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RISS 인기검색어
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- 저자 : 성세라(Se Ra Seong) (검색결과 2 건)
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SV40 T 항원의 온도조건부 변이형 유전자가 포함된 Amphotropic Retrovirus 에 의한 사람 태아 간세포의 불멸화
이정일(Joung Il Lee),동석호(Seok Ho Dong),김효종(Hyo Jong Kim),김병호(Byung Ho Kim),장영운(Young Woon Chang),장린(Rin Chang),성세라(Se Ra Seong),박재경(Jae Kyung Park),김승보(Seung Bo Kim),이상목(Sang Mok Lee) 대한내과학회 1999 대한내과학회지 Vol.57 No.1
N/A Human cells are almost never spontaneously immortalized in vitro. We tried to immortalize human fetal hepatocytes (h-FH) and evaluate the differentiational status and its change. Methods : Hepatocytes were isolated from a liver fragment of 20 week old fetus and infected with amphotropic recombinant retrovirus containing a temperature- sensitive mutant of SV40 large T antigen and neomycin phosphotransferase gene. G418 resistant colonies were cloned and expanded. The cells which were able to divide more than 30 times were used to analyze various functions. Results : The immortalization rate was 3.3 x 10-8 and two cell lines (C11, D21) were established. C11-60, C11-80, D21-30 and D21-60 (suffix number means the cell division counts) were evaluated. D21-30 was thougt to be imcompletely immortalized because a considerable portion of cells died during culture. The morphology was similar to that of epithelial cells except for D21-30 which looked like fibroblast. The cells grew rapidly at 33oC but stopped growing at 39oC. T antigen and p53 was expressed at 33oC but disappeared at 39oC, which suggest that T antigen binds to p53. Chromosomal changes were so marked that it was impossible to discriminate exact number. Albumin was secreted as about 1/10 as that of h-FH, but alpha-fetoprotein secretion stopped after immortalization. Telomerase was activated in both cell lines except for the incompletely immortalized cells D21-30. Telomere was elongated in competely immortalized cell lines, but it was rather shortened in D21-30 compared to that of h-FH. Macroscopic colonies did not develop in soft agar assay. Conclusions : We successfully immortalized human fetal hepatocytes. Although the cells are not likely to have oncogenicity, the functions are not so good, possibly due to marked chromosomal changes which are thought to occur before telomerase is activated during immortalization step.
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SV40 T항원에 의한 불멸화 간세포의 장기 계대 배양이 분화능 및 암성 변화에 미치는 영향
김병호,김효종,장린,이정일,동석호,장영운,성세라,이우인 대한소화기학회 1999 대한소화기학회지 Vol.34 No.1
Background/Aims: The transcription of liver specific genes decreases dramatically in primary hepatocyte culture. In this study, we evaluated the long-term differentiation of immortalized hepatocytes. Methods: Immortalized rat hepatocytes were established with a temperature-sensitive mutant of SV40 T antigen and were cultured until they divided more than 200 doublings. We measured the change of various differentiations and investigated its mechanism. Results: The epithelial cell morphology was not changed, but their size were decreased about 20% in the later passage, in which chromosomal changes were marked. In the early passage, albumin secretion and urea synthesis were reduced to 1/6 and 1/5 level of primary hepatocytes, respectively, and they were decreased to the undetectable level in the later passage. The expression of T antigen was increased about 29 times in the later passage. Telomerase activity was progressively increased according to the passage. In immortalized hepatocytes the mean telomere length was not changed, but the length variation increased. Conclusions: The differentiation of immortalized hepatocytes disappears after long-term passage possibly due to marked chromosomal changes that occur with compulsive and continuous replications by the increased expression of T antigen which inhibits sequentially the function of p53.
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