Intracellular Trafficking and Unpacking of siRNA/Quantum Dot-PEI Complexes Modified with and without Cell Penetrating Peptide: Confocal and Flow Cytometric FRET Analysis
| Autor: | Tg Park Tae Gwan Park, Ik Kim In-Kyoung Kim, Hyukjin Lee |
|---|---|
| Rok vydání: | 2010 |
| Předmět: |
Vascular Endothelial Growth Factor A
Small interfering RNA Time Factors Molecular Sequence Data Intracellular Space Biomedical Engineering Pharmaceutical Science Bioengineering Gene delivery chemistry.chemical_compound Transduction (genetics) Cell Line Tumor Quantum Dots Fluorescence Resonance Energy Transfer Animals Humans Polyethyleneimine Gene silencing Amino Acid Sequence RNA Small Interfering Pharmacology Drug Carriers Polyethylenimine Microscopy Confocal Base Sequence Organic Chemistry technology industry and agriculture Biological Transport Flow Cytometry Förster resonance energy transfer chemistry Biochemistry Biophysics Cell-penetrating peptide Peptides Intracellular Biotechnology |
| Zdroj: | Bioconjugate Chemistry. 21:289-295 |
| ISSN: | 1520-4812 1043-1802 |
| Popis: | Cationic quantum dots (QDs) were utilized to complex small interfering RNA (siRNA) for studying intracellular trafficking, unpacking, and gene silencing. Positively charged polyethylenimine (PEI) was covalently conjugated on the surface of QDs to complex with cyanine dye labeled vascular endothelial growth factor siRNA (cy5-VEGF siRNA) for the formation of nanosized polyelectrolyte complexes (PEC). Fluorescence resonance energy transfer (FRET) was achieved between cy5-VEGF siRNA and PEI conjugated QDs (QD625) in the complex. From confocal microscopic analysis, intracellular uptake and release of siRNA from the PEC were visualized as a function of incubation time. The extent of cy5-siRNA release from the PEC was quantitatively evaluated by flow cytometric analysis. In addition, PEI conjugated QDs were further modified with a protein transduction domain (PTD) from human transcriptional factor, Hph-1. The two siRNA/QD-PEI complexes with and without Hph-1 have shown markedly different intracellular uptake behaviors and unpacking kinetics of cy5-siRNA. However, they exhibited similar extent of VEGF gene knockout regardless of Hph-1, but showed much higher gene silencing efficiency than siRNA/PEI complexes. The present study demonstrates that PEI conjugated QDs can be utilized as a useful siRNA carrier to analyze intracellular trafficking and unpacking pathway as well as to effectively silence a target gene. |
| Databáze: | OpenAIRE |
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