Rapid release of plasmid DNA from surfaces coated with polyelectrolyte multilayers promoted by the application of electrochemical potentials

Abstract

We report an approach to the rapid release of DNA based on the application of electrochemical potentials to surfaces coated with polyelectrolyte-based thin films. We fabricated multilayered polyelectrolyte films (or "polyelectrolyte multilayers", PEMs) using plasmid DNA and a model hydrolytically degradable cationic poly(β-amino ester) (polymer 1) on stainless steel substrates using a layer-by-layer approach. The application of continuous reduction potentials in the range of -1.1 to -0.7 V (vs a Ag/AgCl electrode) to film-coated electrodes in PBS at 37 °C resulted in the complete release of DNA over a period of 1-2 min. Film-coated electrodes incubated under identical conditions in the absence of applied potentials required 1-2 days for complete release. Control over the magnitude of the applied potential provided control over the rate at which DNA was released. The results of these and additional physical characterization experiments are consistent with a mechanism of film disruption that is promoted by local increases in pH at the film/electrode interface (resulting from electrochemical reduction of water or dissolved oxygen) that disrupt ionic interactions in these materials. The results of cell-based experiments demonstrated that DNA was released in a form that remains intact and able to promote transgene expression in mammalian cells. Finally, we demonstrate that short-term (i.e., non-continuous) electrochemical treatments can also be used to promote faster film erosion (e.g., over 1-2 h) once the potential is removed. Past studies demonstrate that PEMs fabricated using polymer 1 can promote surface-mediated transfection of cells and tissues in vitro and in vivo. With further development, the electrochemical approaches reported here could thus provide new methods for the rapid, triggered, or spatially patterned transfer of DNA (or other agents) from surfaces of interest in a variety of fundamental and applied contexts.

Figure 1

Plots showing the release of DNA from polymer 1/DNA films (A) in the absence of an applied electrochemical potential, and (B) in the presence of applied electrochemical potentials of -1.1 V (●), -0.9 V (○), -0.7 V (▲), and -0.2 V (■). Error bars represent the standard deviation of the calculated percentages of DNA released obtained from multiple samples collected from experiments using two different film-coated electrodes for each condition. In some cases, error bars are smaller than the symbols used to represent the data.

Figure 2

Fluorescence microscopy images (40× magnification; 730 μm × 510 μm) of the edge of a stainless steel substrate coated with a polymer 1/DNA film 8-bilayers thick, incubated in an aqueous ethidium bromide solution that intercalates DNA and labels it with a red color (A) before electrochemically-induced erosion, and (B) after applying a potential of -1.1 V for 1 min in PBS.

Figure 3

AFM images of polymer 1/DNA films (5 μm × 5 μm) (A) before erosion, (B) after 5 sec of erosion at -1.1 V in PBS, and (C) after 30 sec of erosion at -0.7 V in PBS. The scale in the z-direction, shown at the far right, is 250 nm.

Figure 4

(A) Agarose gel electrophoresis characterization of samples of DNA released from stainless steel substrates coated with polymer 1/DNA films and incubated with an applied potential of -1.1 V. Lane labels indicate the time at which samples were collected for analysis. The lane labeled ‘C’ corresponds to a pEGFP control. (B) Representative fluorescence micrographs (100× magnification; 1194 μm × 895 μm) of confluent monolayers of COS-7 cells showing levels of EGFP expression mediated by samples of released DNA collected after the electrochemically promoted erosion of polymer 1/DNA films (at an applied potential of -1.1 V) for 30 sec, 1 min, 2 min, and 3 min.

Figure 5

Plot showing the release of DNA from polymer 1/DNA films incubated at an applied potential of -1.1 V for 5 sec and then subsequently incubated in PBS in the absence of an applied potential (see text). The arrow indicates the point at which the potential was removed. Error bars show the standard deviation of the percentage of DNA released using multiple measurements of samples during experiments using two different film-coated electrodes.

Polymer 1