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Tracking in vitro and in vivo siRNA electrotransfer in tumor cells

Research Article

J RNAi Gene Silenc (May 2008), 4(1), 281-288

doi: jrgsxx

Published online: 27 May 2008

Full Text: (html | pdf ~739kb | refs)

Tracking in vitro and in vivo siRNA electrotransfer in tumor cells

Aurelie Paganin-Gioannii †, Elisabeth Bellard †, Bettina Couderc ‡, Justin Teissié †* and Muriel Golzio †*

† IPBS CNRS (UMR 5089 Université de Toulouse III, CNRS), 205 Route de Narbonne, 31077 Toulouse France

‡ Université de Toulouse III ; Institut C. Regaud, 20-24 rue du Pont ST Pierre, 31052 Toulouse France

*Correspondence to: Muriel Golzio and Justin Teissié, Email: muriel.golzio@ipbs.fr (MG) or justin.teissie@ipbs.fr (JT), Tel: +33 561 175812/13; Fax: +33 561 175994

Received: 29 April 2008, Revised: 15 May 2008, Accepted: 20 May 2008

© Copyright The Authors

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ABSTRACT

RNA interference-mediated gene silencing offers the potential of targeted inhibition of disease-relevant genes. In vivo delivery of RNAi reagents can be obtained by a variety of approaches. Physical delivery methods appear safer and lack side effects. Electro-permeabilization is one of the non-viral methods successfully used to transfer small interfering RNAs (siRNAs) in vitro and in vivo. A promising approach may be, very little is known about the fundamental processes mediating siRNA transfer. In this study, we have investigated cellular delivery pathways involved in electro-delivery of siRNAs by a direct fluorescence imaging method. An Alexa-labeled siRNA was electro-transferred into murine melanoma cells stably-expressing the enhanced green fluorescent protein (eGFP) target reporter gene. The silencing of eGFP gene expression was quantified by time-lapsed fluorescence microscopy. Fluorescently-labeled siRNAs were found distributed homogeneously in cytoplasm 48 hours after electro-transfer, apparently by diffusion. Furthermore, siRNAs showed homogeneous distribution in vivo 48 hrs after intra-tumoral injection followed by electro- permeabilization. Histological fluorescence microscopy showed that siRNAs were mostly localized in the cytoplasm. Overall, this study shows that electro-permeabilization facilitates cytoplasmic distribution of siRNA, both in cultured cells and in vivo. This method offers a potential therapeutic tool to facilitate direct siRNA penetration into solid tumors.

KEYWORDS: Electro-permeabilization, electro-poration, RNAi, tumors, fluorescence microscopy

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