Involvement of specific mechanism in plasmid DNA uptake by mouse peritoneal macrophages.
The binding and uptake of plasmid DNA encoding luciferase reporter gene (pCMV-Luc) were studied in vitro using cultured mouse peritoneal macrophages. A significant and time-dependent cellular association of [32P]pCMV-Luc with resident macrophages was observed at 37 degrees C and this decreased at 4 degrees C. The binding at 4 degrees C was saturable and a Scatchard plot gave a maximum binding capacity of 0.81 microgram/mg-protein and a dissociation constant of 0.30 microgram/ml. The binding of [32P]-pCMV-Luc was inhibited by polyinosinic acid, dextran sulfate and salmon sperm DNA, but not by polycytidylic acid, dextran and EDTA. A confocal microscopic study demonstrated that fluorescein-labeled pCMV-Luc was internalized at 37 degrees C while only cell surface binding occurred at 4 degrees C. No significant luciferase gene expression was obtained after incubation with a high concentration (100 micrograms/ml) of pCMV-Luc. These data suggest that plasmid DNA is taken up by macrophages via a mechanism mediated by a receptor like the macrophage scavenger receptor.
Takagi
T
T
Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Japan.
Hashiguchi
M
M
Mahato
R I
RI
Tokuda
H
H
Takakura
Y
Y
Hashida
M
M
D000818:Animals; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D016264:Dextran Sulfate / Q000494:pharmacology; D017930:Genes, Reporter; D008156:Luciferases / Q000235:genetics; D017737:Macrophages, Peritoneal / Q000378:metabolism*; D051379:Mice; D010957:Plasmids / Q000493:pharmacokinetics*; D011069:Poly I / Q000494:pharmacology
D011069:Poly I; D016264:Dextran Sulfate; D008156:Luciferases