Comparison of the gene transfer efficiency of mRNA/GL67 and pDNA/GL67 complexes in respiratory cells.
Complexes between mRNA and GL67:DOPE:DMPE-PEG5000 (GL67) liposomes were formulated and characterized. Subsequently, the in vitro and in vivo expression characteristics of mRNA/GL67 complexes and pDNA/GL67 complexes, each produced at their optimal ratio, were compared in respiratory cells. Transfection of A549 cells with mRNA/GL67 complexes resulted in a much faster expression than after transfection with pDNA/GL67 complexes. The percentage of GFP-positive cells after mRNA and pDNA transfection peaked after 8 and 24 h, respectively. At these time points the percentage of GFP-positive cells was two times higher after mRNA transfection than after pDNA transfection. Furthermore, the efficacy of mRNA/GL67 complexes was independent of the cell cycle. This was in sharp contrast with pDNA/GL67 complexes that caused only a weak expression in nondividing cells. This confirms that the nuclear barrier is a crucial obstacle for pDNA but not for mRNA. Finally, mRNA/GL67 and pDNA/GL67 complexes encoding luciferase were administered intranasally to the lungs of mice. The mRNA/GL67 complexes did not give rise to a measurable luciferase expression in the murine lungs. In contrast, a detectable bioluminescent signal was present in the lungs of mice that received the pDNA/GL67 complexes. We showed that mRNA/GL67 complexes have a lower stability in biological fluids. Consequently, this may be an explanation for their lower performance in vivo.
Laboratory of Gene Therapy, Department of Nutrition, Genetics and Ethology, Faculty of Veterinary Medicine, Ghent University, Heidestraat 19, B-9820 Merelbeke, Belgium.
Oliwia
O
Andries
Marina
M
De Filette
Joanna
J
Rejman
Stefaan C
SC
De Smedt
Jo
J
Demeester
Mario
M
Van Poucke
Luc
L
Peelman
Cindy
C
Peleman
Tony
T
Lahoutte
Niek N
NN
Sanders
D000818:Animals; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D018014:Gene Transfer Techniques; D006801:Humans; D008081:Liposomes / Q000737:chemistry*; D051379:Mice; D008807:Mice, Inbred BALB C; D012333:RNA, Messenger / Q000737:chemistry* / Q000235:genetics*; D014162:Transfection / Q000379:methods
D008081:Liposomes; D012333:RNA, Messenger