Temperature-responsive polymeric carriers incorporating hydrophobic monomers for effective transfection in small doses.
A series of thermoresponsive ternary random copolymers, poly[N-isopropylacrylamide (PIPAAm)-co-(dimethylamino)ethylmethacrylate (DMAEMA)-co-butylmethacrylate (BMA)], was synthesized and their in vitro gene transfection efficiency in cell culture was evaluated. A control copolymer containing 20 mol% DMAEMA units, IP-20D (mole ratio of IPAAm/DMAEMA/BMA=80/20/0 in feed, no BMA units) was inert in transfection. In contrast, copolymer IP-20D-10B (IPAAm/DMAEMA/BMA=70/20/10 in feed) effectively transfected plasmid DNA into COS-1 cell cultures even under small dosing conditions of 0.1 microg of plasmid DNA per well in a 96-well plate, suggesting that incorporation of the appropriate amount of hydrophobic unit is crucial to transfection efficiency. Gene expression was much more significant when transfected by the IP-20D-10B carrier in comparison with control homopolymer poly-DMAEMA, and almost equal to that of the highly competent lipid carrier, LipofectAMINE PLUS trade mark. Furthermore, the transfection efficiency of IP-20D-10B is altered in a thermally responsive manner. By temporarily lowering the cell culture incubation temperature to 20 degrees C in the posttransfection period, gene expression doubled over that for incubation temperature at 37 degrees C. The DNA EtBr intercalation assay suggested that DNA affinity for IP-20D-10B is decreased by lowering incubation temperature, implying that the thermally regulated gene expression could provide more efficient DNA release from the polymeric carrier.
Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku, Tokyo 162-8666, Japan.
Naoya
N
Takeda
Emiko
E
Nakamura
Masayuki
M
Yokoyama
Teruo
T
Okano
D000180:Acrylic Resins / Q000138:chemical synthesis; D000818:Animals; D019556:COS Cells; D002478:Cells, Cultured; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002522:Chlorocebus aethiops; D004247:DNA / Q000008:administration & dosage / Q000235:genetics; D004337:Drug Carriers; D004996:Ethidium; D005456:Fluorescent Dyes; D015870:Gene Expression; D007364:Intercalating Agents / Q000737:chemistry; D008027:Light; D008970:Molecular Weight; D010316:Particle Size; D010957:Plasmids; D011721:Pyrenes / Q000737:chemistry; D012542:Scattering, Radiation; D013696:Temperature; D014162:Transfection / Q000379:methods*
D000180:Acrylic Resins; D004337:Drug Carriers; D005456:Fluorescent Dyes; D007364:Intercalating Agents; D011721:Pyrenes; C492295:poly(N-isopropylacrylamide-co-(dimethylamino)ethylmethacrylate-co-butylmethacrylate); D004247:DNA; D004996:Ethidium