Calcium enhances the transfection potency of stabilized plasmid-lipid particles.
Previous work from this laboratory has shown that plasmid DNA can be encapsulated in small (70-nm-diameter) stabilized plasmid-lipid particles (SPLP) that consist of a single plasmid encapsulated within a bilayer lipid vesicle. SPLP preferentially transfect tumor tissue following intravenous administration. Although the levels of transgene expression in vivo are greater for SPLP than can be achieved with naked DNA or complexes, they are lower than may be required for therapeutic benefit. In the present work we examine whether Ca2+ can enhance the transfection potency of SPLP. It is shown that Ca2+ can enhance SPLP transfection potency in bovine hamster kidney cells by 60- to 100-fold when treated in serum containing medium and an additional 60-fold when serum is absent for the initial 10 min of the transfection period. When cells are treated with SPLP in the presence of Ca2+, there is a fivefold increase in intact plasmid in the cell. It is also shown that this Ca2+ effect involves the formation of calcium phosphate precipitates; however, these precipitates are not directly associated with the SPLP plasmid DNA. The ability of calcium phosphate to facilitate delivery of other macromolecules without direct association is also demonstrated by the release of large-molecular-weight dextrans from endosomal/lysosomal compartments in the presence of calcium phosphate. Finally, it is shown that, unlike naked DNA, SPLP transfection potency in the presence of calcium phosphate is not affected by nuclease activity.
Sandhu
Ammen P
AP
Department of Biochemistry and Molecular Biology, 2146 Health Sciences Mall, University of British Columbia, Vancouver, BC, Canada V6T 1Z3. ammen@interchange.ubc.ca
Lam
Angela M I
AM
Fenske
David B
DB
Palmer
Lorne R
LR
Johnston
Michael
M
Cullis
Pieter R
PR
D000818:Animals; D002118:Calcium / Q000737:chemistry* / Q000502:physiology; D002130:Calcium Phosphates / Q000737:chemistry; D002417:Cattle; D002460:Cell Line; D006224:Cricetinae; D011992:Endosomes / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology; D007424:Intracellular Fluid / Q000737:chemistry / Q000378:metabolism; D050356:Lipid Metabolism / Q000235:genetics; D008055:Lipids / Q000737:chemistry* / Q000235:genetics; D008247:Lysosomes / Q000378:metabolism; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism*; D014162:Transfection
D002130:Calcium Phosphates; D008055:Lipids; C020243:calcium phosphate; D002118:Calcium