Small interfering RNA (siRNA) delivery into murine bone marrow-derived dendritic cells by electroporation
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- 作者：Jonathan Jantsch ; Nadine Turza ; Melanie Volke ; Kai-Uwe Eckardt ; Michael Hensel ; Alex ; er Steinkasserer ; Carsten Willam ; Alex ; er T. Prechtel
- 关键词：Dendritic cells ; siRNA transfer ; Electroporation ; Lipofection ; MAPK1
- 刊名：Journal of Immunological Methods
- 出版年：2008
- 出版时间：20 August 2008
- 年：2008
- 卷：337
- 期：1
- 页码：71-77
- 全文大小：720 K
文摘
Selective gene silencing by RNA interference (RNAi) has been shown to be an efficient method for the targeted manipulation of cellular functions. In this study we describe for the first time electroporation as a suitable and efficient method for the delivery of small interfering RNA (siRNA) into murine bone marrow-derived dendritic cells (BM-DC). Using a fluorescein-labeled non-silencing siRNA duplex, we established an electroporation protocol yielding routinely > 90 % positive cells. We investigated the effects of siRNA electroporation on BM-DC viability, phenotype and ability to induce allogeneic T cell proliferation. Finally, using siRNAs directed against MAPK1 and the transcription factor HIF-1α we were able to demonstrate an efficient knock down of cellular mRNA- and protein level in electroporated BM-DC. Furthermore, knocking down the transcription factor HIF-1α impeded hypoxic induction of HIF-1α target genes. We therefore propose siRNA electroporation into murine BM-DC as an efficient method to manipulate BM-DC function without the use of chemical transfection reagents. This new approach is superior to lipofection regarding detrimental effects of lipid-based transfection agents on BM-DC immunobiology.