• Korean Journal of Environmental Biology
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• v.22 no.2
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• pp.329-335
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• 2004

To develop a biomarker for the monitoring of the contamination of estrogenic endocrine disrupters in the aquatic environment, reverse transcription -polymerase chain reaction (RT-PCR) analysis of vitellogenin (Vg) mRNA expression was optimized in Bombina orientalis, a Korean red bellied toad species. Based on partial cDNA sequences of both Vg and beta actin genes of B. orientalis, specific primers for RT-PCR of Vg and beta actin mRNAs were developed. Semiquantitative RT-PCR of the Vg mRNA in liver was optimized using a beta actin mRNA as an internal control in both sexes. In female RT-PCR using $1\;\mu{g}$ of the liver cDNA resulted in a linear increment in the PCR product of Vg from 18 to 34 cycles of amplification. In male, on the contrary, the RT- PCR product was first detected at 30 cycles of amplification and a linear increment was observed from 30 to 40 cycles of amplification, suggesting that male B. orientalis expresses minute amount of Vg mRNA which is a $2^{-12}$ equivalent of female. In conclusion, the optimized protocol for semiquantitative RT-PCR analysis of Vg mRNA level in B. orientalis male liver will be useful for the environmental monitoring the xenoestrogen contamination in the freshwater environment in Korea.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.40 no.1
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• pp.16-23
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• 2007

Nonylphenol (NP) is an estrogen-mimicking compound or xenoestrogen. This study investigated the effects of nonylphenol on the reproductive status of the equilateral venus Gomphina veneriformis. The experiment lasted 24 weeks, Experimental groups consisted of a control and three nonylphenol exposures ($1.0,\;2.5,\;and\;5.0\;{\mu}g\;NP/L$). Mortality did not differ significantly between the control and the exposure groups. The sex ratio (F:M) was 1:1 in nature and 1:1.03 in the control group. However, it changed to 1:3.5 with $5.0\;{\mu}g\;NP/L$ exposure. Gonad maturity in females was higher in the nonylphenol exposure groups than in the control group. By contrast, in males, it was lower in the nonylphenol exposure groups. Intersex individuals constituted 0% in nature, 3.08% in the control group, and 23.6% in the group exposed to nonylphenol, with female characteristics more prevalent than male. As the concentration of nonylphenol increased, the accumulation of lipofuscin increased in the mid-gut gland.

• Environmental Mutagens and Carcinogens
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• v.22 no.3
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• pp.175-182
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• 2002

Endocrine disruptors (EDs) are the chemicals that affect endocrine systems through activation or inhibition of steroid hormone response. It is necessary to have a good system to evaluate rapidly and accurately endocrine-disrupting activities of suspected chemicals and their degradation products. The key targets of EDs are nuclear hormone receptors, which bind to steroid hormones and regulate their gene transcription. We constructed a co-expression system of Gal4p DNA binding domain (DBD)- ligand binding domain of human estrogen receptor $\alpha$ or $\beta$, and Gal4p transactivation domain (TAD)-co-activator AIB-1, SRC-1 or TIF-2 in Saccharomyces cerevisiae with a chromosome-integrated lacZ reporter gene under the control of CYC1 promoter and Gal4p binding site (GAL4 upstream activating sequence, GAL4$_{UAS}$). Expression of this reporter gene was dependent on the presence of estrogen or EDs in the culture medium. We found that the two-hybrid system with combination of the hER$\beta$ LBD and co-activator SRC-1 was most effective in the xenoestrogen-dependent induction of reporter activity. The extent of transcriptional activation by those chemicals correlated with their estrogenic activities measured by other assay systems, indicating that this assay system is efficient and reliable for measuring estrogenic activity. The data in this research demonstrated that the yeast detection system using steroid hormone receptor and co-activator is a useful tool for identifying chemicals that interact with steroid receptors.s.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.2
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• pp.131-140
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• 2001

Objective: To know the effects of xenoestrogen on spermatogenesis, we investigated the expression of cytochrome P450s enzymes (CYPscc, $CYP_{17{\alpha}}$, CYP19) and $3{\beta}$-HSD genes involved in steroidogenesis. Methods: Mouse testicular cells were prepared from 15-day-old ICR mice which had only pre-meiotic germ cells by enzyme digestion using collagenase and trypsin. Testicular cells were cultured in DMEM supplemented with FSH (0.1 IU/ml) and 10% FBS or medium with estrogen ($E_2$), bisphenol-A (BPA), octylphenol (OP; $10^{-9},\;10^{-7},\;10^{-6},\;10^{-5},\;10^{-4}M$, respectively) and aroclor 1254 (A1254) known as PCBs for 48 hours. The gene expression of cytochrome P450 enzymes were examined by semi-quantitive RT-PCR. The production of estrogen and testosterone was examined by RIA. Results: As results, expression of CYPscc mRNA was not significantly decreased, but $3{\beta}$-HSD and $CYP_{17{\alpha}}$. mRNA were significantly dose-dependent decreased. And production of testosterone and estrogen were not different except BPA and OP group ($10^{-5}M$). Conclusion: BPA, OP and A1254 might inhibit steroidogenesis by decreasing CYPscc, $3{\beta}$-HSD and $CYP_{17{\alpha}}$. mRNA expression in the mouse testis. These results suggest that BPA, OP and PCBs like as an endocrine disruptors inhibit the productions of steroidogenic enzymes and decrease the production of T and E by negative feedback mechanism. Therefore, these might disrupt steroidogenesis in Leydig cells of testis and would disturb testicular function and subsequently impair spermatogenesis.

• Korean Journal of Ecology and Environment
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• v.37 no.3 s.108
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• pp.313-318
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• 2004

Di-2-ethylhexyl phthalate (DEHP) is a widely used plasticizer known to be a suspected xenoestrogen and a causative agent of oxidative damage to the RBC cell membrane ir vitro. We evaluated the toxic effects of a scarcely documented aquatic environmental pollutant, DEHP, on selected haematological endpoints in the bagrid catfish, Pseudobagrus fulvidraco. Bagrid catfish were exposed to DEHP (300, 1,000 mg DEHP kg body weight$^{-1}$) through thrice intraperitoneal injection and effects were assessed in blood of the exposed organisms. Haematological property, serum organic and inorganic chemistry were monitored in blood of Bagrid catfish. DEHP exposed-fish showed erythropenia; low hematocrit (Ht) and hemoglobin (Hb) content and red blood cell count showed a significantly higher than in that of the control group. The treatment group showed a significantly lower concentration of serum total protein, cholesterol and triglyceride compared with those in the control group. The value of calcium and osmolality were significantly decreased in the DEHP treatment group, compared with the control group.