title: Telomerase-mediated life-span extension of human primary fibroblasts by human artificial chromosome (HAC) vector.
abstract: Telomerase-mediated life-span extension enables the expansion of normal cells without malignant transformation, and thus has been thought to be useful in cell therapies. Currently, integrating vectors including the retrovirus are used for human telomerase reverse transcriptase (hTERT)-mediated expansion of normal cells; however, the use of these vectors potentially causes unexpected insertional mutagenesis and/or activation of oncogenes. Here, we established normal human fibroblast (hPF) clones retaining non-integrating human artificial chromosome (HAC) vectors harboring the hTERT expression cassette. In hTERT-HAC/hPF clones, we observed the telomerase activity and the suppression of senescent-associated SA-beta-galactosidase activity. Furthermore, the hTERT-HAC/hPF clones continued growing beyond 120days after cloning, whereas the hPF clones retaining the silent hTERT-HAC senesced within 70days. Thus, hTERT-HAC-mediated episomal expression of hTERT allows the extension of the life-span of human primary cells, implying that gene delivery by non-integrating HAC vectors can be used to control cellular proliferative capacity of primary cultured cells.
authors.affiliation: Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Sciences, Tottori University, 86 Nishimachi, Yonago, Tottori 683-8503, Japan.
authors.forename: Shingo
authors.initials: S
authors.lastname: Shitara
authors.forename: Minoru
authors.initials: M
authors.lastname: Kakeda
authors.forename: Keiko
authors.initials: K
authors.lastname: Nagata
authors.forename: Masaharu
authors.initials: M
authors.lastname: Hiratsuka
authors.forename: Akiko
authors.initials: A
authors.lastname: Sano
authors.forename: Kanako
authors.initials: K
authors.lastname: Osawa
authors.forename: Akiyo
authors.initials: A
authors.lastname: Okazaki
authors.forename: Motonobu
authors.initials: M
authors.lastname: Katoh
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kazuki
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Oshimura
authors.forename: Kazuma
authors.initials: K
authors.lastname: Tomizuka
mesh_terms: D000818:Animals; D016466:CHO Cells; D002455:Cell Division; D049109:Cell Proliferation; D016922:Cellular Senescence; D022222:Chromosomes, Artificial, Human / Q000235:genetics*; D006224:Cricetinae; D003412:Cricetulus; D005347:Fibroblasts / Q000201:enzymology / Q000502:physiology; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D019098:Telomerase / Q000032:analysis / Q000235:genetics*
chemical_list: C509186:TERT protein, human; D019098:Telomerase