Biophysical and Structural Characterization of Polyethylenimine-Mediated siRNA Delivery in Vitro
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The goals of this study were as follows: 1) to evaluate the efficacy of different polyethylenimine (PEI) structures for siRNA delivery in a model system, and 2) to determine the biophysical and structural characteristics of PEI that relate to siRNA delivery.
Materials and Methods
Biophysical characterization (effective diameter and zeta potential), cytotoxicities, relative binding affinities and in vitro transfection efficiencies were determined using nanocomplexes formed from PEI's of 800, 25,000, (both branched) and 22,000 (linear) molecular weights at varying N:P ratios and siRNA concentrations. The HR5-CL11 cell line stably expressing luciferase was used as a model system in vitro.
Successful siRNA delivery was observed within a very narrow window of conditions, and only with the 25,000 branched PEI at an N:P ratio of 6:1 and 8:1 and with 200 nM siRNA. While the zeta potential and size of PEI:siRNA complexes correlated to transfection efficacy in some cases, complex stability may also affect transfection efficacy.
The ability of PEI to transfer functionally active siRNA to cells in culture is surprisingly dependent on its biophysical and structural characteristics when compared to its relative success and ease of use for DNA delivery.
Key Wordsluciferases polyethylenimine RNA interference small interfering RNA transfection
4-(2-hydroxyethyl) piperazine-1-ethanesulfonic acid
small interfering RNA
- 22 K L-PEI
22,000 molecular weight linear PEI
- 25 K B-PEI
25,000 molecular weight branched PEI
- 800 B-PEI
800 molecular weight branched PEI
Research described in this article was supported in part by Philip Morris USA Inc. and Philip Morris Internationaland in part by the Ladies Auxiliary to the Veterans of Foreign Wars Cancer Research Fellowship. The expert help and assistance of Mr. Peter Zawaneh is greatly appreciated.
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