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Insight into the relationship between the cell culture model, cell trafficking and siRNA silencing efficiency

Capel, Victoria; Vllasaliu, Driton; Watts, Peter; Stolnik, Snow

Authors

Victoria Capel

Driton Vllasaliu

Snow Stolnik snow.stolnik@nottingham.ac.uk



Abstract

Despite research efforts, cell uptake processes determining siRNA silencing efficiency remain unclear. Here, we examine the relationship between in vitro cell culture models, cellular trafficking and siRNA silencing efficiency to provide a mechanistic insight on siRNA delivery system design. Model siRNA-polyplexes, based on chitosan as a ‘classical’ condensing agent, were applied to a panel of lung epithelial cell lines, H1299, A549 and Calu-3 and cell internalization levels, trafficking pathways and gene silencing assessed on exposure to pharmacological inhibitors. The data reveal striking differences in the internalization behaviour and gene silencing efficiency in the tested cell lines, despite their common lung epithelial origins. The model system’s silencing was lower where clathrin internalization pathway predominated in Calu-3, relative to silencing in H1299 cells where a non-clathrin internalization appears dominant. Increased silencing on endosomal disruption was apparent in Calu-3 cells, but absent when cellular internalization was not predominantly clathrin-mediated in A549 cells. This highlights that identifying cell trafficking pathways before incorporation of functional components to siRNA delivery systems (e.g. endosomolytic compounds) is crucial. The study hence stresses the importance of selection of appropriate cell culture model, relevant to in vivo target, to assess the gene silencing efficiency and decide which functionalities the ‘stratified siRNA silencing vector’ requires.

Citation

Capel, V., Vllasaliu, D., Watts, P., & Stolnik, S. (2016). Insight into the relationship between the cell culture model, cell trafficking and siRNA silencing efficiency. Biochemical and Biophysical Research Communications, 477(2), 260-265. https://doi.org/10.1016/j.bbrc.2016.06.054

Journal Article Type Article
Acceptance Date Jun 9, 2016
Online Publication Date Jun 24, 2016
Publication Date Aug 19, 2016
Deposit Date Jul 22, 2016
Publicly Available Date Jul 22, 2016
Journal Biochemical and Biophysical Research Communications
Print ISSN 0006-291X
Electronic ISSN 0006-291X
Publisher Elsevier
Peer Reviewed Peer Reviewed
Volume 477
Issue 2
Pages 260-265
DOI https://doi.org/10.1016/j.bbrc.2016.06.054
Keywords Endocytosis pathways; Lung siRNA therapy; Pharmacological inhibitors; siRNA polyplexes; siRNA silencing
Public URL http://eprints.nottingham.ac.uk/id/eprint/35333
Publisher URL http://www.sciencedirect.com/science/article/pii/S0006291X1630972X?via%3Dihub
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0
Additional Information This article is maintained by: Elsevier; Article Title: Insight into the relationship between the cell culture model, cell trafficking and siRNA silencing efficiency; Journal Title: Biochemical and Biophysical Research Communications; CrossRef DOI link to publisher maintained version: https://doi.org/10.1016/j.bbrc.2016.06.054; Content Type: article; Copyright: © 2016 The Authors. Published by Elsevier Inc.

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0





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