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|標題:||Sustained baculovirus-mediated expression in myogenic cells||作者:||Shen, H.C.
|關鍵字:||baculovirus;C2C12;gene transfer;long-term;sustained transgene;expression;long-term expression;gene delivery vector;mammalian-cells;adenoassociated virus;transgene expression;in-vivo;recombinant;baculovirus;systemic delivery;human myoblasts;stem-cells||Project:||Journal of Gene Medicine||期刊/報告no：:||Journal of Gene Medicine, Volume 10, Issue 11, Page(s) 1190-1197.||摘要:||
Background Baculovirus has emerged as a promising gene delivery vector due to its low cytotoxicity and nonreplication nature in mammalian cells. However, baculovirus-mediated expression is transient and generally lasts less than 14 days, which could restrict its application in the treatment of diseases requiring stable transgene expression. Methods We transduced myoblast cell lines C2C12, Sol 8 and primary myoblasts with a baculovirus expressing the enhanced green fluorescent protein (EGFP) under the control of cytomegalovirus immediate-early prornoter and measured the transduction efficiency by flow cytometry. Myogenic differentiation was induced after transduction and the longevity of EGFP expression Was monitored by fluorescence microscopy. The myogenic differentiation was confirmed by reverse transcription-polymerase chain (RT-PCR). The persistence of the egfp DNA and transcripts was reaction (RT-PCR). The persistence of the egfp DNA and transcripts was monitored by real-time PCR and quantitative real-time RT-PCR. Results Baculovirus efficiently transduced C2C12, Sol 8 and the primary myoblasts. The transgene expression persisted for a prolonged period of time (at least 63 days) in the cells differentiating into myotubes, but was transient in HeLa cells (<7 days). The sustained expression paralleled the myogenic differentiation and stemmed from the intracellular persistence of egfp DNA and mRNA. Conclusions The transgene delivered by baculovirus persists in the myotubes and endows sustained expression, which is distinct from its rapid degradation and transient expression in other cell types. These findings justify the future use of baculovirus for muscle-based gene therapy. Copyright (C) 2008 John Wiley & Sons, Ltd.
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