• 한국환경독성학회:학술대회논문집
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• 한국환경독성학회 2003년도 추계국제학술대회
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• pp.159-159
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• 2003

Comet assay is a potential monitoring tool because DNA strand breakage may be produced by a wide range of agents. The comet assay, also called the single-cell gell electrophoresis (SCGE) assay, is rapid and sensitive method for the detection of DNA damage in cells. This study was performed for the identification of DNA damage in the cells from flounders and clams exposed to PAHs. As a control experiments, flounder and clam cells were exposed to $H_2O$$_2$. The cells exposed to $H_2O$$_2$ were displayed a typical nuclei movement DNA damage of cells were significantly increased when the isolated cells from the blood of flounders and the tissue of clams were in vitro exposed to the different concentrations (5, 10, 50, 100 ppb) of five kinds of PAHs (benzo[a]pyrene, pyrene, fluoranthene, anthrancene, and phenanthrene). For the in vivo test, flounders and clams were exposed to the different concentrations of BaP for 4 days. The results showed that DNA strand breakage was effected by the concentration of BaP and the duration of exposure. In high concentration of BaP, the mean tail lengths of nuclei was longer than it In low concentration, while the mean size of head DNA decreased. In this research, both in vitro and in vivo genotoxicity of PAHs could be biomonitored by the comet assay. Especially, clams and flounders seem to be useful as materials for monitoring genotoxic damage by comet assay.

• Fisheries and Aquatic Sciences
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• 제13권4호
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• pp.284-293
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• 2010

To reduce anti-nutritional factors in plant protein sources for fish meal replacement in fish feeds, cottonseed and soybean meal (CS) were fermented with Aspergillus oryzae. A feeding trial was conducted to verify the effects of fermented CS (FCS) with phytase supplementation on gossypol detoxification, phosphorus digestibility, antioxidant activity, and growth performance of juvenile olive flounder over 10 weeks. Four diets were formulated to replace 0, 30, or 40% fish meal protein with CS or FCS (designated as CS0, CS30, FCS30P, and FCS40P). Phytase (1,000 FTU/kg) was added to FCS30P and FCS40P. The microbial fermentation significantly increased dietary total polyphenols and consequently led to higher DPPH radical-scavenging activities in fish feed and fish tissue. Dietary and liver gossypol concentrations were dramatically decreased by the fermentation process. Phosphorus digestibility was significantly increased in fish fed the FCS40P diet. However, growth performance decreased in fish fed FCS diets. This study demonstrates that the fermentation process and phytase supplementation can improve the phosphorus availability of plant protein sources in fish. The fermentation of CS by A. oryzae could increase antioxidant activities in feed and fish and effectively degrade toxic gossypol in cottonseed meal.

• 한국수산과학회지
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• 제51권1호
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• pp.42-46
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• 2018

Fetal bovine serum (FBS) is essential for cell culture and is used in the determination of infectivity titer and propagation of viruses. To clarify the effects of FBS on the propagation of viral haemorrhagic septicaemia virus (VHSV), which is a causative agent of mass mortalities of olive flounder Paralichthys olivaceus in Korea, VHSV was inoculated into an EPC (epithelioma papulosum cyprinid) cell line supplemented with MEMs (minimal essential medium) with FBS concentrations of 0%, 2%, 5%, and 10% (MEM0, MEM2, MEM5, and MEM10), respectively, and infectivity titers were compared. Cytopathic effects were observed in all experimental groups at 2 days post virus inoculation (dpi) and all cells were detached from cell culture flasks at 7 dpi. Infectivity titers increased to 3 dpi, persisted to 7 dpi, and decreased when cells were detached. The titer of VHSV in EPC cells in MEM0 was the lowest while those in the other experimental groups showed similar levels. In conclusion, 2% (v/v) of FBS was sufficient to propagate VHSV in EPC cells and the withdrawal of VHSV from cell culture flasks should be performed before cell detachment.

• 한국수산과학회지
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• 제51권1호
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• pp.15-22
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• 2018

Alpha-pinene is an organic compound that possesses antibiotic and anti-parasitic activities. In this study, we developed a layered nanocomposite to combat against Miamiensis avidus which causes scuticociliatosis in the olive flounder Paralichthys olivaceus. We used a solid-solid reaction to develop the layered nanocomposite, incorporating-pinene ($C_{10}H_{16}$) into organically modified montmorillonite. We used cetyltrimethylammonium cations as the interlayered modifier for the adsorption of hydrophobic pinene molecules. The X-ray diffraction patterns of the nanocomposite structure showed that the basal spacing increased from $9.6{\AA}$ to $30.4{\AA}$. Interestingly, the fraction of ${\alpha}$-pinene released remained constant for a long period of time (228 h) due to the layered nature of the nanocomposite. Additionally, optical microscopic images of the treated scuticociliatids revealed that their cells were lysed, and this effect increased with the increasing concertration of ${\alpha}$-pinene. Histopathological assessment of ${\alpha}$-pinene nanocomposite-treated olive flounder gills revealed no significant morphological changes, even at the highest concentration of the ${\alpha}$-pinene. The nanocomposite has several advantages, including easy handling, high solubility, low toxicity, and the easy formulation of granules or powder, which improve the pesticidal activity of ${\alpha}$-pinene. Collectively, our results suggest that ${\alpha}$-pinene nanocomposite may be a useful treatment against scuticociliatosis.

