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마우스 배아줄기세포를 이용한 심근세포 분화촉진 물질 탐색
남기환 ( Ki Hoan Nam ),김은경 ( Eun Kyoung Kim ),원영석 ( Young Suk Won ),문옥성 ( Ok Seong Moon ),안병우 ( Byeong Woo Ahn ),김형진 ( Hyoung Chin Kim ) 한국동물실험대체법학회 2007 동물실험대체법학회지 Vol.1 No.2
Embryonic stem cells are derived from inner cell mass of blastcyst. They are pluripotent in differentiation and show self-renewal ability. Therefore, the cells can differentiate into cell types which can be derived from all the three germ layers. In this report, we screened 100 compounds for their potential effects on the differentiation of KH2 mouse embryonic stem cells to cardiomyocytes. KH2 cells are developed in our lab and express enhanced green fluorescence protein under the direction of α-myosin heavy chain gene promoter. This cell line shows no fluorescence in undifferentiated condition. However, cardiomyocytes derived from the cells show green fluorescence. Test compound was added during the differentiation period of the cells. After 15 days of differentiation, the cells were trypsinized and measured by flow cytometry for their fluorescence protein expression. All tested compound did not show any significant increase in the rate of fluorescent cells compared with that of their control group. However, we could confirm that our cell screening system is very stable.
연구논문 : 마우스 배아줄기세포를 이용한 심근세포 분화촉진 물질 탐색
남기환 ( Ki Hoan Nam ),김은경 ( Eun Kyoung Kim ),원영석 ( Young Suk Won ),문옥성 ( Ok Seong Moon ),안병우 ( Byeong Woo Ahn ),김형진 ( Hyoung Chin Kim ) 한국동물실험대체법학회 2007 동물실험대체법학회지 Vol.1 No.2
Embryonic stem cells are derived from inner cell mass of blastcyst. They are pluripotent in differentiation and show self-renewal ability. Therefore, the cells can differentiate into cell types which can be derived from all the three germ layers. In this report, we screened 100 compounds for their potential effects on the differentiation of KH2 mouse embryonic stem cells to cardiomyocytes. KH2 cells are developed in our lab and express enhanced green fluorescence protein under the direction of α-myosin heavy chain gene promoter. This cell line shows no fluorescence in undifferentiated condition. However, cardiomyocytes derived from the cells show green fluorescence. Test compound was added during the differentiation period of the cells. After 15 days of differentiation, the cells were trypsinized and measured by flow cytometry for their fluorescence protein expression. All tested compound did not show any significant increase in the rate of fluorescent cells compared with that of their control group. However, we could confirm that our cell screening system is very stable.
A Potential Demerit of the Pronuclear Microinjection Technique
왕애국,김선욱,문형배,현병화,남기환,서준교,김남순,유대열,이동석,Wang, Ai-Guo,Kim, Sun-Uk,Moon, Hyung-Bae,Hyun, Byung-Hwa,Nam, Ki-Hoan,Suh, Jun-Gyo,Kim, Nam-Soon,Yu, Dae-Yeul,Lee, Dong-Seok Korean Society of Life Science 2006 생명과학회지 Vol.16 No.4
Pronuclear microinjection (PMI) is a primary method for producing transgenic mice and offers a powerful tool for investigating gene function in vivo. The method has several reported advantages and disadvantages. Here, we report another potential shortcoming. The survival rate of fertilized one cell-stage embryos was significantly reduced after PMI procedure (65.4% (1202/1838)). In addition, the proportion of embryos developing to full-term was also significantly lower than that of embryos not undergoing PMI (26.5% (319/1202) vs 41.9% (57/136)). Moreover, 3 out of 21 (14.3%) founder control mice which were non-transgene-carrying littermates of transgenic founders showed histopathological changes in their liver, which was comparable to that in of transgenic lineages (4 out of 27 (14.8%)). In conclusion, the mechanical damages in chromosomes occurring during PMI procedure may be a potential factor influencing phenotypes of transgenic mice. 현재, 유전자의 in vivo 기능을 연구하기 위해 가장 많이 이용되고 있는 transgenic mice를 생산하기 위한 기본적으로 이용되고 있는 방법이 one cell-stage embryo에 pronuclear microinjection (PMI)이다. 그러나, 이 PMI 후에 one cell-stage embryo들의 생존율은 현저히 감소 (65.4%)할 뿐만 아니라 PMI 후의 embryo의 출생률(26.4%)이 PMI 처리를 하지 않은 것 (41.9%) 보다 현저히 낮다. 더욱이, PMI 방법에 의해 태어난 transgenic founder들의 간 조직에 병리학적 변화가 14.8% 정도에 대해서 같은 한배의 새끼 non-transgenic founder들의 경우도 간 조직에 병리학적 변화가 14.3%로 나타났다. 결론적으로, 이 PMI 방법에 의한 염색체에 물리적 손상은 형질전환 마우스의 생산 및 표현형에 영향을 미치는 잠재적 요소로 생각된다.
