Nonviral and Viral Gene Transfer Into Different Subsets of Human Dendritic Cells Yield Comparable Efficiency of Transfection
作者:
Andreas Lundqvist,
Gabriele Noffz,
Maxim Pavlenko,
Stein Sæbøe-Larssen,
Timothy Fong,
Norman Maitland,
Pavel Pisa,
期刊:
Journal of Immunotherapy
(OVID Available online 2002)
卷期:
Volume 25,
issue 6
页码: 445-454
ISSN:1524-9557
年代: 2002
出版商: OVID
关键词: Dendritic cells;Electroporation;Gene transfer;Transfection efficiency
数据来源: OVID
摘要:
Among the many promising cancer immunotherapeutic strategies, dendritic cells (DC) have become of particular interest. This study aims to optimize a clinical grade protocol for culture and transfection of human DC. Monocytes and CD34+hematopoietic stem cells (HSC) from same donor were differentiated under serum-free conditions and analyzed for their susceptibility to several recently described nonviral transfection methods as compared with established virally mediated gene transfer. Nonviral gene transfer methods studied were square-wave electroporation, lipofection, and particle-mediated transfer of plasmid DNA or in vitro transcribed mRNA. We conclude that DNA is not suitable for transduction of DC using nonviral methods. In contrast, mRNA and square-wave electroporation reproducibly yields 60% and 50% transfected monocyte- and CD34+-derived DC, respectively, measured at protein level, without affecting the cell viability. Thus, the transfection efficiency of this method is comparable with the 40–90% transgene expression obtained using retroviral (RV) or adenoviral (AdV) vectors in CD34+- and monocyte-derived DC, respectively. In monocyte-derived DC, however, the amount of protein expressed per-cell basis was higher after AdV (MOI = 1000) compared with mRNA electroporation-mediated transfer. This is the first study directly demonstrating side-by-side that mRNA electroporation into DC of different origin indeed results in a comparable number of transduced cells as when using virus-mediated gene transfer.
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