• Toxicological Research
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• 제28권3호
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• pp.159-164
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• 2012

Allergic skin inflammation such as atopic dermatitis (AD) is characterized by edema and infiltration with various inflammatory cells such as mast cells, basophils, eosinophils and T cells. Thymic stromal lymphopoietin (TSLP) is produced mainly by epidermal keratinocytes, as well as dermal fibroblasts and mast cells in the skin lesions of AD. Omega-3 polyunsaturated fatty acids in fish oil can reduce inflammation in allergic patients. Fermentation has a tremendous capacity to transform chemical structures. The antiinflammatory effects of fish oil have been described in many diseases, but the beneficial effects by which fermented olive flounder oil (FOF) modulates the allergic response is poorly understood. In this study, we produced FOF and tested its ability to suppress the various allergic inflammatory responses. The ability of FOF to modulate the immune system was investigated using a mouse model of AD. The FOF-treated group showed significantly decreased immunoglobulin E (IgE) and histamine in serum. Also, the increased TSLP expression was significantly inhibited in the FOF group; the FOF-treated group was not appreciably different from the hydrocort cream treatment group. In addition, FOF treatment resulted in a smaller spleen size with reduced the thickness and length compared to the induction group. Splenocytes from mice treated with FOF produced significantly less IFN-${\gamma}$, IL-4, T-box transcription factor (T-bet) and GATA binding protein 3 (GATA3) expression compared with the induction group. These results suggest that FOF may be effective in treating the allergic symptoms of AD. 5.

• 한국발생생물학회지:발생과생식
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• 제18권4호
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• pp.233-240
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• 2014

The early growth response protein 1 (Egr-1) is a widely reported zinc finger protein and a well known transcription factor encoded by the Egr-1 gene, which plays key roles in many aspects of vertebrate embryogenesis and in adult vertebrates. The Egr-1 expression is important in the formation of the gill vascular system in flounders, which develops during the post-hatching phase and is essential for survival during the juvenile period. However, the complete details of Egr-1 expression during embryo development in olive flounder are not available. We assessed the expression patterns of Egr-1 during the early development of olive flounders by using reverse transcription polymerase chain reaction (RT-PCR) analysis. Microscopic observations showed that gill filament formation corresponded with the Egr-1 expression. Thus, we showed that Egr-1 plays a vital role in angiogenesis in the gill filaments during embryogenesis. Further, Egr-1 expression was found to be strong at 5 days after hatching (DAH), in the development of the gill vascular system, and this strong expression level was maintained throughout all the development stages. Our findings have important implications with respect to the biological role of Egr-1 and evolution of the first respiratory blood vessels in the gills of olive flounder. Further studies are required to elucidate the Egr-1-mediated stress response and to decipher the functional role of Egr-1 in developmental stages.

• 한국어병학회지
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• 제17권2호
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• pp.91-98
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• 2004

본 연구의 목적은 국내 어류 연쇄구균증 병원체 동정에 있어서 Streptococcus iniae에 대한 구체적인 국내 연구 보고가 없어, 기존에 수행되고 있는 동정방법을 근간으로 16S-23S intergenic spacer region (ISR)의 염기서열을 분석하여 보다 명확한 동정을 실시하고자 하였다. API 20 strep system에 의한 생화학적 성상을 분석해본 결과 정확한 종 동정은 이루어지지 않았지만 기존의 연구에서 보고된 API 20 strep system에 의한 S. iniae의 생화학적 성상과 높은 유사성을 보이는 10균주를 분리할 수 있었다. S. iniae 16S rRNA gene 서열 유래 종 특이적 primer Sin-1 5'-CTAGAGTACACATGTACT(AGCT)AAG-3'와 Sin-2 5'-GGATTTTC CACTCCCATTAC-3'에 의한 PCR-assay 결과 시험균주 10 균주 모두 동일하게 약 300bp의 증폭산물이 관찰되었고, 비 특이적인 반응은 관찰되지 않았다. 시험균주 모두 3개의 16S-23S ISR operon 구조가 관찰되었으나 S. iniae 표준균주 (KCTC 3657)의 경우 단일 구조가 관찰되었다. 16S-23S intergenic spacer region (ISR)의 염기서열을 분석해본 결과 기존에 보고된 S. iniae (Genbank accession number AF 048773)와 96%의 상동성을 보였으며 전체서열에서 상동성을 보이는 근연종이나 근연속은 검색되지 않아 최종적으로 S. iniae로 동정하였다.

