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Tissue Engineering and Regenerative Medicine International Society Asia Pacific Meeting 2011
Dual Delivery Of TGF-Β3 And Type I Collagen-Targeting Shrna With Viral Vectors For Engineered Articular Chondrogenesis
F Zhang , YC Yao , DA Wang
Background: Despite the advances in therapies for cartilage regeneration, articular cartilage degeneration remains a tough problem worldwide. In cartilage tissue engineering, synovium-derived mesenchymal stem cells have proved to be a promising cell type due to their proneness to chondrogenic differentiation. Transforming growth factor (TGF) β3 is a potent growth factor that can promote chondrogenic differentiation of stem/progenitor cells. On the other hand, type I collagen is inherently expressed in synovium-derived mesenchymal stem cells. The situation is further complicated by the fact that TGF-β3 would elevate the expression of type I collagen in some types of cells. The existence of type I collagen will induce the formation of fibrous cartilage and alter its mechanical strength. Therefore, it is necessary to block the expression of type I collagen, whereas RNA interference (RNAi) has provided a powerful strategy.
Methods: In our studies, we have constructed a series of single functioning and dual-functioning adenoviral or lentiviral vectors that can express TGF-β3 and/or type I collagen-targeting small hairpin RNA (shRNA). As adenoviral vector induces a transient expression whilst lentiviral vector mediates a more sustained expression, the use of these viral vectors would endow differential expression profiles of TGF-β3 and shRNA. Combinatorial use of them was practices in our studies in order to simultaneously deliver both TGF-β3 and type I collagen-targeting shRNA to synovium-derived mesenchymal stem cells to promote type I collagen-suppressed chondrogenesis. Real time PCR and biochemical assays were used to assess the effect of these delivery modes on type I collagen-suppressed chondrogenesis.
Results: Best induction of aggrecan, cartilage oligomeric matrix protein, glycosaminoglycans and total collagen was observed in the combination of TGF-β3-encoding lentiviral vector and shRNA-encoding adenoviral vector, while the combination of TGF-β3-encoding adenoviral vector and shRNA-encoding lentiviral vector witnessed the highest induction of type II collagen. No significant difference was shown between the combinatorial groups regarding type I collagen expression.
Conclusions: Relatively optimal chondrogenesis with type I collagen suppression was witnessed in the combination of TGF-β3-encoding lentiviral vector and shRNA-encoding adenoviral vector during a 30-day 3-dimensional culture. However, more work is to be done to investigate the long-term effect of the combinatorial applications.
Date: 3th August 2011 Time:1200-1215
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