Silicon Quantum Dots Functionalized for siRNA Delivery
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1257-O07-06
Silicon Quantum Dots Functionalized for siRNA Delivery
Stefanie Klein1, Oliver Zolk2, Falk Schrödl3,4 and Carola Kryschi1 1
Department Chemistry and Pharmacy, Physical Chemistry I and ICMM, Friedrich-Alexander University of Erlangen-Nuremberg, Egerlandstr. 3, D-91058 Erlangen, Germany.
2
Institute of Experimental and Clinical Pharmacology and Toxicology, Friedrich-Alexander University of Erlangen-Nuremberg, Fahrstr. 17, D-91054 Erlangen, Germany.
3
Institute of Anatomy I, Friedrich-Alexander University of Erlangen-Nuremberg, Krankenhausstr. 9, D-91054 Erlangen, Germany.
4
Deptartments of Anatomy/Ophthalmology, Paracelsus Medical University, Strubergasse 21, A5020 Salzburg, Austria.
ABSTRACT Biocompatible, water-soluble, green luminescent silicon quantum dots (SiQDs) were developed as transfection tool for small interfering RNA (siRNA). The research goal was to down-regulate via the RNA interference (RNAi) mechanism the P-glycoprotein expression of the multidrug resistant gene 1 (MDR1) in a human colon carcinoma cell line (Caco-2). The internalization of 2-vinylpyridine terminated SiQDs (2-vipySiQDs) by Caco-2 cells as observed in confocal laser scanning microscopy imaging studies occurs via endocytosis. Experiments employing agarose gel electrophoresis revealed that 2-vipySiQD-siRNA complexes are formed through electrostatic interactions. The release of siRNA in the cytosol with subsequently RNAi induced down-regulation of the P-glycoprotein translation was verified by detecting a reduced ABCB1 mRNA level in transfected Caco-2 cells employing real-time PCR. Additional evidence for successful ABCB1 gene silencing was obtained by measuring a significant decrease of the Pglycoprotein transporter efficiency for the fluorescent substrate Rhodamine 123 (Rh123).
INTRODUCTION RNAi is a post-transcriptional gene silencing mechanism for regulating gene expression during development [1-3]. Prospect of utilizing this gene-silencing mechanism for biomedical research and therapy has been raised, when Tuschl et al. [4] verified that RNAi in mammalian cells may be mediated by 21- and 22-nucleotide RNAs serving as effector molecules of sequence-specific gene silencing. Therefore the antisense (guide) strand of the short interfering, double stranded RNA (siRNA) is incorporated into a RNA-induced silencing complex (RISC). Gene silencing is achieved when the antisense strands of the siRNA guide the RISCs to homologous mRNA in the cell. The RNA endonucleases in the RISCs provide sequence-specific cleavage of mRNAs and thereupon down-regulation of protein expression. Therapeutic applications of siRNAs require their in-vivo delivery into the target cell. This remains a major hurdle for RNAi therapy, since naked siRNA may not cross the mammalian cell membrane. In
addition, many of the transfection methods used for in vitro studies cannot be used in most invivo settings. One promising strategy for in-vivo delivering siRNAs into cells is based on luminescent semiconductor quantum dots (QDs) as allowing for tracking t
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