title: Establishment of ultra long-lived cell lines by transfection of TERT into normal human fibroblast TIG-1 and their characterization.
abstract: To establish useful human normal cell lines, TERT (telomerase reverse transcriptase) cDNA was transfected into normal female lung fibroblast, TIG-1. After long-term-sub-cultivation of 74 individual clones selected for resistance to G418, we obtained 55 cultures with normal range of life span [75 PDL (population doubling level)], 16 cultures with extended life span (75-140 PDL). In addition, 3 immortal cell strains and unexpectedly, one ultra long-lived cell line (ULT-1) with life span of 166 PDL were established. IMT-1, one of the immortal cell strains was confirmed to maintain long telomere length, high telomerase activity and an extremely low level of p16INK4A. They also showed moderate p53 and p21CIP1 expression, keeping vigorous growth rate even at 450 PDL. High level of fibronectin and collagen 1α expression confirmed IMT-1 as normal fibroblasts, although one X chromosome had been lost. ULT-1, however, kept a near normal karyotypes and had shortening of telomere length, high expression of p16INK4A, moderate levels of senescence associated-β-galactosidase positive cells and decreased growth rate only after 150 PDs (population doublings), and finally reached senescence at 166 PDL with morphology of normal senescent fibroblasts. As resources of standard normal human cell, abundant vials of early and middle passages of ULT-1 have been stocked. The use of the cell line is discussed, focusing on isograft of artificial skin and screening of anti-aging or safe chemical agents.
authors.affiliation: Laboratory of Physiological Chemistry, Faculty of Pharmaceutical Sciences at Kagawa, Tokushima Bunri University, 1314-1 Shido, Sanuki, Kagawa, 769-2193, Japan.
authors.forename: Mizuna
authors.initials: M
authors.lastname: Kamada
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Kumazaki
authors.forename: Taira
authors.initials: T
authors.lastname: Matsuo
authors.forename: Youji
authors.initials: Y
authors.lastname: Mitsui
authors.forename: Tomoko
authors.initials: T
authors.lastname: Takahashi
mesh_terms: D059786:Abnormal Karyotype; D000818:Animals; D002460:Cell Line; D048430:Cell Shape; D002470:Cell Survival; D002471:Cell Transformation, Neoplastic; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism / Q000502:physiology* / Q000637:transplantation; D005839:Gentamicins / Q000494:pharmacology; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D019098:Telomerase / Q000096:biosynthesis* / Q000235:genetics; D059505:Telomere Homeostasis; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
chemical_list: C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D005839:Gentamicins; D011994:Recombinant Proteins; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C010680:antibiotic G 418; C509186:TERT protein, human; D019098:Telomerase