Delivery of pDNA Polyplexes to Bronchial and Alveolar Epithelial Cells Using a Mesh Nebulizer
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In this study, a cell penetrating peptide was used as an uptake enhancer for pDNA delivery to the lungs.
Polyplexes were prepared between pDNA and CPP. Intracellular delivery of pDNA was assessed in both alveolar (A549) and bronchial (Calu-3) epithelial cells. Aerosol delivery was investigated using a mesh nebulizer.
Efficient intracellular delivery of pDNA occurs in both A549 and Calu-3 cells when delivered as polyplexes. Protection against nucleases and endosomal escape mechanism occurs when pDNA is formulated within the polyplexes. For aerosol delivery, 1% (w/v) mannitol was able to protect naked DNA structure during nebulization with a significant increase in fine particle fraction (particles <5 μm). The structure of polyplexes when delivered via a mesh nebulizer using 1% (w/v) mannitol could partially withstand the shear forces involved in aerosolization. Although some loss in functionality occurred after nebulization, membrane-associated fluorescence was observed in A549 cells. In Calu-3 cells mucus entrapment was a limiting factor for polyplex delivery.
The presence of CPP is essential for efficient intracellular delivery of pDNA. The polyplexes can be delivered to lung epithelial cells using mesh nebulizer. The use of different excipients is essential for further optimization of these delivery systems.
Key Wordsaerosol cell-penetrating peptide gene delivery lung nebulization
Dynamic light scattering
Double stranded DNA
Enhanced green fluorescent protein
Electrophoretic Mobility Shift Assay
Fine particle fraction
Next Generation Impactor
Phosphate buffered saline
Regions of Interest
Scanning electron microscopy
Transepithelial Electrical Resistance
Twin Stage Impinger
Acknowledgments and Disclosures
This work was supported by an Australian Research Council Linkage Grant LP140100181. Larissa Gomes dos Reis is financially supported by the Brazilian Research Funding CAPES.
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