Immortalized hepatocytes established with a temperature-sensitive SV40 T antigen could be independent of T antigen.
- Author:
Seok Ho DONG
1
;
Byung Ho KIM
;
Gi Deog NAM
;
Jae Young JANG
;
Nam Hoon KIM
;
Sang Kil LEE
;
Kwang Ro JOO
;
Hyo Jong KIM
;
Young Woon CHANG
;
Joung Il LEE
;
Rin CHANG
Author Information
1. Department of Internal Medicine, School of Medicine, Kyung Hee University, Seoul, Korea. kimbh@khu.ac.kr
- Publication Type:Original Article
- Keywords:
Conditional immortalization;
Hepatocytes;
SV40 large T antigen;
Temperature-sensitive
- MeSH:
Antigens, Viral, Tumor*;
Apoptosis;
Cell Death;
Cell Line;
Clone Cells;
Hepatocytes*;
Point Mutation;
Telomerase
- From:Korean Journal of Medicine
2005;68(3):268-276
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
BACKGROUND: Conditionally immortalized hepatocytes (CIH) can be cultured almost indefinitely at permissive temperatures (33 degrees C), but they undergo apoptosis at nonpermissive temperatures (37~39 degrees C) by the release of p53 through inactivation of T antigen, which is called T antigen dependency. This study was aimed at examining if T antigen-independent clones can develop from CIH. METHODS: CIH established with a temperature-sensitive T antigen (WA1) were cultured continuously at 39 degrees C. Three clones (W39B, W39C, and W39J) survived at this temperature and was subject to following analyses: the morphology, growth, apoptosis, the expression of T antigen and p53, telomerase, and the T antigen gene sequence. RESULTS: WA1 proliferated at 33 degrees C with the population doubling time of 30.8 +/- 1.7 hours, but they underwent cell death at 39 degrees C. However, T antigen-independent clones (W39B, W39C, and W39C) proliferated at 39 degrees C without undergoing apoptosis, suggesting they lost the temperature-sensitive characteristics. WA1 expressed the T antigen at 33 degrees C, but not at 39 degrees C, and this temperature-sensitive pattern was maintained in T antigen-independent clones. In p53 expression, however, T antigen-independent clones revealed a different pattern. p53 was detected even at 39 degrees C where it normally would not be detected. Telomerase was activated in all the analyzed cell lines. A temperature-sensitive point mutation at nucleotide position 3505 of the WA1 was retained in all T antigen-independent clones. CONCLUSION: CIH can lost temperature-sensitive characteristics and acquire an ability to proliferate at nonpermissive temperatures. These changes might be related to the change of p53 rather than the change of T antigen itself in these cell lines.
