• 한국수정란이식학회지
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• 제33권4호
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• pp.343-348
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• 2018

Although the efforts to establish fish embryonic stem cells (ESCs) have been made for a long time, derivation of authentic ESCs that possess pluripotency is still difficult suggesting a need for the stepwise optimization of the methods to establish fish ESCs. Primary culture of the blastomeres from the embryos at blastula stage is a critical step for establishing continuous ESC lines. Here, we evaluated the effects of temperatures and basal media on primary culture of blastula embryo-derived blastomeres in marine medaka (Oryzias dancena). The blastomeres were isolated from the blastula embryos and cultured in various conditions designed by the combination of 4 temperatures including $28^{\circ}C$, $31^{\circ}C$, $34^{\circ}C$, and $37^{\circ}C$ and 2 basal media including Dulbecco's modified eagle's medium (DMEM) and Leibovitz's L-15 medium (L15). With the exception of a case cultured in L15 at $31^{\circ}C$, the rate of primary cell adherence reached 100% when the blastomeres were cultured over $31^{\circ}C$. The period for primary adherence was significantly shorter in the groups cultured in $34^{\circ}C$ and $37^{\circ}C$ than in the ones in $28^{\circ}C$ and $31^{\circ}C$. The proportion of subculture was significantly high in the group cultured in DMEM at $31^{\circ}C$ compared to the other groups. Collectively, we demonstrated that the culture in DMEM at $31^{\circ}C$ was effective to primary culture of the blastomeres derived from blastula embryos.

• Fisheries and Aquatic Sciences
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• 제5권1호
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• pp.32-35
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• 2002

In this study, we investigated the mechanism of cell death in rhabdovirus-infected cells, chinook salmon embryonic cell line (CHSE-2l4) infected with hirame rhabdovirus (HIRRV). Studies using light microscopy, fluorescence microscopy, TUNEL method, electron microscopy, and agarose gel electrophoresis revealed changes in the cell morphology and DNA fragmentation indicative of apoptosis in early infection. It was observed that HIRRV induced apoptosis as well as necrosis in infected cells.

• Fisheries and Aquatic Sciences
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• 제3권1호
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• pp.49-51
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• 2000

This study was performed to confirm the relationship between viral propagation and apoptosis by the infection of marine birnavirus strain (MABV NF-4) on chinook salmon embryo (CHSE-214) cells. After 6 hr viral infection, MABV was detected by PCR method. Also, as a result of DNA assay on the cells, MABV infection resulted in a typical feature of apoptosis, DNA fragmentation. The results suggest that MABV replicated to high concentrations during the early stage of infection induces apoptosis.

• 한국어병학회지
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• 제5권1호
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• pp.1-7
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• 1992

CHSE-214 무지개송어 세포주는 10% fetal bovine serum과 2mM-glutamine이 첨가된 Eagle's 기본배지에서 성장하였다. CHSE-214 세포주의 최적온도는 $20^{\circ}C$였다. IPN바이러스는 CHSE-214 세포주에서 복제되었으며, 세포변성효과를 나타내었다. 또한 IPN 바이러스의 감염시간에 따른 감염증상을 역위상차 현미경으로 관찰하였다. 감염 6-12시간 후 세포는 정상세포와 비슷하였다. 감염 18-24시간 후 세포는 일부가 정상 세포보다 더 둥근형태로 되었고, 일부세포는 세포배양용 플라스크 바닥에서 떨어져 부유상태로 되었다. 감염 30시간 후 세포는 더더욱 비정상적인 형태로 되었다. 감염 48~68시간 후 감염된 세포는 파열되었고, 아주 높은 세포 병변 효과를 나타내었다.

• Membrane and Water Treatment
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• 제14권3호
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• pp.141-146
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• 2023

Algal organic matters (AOMs) are challenging to remove using traditional water treatment methods. Additionally, they are recognized as disinfection by product (DBP) precursors during the chlorination process. These compounds have the potential to seriously harm aquatic creatures. Despite the fact that AOMs and DBPs formed from algae can harm aquatic species by impairing their cognitive function and causing behavioral problems, only a few studies on the effects of AOMs and associated DBPs have been conducted. To assess the impact of extracellular organic materials (EOMs) produced by three different hazardous algal species and the chlorinated EOMs on zebrafish, this study used fish acute embryo toxicity (FET) and cognitive function tests. With rising EOM concentrations, the embryo's survival rate and mental capacity both declined. Of the three algal species, the embryo exposed to Microcystis aeruginosa EOM exhibited the lowest survival rate. On the other hand, the embryo exposed to EOMs following chlorination demonstrated a drop in CT values in both the survival rate and cognitive ability. These findings imply that EOMs and EOMs treated with chlorine may have detrimental effects on aquatic life. Therefore, an effective EOM management is needed in aquatic environment.

• 한국어병학회지
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• 제20권3호
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• pp.257-267
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• 2007

The hypo-salinity effects on fertilized eggs, embryos and larvae were investigated in olive flounder (Paralichthys olivaceus) obtained from hatcheries in Cheju-Island, Yeosu and Chungnam. Those were treated to eight concentration; 0, 3.4, 6.7, 10.1, 13.4, 20.2, 27.4 and 33.6 ‰. It was not discrepancy in the survival rate and hatching success rate of fertilized embryos obtained from different regions. Also, in the larvae, the regional difference was not appeared. The survival rate and hatching ability of embryos significantly diminished in the lower groups than 13.4 ‰ compared to 33.6 ‰. After fertilization, namely embryos are tolerant of a wide range of salinity (13.4 - 33.6 ‰). Reduced salinity induced an increase of the malformed embryo and larvae including various deformities; irregular embryos membrane (or yolk sac depression), fin erosion and swim bladder inflation in the flounder embryo. The hatching success of embryos was significantly reduced in lower salinity than 13.4 ‰. Notably, the reduction of larval survival rate significantly was observed in ≤10.1 ‰ treated groups with the same manner of survival rates of the embryos. Additionally, olive flounder was found to be adequate model for measuring external impulses because there are no the regional differences.