• 한국어병학회지
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• 제17권2호
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• pp.131-137
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• 2004

본 실험은 넙치 (Paralichthys olivaceus) 간으로부터 membrane부분에 존재하는 PC 가수분해 효소의 특성에 대해 조사하였다. 먼저 간 조직을 초고속 원심 분리기와 nonion-detergent인 1% triton X - 100을 이용하여 membrane 부분을 분리하였으며, Heprain-Sepharose CL-6B 칼럼과 Heparin-5PW 칼럼을 이용하여 분리정제 하였다. 얻어진 PC 가수분해효소에 대한 반응 ․ 생성물을 확인하기 위해 autoradiography를 실시하였다. 지용성 부분의 결과에서 transphosphatidylation 반응의 결과물인 PEtOH을 형성하는 것으로 보아 PC-PLD임을 알 수 있었다. 얻어진 PC 가수분해효소에 대한 생화학적 특성을 조사한 결과 적정 pH가 6.5이하인 산성 조건 및 $37^\circ{C}$의 배양온도에서 최고 활성을 나타내었으며, 이가 이온들에 대한 영향의 경우 칼슘은 1.67mM 농도에서 최고 활성을 나타냈으나, 마그네슘은 활성에 영향을 미치지 않았다. 각종 세포막 기질에 대한 영향을 조사한 결과 PC는 $0.75\mu{M}$, PIP2는 $2.35\mu{M}$, PE는 $26.8\mu{M}$ 농도에서 최고 활성을 나타내었다. 이상의 결과부터 넙치 간조직의 막층부분에 존재하는 PC를 가수분해효소는 PC-PLD가 존재함을 알 수 있었다.

• 한국어병학회지
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• 제21권3호
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• pp.247-257
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• 2008

To understand the activity of non-specific defence factors in cultured Sebastes, the antibacterial effect of the serum, skin mucus and homogenate of various organs from cultured oblong rockfish (Sebastes oblongus) and rockfish(Sebastes schlegeli) against pathogenic bacteria, Aeromonas hydrophila, Edwardsiella tarda, Vibrio anguillarum, and Streptococcus sp. was compared with that of flounder(Paralichthys olivaceus) and seabass(Leteolabrax japonicus). And the activities of proteolytic enzyme, chitinolytic enzyme and haemolycin as non-specific defence factor were investigated on the oblong rockfish and rockfish. Samples from oblong rockfish showed the highest antibacterial activity by lysoplate assay on agar plate mixed with pathogens, followed in descending order by rockfish, seabass, and flounder. Turbidimetric assay was carried to evaluate the lysozyme activity of fish samples against lyophilized cells of Micrococcus lysodeiktikus. The serum, kidney, liver, stomach, intestine and eyeball of oblong rockfish and the mucus and gill of rockfish appeared to have the highest lysozyme activity among the fish strains investigated. All samples except skin mucus, liver, and eyeball of oblong rockfish and rockfish showed proteolytic enzyme activity. Chitinolytic enzyme activity was showed in random sampling and haemolytic activity was remarkable in oblong rockfish. Therefore, Sebastes strain was proved to have effective defense mechanisms based on the antibacterial activities, and lysozyme, proteolytic enzyme, chitinolytic enzyme, and haemolycin were considered to act as the non-specific defence factor of Sebastes.

• 한국어병학회지
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• 제29권1호
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• pp.13-23
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• 2016

Lectins play significant roles in the innate immune responses through binding to pathogen-associated molecular patterns (PAMPs) on the surfaces of microorganisms. In the present study, tissue distribution and expression analysis of olive flounder lectins were performed after viral hemorrhagic septicemia virus (VHSV) challenge. Fish egg lectin and serum lectin were found to be predominantly expressed in the gills and liver, these results indicate that the transcript expression of olive flounder lectins is concentrated in immune-related tissues. Following a VHSV challenge, an overall increase in the transcript levels of the genes was observed and the expression patterns were distinctly divided into early and later responses during VHSV infection. In conclusion, olive flounder lectins are specifically expressed in immune-related organs and induced in both the immediate and long-lasting immune responses to VHSV in the olive flounder. These results indicate that lectins may be play important roles in the host defense mechanism and involved in the innate and adaptive immune response to viruses in fish.