• 생명과학회지
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• 제12권4호
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• pp.490-495
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• 2002

병원성 미생물인 Vibrio fluvialis로부터 hemolysin을 분리정제 하였다. 즉, hemolysin의 정제는 균 배양액을 황산암모늄, ion exchange chromatography를 행하였으며 SDS-PAGE를 통해 그 단백질의 크기가 약 79kDa임을 알게되었다. 정제된 hemolysin (VFH)는 35$^{\circ}C$에서 최적활성을 나타내었고, 4$0^{\circ}C$에서는 그 활성이 감소되었다. RTG-2 어류유래 세포주에 hemolysin (VFH)의 처리로 세포독성을 측정한 결과 50$\mu\textrm{g}$의 VFH가 약 80%의 cell line을 사멸시켰다. 또한 현미경을 통한 관찰에서도 세포의 형태변화를 관찰할 수 있었다.

• 한국해양생명과학회지
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• 제8권2호
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• pp.186-189
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• 2023

Uncoupling protein 1 (UCP1) is a unique mitochondrial membranous protein expressed in brown adipose tissue (BAT) in mammals. While its expression in response to cold temperatures and adipogenic inducers is well-characterized in mammals and human infants, the molecular characterization and expression of UCP1 in fish remain unexplored. To address this gap, we analyzed UCP1 expression in response to adipogenic inducers in a fish cell line, rainbow trout gonadal cells (RTG-2), and compared it with UCP1 expression in three mammalian preadipocytes, 3T3-L1, T37i, and WT1 exposed to the Peroxisome proliferator-activated receptor gamma (PPARγ) agonists, rosiglitazone (Rosi). In mammalian preadipocytes, UCP1 protein was highly expressed by Rosi, with an induction of adipogenesis observed in a time-dependent manner. This suggests that UCP1 plays a significant role in adipogenesis in mammals. However, RTG-2 cells showed no response to adipogenic inducers and exhibited only marginal expressions of UCP1. These results imply that RTG-2 cells may lack crucial responsive mechanisms to adipogenic signals or that the adipogenic response is regulated by other mechanisms. Further studies are needed to confirm these phenomena in fish preadipocytes when an appropriate cell line is established in future research.

• 한국어병학회지
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• 제16권2호
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• pp.69-73
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• 2003

In this study, GFP reporter gene was transfected into a fibroblast cell line RTG-2 using a polycationic transfection reagent (Superfect) and showed a successful expression of GFP. The transfection efficiency by Superfect was compared to the commonly used transfection method, i.e. DNA-calcium phosphate coprecipitatlon. Transfection by Superfect was more effective than calcium phosphate coprecipltation method (frequency of cell expressing orr was 11.3% and 3.5%, respectively). The optimal expression of GFP and {\beta}-galactosidase was observed when $5-6\;{\mu}{\ell}$ of Superfect per ${\mu}g$ DNA was used for transfcction, 1:5-6 ratio between DNA(${\mu}g$) and Superfect ($\mu\ell$).

• 한국어병학회지
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• 제11권2호
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• pp.89-96
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• 1998

해양버나바이러스(MABV)는 여러 종의 해양생물에 감염되며 숙주역이 넓다. 다양한 종의 숙주에 감염되었을 때의 MABV의 감염 특성을 규명하기 위하여, 주화세포 내에서 바이러스를 10대 계대 배양하여 in vitro로 연구했다. CHSE-214, RTG-2와 RSBK-2세포에서는 전형적인 CPE를 보이며 많은 양의 바이러스가 생산되었고, 높은 바이러스 단백질의 발현도 관찰되었다. 이에 반하여, EPC, FHM과 BF-2세포에서는 형광항체법에 의하여 바이러스단백질은 검출되었으나 CPE는 나타나지 않았다. EPC와 FHM 세포에서는 계대를 할수록 바이러스의 역가가 높아져, 바이러스의 숙주세포에의 적응이 일어난 것으로 보인다. 플라크의 크기는 CHSE-214, RTG-2와 RSBK-2세포에서 계대한 것이 다른 세포에서 계대한 것보다 커, 숙주세포의 종류에 따른 변이가 바이러스에 일어난 것으로 추측되었다. 게놈분절 A에 존재하는 VP2/NS 경계영역의 염기배열에서는 195번째의 염기에 특이적인 변이가 보였다. 숙주세포의 종류에 따라 다른 MABV의 감염특성은 자연계에서 다양한 숙주종에서 일어나는 in vivo에서의 감염특성을 반영하는 것으로 생각된다.

