Pore forming polyalkylpyridinium salts from marine sponges versus synthetic lipofection systems: distinct tools for intracellular delivery of cDNA and siRNA

Debra McLaggan, Noppadon Adjimatera, Kristina Sepcic, Marcel Jaspars, David J MacEwan, Ian S Blagbrough, Roderick H Scott

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Abstract

Background: Haplosclerid marine sponges produce pore forming polyalkylpyridinium salts ( poly-APS), which can be used to deliver macromolecules into cells. The aim of this study was to investigate the delivery of DNA, siRNA and lucifer yellow into cells mediated by poly-APS and its potential mechanisms as compared with other lipofection systems ( lipofectamine and N-4, N(9-)dioleoylspermine (LipoGen)). DNA condensation was evaluated and HEK 293 and HtTA HeLa cells were used to investigate pore formation and intracellular delivery of cDNA, siRNA and lucifer yellow.

Results: Poly-APS and LipoGen were both found to be highly efficient DNA condensing agents. Fura-2 calcium imaging was used to measure calcium transients indicative of cell membrane pore forming activity. Calcium transients were evoked by poly-APS but not LipoGen and lipofectamine. The increases in intracellular calcium produced by poly-APS showed temperature sensitivity with greater responses being observed at 12 C compared to 21 C. Similarly, delivery of lucifer yellow into cells with poly-APS was enhanced at lower temperatures. Transfection with cDNA encoding for the expression enhanced green fluorescent protein was also evaluated at 12 C with poly-APS, lipofectamine and LipoGen. Intracellular delivery of siRNA was achieved with knockdown in beta-actin expression when lipofectamine and LipoGen were used as transfection reagents. However, intracellular delivery of siRNA was not achieved with poly-APS.

Conclusion: Poly-APS mediated pore formation is critical to its activity as a transfection reagent, but lipofection systems utilise distinct mechanisms to enable delivery of DNA and siRNA into cells.

Original languageEnglish
Article number6
Number of pages12
JournalBMC Biotechnology
Volume6
DOIs
Publication statusPublished - 16 Jan 2006

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Keywords

  • gene delivery
  • Reniera-sarai
  • membrane-properties
  • Haliclona-vicosa
  • ethidium-bromide
  • mammalian-cells
  • DNA
  • spermine
  • condensation
  • transfection

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