Baculovirus transduction of rat articular chondrocytes: roles of cell cycle.
BACKGROUND
We have previously demonstrated highly efficient baculovirus transduction of primary rat articular chondrocytes, thus implicating the possible applications of baculovirus in gene-based cartilage tissue engineering. However, baculovirus-mediated gene expression in the chondrocytes is transient.


METHODS
In this study, we attempted to prolong the expression by supertransduction, but uncovered that after long-term culture the chondrocytes became more refractory to baculovirus transduction. Therefore, the correlation between baculovirus-mediated enhanced green fluorescent protein (EGFP) expression and cell cycle was investigated by comparing the cycling chondrocytes and chondrocytes rich in quiescent cells, in terms of EGFP expression, virus uptake, cell cycle distribution, nuclear import and methylation of viral DNA.


RESULTS
We demonstrated, for the first time, that baculovirus-mediated transduction of chondrocytes is correlated with the cell cycle. The chondrocytes predominantly in G2/M phase were approximately twice as efficient in EGFP expression as the cycling cells, while the cells in S and G1 phases expressed EGFP as efficiently as the cycling cells. Notably, the chondrocyte populations rich in quiescent cells resulted in efficient virus uptake, but less effective nuclear transport of baculoviral DNA and higher degree of methylation, and hence poorer transgene expression.


CONCLUSIONS
These findings unravel the practical limitations when employing baculovirus in cartilage tissue engineering. The implications and possible solutions are discussed.
Lee
Hsiao-Ping
HP
Department of Chemical Engineering, National Tsing Hua University, Hsinchu, Taiwan 300.
Chen
Yen-Lin
YL
Shen
Heng-Chun
HC
Lo
Wen-Hsin
WH
Hu
Yu-Chen
YC
D021581:Active Transport, Cell Nucleus; D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D002358:Cartilage, Articular / Q000166:cytology*; D002453:Cell Cycle; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000502:physiology*; D019175:DNA Methylation; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D023822:Tissue Engineering; D014161:Transduction, Genetic; D019076:Transgenes
D049452:Green Fluorescent Proteins