Passaged Chondrocytes: a source of cells for cartilage tissue engineering

The interest in bioengineered cartilage for the replacement of damaged articular cartilage has gained momentum in recent years. One of the issues limiting clinical translation has been identifying an appropriate cell source. To address this we identified that hyaline cartilage can be formed through the co-culture of a small number of primary articular chondrocytes with twice passaged chondrocytes. Not only were passaged human cells able to redifferentiate, but after separation from the primary cells, maintained their differentiation and formed cartilagenous tissue. This tissue was rich in collagen type II and aggrecan, the major molecules present in hyaline cartilage. There was little collagen type I. Tissue characterization studies demonstrated that redifferentiation was occurring during the first week of culture. Co-culture of primary and passaged cells in side-by-side culture indicated that a soluble factor was responsible for redifferentiation and that direct cell-cell contact was not essential. A serum-free culture system was developed and tissue characterized to confirm that the cells maintained phenotype, in order to be able to examine the secretome. Protein analysis through mass spectrometry and immunohistochemical staining of tissue sections indicated that versican, aggrecan, and types II and XII collagens were accumulated early within the first 3 days, by passaged chondrocytes were redifferentiating and formed cartilage tissue in vitro. This suggested that these molecules may be necessary to provide a microenvironment that supports hyaline cartilage formation. Further study is required to determine if these molecules are also accumulated by passaged human chondrocytes and their role in promoting hyaline cartilage formation. In addition these molecules also may then serve as markers that can be used to identify culture conditions and/or scaffolds that will favour articular cartilage formation by passaged chondrocytes and facilitate rapid screening of culture conditions for chondrogenic inducing potential. Together these studies facilitate the clinical transition of regenerative medicine approaches to cartilage repair.