ArticlePharmaceutical research2002
Intracellular processing of poly(ethylene imine)/ribozyme complexes can be observed in living cells by using confocal laser scanning microscopy and inhibitor experiments.
Article in Pharmaceutical research, 2002. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 30 papers.
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30 citing papers in PubMed, 149 citations in OpenAlex.
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- BMP-2 gene modified canine bMSCs promote ectopic bone formation mediated by a nonviral PEI derivative.Annals of biomedical engineering · 2011Article
- Phospholipid-polyethylenimine conjugate-based micelle-like nanoparticles for siRNA delivery.Drug delivery and translational research · 2011Article
- Understanding the protonation behavior of linear polyethylenimine in solutions through Monte Carlo simulations.Biomacromolecules · 2010Article
- Missing pieces in understanding the intracellular trafficking of polycation/DNA complexes.Journal of controlled release : official journal of the Controlled Release Society · 2009Article
- Molecular dynamics simulations of DNA-polycation complex formation.Biophysical journal · 2009Article
- Transient transfection methods for clinical adeno-associated viral vector production.Human gene therapy · 2009Review
- Facile synthesis of multivalent folate-block copolymer conjugates via aqueous RAFT polymerization: targeted delivery of siRNA and subsequent gene suppression.Biomacromolecules · 2009Article
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Authors and funding
5 authors at 2 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
purposeCritical steps in the subcellular processing of poly(ethylene imine)/nucleic acid complexes, especially endosomal/lysosomal escape, were visualized by using living cell confocal laser scanning microscopy (CSLM) to obtain an insight into their mechanism.
methodsLiving cell confocal microscopy was used to examine the intracellular fate of poly(ethylene imine)/ribozyme and poly(L-lysine)/ribozyme complexes over time, in the presence of and without bafilomycin Al, a selective inhibitor of endosomal/lysosomal acidification. The compartment of complex accumulation was identified by confocal microscopy with a fluorescent acidotropic dye. To confirm microscopic data, luciferase reporter gene expression was determined under similar experimental conditions.
resultsPoly(ethylene imine)/ribozyme complexes accumulate in acidic vesicles, most probably lysosomes. Release of complexes occurs in a sudden event, very likely due to bursting of these organelles. After release, poly(ethylene imine) and ribozyme spread throughout the cell, during which slight differences in distribution between cytosol and nucleus are visible. No lysosomal escape was observed with poly(L-lysine)/ribozyme complexes or when poly(ethylene imine)/ ribozyme complexes were applied together with bafilomycin A1. Poly(ethylene imine)/plasmid complexes exhibited a high luciferase expression, which was reduced approximately 200-fold when lysosomal acidification was suppressed with bafilomycin A1.
conclusionsOur data provide, for the first time, direct experimental evidence for the escape of poly(ethylene imine)/nucleic acid complexes from the endosomal/lysosomal compartment. CLSM, in conjunction with living cell microscopy, is a promising tool for studying the subcellular fate of polyplexes in nucleic acid/gene delivery.
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