Co-transfection of star-shaped PDMAEMAs enhance transfection efficiency of protamine/pDNA complexes in the presence of serum. (June 2018) - Chen, Baizhu
Graphical abstract: Highlights: Free star-shaped PDMAEMAs enhance the transfection efficiency of protamine/pDNA complexes and the result is even better than bPEI in complete medium. Free stars lead to a faster cellular uptake of protamine/pDNA complexes. Free stars mainly facilitate cellular internalization stage during transfection. Abstract: In previous studies, we have found that un-complexed free cationic polymers greatly promote gene transfection efficiency of pDNA complexes. Dividing the gene transfection system into two parts, bound chains and free chains, we exploited mixing different biomaterials as the second component of our designed system to improve gene transfection outcome. In this study, we investigated the effectiveness of star-shaped poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) with different arm numbers as free chains (i.e., co-transfection agent) while using natural salmon protamine as bound chains. We explored the gene transfer ability of the system in both serum free and complete cell culture medium. Our results reveal that the star-shaped PDMAEMAs enhance gene transfection efficiency of protamine/pDNA complexes, in the same range and even better than the gold standard polyethyleneimine (PEI, branched type with 25 kg/mol molar mass) in serum-free and serum containing medium conditions, respectively. From 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and Lactate Dehydrogenase (LDH) release data, it was found that the PDMAEMA stars have tolerable cytotoxicity but with significant membrane disruption level, especially when increasing the arm number. Flow cytometry experiments showed that PDMAEMA stars clearly improve the uptake rate as well as the number of protamine/pDNA complexes taken up by the target cells. Using Taqman Real-time Polymerase Chain Reaction (PCR), further exploration of the effect of PDMAEMAs during different gene transfection stages reflects that these un-bound star-shaped polymers mainly act upon the cellular internalization step. Our results indicate that the star-shaped PDMAEMAs are promising vectors for gene delivery.