Efficient gene transfer to periodontal ligament cells and human gingival fibroblasts by adeno-associated virus vectors.
OBJECTIVE
We explored for the first time the possibility to deliver a reporter gene (Green Fluorescence Protein) to human primary periodontal ligament (PDL) cells and human gingival fibroblasts (HGF) using shuttle vectors derived from adeno-associated virus (AAV). Since AAV transduction rates on other human primary fibroblasts have been previously shown to depend on the particular cell lineage and on the employed viral serotype, we determined the most effective AAV variant for periodontal cells comparing different vector types.


METHODS
AAV serotypes 1-5 encoding GFP in single stranded (ss) and self-complementary (sc) vector genome conformations were used to infect primary HGF and PDL cells. Two days post-infection, the percentage of GFP expressing cells was determined by flow cytometry.


RESULTS
Highest transduction rates for both cell types were achieved with self-complementary vectors derived from AAV-2, resulting in GFP expression in up to 86% of PDL cells and 50% of HGF. Transgene expression could be observed by optical microscopy for 2 months after infection. Lower but detectable rates were obtained with serotypes 1, 3 and 5.


CONCLUSIONS
The efficacy demonstrated here and the safety and versatility of AAV technology indicated in previous studies clearly suggest the potential of AAV vectors as tools for gene transfer to periodontal tissues.
Center of Dental Medicine, Department of Operative Dentistry and Periodontology, University of Cologne, Germany.
Melanie
M
Kunze
Anke
A
Huber
Anna
A
Krajewski
Esther
E
Lowden
Natascha
N
Schuhmann
Hildegard
H
Buening
Michael
M
Hallek
Michael
M
Noack
Luca
L
Perabo
D000293:Adolescent; D000328:Adult; D002478:Cells, Cultured; D004277:DNA, Single-Stranded / Q000235:genetics; D000229:Dependovirus / Q000145:classification / Q000235:genetics*; D004847:Epithelial Cells / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology*; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000235:genetics; D018014:Gene Transfer Techniques / Q000145:classification*; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors / Q000145:classification / Q000235:genetics*; D016679:Genome, Viral / Q000235:genetics; D005881:Gingiva / Q000166:cytology*; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D010513:Periodontal Ligament / Q000166:cytology*; D010957:Plasmids / Q000235:genetics; D012703:Serotyping; D013997:Time Factors; D014161:Transduction, Genetic; D019076:Transgenes / Q000235:genetics; D055815:Young Adult
D004277:DNA, Single-Stranded; D049452:Green Fluorescent Proteins