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Related Collections Gene Therapy Social Bookmarking                   What's this?  Previous Article  |  Table of Contents  |  Next Article  Blood, 1 July 2001, Vol. 98, No. 1, pp. 49-56 GENE THERAPY Highly efficient gene delivery by mRNA electroporation in human hematopoietic cells: superiority to lipofection and passive pulsing of mRNA and to electroporation of plasmid cDNA for tumor antigen loading of dendritic cells Viggo F. I. Van Tendeloo, Peter Ponsaerts, Filip Lardon, Griet Nijs, Marc Lenjou, Christine Van Broeckhoven, Dirk R. Van Bockstaele, and Zwi N. Berneman From the Laboratory of Experimental Hematology, Antwerp University Hospital, University of Antwerp; the Laboratory of Molecular Genetics, Department of Molecular Genetics, Flanders Interuniversity Institute for Biotechnology (VIB), University of Antwerp; and the Laboratory of Cancer Research and Clinical Oncology, University of Antwerp, Antwerp, Belgium. Designing effective strategies to load human dendritic cells (DCs) with tumor antigens is a challenging approach for DC-based tumor vaccines. Here, a cytoplasmic expression system based on mRNA electroporation to efficiently introduce tumor antigens into DCs is described. Preliminary experiments in K562 cells using an enhanced green fluorescent protein (EGFP) reporter gene revealed that mRNA electroporation as compared with plasmid DNA electroporation showed a markedly improved transfection efficiency (89% versus 40% EGFP+ cells, respectively) and induced a strikingly lower cell toxicity (15% death rate with mRNA versus 51% with plasmid DNA). Next, mRNA electroporation was applied for nonviral transfection of different types of human DCs, including monocyte-derived DCs (Mo-DCs), CD34+ progenitor-derived DCs (34-DCs) and Langerhans cells (34-LCs). High-level transgene expression by mRNA electroporation was obtained in more than 50% of all DC types. mRNA-electroporated DCs retained their phenotype and maturational potential. Importantly, DCs electroporated with mRNA-encoding Melan-A strongly activated a Melan-A-specific cytotoxic T lymphocyte (CTL) clone in an HLA-restricted manner and were superior to mRNA-lipofected or -pulsed DCs. Optimal stimulation of the CTL occurred when Mo-DCs underwent maturation following mRNA transfection. Strikingly, a nonspecific stimulation of CTL was observed when DCs were transfected with plasmid DNA. The data clearly demonstrate that Mo-DCs electroporated with mRNA efficiently present functional antigenic peptides to cytotoxic T cells. 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	Copyright © 2001 by American Society of Hematology         Online ISSN: 1528-0020