Pore forming polyalkylpyridinium salts from marine sponges versus synthetic lipofection systems: distinct tools for intracellular delivery of cDNA and siRNA.
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 N4,N9-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 degrees C compared to 21 degrees 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 degrees 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.


CONCLUSIONS
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.
McLaggan
Debra
D
School of Medical Sciences, College of Life Sciences and Medicine, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, UK. debbie@cabernet.demon.co.uk
Adjimatera
Noppadon
N
Sepcić
Kristina
K
Jaspars
Marcel
M
MacEwan
David J
DJ
Blagbrough
Ian S
IS
Scott
Roderick H
RH
D000818:Animals; D002460:Cell Line; D004247:DNA / Q000008:administration & dosage / Q000493:pharmacokinetics*; D004337:Drug Carriers / Q000737:chemistry*; D006801:Humans; D007668:Kidney / Q000378:metabolism; D008055:Lipids / Q000737:chemistry*; D008081:Liposomes / Q000737:chemistry; D011161:Porifera / Q000737:chemistry*; D016062:Porosity; D011726:Pyridinium Compounds / Q000737:chemistry*; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000493:pharmacokinetics*; D012492:Salts / Q000737:chemistry; D014162:Transfection / Q000379:methods*
D004337:Drug Carriers; D008055:Lipids; C086724:Lipofectamine; D008081:Liposomes; D011726:Pyridinium Compounds; D034741:RNA, Small Interfering; D012492:Salts; D004247:DNA