The transfection of Jurkat T-leukemic cells by use of pH sensitive immunoliposomes.
A gene transfer vector has been developed utilising anionic liposomes as a carrier of plasmid DNA (pEGlacZ, 7.6 kb) to transfect CD3+ T lymphocytes (Jurkat cells). The plasmid DNA that contained the Escherichia coli beta-galactosidase reporter gene was condensed using poly-l-lysine of molecular mass 20,700 (PLK99) to form a polyplex which was interacted with several anionic liposome formulations to form lipopolyplexes. The liposome formulations where based on dioleoylphosphatidylethanolamine (DOPE) in combination with cholesterol and dioleoylphosphatidylcholine (DOPC) and oleic acid, or dimyristoylphosphatidylethanolamine (DMPE). For targeting to the Jurkat cells distearoylphosphatidylethanolamine (DSPE) linked to poly (ethylene glycol) molecular mass 2,000 and coupled to anti-CD3 antibody was incorporated. The polyplexes and lipopolyplexes were characterised in terms of size, zeta potential, agarose gel electrophoresis and electron microscopy and the permeability of the lipopolyplexes to liposome-encapsulated glucose was determined. The polyplexes consisted of a mixed population of rod-like structures (53-160 nm long and 23-31 nm diameter) and spheres (18-30 nm diameter). The lipopolyplexes retained a permeability barrier although were more permeable to glucose than their component liposomes. The poly-l-lysine condensing agent was still susceptible to pronase digestion suggesting that the polyplex was associated with the outer surface of the liposome. The lipopolyplexes with lipid composition DOPE/cholesterol/OA/DSPE-PEG2000 anti-CD3+ PLK99-plasmid DNA had significant gene transfer activity, as monitored by beta-galactosidase expression, that depended on the charge ratio of the component polyplex and the lipid/DNA weight ratio. The anti-CD3 antibody, the liposomal lipid and pH sensitivity were essential for transfection activity.
Turner
Christopher
C
School of Biological Sciences, University of Manchester, UK.
Weir
Neil
N
Catterall
Catherine
C
Baker
Terry S
TS
Carrington
Bruce
B
Jones
Malcolm N
MN
D000906:Antibodies; D004247:DNA / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D005822:Genetic Vectors / Q000235:genetics / Q000648:ultrastructure; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019169:Jurkat Cells; D008081:Liposomes / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D010714:Phosphatidylethanolamines; D011107:Polylysine / Q000378:metabolism; D011402:Pronase / Q000378:metabolism; D014162:Transfection / Q000379:methods*
D000906:Antibodies; D008081:Liposomes; D046911:Macromolecular Substances; D010714:Phosphatidylethanolamines; C038089:1,2-distearoylphosphatidylethanolamine; C094877:dioleoyl phosphatidylethanolamine; D011107:Polylysine; D004247:DNA; D011402:Pronase; C018525:1,2-dimyristoylphosphatidylethanolamine