• 한국어병학회지
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• 제16권3호
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• pp.147-152
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• 2003

The CMV promoter driven luciferase reporter gene coding plasmid (pcDNA-luc) was constructed and used as a model for DNA immunization study. Expression of the recombinant luciferase protein was confirmed in vitro in RTG-2 cell line before using in vivo study in olive flounder. In dose response study, the maximum expression of the luciferase gene was found in the group injected with 10-15μg of plasmid DNA. The kinetic study showed that the luciferase gene expression was reached at the maximum level at one day after injection and slightly decreased after then but significantly high level of expression was sustained until the conducted experiment of 7 days. In the study of tissue distribution of gene expression, it was found that luciferase gene was expressed at the significant level in immune organs such as gill and spleen, located far from the injected site, suggesting the systemic distribution of the intramuscularly injected DNA in olive flounder.

• 한국어병학회지
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• 제15권1호
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• pp.9-16
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• 2002

The objective of the study was to clarify the mechanism of persistent infection of marine birnavirus (MABV) in various nonpermissive cell lines. It was observed in CHSE-214, RTG-2 and RSBK-2 that the virus produced at high yield with typical cytopathic effect (CPE). On the contrary, the CPE was not produced in EPC, FHM and BF-2 cells. However amount of virus protein in both permissive and nonpermissive cell lines detected by ELISA was almost the same. Electron microscopy showed virions in permissive cells but not in nonpermissive cells. From the results, it is clear that virus protein and RNA were produced in nonpermissive cells as observed in permissive cells; however, assembly of the virus particles did not occur in nonpermissive cells.

• Journal of Microbiology
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• 제34권3호
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• pp.253-269
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• 1996

Infection of fish cells with IHNV resulted in gradual increase in cytosolic free Ca$\^$2+/ concentration ([Ca$\^$2+/)] in CHSE, gradual decrease in [Ca$\^$2+/] in FHM, and no significant change in RTG cells. The degree of [Ca$\^$2+/] increase or decrease was dependent on the amount of infectious virus, and these [Ca$\^$2+/] variations were maximal at 16 hours after virus infection (p. i.) in both cell lines. When the fish cells were infected with inactivated IHNV, evident variation in [Ca$\^$2+/] was not observed. Thus, infectivity of IHNV appears to correlate with changes in [Ca$\^$2+/] in virus-infected cells. These IHNV-induced [Ca$\^$2+/] changes were partially blocked by cycloheximide, but not affected by cordycepin. It seems to be that virus-induced Ca$\^$2+/ variations were more related with protein synthesis than RNA synthesis. Various Ca$\^$2+/ related drugs were used in search for the mechanisms of the [Ca$\^$2+/], changes following IHNV infection of CHSE cells. Decreasing extracellular Ca$\^$2+/ concentration or blocking Ca$\^$2+/ influx from extracellular media inhibited the IHNV-induced increase in [Ca$\^$2+/], in CHSE cells. Similar results were obtained with intracellular Ca$\^$2+/ sources are important in IHNV-induced [Ca$\^$2+/] increase in CHSE cells.

• 한국어병학회지
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• 제11권2호
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• pp.129-136
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• 1998

