• Toxicological Research
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• 제20권3호
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• pp.251-261
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• 2004

A large number of chemical pollutants including phthalates, alkylphenolic compounds and organochlorine pesticides have the ability to disrupt endocrine function in animals, and alter cog-nitive function. Because hormone mediated events play an important role in central nervous system development and function, the changes in cognitive function seem to be mediated by the endocrine-like action of these chemicals. The present study therefore was designed to investigate effect of bisphenol A (BPA), an endocrine disrupting chemical on neuro-behavial patterns, and expression of estrogen receptors and tyrosine hydroxylase, a limiting enzyme of dopamine synthesis pathway. BPA was treated orally for 3 weeks into 3 week old mice, and then the neuro-behavial patterns (stereo-type behaviors such as jumping rearing and forepaw tremor, climbing behavior, tail flick, rotarod and locomotor activity), and the expression of estrogen receptors and tyrosine hydroxylase were deter-mined every 3 week for 9 weeks. During the treatment of BPA, the food uptake and body weight increase were not significantly changed. BPA resulted in the increased stereotype behaviors (jump-ing, rearing and forepaw tremor) 6 or 9 weeks after treatment. The time response to tail flick and locomotor activity were decreased by the treatment of BPA, whereas the time for rotarod was increased by the treatment of BPA. The expression of estrogen receptor alpha and beta was increased in the brain and pituitary gland. Maximum expression was found in the brain after 9 week of 100 mg/kg BPA treatment and in the pituitary gland after 6 week of 100 mg/kg BPA treatment. Tyrosine hydroxylase was increased in dose and time dependent manners in the brain but no change was found in the pituitary gland. The present data show that exposure of BPA in the young mice could alter expression of estrogen receptors and dopamine synthesis pathway, thereby modulate neuro-behavial patterns (increase of stereotype behaviors but decrease locomotor activity).

• 한국발생생물학회지:발생과생식
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• 제10권4호
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• pp.247-254
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• 2006

내분비 교란물질들이 연안 어류의 정상적인 성적 발달을 위협할 수 있다는 사실은 잘 알려져 있지만, 해산 어종에 대한 이러한 위협을 파악하는 데 이용할 수 있는 적절한 시험 어종은 아직 개발되지 못했다. 그 이유 중의 하나는 성분화 초기에 어류의 생식소가 외형으로나 조직학적으로 암수 구분이 불분명한 경우가 많기 때문이다. 본 연구에서는 조피 볼락 Sebastes schlegeli의 자연 성비, 생식소 구조, 그리고 외인성 성스테로이드 호르몬에 의한 성분화 교란 여부 등을 조사하여 이 종이 시험 어종으로 개발될 수 있는 지를 조사하였다. 1년생 조피볼락 240마리의 생식소를 조사한 결과, 이들의 성비(암수 비율)는 통계학적으로 1:1을 벗어나지 않았으며, 암컷은 유백색의 두터운 생식소를, 수컷은 검은색의 가느다란 생식소를 가지고 있었다. 성분화 시기의 조피볼락 치어를 $estradiol-17\;{\beta}(E_2)$ 또는 $17\;{\alpha}-methyltestosterone$ (MT)으로 처리하여 성분화 교란 여부를 조사한 결과, $E_2$ 처리는 93%의 암컷 유도율을 보였으나, MT 처리의 경우 수컷은 53%였고, 38%가 암수의 생식소 구조를 모두 갖는 intersex로 나타났다. 본 연구에 의하면 조피볼락은 미성숙 시기에도 암수 생식소의 형태가 뚜렷이 다르고, 외인성 성스테로이드 호르몬에 의해 뚜렷한 성분화 교란 현상이 초래되어 시험 어종으로 개발될 수 있는 가능성 있다.

