Relative potencies of natural estrogens on vitellogenin and choriogenin levels in the Indian freshwater spotted snakehead, Channa punctata: in vivo and in vitro studies

摘要

The relative efficacies of three natural estrogens viz., estrone (E-1), estradiol-17 beta (E-2) and estriol (E-3) to induce synthesis of vitellogenin (Vg) and choriogenin (Chg) were assessed in primary hepatocyte cultures of the Indian freshwater spotted snakehead, Channa punctata. Hepatocytes were isolated from the spotted snakehead liver by a non-enzymatic protocol. Optimum culture conditions were standardized for ensuring their viability and functioning. Isolated hepatocytes were cultured for 48 h for monolayer formation and then exposed to various concentrations (0.001-10 mu M) of the three estrogens. Competitive homologous ELISAs, developed and validated for spotted snakehead Vg and Chg were employed to determine the amounts of these two proteins secreted into the culture medium after 48 h of incubation. The results reveal that although all the three estrogens were effective in inducing the production of Vg and Chg in a dose-dependent manner, there were differences in their relative potencies. Of three estrogens, E-1 was the least potent and could induce synthesis of Vg and Chg only at a minimum concentration of 0.5 mu M; whereas significant levels of both the proteins were quantified in culture medium by exposing the hepatocytes to E-2 or E-3 even at a concentration of 0.001 mu M. All three estrogens were effective in inducing synthesis of Vg and Chg in vivo also. These results suggest the possibility of employing the above in vitro experimental design to monitor the presence of estrogens/estrogen-like chemicals in natural waters, which could interfere with the estrogen receptor system of fish. This study further points to the possibility of using Chg, in addition to Vg, as a parameter for screening various chemicals for their estrogenic activity.