| Summary: | 博士 === 國立清華大學 === 化學工程學系 === 98 === Baculovirus/insect cell has been used as protein expression system expressing the protein which needs post-translational modifications. In recent years, baculovirus was reported to be capable of transducing mammalian cells and expressing the foreign genes, implying the potential of baculovirus as a gene delivery vector. In this study, we explored the feasibility that baculovirus as a gene delivery vector transduced into chondrocytes cultured with rotating-shaft bioreactor (RSB) could stimulate cartilage formation.
We seeded the baculovirus-transduced chondrocytes into PLGA scaffold and cultured the chondrocytes/scaffold in the RSB for examining the effects of baculovirus transduction on engineered cartilage growth. Also, we constructed a recombinant baculovirus encoding BMP-2 (Bac-CB) to transduce chondrocytes, thus inducing chondrocytes to produce BMP-2 and stimulating themselves to form cartilage. Finally, we combined RSB and Bac-CB-transduced chondrocytes to form engineered cartilage with different qualities (according to cultured time) and implanted the engineered cartilage in the full-thickness defect of New Zealand rabbit’s knee to explore which quality in the engineered cartilage can repair the full-thickness defect well.
The results showed that baculovirus-transduced and mock-transduced chondrocytes both grew with high viability (90%) and form cartilage similar to native cartilage, indicating that baculovirus transduction did not hinder cartilage formation. Also, the Bac-CB transduced chondrocytes expressed high level of BMP-2, stimulating chondrocytes to secrete large amount of extracellular matrix (ECM) and form engineered cartilage. Moreover, the engineered cartilage with different quality produced by RSB showed different repair. The in vitro culture time dictates the construct properties, cell differentiation status, and subsequent in vivo repair, and constructs derived from 3-week culture result in the best repair. Since baculovirus is generally regarded as a safe vector and no adverse effects were observed in this study, and RSB provides a defined dynamic environment for the maturation of cartilaginous constructs, the combination of baculovirus-mediated gene transfer and RSB culture represents a promising, cost-effective method to produce tissue engineered cartilages for in vivo cartilage repair.
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