• 미생물학회지
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• 제46권1호
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• pp.93-95
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• 2010

리그닌 분해효소 군을 가진 백색부후균들은 다핵방향족 화합물을 포함하여 염료와 폭약 및 내분비장애 물질의 분해 등 다양한 난분해성 물질을 분해할 수 있다. 리그닌 분해효소 중 laccase와 manganese peroxidase (MnP)를 각각 발현하는 벡터를 국내에서 분리한 백색부후균류의 하나인 아교버섯(Phlebia tremellosa)에 형질전환 방법으로 도입시킨 형질전환체를 사용하여 염료의 탈색능력을 분석하였다. Methyl green의 경우 3일 후 약 50%의 탈색을 보인 야생형에 비하여 laccase 형질전환체(TF2-1)와 MnP 형질전환체(T5) 모두 90% 이상의 탈색을 보였다. Remazol brilliant blue R (RBBR)에서는 야생형이 약 67%의 탈색을 보인 반면 두 가지 형질전환체들은 약 85%의 탈색을 보였다. 각각의 형질전환체들은 laccase와 MnP의 활성 및 유전자 발현도 활발한 것으로 확인되었다.

• Environmental Analysis Health and Toxicology
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• 제29권
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• pp.2.1-2.7
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• 2014

Objectives Heat shock protein 90 (HSP90) is a highly conserved molecular chaperone important in the maturation of a broad spectrum of protein. In this study, an HSP90 gene was isolated from Asian paddle crab, Charybdis japonica, as a bio-indicator to monitor the marine ecosystem. Methods This work reports the responses of C. japonica HSP90 mRNA expression to cellular stress by endocrine disrupting chemicals (EDCs), such as bisphenol A (BPA) and 4-nonylphenol (NP) using real-time. reverse transcription polymerase chain reaction. Results The deduced amino acid sequence of HSP90 from C. japonica shared a high degree of homology with their homologues in other species. In a phylogenetic analysis, C. japonica HSP90 is evolutionally related with an ortholog of the other crustacean species. The expression of HSP90 gene was almost distributed in all the examined tissues of the C. japonica crab but expression levels varied among the different body parts of the crabs. We examined HSP90 mRNA expression pattern in C. japonica crabs exposed to EDCs for various exposure times. The expression of HSP90 transcripts was significantly increased in C. japonica crabs exposed to BPA and NP at different concentrations for 12, 24, 48 and 96 hours. The mRNA expression of HSP90 gene was significantly induced in a concentration- and time-dependent manner after BPA or NP exposures for 96 hours. Conclusions Taken together, expression analysis of Asian paddle crab HSP90 gene provided useful molecular information about crab responses in stress conditions and potential ways to monitor the EDCs stressors in marine environments.

• Fisheries and Aquatic Sciences
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• 제14권1호
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• pp.22-30
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• 2011

Vitellogenin (Vg) is the precursor of vitellin (Vn), which is the major yolk protein in nearly all oviparous species, including fish, amphibians, reptiles, and most invertebrates. It is one of the most important factors during reproduction, and numerous studies have shown that Vg genes are markers of the reproductive cycle and effecter genes induced by endocrine-disrupting chemicals (EDCs). Previously, we isolated two distinct cDNAs encoding vitellogenin homologs Pj-Vg1 and Pj-Vg2 from Pandalus shrimp Pandalopsis japonica. In this study, full-length genomic sequences of Pj-Vg1 and Pj-Vg2 were determined using a PCR-based genome walking strategy. Isolated Pj-Vg1 and Pj-Vg2 genes were 11,910 and 11,850 bp long, respectively. Both Pj-Vg genes had 15 exons and 14 introns, and the splicing sites were also the same, suggesting that they arose via gene duplication. The similar structural characteristics of decapod Vg genes suggest that they are all orthologs that evolved from the same ancestral gene. Analysis of Pj-Vg1 and Pj-Vg2 expression revealed that the relative copy numbers of Pj-Vg1 and Pj-Vg2 were similar in the hepatopancreas, whereas Pj-Vg2 transcripts were also detected in the ovary. Expression of both Pj-Vg genes was induced in hepatopancreas of mature individuals, whereas only Pj-Vg2 transcripts were upregulated in the ovaries from mature animals, suggesting that both Pj-Vgs are important for oocyte development. A strong positive correlation was found between Pj-Vg1 and Pj-Vg2 transcripts in the same individual, indicating they are under the same control mechanisms. Additionally, a positive correlation was found between ovarian and hepatopancreatic Pj-Vg2 transcripts, suggesting that its dual expression is regulated by similar physiological conditions. Knowledge of the similarities and differences between the two vitellogenin-like genes, Pj-Vg1 and Pj-Vg2, would help us to understand their roles in reproduction and other physiological effects.

