Toxicological Sciences 67, 75-80 (2002)
Copyright © 2002 by the Society of Toxicology
ENVIRONMENTAL TOXICOLOGY |
Effects of Primary Exposure to Environmental and Natural Estrogens on Vitellogenin Production in Carp (Cyprinus carpio) Hepatocytes
* Institute for Risk Assessment Sciences (IRAS), Utrecht University, P. O. Box 80176, 3508 TD, Utrecht, The Netherlands;
Great Lakes Institute for Environmental Research (GLIER), University of Windsor, Windsor, Ontario, Canada; and
Department of Zoology, National Food Safety and Toxicology Center, Institute of Environmental Toxicology, Michigan State University, East Lansing, Michigan 48824
Vitellogenin (vtg) is a precursor of the yolk proteins lipovitelline and phosvitin and is synthesized as a consequence of estrogen-dependent gene expression in female and male hepatocytes of egg-laying vertebrates. Freshly isolated carp hepatocytes of a genetically uniform strain of adult male carp (Cyprinus carpio) were used to investigate the effects of primary exposure to estrogenic compounds on the vitellogenic response to xenoestrogens. Isolated carp hepatocytes were first exposed (primary exposure) to 50 or 100 µM of either methoxychlor (MXCL) or bisphenol A (BPA), different concentrations of estrone (E1; 1 or 10 nM) or 17ß-estradiol (E2; 0.1 or 1 nM) for 2 days. Hepatocytes were exposed to xenoestrogens (secondary exposure) at both 2 and 5 days after isolation. Hepatocytes were cultured for a total period of 8 days. A competitive indirect ELISA was used to determine the level of vtg after 8 days. The concentration of chemicals used for the primary exposure induced vtg to a level that was less than 10% of the response elicited by E2 (1000 nM). A cytotoxic response, measured by MTT, was not observed after primary exposure to any of the xenoestrogens. After primary exposure to MXCL, vtg production in response to E2 was increased by 4-fold, and vitellogenesis in response to E1 treatment was doubled compared with vitellogenesis without pretreatment. No significant differences were observed between primary exposure to 50 and 100 µM MXCL. Primary exposure to 50 and 100 µM BPA increased the maximum vtg production in response to secondary E2 exposure by about 5- and 7-fold, respectively. Primary exposure to BPA (50 and 100µM) followed by secondary exposure to E1 showed a 4- and 5-fold increase of the vtg synthesis in comparison to E1 exposure alone. Primary exposure to the endogenous estrogens had no significant influence on the vtg synthesis in response to secondary exposure to E1 or E2. Compared to hepatocytes exposed only to MXCL or BPA, primary exposure to E2 increased the vtg synthesis in hepatocytes induced by MXCL or BPA by almost a factor of 2. Primary exposure to E1 increased vitellogenesis after secondary exposure to MXCL only marginally. The present results indicate that weakly estrogenic environmental chemicals such as MXCL and BPA can increase the sensitivity of carp hepatocytes towards endogenous estrogens with respect to VTG synthesis.
Key Words: carp hepatocytes; vitellogenin; in vitro; primary exposure; methoxychlor; bisphenol A.
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