Polylysine and polyornithine gene transfer complexes: a study of complex stability and cellular uptake as a basis for their differential in-vitro transfection efficiency.
Gene transfer vectors formed between the cationic polyamino acid, poly-(L)-omithine (PLO) and plasmid DNA (pDNA) have demonstrated superior transfection efficiency (up to x 10-fold) compared to equivalent polylysine-based systems in-vitro. The mechanism(s) underlying this observation remains to be elucidated. We previously reported no significant difference in colloidal particle size or zeta potential of polycation/pDNA complexes formed with poly-(L)-lysine (PLL), poly-(D)-lysine (PDL) or PLO. Here we report spectrofluorometric analysis indicating that PLO condenses pDNA at lower charge (+/-) ratios than PLL or PDL (cf. 0.8:1, 1.2:1 and 1.5:1). Moreover, PLO/pDNA complexes proved more stable to disruption by the polyanions, poly-(L)-aspartic acid (PAA) and heparin. There were no qualitative differences in the ability of the polycations to protect complexed pDNA from enzymatic degradation both in the presence and in the absence of polyanions. The superior transfection efficiency of PLO/pDNA complexes did not appear to be mediated by an increased cellular delivery of pDNA. The data suggests a greater affinity of PLO for pDNA as an important parameter for the observed superior in-vitro transfection efficiency.
Ramsay
E
E
Welsh School of Pharmacy, Cardiff University, UK.
Gumbleton
M
M
D002460:Cell Line; D004247:DNA / Q000008:administration & dosage* / Q000737:chemistry; D003850:Deoxyribonuclease I / Q000737:chemistry; D004337:Drug Carriers; D004587:Electrophoresis, Agar Gel; D004996:Ethidium; D005434:Flow Cytometry; D019793:Fluorescein; D005456:Fluorescent Dyes; D006801:Humans; D010455:Peptides; D011107:Polylysine; D013050:Spectrometry, Fluorescence; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured
D004337:Drug Carriers; D005456:Fluorescent Dyes; D010455:Peptides; C008973:polyornithine; D011107:Polylysine; D004247:DNA; D003850:Deoxyribonuclease I; D004996:Ethidium; D019793:Fluorescein