• Environmental Analysis Health and Toxicology
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• 제24권4호
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• pp.287-292
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• 2009

Over the a few decades, the incidences of obesity and associated metabolic syndrome diseases have been increased dramatically and resulted in a global health crisis. Recent findings suggest endocrine disrupting chemicals (EDCs) as an obesogen, because they disrupt normal development and homeostatic controls over adipogenesis and energy balance in obesity. Furthermore, risk of cardiovascular disease and mortality is elevated among those who were obese during childhood. Thus, we focused on etiology of obesity in children and performed biological monitoring of bisphenol A (BPA), which is a broadly exposed EDC in environment. Study subjects were age and sex-matched obese and normal children in Seoul (N=52; age, $8.67{\pm}1.46$ years). Exposure levels of BPA were analyzed with HPLC/FLD as a conjugated form in urine. As results, ranges of urinary BPA were 0~54.38 ${\mu}g/g$ creatinine (median, 4.57 ${\mu}g/g$ creatinine). Levels of urinary BPA were 1.7 fold higher in the obese children than those in the controls (medians of obese and control children, 7.31 and 4.25 ${\mu}g/g$ creatinine, respectively, p=0.22). In the near future, enlarge scaled studies should be performed to confirm the risk of BPA for obesity.

• Fisheries and Aquatic Sciences
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• 제12권4호
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• pp.276-285
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• 2009

Despite great commercial interest, relatively little has been described about molecular mechanism of bivalve reproduction. We investigated genes involved in ovarian maturation of the Yesso scallop, Patinopecten yessoensis. GSI index and histological analysis revealed that maturation of ovary begin in February and spawning period is from April to June which is similar to the previous study in the East Sea. As result of combination analysis of differential display RTPCR (DDRT-PCR) and histological examination, vitellogenin (Vg), ferritin (Ft) and ADT/ATP carrier protein (ACC) were identified as differently expressed genes in maturating ovary. Endpoint RT-PCR results showed that Vg is ovary-specific genes whereas Ft and ACC are expressed ubiquitously suggesting that Vg can be good molecular markers for ovarian development and sex determination in bivalves. Quantitative PCR results revealed that Vg were expressed highest during growth stage and appears to play a major role in oocyte maturation. On the contrary, expression of Ft was highest after spawning stage, which suggests that up-regulation may be involved in spawning and inactive stages in which the scallops recover from spawning. In addition, high level of the mitochondrial gene, ACC, may play a role in energy metabolism in maturating oocytes. Isolation and molecular studies of these key genes will expand our knowledge of the physiological changes from various exogenous factors including temperature, salinity, pH, even or numerous endocrine disrupting chemicals (EDCs) during reproductive cycle. In addition, further study of these genes implicates various industrial applications including the stable seed production, increased food quality, or economic aquaculture system.

• Biomolecules & Therapeutics
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• 제8권1호
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• pp.78-83
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• 2000

The objective of this study was to prevalidate the Organization for Economic Cooperation and Development's (OECD) rodent uterotrophic assay as a test method for screening of potential endocrine disrupting chemicals (EDCs). This study was conducted exactly as described in the OECD protocol documents. A positive control substance, 17$\alpha$-ethinyl estradiol (EE), was administered daily for three days to ovariectomized (OVX) Sprague-Dawley rats at various doses for determine the dose-response curve. Additionally, a pure antiestrogenic chemical, ZM189, 154 was administered to OVX rats at the same time EE to determine the effectiveness of the material against blocking the estrogenic effects of EE. At higher concentration of EE (10 $\mu\textrm{g}$/kg), a statistically significant difference in body weight gain and food consumption was observed compared to vehicle controls. In uterine responses, EE produced a dose-related increase in uterus weights compared to vehicle control. These increases were statistically significant at the >1.0 $\mu\textrm{g}$/kg doses. However, a similar dose-response relationship was not observed in vagina weight. A comparison of the two groups receiving ZM189,154 (0.1 and 1.0 mg/kg) with 0.3 $\mu\textrm{g}$/kg of EE and the group receiving only 0.3 $\mu\textrm{g}$/kg of EE showed dose-related decreases in uterus weights. However, statistical significance was shown in 1.0 mg/kg of ZM189,154. In conclusion, administration of EE produced a dose-related increase in uterine (wet and blotted) weights. Additionally, the 1.0mg/kg dose of ZM189,154 was effective in blocking the estrogenic activity of EE. These data suggest 3-day uterotrophic assay using OVX rats may serve as a good tool for EDCs screening.

