title: The escape of temperature-sensitive T antigen immortalized rat hepatocytes from conditional immortalization.
abstract: Conditionally immortalized hepatocytes (CIH) established with a gene for the temperature-sensitive mutant of the T antigen (tsT) have characteristics to stop proliferating and to differentiate at nonpermissive temperatures (37-39 degrees C) due to inactivation of the T antigen. Therefore, they may be a good alternative to primary hepatocytes for experimental investigations or clinical applications. Deinduction of the T antigen results in a transient increase of p53 in these cells, leading to reexpression of normal senescence because of the telomere attrition occurring during the early stages of immortalization. To determine this T antigen dependency for the maintenance of immortality, a type of rat CIH was cultured continuously at 39 degrees C. The frequency of occurrence of T-antigen-independent clones ranged from 0.053% to 0.093%. These clones maintained the temperature-sensitive property of the T antigen; nevertheless, they were able to progress to the S phase and proliferate without undergoing apoptosis at 39 degrees C as at 33 degrees C, a permissive temperature. The temperature-sensitive point mutation of tsT was not affected in these clones and the T antigen was functioning properly. The integrity of the p53 pathway was also maintained from the point of Western blot analysis of p21. Although the telomerase continued to be expressed and the telomere length was maintained, marked chromosomal damage could not be avoided in these cells. It is a plausible explanation that this escape phenomenon from conditional immortalization may be related to the change of other genes involved in cell cycles, which have yet to be elucidated. In conclusion, CIH could lose their temperature-sensitive characteristics without the change of tsT, itself, and the T antigen is not always necessary to maintain their immortality. Therefore, the results obtained from experimental investigations using these cells should be interpreted carefully, and unpredictable phenotypic changes should also be taken into consideration when using them in clinical applications.
authors.affiliation: Department of Internal Medicine, Kyung Hee University College of Medicine, Seoul, Korea. kimbh@khu.ac.kr
authors.forename: Byung-Ho
authors.initials: BH
authors.lastname: Kim
authors.forename: Yo Seb
authors.initials: YS
authors.lastname: Han
authors.forename: Bong-Keun
authors.initials: BK
authors.lastname: Choe
authors.forename: Hyeseong
authors.initials: H
authors.lastname: Cho
authors.forename: Gi Deog
authors.initials: GD
authors.lastname: Nam
authors.forename: Jin Woo
authors.initials: JW
authors.lastname: Lee
authors.forename: Young Il
authors.initials: YI
authors.lastname: Kim
authors.forename: Jai Kyung
authors.initials: JK
authors.lastname: Park
authors.forename: Seok Ho
authors.initials: SH
authors.lastname: Dong
authors.forename: Hyo Jong
authors.initials: HJ
authors.lastname: Kim
authors.forename: Young Woon
authors.initials: YW
authors.lastname: Chang
authors.forename: Joung Il
authors.initials: JI
authors.lastname: Lee
authors.forename: Rin
authors.initials: R
authors.lastname: Chang
mesh_terms: D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics*; D017209:Apoptosis; D002453:Cell Cycle / Q000502:physiology; D002461:Cell Line, Transformed; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D002478:Cells, Cultured; D043171:Chromosomal Instability / Q000502:physiology; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology; D022781:Hepatocytes / Q000166:cytology*; D017354:Point Mutation / Q000235:genetics*; D051381:Rats; D011917:Rats, Inbred Lew; D019098:Telomerase / Q000378:metabolism; D013696:Temperature; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
chemical_list: D000952:Antigens, Polyomavirus Transforming; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016159:Tumor Suppressor Protein p53; D019098:Telomerase