title: In vitro gene expression by cationized derivatives of an artificial protein with repeated RGD sequences, Pronectin.
abstract: The objective of this study is to investigate the efficiency of a non-viral gene carrier with RGD sequences, Pronectin F(+) for gene transfection. The Pronectin F(+) was cationized by introducing ethylenediamine (Ed), spermidine (Sd), and spermine (Sm) to the hydroxyl groups while the corresponding gelatin derivative was prepared similarly because gelatin also has one RGD sequence per molecule. The zeta potential and molecular size of Pronectin F(+) and gelatin derivatives were examined before and after polyion complexation with a plasmid DNA of luciferase. When complexed with the plasmid DNA at the Pronectin F(+)/plasmid DNA mixing ratio of 50, the complex exhibited a zeta potential of about 10 mV, which is similar to that of the gelatin derivative-plasmid DNA complex. Irrespective of the type of Pronectin F(+) and gelatin derivatives, their complexation enabled the apparent molecular size of plasmid DNA to reduce to about 200 nm, the size decreasing with the increased derivative/plasmid DNA weight mixing ratio. The rat gastric mucosal (RGM)-1 cells treated with both complexes exhibited significantly stronger luciferase activities than free plasmid DNA although the enhanced extent was significant for the Sm derivative compared with the corresponding Ed and Sd derivatives. Cell attachment was enhanced by the Pronectin F(+) derivative to a significant high extent compared with the gelatin derivative. The amount of plasmid DNA internalized into the cells was enhanced by the complexation with every Pronectin F(+) derivative compared with the gelatin derivative. For both of Pronectin F(+) and gelatin carriers, the buffering capacity of Sm derivatives was higher than that of Ed and Sd derivatives and comparable to that of polyethyleneimine. It is likely that the high efficiency of gene transfection for the Sm derivative is due to the superior buffering effect. We conclude that the Sm derivative of Pronectin F(+) is promising as a non-viral vector of gene transfection.
authors.affiliation: Department of Biomaterials, Field of Tissue Engineering, Institute for Frontier Medical Sciences, Kyoto University, 53 Kawara-cho Shogoin, Sakyo-ku, Kyoto 606-8507, Japan.
authors.forename: Hossein
authors.initials: H
authors.lastname: Hosseinkhani
authors.forename: Yasuhiko
authors.initials: Y
authors.lastname: Tabata
mesh_terms: D000818:Animals; D002412:Cations; D002478:Cells, Cultured; D001517:Coleoptera; D005353:Fibronectins / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D009842:Oligopeptides / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D010957:Plasmids; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D020449:Repetitive Sequences, Amino Acid; D014162:Transfection
chemical_list: D002412:Cations; D005353:Fibronectins; D009842:Oligopeptides; D011994:Recombinant Proteins; C070367:ProNectin F; C047981:arginyl-glycyl-aspartic acid