• 환경생물
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• 제22권1호
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• pp.171-176
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• 2004

비이온성 계면활성제로 많이 쓰이는 알킬페놀류의 하나인 노닐페놀(nonylphenol)이 해산어류에게 미치는 영향을 조사하려고 주요 양식어종인 조피볼락에게 복강주사로 10 및 25 mg $kg^{-1}$을 1회 투여하였다. 한편 용제인 DMSO만을 주사한 sham구를 설정하여 비교하였다. 주사 후 7일간 간중량지수(hepatosomatic index)의 변화를 조사하였고, 또한 간장 미크로좀 중 대표적인 약물대사효소인 cytochrome P450 (CYP) 농도와 ethoxyresorufin deethylase (EROD) 활성의 변화도 측정하였다. 그 결과, 고농도(25 mg $kg^{-1}$) 주사구에서는 HSl가 감소하였는데, 이것은 NP가 간장에 독성이 있다는 것을 보여준다. 한편, 약물대사효소 중 CYP 농도는 유도되었지만 EROD 활성은 저해되는 경향을 보였다. 이번에 주사한 농도는 우리나라 연근해 해수 중의 NP 농도에 비해 매우 높은 수준이지만, 본 실험에서는 NP가 조피볼락에 게간장 독성을 보였고, 간장의 약물대사효소에도 영향을 미친다는 것을 확인할 수 있었다.

• Toxicological Research
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• 제16권3호
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• pp.201-209
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• 2000

In association with the international validation program to establish a rodent uterotrophic assay, we conducted preliminary uterotrophic assay proposed by GECD using immature female rats. In the present study, oral and subcutaneous routes were chosen to compare the effects of estrogenic com-pounds in the two dosing regimens. The reference compound ethinyl estradiol (EE) and the antagonist ZM189154(ZM) were administered by gavage or subcutaneously (s.c.) to immature female SD rats from 20 to 22 days of age. For each study, sixty-six female rats were randomly assigned to eleven groups: Untreated control, EE 0,0.01, 0.03, 0.1, 0.3, 1.0,3.0 and 10.0 $\mu\textrm{g}$/kg, EE 3.0 $\mu\textrm{g}$/kg(gavage)/0.3 $\mu\textrm{g}$/kg(s.c) & ZM 0.1 mg/kg, and EE 3.0 $\mu\textrm{g}$/kg(gavage)/0.3 $\mu\textrm{g}$/kg (s.c) & ZM 1.0 mg/kg. There were no treatment-related changes in clinical signs, body weights, food consumption, and necropsy findings in any groups of two studies. The wet and blotted uterus weights increased dose-dependently. Histopathological examination revealed that diameter of uterine duct, height of uterine luminal epithelium. and height oj vaginal epithelium increased dose-dependently. The proliferating cell nuclear antigen (PCNA) immunoreactive cells were increased in number dose-dependently. The estrogenic effects observed in the present studies occurred at $\geq$ 0.3 $\mu\textrm{g}$/kg of oral dose and $\geq$ 0.1 $\mu\textrm{g}$/kg of s.c. dose. An antagonistic effect of ZM against EE was found in both uterus weight and histopathological parameters. From the results obtained, it can be concluded that dose-dependence of the uterotrophic assay using EE and ZM was well demonstrated by gavage and subcutaneous administration and that the estrogenic effects of EE by s.c. dose were higher than those by gavage administration. In addition, blotted uterus weight was more sensitive than wet uterus weight and vaginal epithelial height was found to be the most sensitive parameter among the parameters examined.