phthalate (DBP) and potent estrogen mimics (ethynylestradiol, EE2) were all able to induce an up-regulation of total sex-steroid binding capacity and SBP gene expression in vitro using a culture of rainbow trout (Oncorhynchus mykiss) hepatocytes. This increase was most likely due to the induction of the sex steroid-binding protein (SBP) gene and protein itself, although other non-identified proteins probably contribute to sex steroid-binding as well. The roles of non-specific binding proteins were assessed using albumin and rainbow trout vitellogenin which both showed a very low capacity for binding sex steroids. Quantification of SBP protein expression in cell media using western blot and zebrafish SBP antibodies was not possible due to low protein concentrations. Exposing hepatocytes to EE2 induced the strongest response in both total sex steroid-binding activity and SBP gene-expression followed by E2 and DBP. Exposure to endogenous sex steroids and environmental estrogens increased SBP gene-expression after 24 hours, while an increase in total sex-steroid binding capacity was seen after 48 hours of exposure, indicating an up-regulation of the SBP gene before secretion of SBP into the cell medium. A concentration-response relationship, most likely due to increased SBP secretion and gene expression was seen after 96 hours of exposure for both total sex-steroid binding capacity and SBP gene expression.