• Fisheries and Aquatic Sciences
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• 제16권3호
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• pp.177-185
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• 2013

The development of species-specific fish cell lines has become a valuable tool for biological research. In recent years, marine medaka Oryzias dancena has been recognized as a good experimental model fish but there are no reports of establishment of cell lines from this fish. In this study, two cell lines from O. dancena blastula embryos were established from 41 total trials (4.9%). The two cell lines displayed typical in vitro morphology and have been cultured for >121 passages, which corresponds to 293 days. The doubling times of the cell lines were 29.84 and 28.59 h, respectively, and both possessed the potential to expand in a clonal manner, albeit with significant differences between the two cell lines. The absence of any of the four main medium supplements; i.e., fish serum, fetal bovine serum, basic fibroblast growth factor, and medaka embryo extract, significantly inhibited growth. The proportion of cells possessing normal chromosome number was 45% and 46.7% of the cell lines, respectively. Taken together, two cell lines that proliferate continuously were established from marine medaka and these cell lines may provide a basic tool for characterizing the unique features of this fish species.

• 한국가축번식학회지
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• 제18권3호
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• pp.183-189
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• 1994

We examined early development in loach(Misgurnus mizolepis) embryos with parthenogenetic agents well-known in mammals. Female loach was superovulated with an intraperitoneal injection of 15 IU human chorionic gonadotrophin (hCG) per gram body weight. After 13 h of hCG injection, the oocytes were obtained from the abdomen. The oocytes were activated with 10% ethanol in tap water or fish Ringer's solution for 5, 10 and 15 minutes(eTW5, 10, 15 and eFRS5, 10, 15), respectively. The activation rates were 29% and 10% in eFRS10 and eFRS15, 5% and 6% in eTW10 and eTW15 by judging the cleaved blastomeres. Whereas, no parthenogenetic embryo was produced by tap water or fish Ringer's solution alone. The activation rate with the fish Ringer's solution was higher than that of tap water. No embryonic development was observed by calcium ionophore, A23187, at concentrations of 10, 20, 40 and 100$\mu$M when treated for 1, 2.5 and 5 minutes, respectively. The activation agents did not cause early development as in mammalian eggs. Therefore, the results suggest that fresh water fish may have a different egg activation pathway from that of mammals.

• 한국발생생물학회지:발생과생식
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• 제18권3호
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• pp.139-143
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• 2014

Different with other fishes, the guppies (Poecilia reticulata) is ovoviviparity, which retain their fertilized eggs within the follicle throughout gestation. The synchronously growing diplotene oocytes store nutrients in droplets and yolk, before their maturation and fertilization. The lecithotrophic strategy of development entails the provisioning of embryos with resources from the maternal yolk deposit rather than from a placenta, it allows the extracorporeal culture of guppy embryo. Studies on their early development of live bearers like the guppy including lineage tracing and genetic manipulations, have been limited. Therefore, to optimize conditions of embryo in vitro culture, explanted embryos from pregnant females were incubated in embryo medium (L-15 medium, supplemented with 5, 10, 15, 20% fetal bovine serum, respectively). We investigated whether the contents of FBS in vitro culture medium impact the development of embryos, and whether they would hatch in vitro. Our study found that in 5% of FBS of the medium, although embryos developed significantly slower in vitro than in the ovary, it was impossible to exactly quantify the developmental delay in culture, due to the obvious spread in developmental stage within each batch of eggs, and embryos can only be maintained until the early-eyed. And although in culture with 20% FBS the embryos can sustain rapid development of early stage, but cannot be cultured for the entire period of their embryonic development and ultimately died. In the medium with 10% and 15% FBS, the embryos seems well developed, even some can continue to grow after follicle ruptures until it can be fed. We also observed that embryonic in these two culture conditions were significantly different in development speed, in 15% it is faster than 10%. But 10% FBS appears to be more optimizing condition than 15% one on development process of embryos and survival rate to larvae stage.

• 한국해양바이오학회지
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• 제14권1호
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• pp.33-42
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• 2022

Tacrolimus, a type of macrolide produced by Streptomyces tsukubaensis, is widely used as an immunosuppressant. However, continuous exposure to tacrolimus causes oxidative stress in normal cells, ultimately inducing cell injury. Therefore, this study investigated whether luthione, a reduced glutathione, could inhibit tacrolimus-induced cytotoxicity in olive flounder (hirame) natural embryo (HINAE) cells. According to the results, luthione significantly inhibited tacrolimus-induced reduction in cell viability in a concentration-dependent manner. Additinally, although luthione unaffected autophagy by tacrolimus, tacrolimus-induced apoptosis was significantly suppressed in the presence of luthione. Luthione also markedly blocked DNA damage in tacrolimus-treated HINAE cells, associated with the inhibition of reactive oxygen species (ROS) generation. Additionally, tacrolimus cytotoxicity in HINAE cells was correlated with increased inflammatory response, also attenuated by luthione. Collectively, these results show that at least luthione protects HINAE cells against tacrolimus-induced DNA damage, apoptosis, and inflammation, but not autophagy, by scavenging ROS. Although additional in-vivo studies are required, this study's results can be used as a basis for utilizing luthione to reduce the toxicity of fish cells caused by excessive immune responses.