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Evidence for plasmid DNA exchange after polyplex mixing
L Pigeon1, C Gonçalves1, C Pichon1
1Centre de Biophysique Moléculaire, CNRS UPR4301, Inserm and University of Orléans, 45071 Orléans cedex 02, France. patrick.midoux@cnrs-orleans.fr.
Soft Matter
|July 28, 2016
Summary
When two different fluorescently labeled plasmid DNA (pDNA) polyplexes are mixed, pDNA exchange occurs, indicating polyplex unpackaging and reassembly. This exchange is dependent on the polymer type and pDNA interaction strength.
Area of Science:
- Biochemistry
- Materials Science
- Nanotechnology
Background:
- Cationic polymers and liposomes self-assemble with plasmid DNA (pDNA) to form nanostructures like polyplexes and lipoplexes.
- These nanostructures are crucial for gene delivery applications.
- Understanding their stability and dynamics is key to optimizing delivery efficiency.
Purpose of the Study:
- To investigate the exchange of plasmid DNA (pDNA) between pre-formed polyplexes.
- To determine the factors influencing pDNA exchange dynamics.
- To elucidate the mechanism of pDNA exchange in polyplexes.
Main Methods:
- Preparation of two distinct fluorescently labeled pDNA/histidinylated lPEI polyplexes (Flu-pDNA/His-lPEI and Cy5-pDNA/His-lPEI).
- Mixing of the two polyplex preparations.
- Analysis of dual fluorescence and Förster Resonance Energy Transfer (FRET) to detect pDNA exchange.
- Comparison with other polyplexes (polylysine, caged His-lPEI) and other nanostructures (lipoplexes, lipopolyplexes).
Main Results:
- Mixing Flu-pDNA/His-lPEI and Cy5-pDNA/His-lPEI polyplexes resulted in a high proportion of dual-fluorescent polyplexes.
- FRET analysis confirmed the presence of both types of fluorescent pDNA within single polyplexes, indicating successful pDNA exchange.
- pDNA exchange was observed to be dependent on the polymer type and the strength of pDNA/polymer interactions.
- No pDNA exchange was detected with polylysine polyplexes, caged His-lPEI polyplexes, lipoplexes, lipopolyplexes, or when His-lPEI polyplexes were mixed with lipoplexes.
Conclusions:
- Polyplexes formed with histidinylated lPEI facilitate pDNA exchange between distinct polyplexes.
- The observed pDNA exchange suggests a mechanism involving polyplex aggregation, unpackaging, and subsequent reassembly.
- The findings highlight the dynamic nature of certain polyplex structures and are critical for understanding gene delivery vector stability.
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