Cationic quantum dots for intracellular delivery of siRNA and quantitative analysis of polyelectrolyte unpacking via FRET

Abstract

A new class of nanoparticle based gene delivery carrier assembled from semiconductor quantum dots (QDs) and polyethyleneimine (PEI) has been developed and utilized for the intracellular trafficking of small interfering RNA (siRNA) as well as the gene silencing. Positively charged PEI was covalently conjugated on the surface of QDs to induce spontaneous charge interactions with cyanine dye labeled vascular endothelial growth factor siRNA (cy-VEGF siRNA) for the formation of nano-sized polyelectrolyte complexes (PECs). Due to the condensed structure of QD labeled PECs, fluorescence resonance energy transfer (FRET) was achieved between cy-VEGF siRNA and PEI conjugated QDs. The resultant PECs offered valuable information about the intracellular pathways of gene delivery carrier and the cytosol specific release of siRNA by the FRET mechanism. From confocal microscopic analysis, intracellular uptake and release of siRNA from the PECs were visualized as a function of incubation time. In addition, the extent of siRNA release from the PECs was quantitatively evaluated by a flow cytometric analysis. An alternative intracellular pathway of siRNA was explored using PEI conjugated QDs modified with the protein transduction domain (PTD) from human transcriptional factor Hph-1. Two kinds of siRNA/QDs PECs have shown different intracellular uptake mechanisms and trafficking pathways of siRNA. Both siRNA/QDs PECs offered superior VEGF gene silencing efficiency in serum condition compared to the conventional gene delivery agents such as lipofectamine and PEI. The present study demonstrates that the PEI conjugated QDs can be utilized not only as a versatile tool to analyze intracellular trafficking pathway but also as an efficient gene delivery carrier for therapeutic RNA.