Omar D Subedar1,2, Loraine L Y Chiu1,2, Stephen D Waldman1,2. 1. Department of Chemical Engineering, Ryerson University, Toronto, Ontario, Canada. 2. Li Ka Shing Knowledge Institute, St. Michael's Hospital, Toronto, Ontario, Canada.
Abstract
OBJECTIVE: Although tissue engineering is a promising option for articular cartilage repair, it has been challenging to generate functional cartilaginous tissue. While the synthetic response of chondrocytes can be influenced by various means, most approaches treat chondrocytes as a homogeneous population that would respond similarly. However, isolated cells heterogeneously progress through the cell cycle, which can affect macromolecular biosynthesis. As it is possible to synchronize cells within discrete cell cycle phases, the purpose of this study was to investigate the effects of cell cycle synchronization on the chondrogenic potential of primary articular chondrocytes. DESIGN: Different methods of cell synchronization (serum starvation, thymidine, nocodazole, aphidicolin, and RO-3306) were tested for their ability to synchronize primary articular chondrocytes during the process of cell isolation. Cells (unsynchronized and synchronized) were then encapsulated in alginate gels, cultured for 4 weeks, and analyzed for their structural and biochemical properties. RESULTS: The double-thymidine method yielded the highest level of cell purity, with cells synchronized in S phase. While the cells started to lose synchronization after 24 hours, tissue constructs developed from initially S phase synchronized cells had significantly higher glycosaminoglycan and collagen II amounts than those developed using unsynchronized cells. CONCLUSIONS: Initial synchronization led to long-term changes in cartilaginous tissue formation. This effect was postulated to be due to the rapid auto-induction of TGF-βs by actively dividing S phase cells, thereby stimulating chondrogenesis. Cell synchronization methods may also be applied in conjunction with redifferentiation methods to improve the chondrogenic potential of dedifferentiated or diseased chondrocytes.
OBJECTIVE: Although tissue engineering is a promising option for articular cartilage repair, it has been challenging to generate functional cartilaginous tissue. While the synthetic response of chondrocytes can be influenced by various means, most approaches treat chondrocytes as a homogeneous population that would respond similarly. However, isolated cells heterogeneously progress through the cell cycle, which can affect macromolecular biosynthesis. As it is possible to synchronize cells within discrete cell cycle phases, the purpose of this study was to investigate the effects of cell cycle synchronization on the chondrogenic potential of primary articular chondrocytes. DESIGN: Different methods of cell synchronization (serum starvation, thymidine, nocodazole, aphidicolin, and RO-3306) were tested for their ability to synchronize primary articular chondrocytes during the process of cell isolation. Cells (unsynchronized and synchronized) were then encapsulated in alginate gels, cultured for 4 weeks, and analyzed for their structural and biochemical properties. RESULTS: The double-thymidine method yielded the highest level of cell purity, with cells synchronized in S phase. While the cells started to lose synchronization after 24 hours, tissue constructs developed from initially S phase synchronized cells had significantly higher glycosaminoglycan and collagen II amounts than those developed using unsynchronized cells. CONCLUSIONS: Initial synchronization led to long-term changes in cartilaginous tissue formation. This effect was postulated to be due to the rapid auto-induction of TGF-βs by actively dividing S phase cells, thereby stimulating chondrogenesis. Cell synchronization methods may also be applied in conjunction with redifferentiation methods to improve the chondrogenic potential of dedifferentiated or diseased chondrocytes.
Authors: Vanessa J Bianchi; Joanna F Weber; Stephen D Waldman; David Backstein; Rita A Kandel Journal: Tissue Eng Part A Date: 2016-12-19 Impact factor: 3.845