title: Non-viral-mediated gene therapy approaches for bone repair.
abstract: OBJECTIVE
  Bone repair strategies continue to be developed for alternatives to autografting, allogeneic implants of banked bone, and other bone substitutes. Efforts have included the delivery of potent growth and/or differentiation factors and the use of gene therapy. For bone regeneration, gene therapy is the delivery, uptake and expression of DNA that has been localized to a wound bed. The objective of the current study is to investigate methods to enhance non-viral-mediated means of gene uptake and expression for use in bone regeneration.
  
  
  METHODS
  Several types of DNA-polymer complexes, either applied directly to baby hamster kidney (BHK) cells, or released from a porous, resorbable gene-activated matrix (GAM), were evaluated in vitro for their ability to transfect cells with a circular plasmid DNA construct expressing green fluorescent protein. Complexes included conjugates containing a lipophilic reagent, liposomes, poly-ethyl-oxazoline, and poly-ethyleneimine (PEI). Data were subjected to analysis of variance and Fisher's protected least significant difference for multiple comparisons with significance established at p < 0.05.
  
  
  RESULTS
  Transfection efficiencies of the liposome and PEI complexes improved in vitro when released from resorbable GAMs. The lipophilic reagent FuGene 6 demonstrated abundant uptake and expression in the initial 1- and 2-day evaluation periods. In contrast, the DNA-liposome and PEI GAM complexes demonstrated a sustained release, uptake and expression by the BHK cells at the 2-, 4-, and 7-day, and 4- and 7-day evaluation intervals, respectively.
  
  
  CONCLUSIONS
  GAM technology appears to improve the functional stability and release duration of incorporated DNA-polymer complexes in the present in vitro studies. The ongoing objective of our research is to develop a localized treatment to improve the uptake and expression of plasmid DNA by non-viral-mediated gene therapy.
authors: Department of Surgery, Division of Plastic and Reconstructive Surgery, School of Medicine, Oregon Health and Science University, OR 97239, USA. winns@ohsu.edu
authors: S R
authors: SR
authors: Winn
authors: J C
authors: JC
authors: Chen
authors: X
authors: X
authors: Gong
authors: S V
authors: SV
authors: Bartholomew
authors: S
authors: S
authors: Shreenivas
authors: W
authors: W
authors: Ozaki
mesh_terms: D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry; D001861:Bone Regeneration / Q000235:genetics*; D002460:Cell Line; D002784:Cholesterol / Q000737:chemistry; D024042:Collagen Type I / Q000737:chemistry; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D005786:Gene Expression Regulation / Q000235:genetics; D015316:Genetic Therapy; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D008055:Lipids / Q000737:chemistry; D008081:Liposomes; D008164:Luminescent Proteins / Q000235:genetics; D010080:Oxazoles / Q000737:chemistry; D010957:Plasmids / Q000235:genetics; D011091:Polyesters / Q000737:chemistry; D011094:Polyethyleneimine / Q000737:chemistry; D011108:Polymers / Q000737:chemistry; D000644:Quaternary Ammonium Compounds / Q000737:chemistry; D014162:Transfection / Q000379:methods
chemical_list: D001672:Biocompatible Materials; D024042:Collagen Type I; D005229:Fatty Acids, Monounsaturated; D005456:Fluorescent Dyes; C411955:FuGene; D008055:Lipids; D008081:Liposomes; D008164:Luminescent Proteins; D010080:Oxazoles; D011091:Polyesters; D011108:Polymers; D000644:Quaternary Ammonium Compounds; D049452:Green Fluorescent Proteins; C071857:polyethyloxazoline; C033616:poly(lactide); D011094:Polyethyleneimine; D004247:DNA; D002784:Cholesterol; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane