• Development and Reproduction
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• v.16 no.4
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• pp.289-294
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• 2012

Gene expressions of cytochrome P4501A (CYP1A), aryl hydrocarbon receptor (AhR) and vitellogenin (Vg) by endocrine disruptors, benzo[${\alpha}$]pyrene (B[a]P) and tributyltin (TBT) were examined in cultured eel hepatocytes which were isolated from eels treated previously with B[a]P (10 mg/kg) or estradiol-$17{\beta}$ (20 mg/kg) in vivo, and the relationship between CYP1A, AhR and Vg genes were studied. When the cultured eel hepatocytes were treated with B[a]P ($10^{-6}-10^{-5}M$) the gene expressions of CYP1A and AhR were enhanced in a concentration-dependent manner. However, when treated with TBT ($10^{-9}-10^{-5}M$) the gene expressions of CYP1A and AhR were suppressed at high concentrations ($10^{-6}-10^{-5}M$), while having no effects at low concentrations ($10^{-9}-10^{-7}M$). Gene expression of Vg was also suppressed by TBT in a concentration-dependent manner in cultured eel hepatocytes which was previously treated in vivo with estradiol-$17{\beta}$.

• Korean Journal of Animal Reproduction
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• v.20 no.3
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• pp.259-269
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• 1996

To establish whether or not androgens is responsible for the induction of vitellogenin(Vg) synthesis and secretion, primary hepatocytes prepared from immature eels were used. The results are follows: 1. Eel hepatocytes were prepared using a collagenase perfusion technique. The isolated cells attached efficiently to fibronectin-coated dishes and subsequently formed monolayers in serum-free medium. These cultures maintained in medium for 10 days with minimal cell loss. 2. Estradiol-17$\beta$(E2) alone was insufficient to induce Vg synthesis. The combination of E2 with methyltestosterone(MT) markedly stimulated Vg synthesis. High vg production occurred in MT concentration from 10-6~10-5M in the presence of E2 (10-6M). Testosterone and androsterone were also effective, but progesterone was not effective in inducing Vg synthesis. Neither MT alone nor testosterone and androsterone alone had any effect on Vg synthesis. 3. E2-primed hepatocytes showed Vg synthesis in both media with and without hormones 1 day after culture. In the cultures with the vehicle, MT, or progesterone, the rate of synthesis seemed to decrease with time. But the combination of E2 and MT showed an intense increase in Vg synthesis. Hepatocytes isolated from E2-primed eels also required androgens for continuating of Vg synthesis. 4. These results demonstrate that androgens act together with E2 in synthesis and secretion of eel Vg.

• Proceedings of the Korean Society of Fisheries Technology Conference
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• 1995.06a
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• pp.159-164
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• 1995

• Journal of fish pathology
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• v.23 no.2
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• pp.245-254
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• 2010

Effects of heavy metal ions on the gene expression of cytochrome P4501A (CYP1A) were examined in cultured eel hepatocytes. When the expression of CYP1A mRNA was measured by RT-PCR after incubation of eel hepatocytes with benzo[$\alpha$]pyrene (B[$\alpha$]P) at concentrations of 10-8~10-5 M, the CYP1A expression increased with B[$\alpha$]P treatment in a dose dependent manner, showing significant increase at concentrations more than 10-7 M. When the eel hepatocyte was treated with cadmium (10-6 and 10-5 M), the expression of CYP1A was inhibited and especially at higher concentration (10-5 M). The inhibition of CYP1A expression by cadmium was also observed in cells treated with B[$\alpha$]P. In another study, effects of heavy metal ions on the expression of CYP1A were examined in cultured hepatocytes isolated from eel which was treated previously with B[$\alpha$]P in vivo. Hepatocytes isolated from the liver of eel taken at 48 hours after injection of B[$\alpha$]P (10 mg/kg) were cultured for 2 days with cadmium, copper, lead or zinc (10-6 and 10-5 M). The expression of CYP1A was found to be suppressed by the metal ions compared with the control in which CYP1A was induced with previous treatment of B[$\alpha$]P in vivo. The present results may provide an important basic information for studying the effects of heavy metal ions on CYP1A expression in other species of fish and studying toxicological mechanisms of heavy metal ions in aquatic livings.

• Development and Reproduction
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• v.13 no.4
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• pp.217-226
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• 2009

Effects of sera from several fish species on monolayer formation, viability and functions of catfish hepatocytes were investigated to establish a primary hepatocyte culture system for screening endocrine disruptors. Hepatocytes of Korean catfish (Silurus asotus) were attached and formed monolayer using the media supplemented with their own serum or sera from eel and tilapia, but not with fetal bovine serum (FBS). The amount of fish sera (0.5~3%) for monolayer culture of the catfish hepatocytes was less than 1/10 of FBS (5~20%) that is commonly used for primary culture of hepatocytes of other species. The results indicate that FBS can be replaced with sera from some fish species and the fish sera are more effective than FBS in maintaining the shape and functions of the hepatocytes. The primary culture of catfish hepatocytes was maintained monolayer with fish sera for at least 10 days, which makes possible to be used for screening the activities of endocrine disruptors. In conclusion, the primary culture system of hepatocytes with fish sera in the present study could be a useful tool for screening and studying endocrine disruptors.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.30 no.2
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• pp.282-290
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• 1997

Hepatocytes of Anguilla japonica have been prepared using a collagenase perfusion technique. The isolated cells attached efficiently to fibronectin-coated culture dishes and subsequently formed monolayers in serum-free medium. These cultures maintained in appropriate medium at least for 10 days with minimal cell loss. The effects of estradiol and pituitary hormones on vitellogenin (Vg) synthesis were examined in primary hepatocyte culture of the immature eels. In fish, as in other oviparous vertebrates, estrogen is a major inducer of Vg synthesis. However, $estradiol-17\beta(E_2)$ alone was insufficient to induce Vg synthesis in cultures of eel hepatocytes. Combination of $E_2$ with growth hormone (GH) and/or prolactin (PRL) markedly stimulated Vg synthesis. Even in cultures exposed to $E_2$ or precultured without hormones for 8 days, $E_2$ alone could not fully induce Vg synthesis. The synthesis of Vg was dramatically increased when hepatocytes were cultured in medium supplemented with $E_{2}+GH+PRL$ for 6 days. At this point, even though GH and/or PRL were eliminated from the medium, Vg synthesis was not influenced by these factors during culture of further 3 days. These results indicate that pituitary hormones, in particular GH and PRL, play important roles in the regulation of Vg synthesis in primary cultures of eel hepatocytes.

• Journal of fish pathology
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• v.20 no.3
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• pp.315-325
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• 2007

Histological changes in gill and liver of eel, Anguilla japonica (average weight 96±3.6 g) were examined with formalin bath at 0~500 ppm for 1, 6 and 24 h. Hyperplasia, hypertrophy, cell fusion, desquamation and necrosis of epithelial cells at gill lamella and gill filament were observed from 6 h at 200 ppm, 1 h at 300 ppm, 1 h at 400 ppm and 1 h at 500 ppm. In the exposure of formalin 100 ppm for 24 h, epithelial cells arrangement of gill filament and gill lamella showed thinner and more regular order than the control. trophy and pyknosis of hepatocytes, congestion at sinus or central vein, degeneration of cytoplasm were observed in the liver from 24 h at 100 ppm and 200 ppm, 6 h at 300 ppm, 1 h at 400 ppm and 500 ppm. However, there were not any histological changes at liver of 100 ppm-1, 6 h, 200 ppm-1, 6 h and 300 ppm-1 h compared with the tissue of control.