Transgene expression and differentiation of baculovirus-transduced human mesenchymal stem cells.
BACKGROUND
Mesenchymal stem cells (MSCs) have drawn considerable attention as vehicles for cell- or gene-based therapies, yet various problems still exist for current gene delivery vectors. On the other hand, baculovirus has emerged as a novel gene therapy vector, but its transduction of stem cells has not been reported.


METHODS
A recombinant baculovirus expressing the enhanced green fluorescent protein (EGFP) was constructed to transduce human MSCs derived from umbilical cord blood (uMSCs) or bone marrow (bMSCs).


RESULTS
In this study, we demonstrated for the first time that human uMSCs or bMSCs could be transduced by baculovirus with high efficiencies (up to approximately 72.8% and 41.1%, respectively) and significantly elevated transgene (enhanced green fluorescent protein, EGFP) expression upon incubation with unconcentrated virus and phosphate-buffered saline for 4 h at 25 degrees C. The transduction efficiency into bMSCs could be further increased to approximately 72.2% by lowering the cell density. The improved transgene expression was partly attributed to the enhanced virus uptake upon transduction, as determined by quantitative real-time polymerase chain reaction (Q-PCR). MSC growth was not obstructed by baculovirus transduction itself, but was somewhat hampered by EGFP expression. Nonetheless, the baculovirus-transduced cells remained capable of differentiating into adipogenic lineage. The adipogenic progenitors appeared more permissive to baculovirus transduction than the undifferentiated bMSCs, thus allowing for the maintenance and enhancement of transgene expression by repeated transduction after subculture.


CONCLUSIONS
These findings implicate the potential applications of baculovirus as an alternative vector to genetically modify MSCs for ex vivo gene therapy.
Ho
Yi-Chen
YC
Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300, Taiwan.
Chung
Yao-Chi
YC
Hwang
Shiaw-Min
SM
Wang
Kuei-Chun
KC
Hu
Yu-Chen
YC
D017667:Adipocytes / Q000502:physiology; D016367:Baculoviridae / Q000235:genetics*; D001854:Bone Marrow Cells; D002454:Cell Differentiation; D002478:Cells, Cultured; D005434:Flow Cytometry; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism / Q000821:virology*; D014161:Transduction, Genetic / Q000379:methods*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
C088645:IE1 protein, cytomegalovirus; D017874:Immediate-Early Proteins; D014764:Viral Proteins; D049452:Green Fluorescent Proteins