Multilayered polyelectrolyte films promote the direct and localized delivery of DNA to cells

ArticleinJournal of Controlled Release 106(1-2):214-23 · September 2005with6 Reads
DOI: 10.1016/j.jconrel.2005.04.014 · Source: PubMed
Abstract
Multilayered polyelectrolyte films fabricated from plasmid DNA and a hydrolytically degradable synthetic polycation can be used to direct the localized transfection of cells without the aid of a secondary transfection agent. Multilayered assemblies 100 nm thick consisting of alternating layers of synthetic polymer and plasmid DNA encoding for enhanced green fluorescent protein (EGFP) were deposited on quartz substrates using a layer-by-layer fabrication procedure. The placement of film-coated slides in contact with COS-7 cells growing in serum-containing culture medium resulted in gene expression in cells localized under the film-coated portion of the slides. The average percentage of cells expressing EGFP relative to the total number of cells ranged from 4.6% to 37.9%, with an average of 18.6%+/-8.2%, as determined by fluorescence microscopy. In addition to providing a mechanism for the immobilization of DNA at the cell/surface interface, a preliminary analysis of film topography by atomic force microscopy (AFM) demonstrated that polymer /DNA films undergo significant structural rearrangements upon incubation to present surface bound condensed plasmid DNA nanoparticles. These data suggest that the presence of the cationic polymer in these materials may also contribute to the internalization and expression of plasmid. The materials and design principles reported here present an attractive framework for the local or non-invasive delivery of DNA from the surfaces of implantable materials or biomedical devices.
    • "Poly1 was synthesized via a Michael-type addition reaction as described previously [21,22,24]. Briefly, 9 mmol of 4,4 0 -trimethyle nedipiperidine was dissolved in anhydrous THF to form a 500 mg mL À1 solution. "
    [Show abstract] [Hide abstract] ABSTRACT: Statement of significance: Degradable polymers are increasingly important in vaccination, but how the inherent immunogenicity of polymers changes during degradation is poorly understood. Using common rapidly-degradable vaccine carriers, we show that the activation of immune cells - even in the absence of other adjuvants - depends on polymer form (e.g., free, particulate) and the extent of degradation. These changing characteristics alter the physicochemical properties (e.g., charge, size, molecular weight) of polymer particles, driving changes in immunogenicity. Our results are important as many common biomaterials (e.g., PLGA) are now known to exhibit immune activity that alters how vaccines are processed. Thus, the results of this study could contribute to more rational design of biomaterial carriers that also actively direct the properties of responses generated by vaccines.
    Full-text · Article · Dec 2015
    • "Preloading was achieved through electrostatic adsorption of pDNA onto bPEI25 coated PS particles prior to SF deposition such that nucleic acid/bPEI25 complexes were entrapped inside of the microcapsules, where the SF multilayer shell constituted both a protective as well as a diffusion barrier ( [47]. Both strategies have been successfully applied in several studies [31,4849505152 and used to tailor drug release characteristics via the loading process and shell thickness [53]. Plasmid DNA encapsulation efficiency was found to be significantly higher for 4 µm microcapsules after postloading while the loading technique had no significant influence on encapsulation efficiency for 1 µm microparticles (Fig. S3B). "
    [Show abstract] [Hide abstract] ABSTRACT: Herein, we describe the delivery of plasmid DNA (pDNA) using silk fibroin (SF) layer-by-layer assembled microcapsules. Deposition of fluorescently labeled SF onto polystyrene (PS) template particles resulted in increasing fluorescence intensity and decreasing surface charge in correlation to SF layer number. After removal of the PS core, hollow, monodisperse, and structurally stable SF microcapsules of variable size and shell thickness were obtained. Plasmid DNA encoding for enhanced green fluorescent protein (eGFP) was loaded onto 1 or 4 μm capsules, either by incorporation of pDNA within the innermost layer of the shell or by adsorption to the microcapsules surface, and in vitro pDNA release, cytotoxicty and eGFP expression were studied. Sustained pDNA release over 3 days was observed using both loading techniques, being accelerated in the presence of protease. DNA loaded SF microcapsules resulted in efficient cell transfection along with low cytotoxicity after 3 days incubation compared to treatment with pDNA/branched polyethylenimine complexes. Among the tested conditions highest transfection efficiencies were achieved using 1 μm capsules where pDNA was adsorbed to the capsule surface. Our results suggest that SF microcapsules are suitable for the localized delivery of pDNA, combining low cytotoxicity and high transfection efficiency.
    Full-text · Article · Jun 2014
    • "Bioapplications of stimuli-responsive films and capsules have been extensively reviewed focusing on cellular response and film biofunctionalization1213141516171819. The LbL films play the role of reservoirs providing a living organism with the required bioactive molecules such as growth factors, drugs, DNA, peptides or other soluble and insoluble signaling factors [15,2021222324 The reservoir capacity may be extremely high due to the large number of free polymer groups and the mobility of polymer molecules in the film [15,25]. The LbL technique can be considered as one of the very promising approaches to engineer the artificial extracellular matrix (ECM) [26,27]. "
    [Show abstract] [Hide abstract] ABSTRACT: Single cell analysis (SCA) is nowadays recognized as one of the key tools for diagnostics and fundamental cell biology studies. The Layer-by-layer (LbL) polyelectrolyte assembly is a rather new but powerful technique to produce multilayers. It allows to model the extracellular matrix in terms of its chemical and physical properties. Utilization of the multilayers for SCA may open new avenues in SCA because of the triple role of the multilayer film: (i) high capacity for various biomolecules; (ii) natural mimics of signal molecule diffusion to a cell and (iii) cell patterning opportunities. Besides, light-triggered release from multilayer films offers a way to deliver biomolecules with high spatio-temporal resolution. Here we review recent works showing strong potential to use multilayers for SCA and address accordingly the following issues: biomolecule loading, cell patterning, and light-triggered release.
    Full-text · Article · May 2014
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