1997년 3월 전라남도 및 경상남도의 해산어 육상 및 가두리 양식장에서 양성 중이던 넙치가 HRV(hirame rhabdovirus, Rhabdovirus olivaceus) 감염증과 유사한 증상을 나타내어 그 원인을 조사한 결과 새로운 rhabdovirus가 분리 되었다. 분리 바이러스(DF-9708)는 RTG-2 및 EPC 세포주에서 $15^{\circ}C$로 배양하였을 때 랩도바이러스 특유의 세포변성효과(CPE)를 나타내었으나 CHSE-214에서는 증식되지 않았다. 투과전자현미경으로 감염 세포내의 바이러스 입자 형태를 관찰해본 결과 bullet-shape의 크기 $70nm{\times}100\sim150nm$으로 인벨롭을 가진 바이러스였다. 분리바이러스는 pH 3 및 diethyl ether의 처리 조건에서는 감염성을 상실하였고, 열($50^{\circ}C$ 5 min, $60^{\circ}C$ 1 min)에도 약한 성상을 나타내었다. DNA 저해제인 IUdR $10^{-4}$M 처리에 의해서는 바이러스 감염가에 영향을 받지 않았다. 분리 바이러스는 Anti-HRV(8401-H) rabbit serum에 의해서만 중화 되었고, 전염성 조혈기 괴사증 바이러스(IHNV), 연어과 레오바이러스(CSV), 바이러스성 선회병 바이러스(RVS) 및 전염성 췌장괴사증 바이러스(IPNV) 등의 국내에서 분리되어지는 바이러스 항체로는 중화되지 않았다. 초원심법으로 정제한 분리 바이러스를 전기영동 한 결과 polymerase(L), glycoprotein(G), nucleoprotein(N) 및 2개의 matrix proteins(M1 및 M2)으로 구성되어져 있었으며, 각 단백질의 분자량은 L, 160 kDa; G, 55 kDa; N, 45 kDa; M1, 26 kDa; M2, 22 kDa의 크기로 계산되었다. 새롭게 분리된 바이러스는 외부증상으로는 기존의 HRV 감염과 유사한 점을 나타내었으나 그 바이러스학적 특성에 있어 약간의 차이점이 있어 본 분리바이러스를 우선 Nubchi rhabdovirus(NRV)(HRV-like)로 부르기로 한다.

• Journal of Microbiology
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• 제34권3호
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• pp.263-263
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• 1996

Infection of fish cells with IHNV resulted in gradual increase in cytosolic free $Ca^{2+}$ concentration $([Ca^{2+}]_i)$ in CHSE, gradual decrease in $[Ca^{2+}]_i$ in FHM, and no significant change in RTG cells. The degree of $[Ca^{2+}]_i$ increase or decrease was dependent on the amount of infectious virus, and these $[Ca^{2+}]_i$ variations were maximal at 16 hours after virus infection (p. i.) in both cell lines. When the fish cells were infected with inactivated IHNV, evident variation in $[Ca^{2+}]_i$ was not observed. Thus, infectivity of IHNV appears to correlate with changes in $[Ca^{2+}]_i$ in virus-infected cells. These IHNV-induced $[Ca^{2+}]_i$ changes were partially blocked by cycloheximide, but not affected by cordycepin. It seems to be that virus-induced $Ca^{2+}$ variations were more related with protein synthesis than RNA synthesis. Various $Ca^{2+}$ related drugs were used in search for the mechanisms of the $[Ca^{2+}]_i$, changes following IHNV infection of CHSE cells. Decreasing extracellular $Ca^{2+}$ concentration or blocking $Ca^{2+}$ influx from extracellular media inhibited the IHNV-induced increase in $[Ca^{2+}]_i$, in CHSE cells. Similar results were obtained with intracellular $Ca^{2+}$ blockers. Thus it is suggested that both the extracellular and the intracellular $Ca^{2+}$ sources are important in IHNV-induced $[Ca^{2+}]_i$ increase in CHSE cells.

• 대한두경부종양학회지
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• 제14권2호
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• pp.151-155
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• 1998

Background and Objectives: Numerous studies were conducted to develop radiosensitizers to increase antitumor effect and decrease systemic toxicity of ionizing radiation. In current study, the authors tested the synergistic effect of mutant $TNF-{\alpha}(M_3S)$ with radiation therapy on heterotransplanted hypoparyngeal squamous cell carcinoma. Materials and Method: SNU-1041 cell line was heterotransplanted to nude mice. When the tumors grew up to $70mm^3$ or more, the animals were randomly placed into 4 groups(n=10/group). Group I : 0.1ml of normal saline injected intraperitoneally once a day for 5 days. Group II : 10ug of $TNF-{\alpha}$ injected intraperitoneally once a day for 5 days. Group III : a single radiation dose of 10 Gy per animal delivered. Group IV : single radiation dose of 10 Gy was delivered 1 hour after intraperitoneal injection of $TNF-{\alpha}$ 10 ug. Results: Four weeks after treatment, group IV showed the least tumor growth during the 4 weeks follow up and the relative tumor growth rate(RTG) of each groups after 4 weeks were 31, 5.8, 10, and 3.2 respectively(p<0.05). Conclusion: These study suggests that pretreatment with $TNF-{\alpha}$ can significantly enhance the effects of radiation therapy and further studies may be needed for clinical trials of combination treatment of $TNF-{\alpha}$ and radiation.