Abstract
Autologous Chondrocytes Implantation (ACI) has been applied to cartilage repair extensively. According to this technique, one obstacle is that it is hard to take sufficient chondrocytes from individual patient, so chondrocytes must be expanded in vitro until sufficient cells are obtained. Currently, expansion using monolayer culture is widely used. However, it is well known that in vitro multiplication of chondrocytes in monolayer results in dedifferentiation of these cells. They lose their phenotypes and become fibroblast-like cartilage in place of hyaline-like cartilage. To expand sufficient chondrocytes and retain their phenotypes, in this study we use porous three-dimensional alginate scaffolds within perfusion system to culture primary porcine chondrocytes at three cell densities as 5□105 (low), 5□106 (intermediate), and 2□107 (high) cells seeded into 40 alginate scaffolds for 4 weeks to study their proliferation and differentiation compared to monolayer culture. The results show that most cells in the former groups could retain their phenotypes and biochemical properties. Chondrocytes at the low cell density group proliferated more than the other two groups and reached about 150-fold. In contrast chondrocytes in the intermediate and the high cell density groups showed increased capability to produce extracellular matrix such as collagen type II or glycosaminoglycans and cartilage-like tissue were produced from these two groups. Otherwise the large cartilage-like tissue from the low cell density group could be observed when proliferating cells were injected into biphasic scaffolds and cultured in vivo by using nude mice as an animal model for 4 weeks. Thus this system of the low cell density may be a useful tool for chondrocytes expansion and may have the potential for Autologous Chondrocytes Implantation (ACI) research.