title: Potential of D-Octaarginine-Linked Polymers as an in Vitro Transfection Tool for Biomolecules.
abstract: We have been investigating the potential use of cell-penetrating peptide-linked polymers as a novel penetration enhancer. Since previous in vivo studies demonstrated that poly(N-vinylacetamide-co-acrylic acid) bearing D-octaarginine, a typical cell-penetrating peptide, enhanced membrane permeation of biomolecules, its potential as an in vitro transfection tool was evaluated in this study. A plasmid DNA encoding green fluorescent protein (pGFP-C1), β-galactosidase, and bovine serum albumin (BSA) were used as model biomolecules. Anionic pGFP-C1 interacted electrostatically with cationic d-octaarginine-linked polymers. When the ratio of mass concentration of polymers to that of pGFP-C1 reached 2.5, complexes whose size and zeta potential were approximately 200 nm and 15 mV, respectively, were obtained. GFP expression was observed in cells incubated with complexes prepared under conditions in which the polymer/pDNA concentration ratio exceeded 2.5. The expression level elevated with an increase in the concentration ratio, but physicochemical properties of the complexes remained unchanged. Results suggested that free polymers contributed to pGFP-C1 internalization. Another cell study demonstrated that β-galactosidase premixed with polymers was taken up into cells in its active tetrameric form. Similar electrostatic interaction-driven complex formation was observed for BSA charged negatively in neutral solution. However, it appeared that the internalization processes of BSA differed from those of pGFP-C1. A mass concentration-dependent increase in internalized BSA was observed, irrespective of the polymer/protein concentration ratio. Due to frail interactions, polymers that were released from the complexes and subsequently immobilized on cell membranes might also contribute to membrane permeation of BSA.
authors.forename: Kohta
authors.initials: K
authors.lastname: Mohri
authors.affiliation: ‡Life Science Materials Laboratory, ADEKA Co., 7-2-34 Higashiogu, Arakawa-ku, Tokyo 116-8553, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Morimoto
authors.forename: Megumi
authors.initials: M
authors.lastname: Maruyama
authors.forename: Norimasa
authors.initials: N
authors.lastname: Nakamoto
authors.forename: Emi
authors.initials: E
authors.lastname: Hayashi
authors.forename: Kengo
authors.initials: K
authors.lastname: Nagata
authors.affiliation: ‡Life Science Materials Laboratory, ADEKA Co., 7-2-34 Higashiogu, Arakawa-ku, Tokyo 116-8553, Japan.
authors.forename: Kohei
authors.initials: K
authors.lastname: Miyata
authors.affiliation: ‡Life Science Materials Laboratory, ADEKA Co., 7-2-34 Higashiogu, Arakawa-ku, Tokyo 116-8553, Japan.
authors.forename: Kyohei
authors.initials: K
authors.lastname: Ochiai
authors.affiliation: ‡Life Science Materials Laboratory, ADEKA Co., 7-2-34 Higashiogu, Arakawa-ku, Tokyo 116-8553, Japan.
authors.forename: Ken-ichiro
authors.initials: K
authors.lastname: Hiwatari
authors.affiliation: ‡Life Science Materials Laboratory, ADEKA Co., 7-2-34 Higashiogu, Arakawa-ku, Tokyo 116-8553, Japan.
authors.forename: Kazufumi
authors.initials: K
authors.lastname: Tsubaki
authors.affiliation: ‡Life Science Materials Laboratory, ADEKA Co., 7-2-34 Higashiogu, Arakawa-ku, Tokyo 116-8553, Japan.
authors.forename: Etsuo
authors.initials: E
authors.lastname: Tobita
authors.forename: Yuki
authors.initials: Y
authors.lastname: Ishimaru
authors.forename: Sadaaki
authors.initials: S
authors.lastname: Maeda
authors.forename: Shinji
authors.initials: S
authors.lastname: Sakuma
mesh_terms: D000818:Animals; D002417:Cattle; D002463:Cell Membrane Permeability; D004337:Drug Carriers / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D009842:Oligopeptides / Q000737:chemistry*; D010957:Plasmids / Q000008:administration & dosage*; D011108:Polymers / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism*
chemical_list: D004337:Drug Carriers; D009842:Oligopeptides; D011108:Polymers; C448619:octaarginine; D049452:Green Fluorescent Proteins; D012710:Serum Albumin, Bovine; D001616:beta-Galactosidase