| Budget Amount *help |
¥14,700,000 (Direct Cost: ¥14,700,000)
Fiscal Year 2000: ¥6,700,000 (Direct Cost: ¥6,700,000)
Fiscal Year 1999: ¥8,000,000 (Direct Cost: ¥8,000,000)
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| Research Abstract |
Usefulness of three types of cationic polymer, i.e., degraded polyamidoamine (PAMAM) dendrimer (SuperFect Transfection Reagent), linear polyethylenimine (PEI) (ExGen 500) and branched PEI in gene delivery into chondrocytes was comparatively examined. A plasmid vector containing the E.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 the O-Nitrophenyl β-D-Galactopyranoside (ONPG) assay and X-gal staining. The ONPG assay showed that the highest delivery rate was achieved when 2 μg 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-gal-staining cells were also the highest (31, 30, and 8%, respectively). LacZ expression reached the highest level 3days after the dendrimer-mediated transduction, gradually declined and returned 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 non-viral vector for gene delivery to the cells in chondrocyte lineage.
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