Cell-surface glycosaminoglycans inhibit cation-mediated gene transfer.
BACKGROUND
Cationic polymers and liposomes are used to wrap DNA into complexes that promote its cellular uptake. The mechanisms of the uptake and the intracellular fate of these complexes are obscure, as are reasons for an unpredictable and sometimes poor efficiency of the transgene expression. Polyanionic glycosaminoglycans (GAGs) on the cell surface interact with the cationic DNA complexes and influence transfection.


METHODS
The quantities of heparan sulfate (HS), chondroitin sulfate (CS) and hyaluronan (HA) on the cell surface of mutated Chinese hamster ovary (CHO) cells and manipulated (chlorate, xyloside, chondroitinase ABC or Streptomyces hyaluronidase) smooth muscle cells were correlated with the uptake of four different DNA complexes, and the expression of the transgene.


RESULTS
Two CHO mutants, without cell-surface HS and CS, showed a 1.5-6-fold increase in cellular association of DNA, and 3-25-fold increase of transgene expression, as compared with the wild type. A CHO mutant with a 5.7-fold increase of cell-surface CS, but devoid of HS, showed enhanced DNA association, but 20-40% reduction in its expression. The removal of HS, CS, or HA from the cell surface of smooth muscle cells had a minor or insignificant effect on the cell association and transfection of the carriers and only polyethyleneimine showed increased association and expression of the transgene.


CONCLUSIONS
The uptake of DNA complexes varies depending on carrier, cell type and amounts of cell-surface HS, CS, and HA, whereas all GAGs inhibit the transgene expression. This implies that cell-surface GAGs probably direct complexes into intracellular compartments that do not support transcription.
Ruponen
Marika
M
Department of Pharmaceutics, University of Kuopio, P.O. Box 1627, FIN-70211 Kuopio, Finland. Marika.Ruponen@uku.fi
Honkakoski
Paavo
P
Tammi
Markku
M
Urtti
Arto
A
D000818:Animals; D016466:CHO Cells; D002412:Cations; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D002809:Chondroitin Sulfates / Q000378:metabolism; D006224:Cricetinae; D004247:DNA / Q000493:pharmacokinetics; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D006025:Glycosaminoglycans / Q000737:chemistry / Q000378:metabolism*; D006497:Heparan Sulfate / Q000378:metabolism; D006820:Hyaluronic Acid / Q000378:metabolism; D006821:Hyaluronoglucosaminidase / Q000737:chemistry / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009130:Muscle, Smooth / Q000166:cytology / Q000502:physiology; D011817:Rabbits
D002412:Cations; D002809:Chondroitin Sulfates; D004247:DNA; D006025:Glycosaminoglycans; D006497:Heparan Sulfate; D006820:Hyaluronic Acid; D006821:Hyaluronoglucosaminidase; D008081:Liposomes; D008156:Luciferases