Release of plasmid DNA from intravascular stents coated with ultrathin multilayered polyelectrolyte films

Abstract

Materials that permit control over the release of DNA from the surfaces of topologically complex implantable devices, such as intravascular stents, could contribute to the development of new approaches to the localized delivery of DNA. We report the fabrication of ultrathin, multilayered polyelectrolyte films that permit both the immobilization and controlled release of plasmid DNA from the surfaces of stainless steel intravascular stents. Our approach makes use of an aqueous-based, layer-by-layer method for the assembly of nanostructured thin films consisting of alternating layers of plasmid DNA and a hydrolytically degradable polyamine. Characterization of coated stents using scanning electron microscopy (SEM) demonstrated that stents were coated uniformly with an ultrathin film ca. 120 nm thick that adhered conformally to the surfaces of stent struts. These ultrathin films did not crack, peel, or delaminate substantially from the surface after exposure to a range of mechanical challenges representative of those encountered during stent deployment (e.g., balloon expansion). Stents coated with eight bilayers of degradable polyamine and a plasmid encoding enhanced green fluorescent protein (EGFP) sustained the release of DNA into solution for up to four days when incubated in phosphate buffered saline at 37 degrees C, and coated stents were capable of mediating the expression of EGFP in a mammalian cell line without the aid of additional transfection agents. The approach reported here could, with further development, contribute to the development of localized gene-based approaches to the treatment of cardiovascular diseases or related conditions.

Figure 1

Scanning electron microscopy images of stainless steel intravascular stents coated with multilayered films fabricated from eight bilayers of polymer 1 and a plasmid DNA construct (pEGFP-N1) encoding enhanced green fluorescent protein. Stents were precoated with a thin multilayered SPS/LPEI film (ca. 20 nm thick) prior to the deposition of the DNA-containing films (see text). Images correspond to different magnifications and perspectives of a coated stent imaged as-coated on a balloon assembly (A-C) and after balloon expansion (D-F).

Figure 2

A-D) Scanning electron microscopy images of intravascular stents coated with multilayered films fabricated from eight bilayers of polymer 1 and a plasmid DNA. Stents were coated while mounted on a balloon assembly and then passed through a silicone septum and arterial inducer prior to imaging (see text). Images correspond to different magnifications and perspectives. The image shown in D shows a magnified view of the delaminated portion of the film shown in C.

Figure 3

Plot of solution absorbance at 260 nm v. time for two expanded stents coated with either eight (■) or sixteen (●) bilayers of a polymer 1/DNA film incubated in phosphate buffered saline at 37 °C. Error bars are shown but are smaller than the symbols used to represent absorbance values.

Figure 4

Scanning electron microscopy images of intravascular stents coated with eight bilayers of a multilayered polymer 1/DNA film and incubated in PBS buffer at 37 °C for 1.5 hours prior to imaging (see text).

Figure 5

A) Fluorescence microscopy image (20X) showing expression of EGFP in COS-7 cells transfected with DNA released from a multilayered polymer 1/DNA film incubated in PBS at 37 °C. Transfection was conducted by combining released DNA with a commercially available cationic lipid (see text). B) Series of adjacent low magnification (4X) fluorescence microscopy images showing expression of EGFP in a confluent population of COS-7 cells 48 hours after the introduction of a stent coated with eight bilayers of polymer 1 and DNA (total area shown is approximately 1.8 mm²; a portion of the expanded stent is shown for comparison). This experiment was conducted without the use of additional transfection agents (see text). Images C and D show representative higher magnification phase contrast and fluorescence microscopy images (10X) of cells shown in B) used to quantify levels of transfection.