Factors that influence transgene expression and cell viability on DNA-PEI-seeded collagen films.
Gene delivery from tissue-engineering devices has the potential to improve healing, but better regulation of the level and duration of gene expression is needed. We hypothesized that transgene expression could be controlled by varying the fabrication and soaking parameters used in making collagen- based gene delivery scaffolds. Collagen films were made from acid-insoluble type I bovine dermal collagen and seeded with plasmid DNA encoding firefly luciferase, complexed with polyethylenimine. By varying the thickness of the films, the volume of the DNA soak solution, and the pH of the DNA soak solution, and by cross-linking the films, we identified variable combinations that produce significantly different levels of cell number and transgene expression in L-929 cells in vitro. Increasing film thickness or soak volume increased overall reporter gene expression. Decreasing film thickness or soak volume decreased cell number but did not significantly change reporter gene expression per cell. Cross-linking by ultraviolet irradiation (before adding the DNA) significantly decreased transgene expression, probably because of decreased swelling of the collagen film. These results suggest that collagen-based biomaterials may be designed and fabricated to induce, in a controlled fashion, various levels of cellularity and transgene expression.
Katz
Jordan M
JM
Orthopedic Research Laboratories, Department of Orthopedic Surgery, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, New Brunswick, New Jersey 08903, USA.
Roth
Charles M
CM
Dunn
Michael G
MG
D000818:Animals; D001672:Biocompatible Materials / Q000378:metabolism*; D002417:Cattle; D002452:Cell Count; D002470:Cell Survival; D024042:Collagen Type I / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D006863:Hydrogen-Ion Concentration; D007739:L Cells; D008156:Luciferases / Q000032:analysis / Q000378:metabolism; D051379:Mice; D010957:Plasmids / Q000378:metabolism; D011094:Polyethyleneimine / Q000378:metabolism*; D011485:Protein Binding; D023822:Tissue Engineering / Q000295:instrumentation; D019076:Transgenes; D014466:Ultraviolet Rays
D001672:Biocompatible Materials; D024042:Collagen Type I; D011094:Polyethyleneimine; D004247:DNA; D008156:Luciferases