마우스배아줄기세포의 in vitro 시험계 활용을 위한 신경세포 분화프로토콜의 비교
김해림 ( Hae Rim Kim ),남기환 ( Ki Hoan Nam ),김은경 ( Eun Kyoung Kim ),윤원기 ( Won Ki Yoon ),원영석 ( Young Suk Won ),문옥성 ( Ok Seong Moon ),정의배 ( Eu Bae Joung ),안병우 ( Byeong Woo Ahn ),김형진 ( Hyoung Chin Kim ) 한국동물실험대체법학회 2008 동물실험대체법학회지 Vol.2 No.1
Mouse embryonic stem cells are pluripotent stem cells that can be differentiate into all the cell types derived from three germ layers in vitro. We aimed to confirm the neuronal cell types derived from the embryonic stem cells by two different differentiation protocols, which would guide us which protocol is useful for in vitro neuronal toxicity test. The mouse embryonic stem cells derived from 129 mouse strain, TC-1 cells, were differentiated according to 30-day differentiation protocol or 15-day differentiation protocol. At the end of the differentiation period, neuronal cells (neuron, astrocyte and oligodendrocyte) were identified by immunocytochemistry using marker antibodies. According to the results, the numbers of astrocytes and oligodendrocytes were much higher than that of neurons in 30-day differntiation protocol. However, oligodendrocytes were overwhelming compared to astrocytes and neurons in the 15-day differntiation protocol. These results indicated that the neuronal cell types and the cell numbers derived from the embryonic stem cells should be considered when selecting in vitro neuronal cell toxicity models using embryonic stem.
이영순,조재진,강경선,김배환,남기환,서광원,강성근,임윤규,허강준,Lee, Yong-soon,Cho, Jae-jin,Kang, Kyung-sun,Kim, Bae-hwan,Nam, Ki-hoan,Seo, Kwang-won,Kang, Seong-keun,Lim, Yun-kyu,Heo, Kang-jun 대한수의학회 1994 大韓獸醫學會誌 Vol.34 No.3
This study was performed for assessing carcinogenicity of Folpet using medium-term carcinogenicity bioassay. Sprague-Dawley rats aged six weeks divided into four grout's and were initially given an intraperitoneal injection of diethylnirosamine at 200mg/kg body weight. Two weeks later, group 1(negative control) was treated with basal diet. A Folpet was given per oral administration to group 2(100 ppm) and goup 3(1,000 ppm). Group 4 was fed on water containing 0.05% phenobarbital sodium as a promtor for six weeks. At three weeks after beginning of the experiment, partial hepatectomy was performed in all rats. The tumor-promoting effects were examined by the numbers and areas per $cm^2$ of induced glutathion S-tranferase placetal form(GST-P) positive foci in liver, and silver stained nucleolar organizer regions(AgNORs) which have recently introduced as one of the indicators for the cell proliferative activity. As the results, Folpet didn't have tumor-promoting effects on GST-P positive foci developement and AgNORs during promoting stage after initiation, whereas phenobarbital sodium treatment group showed promoting effect. It was concluded that Folpet didn't have promoting effect at 500, 1,000 ppm using this midium-term carcinogenicity bioassay model.
