title: Long-lasting secretion of transgene product from differentiated and filter-grown retinal pigment epithelial cells after nonviral gene transfer.
abstract: OBJECTIVE
  The purpose of this study was to investigate the extent, duration, and direction of transgene expression after nonviral gene transfer to differentiated retinal pigment epithelial (RPE) cells.
  
  
  METHODS
  Polarized human RPE cells (ARPE-19) were transfected with nonviral vectors [DOTAP/DOPE with and without protamine sulfate (PS), DOTAP, PEI (polyethyleneimine), DHP-12] using secreted alkaline phosphatase (SEAP) as a reporter gene. Cellular uptake was studied by flow cytometry.
  
  
  RESULTS
  Up to 80-fold differences were observed in the peak reporter gene expression. The highest peak levels and the longest lifetime of SEAP expression (> 69 days) were obtained with DOTAP/DOPE/PS/pDNA complexes. With PEI, higher expression was seen to the apical side than to the basolateral side.
  
  
  CONCLUSIONS
  In contrast to most differentiated epithelial cells, the differentiated RPE cells can be transfected at high and prolonged levels with selected lipoplexes.
authors: Department of Pharmaceutics, University of Kuopio, Kuopio, Finland.
authors: Eliisa
authors: E
authors: Mannermaa
authors: Seppo
authors: S
authors: Rönkkö
authors: Marika
authors: M
authors: Ruponen
authors: Mika
authors: M
authors: Reinisalo
authors: Arto
authors: A
authors: Urtti
mesh_terms: D000469:Alkaline Phosphatase / Q000235:genetics*; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation; D005229:Fatty Acids, Monounsaturated / Q000378:metabolism; D005434:Flow Cytometry; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000201:enzymology*; D010957:Plasmids; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D019076:Transgenes
chemical_list: C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D005229:Fatty Acids, Monounsaturated; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D000469:Alkaline Phosphatase; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane