title: Transfection of human monocyte-derived dendritic cells with CpG oligonucleotides.
abstract: Monocyte-derived dendritic cells (mDC), the most frequently applied DC subset in clinical studies, which can be obtained easily from peripheral blood monocytes after incubation with GM-CSF and IL-4, have not been clearly demonstrated to be activated by CpG oligodeoxynucleotides (ODN). The development of novel molecular strategies - such as the use of CpG-ODN - to increase immunological functions and thus improve the therapeutic efficiency of mDC vaccines in the treatment of malignant diseases is highly desirable. CpG-ODN need to be internalized into specific intracellular compartments to be active. Therefore, we applied electroporation and lipofection and compared these techniques with incubation to overcome possible defects in localization. Conditions of CpG-ODN transfection of these cells were optimized using fluorescein-marked ODN 2216. We were able to achieve high transfection efficiencies with various methods of delivery. However, we did not observe increased expression of maturation-associated and functionally relevant surface antigens (CD14, HLA-DR, CD40, CD83, CD80 and CD86), significant secretion of IL-12 and IFN-alpha in culture supernatant, or enhanced antitumour activation of cytokine-induced killer cells. In conclusion, our results show that non-viral transfection of CpG-ODN is not sufficient to overcome resistance of mDC to CpG activation.
authors.affiliation: Department of Internal Medicine I, University of Bonn, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Erhardt
authors.forename: Marcus
authors.initials: M
authors.lastname: Gorschlüter
authors.forename: Jens
authors.initials: J
authors.lastname: Sager
authors.forename: Carsten
authors.initials: C
authors.lastname: Ziske
authors.forename: John
authors.initials: J
authors.lastname: Strehl
authors.forename: Marie V
authors.initials: MV
authors.lastname: Lilienfeld-Toal
authors.forename: Ingo G H
authors.initials: IG
authors.lastname: Schmidt-Wolf
mesh_terms: D015703:Antigens, CD / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D018920:Coculture Techniques; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology* / Q000378:metabolism; D018274:Electroporation; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry; D005434:Flow Cytometry; D006133:Growth Substances / Q000494:pharmacology; D006684:HLA-DR Antigens / Q000378:metabolism; D006801:Humans; D016898:Interferon-alpha / Q000378:metabolism; D018664:Interleukin-12 / Q000378:metabolism; D007694:Killer Cells, Natural / Q000276:immunology; D008070:Lipopolysaccharides / Q000494:pharmacology; D009000:Monocytes / Q000166:cytology*; D009838:Oligodeoxyribonucleotides / Q000235:genetics / Q000276:immunology*; D000644:Quaternary Ammonium Compounds / Q000737:chemistry; D014162:Transfection / Q000379:methods*
chemical_list: D015703:Antigens, CD; C408982:CPG-oligonucleotide; D005229:Fatty Acids, Monounsaturated; D006133:Growth Substances; D006684:HLA-DR Antigens; D016898:Interferon-alpha; D008070:Lipopolysaccharides; D009838:Oligodeoxyribonucleotides; D000644:Quaternary Ammonium Compounds; C482199:lipopolysaccharide, E coli O55-B5; D018664:Interleukin-12; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane