Delivery of siRNA from lyophilized polymeric surfaces

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Standard in vitro gene silencing protocols are performed using aqueous formulations of transfection reagents and small interfering RNAs (siRNA) reconstituted immediately prior to use. In this study, we describe a method for producing gene silencing-active lyophilized cationic polymer (chitosan) or lipid (TransIT-TKO) siRNA formulations. We demonstrate specific and efficient knockdown of enhanced green fluorescent protein (EGFP) in H1299 human lung carcinoma cells transfected in plates pre-coated with both TransIT-TKO/siRNA ( approximately 85%) and a chitosan/siRNA formulation containing sucrose as lyoprotectant ( approximately 70%). This method removes the necessity for both siRNA reconstitution immediately prior to use and addition onto cells. Furthermore, silencing activity of the chitosan/siRNA formulation was shown over the period studied ( approximately 2 months) when stored at room temperature. Higher cell viability was observed using the chitosan system compared to the lipid formulation. Silencing of the proinflammatory cytokine tumour necrosis factor (TNF-alpha) was also demonstrated in the RAW macrophage cell line using the lyophilized chitosan/siRNA system suggesting that the coating can improve the biocompatibility of medical implants. This work describes an efficient gene silencing methodology using freeze-dried formulations with potential applications as a high throughput screening tool for gene function, biocompatible medical implant components and longer shelf-life therapeutics.
OriginalsprogEngelsk
TidsskriftBiomaterials
Vol/bind29
Nummer4
Sider (fra-til)506-512
Antal sider7
ISSN0142-9612
DOI
StatusUdgivet - 2008

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