The effect of CpG motifs on gene expression and clearance kinetics of aerosol administered polyethylenimine (PEI)-plasmid DNA complexes in the lung.
Presence of CpG motifs within pDNA is widely reported to influence transgene expression as well as inflammatory response to nonviral gene vector complexes. Here, we analyzed gene expression kinetics and lung clearance after aerosol delivery of polyethylenimine (PEI) complexes with two different plasmid vectors: a first generation plasmid, pCMVLuc, and a plasmid with depleted CpG motifs, pCpG-free-Luc. After aerosol delivery, equal nanogram amounts of PEI-pDNA complexes were deposited in murine lungs. Luciferase expression observed at day one post administration of PEI-pCpG-free-Luc complexes was 60-fold higher than for PEI-pCMVLuc complexes and decreased 16-fold at day 7 post application. In contrast, luciferase expression from PEI-pCMVLuc particles remained at levels comparable to day 1 post application. In agreement with these observations, PEI-pCpG-free-Luc complexes were cleared from the lungs at rates 6-fold faster than those observed for PEI-pCMVLuc particles. A more detailed analysis of pDNA distribution within bronchoalveolar lavage fluid (BALF), BALF cells and lung tissue showed 660-fold higher amounts of pCpG-free-Luc in BALF cells compared to pCMVLuc, whereas the amount of pCpG-free-Luc in lung tissue was 15-fold lower compared to pCMVLuc 1h after administration. Our results demonstrate that complexes of PEI with CpG-motif-free DNA are taken up more extensively by BALF cells, while the clearance of pCMVLuc from the lung tissue is significantly slower than for the CpG-free plasmid. Administration of PEI-pCpG-free-Luc caused transient decrease in number of resident lung cells, while their activation was more pronounced with PEI-pCMVLuc particles. Our results demonstrate that the level of transgene expression is increased with CpG-motif-free pDNA but the longevity of expression correlates with pDNA clearance pattern depending on the presence of CpG motifs within the plasmid.
Lesina
Eugenia
E
Department of Pediatrics, Ludwig-Maximilians University, 80337 Munich, Germany.
Dames
Petra
P
Rudolph
Carsten
C
D000336:Aerosols / Q000737:chemistry; D000818:Animals; D018899:CpG Islands; D004247:DNA / Q000008:administration & dosage*; D005260:Female; D015870:Gene Expression; D018014:Gene Transfer Techniques; D007700:Kinetics; D008156:Luciferases / Q000235:genetics; D008168:Lung / Q000378:metabolism*; D016676:Macrophages, Alveolar / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids / Q000008:administration & dosage; D011094:Polyethyleneimine / Q000737:chemistry*
D000336:Aerosols; D011094:Polyethyleneimine; D004247:DNA; D008156:Luciferases