Cationic polymer-mediated genetic transduction into cultured human chondrosarcoma-derived HCS-2/8 cells

Suzuyo Ohashi, Toshikazu Kubo, Takumi Ikeda, Yuji Arai, Kenji Takahashi, Yasusuke Hirasawa, Masaharu Takigawa, Etsuko Satoh, Jiro Imanishi, Osam Mazda

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27 Citations (Scopus)

Abstract

The usefulness of three types of cationic polymer, i.e., degraded polyamidoamine (PAMAM) dendrimer (SuperFect Transfection Reagent; Qiagen), linear polyethylenimine (PEI: ExGen 500: Euromedex), and branched PEI in gene delivery into chondrocytes was examined comparatively. A plasmid vector containing the Escherichia coli LacZ (pSES.β) was combined with one of the three cationic polymers at various molar ratios and the resultant complex (polyplex) was used to transduce a human chondrocyte-like cell line, HCS-2/8. Gene expression was evaluated by an O-nitrophenyl β-D-galactopyranoside (ONPG) assay and by staining with 0.05% 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside (X-gal: Nacalai Tesque). The ONPG assay showed that the highest delivery rate was achieved when 2μg of pSES.β was combined with either 21 μg of dendrimer, 1.7 μg of linear PEI, or 2.0 μg of branched PEI. At the same DNA/polymer ratios, the proportions of X-galstained cells were also the highest (31.3 ± 7.5%, 30.3 ± 9.0%, and 8.3 ± 3.1%, respectively). LacZ expression reached the highest level 3 days after the dendrimer-mediated transduction, and gradually declined, returning to the background level on day 14. Possible cytotoxicity was examined by trypan blue staining and phase contrast microscopic observations. Neither cytotoxicity nor morphological change was induced at the optimal dose of each polymer. The cationic polymers, particularly the degraded dendrimer and linear PEI, would be a useful nonviral vector for gene delivery to cells of chondrocytes.

Original languageEnglish
Pages (from-to)75-81
Number of pages7
JournalJournal of Orthopaedic Science
Volume6
Issue number1
DOIs
Publication statusPublished - 2001

Keywords

  • Branched polyethylenimine
  • Chondrocytes
  • Degraded polyamidoamine dendrimer
  • Gene delivery
  • Linear polyethylenimine

ASJC Scopus subject areas

  • Surgery
  • Orthopedics and Sports Medicine

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    Ohashi, S., Kubo, T., Ikeda, T., Arai, Y., Takahashi, K., Hirasawa, Y., Takigawa, M., Satoh, E., Imanishi, J., & Mazda, O. (2001). Cationic polymer-mediated genetic transduction into cultured human chondrosarcoma-derived HCS-2/8 cells. Journal of Orthopaedic Science, 6(1), 75-81. https://doi.org/10.1007/s007760170028