Slinkin M A, Shevelev A Ia
Russian Cardiology Research Center, Third Cherepkovskaya ul. 15a, Moscow, 121552 Russia.
Bioorg Khim. 2001 Jul-Aug;27(4):291-9. doi: 10.1023/a:1011308604647.
A new approach to optimization of mixtures for the condensation and introduction of plasmid DNA into eukaryotic cells is proposed, which is based on the formation of ternary interpolyelectrolyte complexes (IPEC) DNA/polycation/polyanion. Polyethyleneimine (PEI) with M 30-40 kDa as polycation and polyacrylic acid (PA) with M 20 kDa or its grafted copolymer with polyethyleneglycol (PEG) as polyanion were used, and ternary complexes with various ratios of the components were prepared. The PA-PEG incorporation into a ternary complex (by itself or as a 1:1 mixture with PA) was shown to confer the solubility onto complexes in a wide range of DNA/PEI ratios. Incorporation of even minute amounts of PA-PEG (as a 1:9 mixture with PA), while not completely preventing the aggregation of ternary IPEC, drastically changed their sorption characteristics. Using a beta-galactosidase-encoding plasmid, efficiencies of transfection of the CHO-AA8 and 293 cells for different IPEC and DNA/lipofectin complex were compared. The maximum efficiency was exhibited by ternary complex DNA/PEI/polyanion where a 1:1 mixture of PA and PA-PEG was used as polyanion. Possible reasons for this effect and further ways of optimization of mixtures for expression of plasmid DNA in the context of the new approach are discussed.
提出了一种优化混合物的新方法，用于将质粒DNA浓缩并导入真核细胞，该方法基于三元聚电解质复合物（IPEC）DNA/聚阳离子/聚阴离子的形成。使用分子量为30 - 40 kDa的聚乙烯亚胺（PEI）作为聚阳离子，分子量为20 kDa的聚丙烯酸（PA）或其与聚乙二醇（PEG）的接枝共聚物作为聚阴离子，并制备了具有不同组分比例的三元复合物。结果表明，将PA - PEG掺入三元复合物中（单独掺入或与PA以1:1的比例混合）可使复合物在广泛的DNA/PEI比例范围内具有溶解性。即使掺入微量的PA - PEG（与PA以1:9的比例混合），虽然不能完全阻止三元IPEC的聚集，但会显著改变其吸附特性。使用编码β - 半乳糖苷酶的质粒，比较了不同IPEC和DNA/脂质体复合物对CHO - AA8和293细胞的转染效率。当使用PA和PA - PEG的1:1混合物作为聚阴离子时，三元复合物DNA/PEI/聚阴离子表现出最高的转染效率。讨论了这种效应的可能原因以及在新方法背景下进一步优化用于质粒DNA表达的混合物的方法。