이영순,강경선,신동진,김형욱,조재진,김배환,남기환,서광원,Lee, Yong-Soon,Kang, Kyung-Sun,Shin, Dong-Jin,Kim, Hyoung-Ook,Cho, Jae-Jin,Kim, Bae-Hwan,Nam, Ki-Hoan,Seo, Kwang-Won 한국독성학회 1992 Toxicological Research Vol.8 No.2
SKI 2053R and SKI 2053R-human serum albumin(HSA) mixture were examined for their antigenicity in Hartley guinea pigs as well as C57BL/6 mice in comparison with distilled water (DW), HSA and DW-HSA conjugate. Several antigenicity tests, including acitive systemic anaphylaxis(ASA), passive systemic anaphylaxis (PSA), passive cutaneous anaphylaxis (PCA) and indirect hamagglutination test (IHA), were performed according to the Established Regulations of National Institute of Safety Research. The results were as follows: 1. When guinea pigs were sensitized with SKI2053R or SKI2053R-HSA emulsified with complete Freund's adjuvant(CFA), these animals showed negative reactions in ASA and PSA. 2.No blue spot was observed on the back skin of guinea pigs in the PCA test. 3. Sera from guinea pigs revealed a negative reaction in IHA. 4.Guinea pigs were sensitized by HSA emulsified with CFA as a positive control, and these animals showed positive reactions in ASA, PSA, PCA, and IHA. As shown above, SKI2053R was considered to possess neither antigenic, nor haptenic properties, and confirmed not to have the haemagglutinating activity.
김형욱,강경선,신동진,조재진,김배환,서광원,남기환,이영순,Kim, Hyoung-Ook,Kang, Kyung-Sun,Shin, Dong-Jin,Cho, Jae-Jin,Kim, Bae-Hwan,Seo, Kwang-Won,Nam, Ki-Hoan,Lee, Yong-Soon 한국독성학회 1992 Toxicological Research Vol.8 No.2
This study was performed to determine the toxic effects of graded dose levels of SKI 2053R after repeated administration. Three groups of Sprague-Dawley rats(10M and 10F per group) were given a total of 25 i.v. injections of SKI 2053R (1.50,3.75,9.38mg/kg/day). In order to compare the toxic effects of SKI 2053R with those of cisplatin, one group of Sprague-Dawley rats (10M and 10F per group) were given a total of 25 i.v.injections of cisplatin (1.70mg/kg/day). The dosing schedule was divided into five courses of 5 consecutive days with 16-day dose-free intervals between each course. No drug-related toxicity occurred in low dose level group (1.50mg/kg/day) of SKI2053R. From the results of hematological examination, peripheral WBC counts, RBC counts and hemoglobin of high dose level group(9.38mg/kg/day)of SKI 2053R were significantly lower than those of no-treated group. Other toxicities including reduced final body weight, proteinuria and hematuria were observed in high dose level group of SKI 2053R. But, no change was detected in serum biochemical values of SKI 2053R treated groups. All of the rats in cisplatin treated group were died between 3 and 13 weeks, while rats treated with SKI2053R survived to the end except one rat of middle dose level group(3.75mg/kg/day). In histopathological examinations, rats that received cisplatin manifested severe tubular damage in kidney and hemosiderosis in spleen, but no critical pathological lesion was observed in rats of other groups. Considering the results of this study, it was concluded that non-toxic dose of SKI 2053R in this treatment schedule was estimated to be 3.75 mg/kg/day and the maximum tolerated dose was to be higher than 9.38mg/kg/day. The toxic profiles fo SKI 2053R were different from those of cisplatin, and its toxicity was considerably lower than that of cisplatin.
Beagle dog 에서 DA-3030 ( G-CSF ) 의 정맥내 4 주간 반복투여 독성
이영순(Yong Soon Lee),조재진(Jae Jin Cho),남기환(Ki Hoan Nam),서광원(Kwang Won Seo),강성근(Sung Keun Kang),박재학(Jae Hak Park),김원배(Won Bae Kim) 한국응용약물학회 1994 Biomolecules & Therapeutics(구 응용약물학회지) Vol.2 No.3
This study was performed to determine the toxic effect of DA-3030(granulocyte-colony stimulating factor, G-CSF) in beagle dogs. DA-3030(G-CSF) was injected intravenously at doses of 115㎍/㎏/day, 11.5 ㎍/㎏/day and 1.15 ㎍/㎏/day seven days per week for 28 days. After completion of the treatments, the dog were necropsied. The number of dead animal was zero in all groups. No specific clinical sign was found, either. In hematological results, WBC was significantly increased dose-dependently in treated groups. In histopathological findings, megakaryocyte and rubricyte were found in the liver and spleen at the dose of 115㎍/㎏/day. Therefore, we could find the extramedullary hematopoiesis was increased. Megaka yocyte and rubricyte were increased in bone marrow, too. In conclusion, those signs were estimated the pharmacological effect of DA-3030(G-CSF). According to the results, non toxic dose of DA-3030(G-CSF) was higher than 115㎍/㎏/day