Background: The fate of transplanted chondrocytes used to elicit the repair of osteochondral defects is unknown. The objective of this study was to examine the fate and the expression of cartilage-specific genes in chondrocytes when the chondrocyte phenotype was maintained preoperatively by alginate suspension culture, the cells were labeled with enhanced green fluorescent protein, and the chondrocytes in alginate were then implanted into full-thickness osteochondral defects in rabbits.

Methods: To determine the effect of alginate on rabbit chondrocytes in vitro, cells were grown in monolayer or in alginate suspension culture, and gene expression for aggrecan, type-I collagen, and type-II collagen was analyzed by reverse transcription-polymerase chain reaction. Cells were genetically labeled with the gene for enhanced green fluorescent protein, and the effect of transfer of the gene for enhanced green fluorescent protein on chondrocyte phenotype was assessed in vitro. Chondrocytes labeled with enhanced green fluorescent protein that were embedded in alginate were implanted into osteochondral defects in rabbit knees, either immediately after creation of the defects or after the cells had been preconditioned in alginate suspension culture for two weeks. The repair tissue within the osteochondral defects was assessed at one to four weeks. Cells labeled with enhanced green fluorescent protein were quantified by confocal microscopy, and the repair tissue was examined histologically with safranin O.

Results: Gene expression by chondrocytes demonstrated a selective upregulation of cartilage-specific genes in alginate suspension culture. This effect was less pronounced in cells that were transduced with enhanced green fluorescent protein. Chondrocytes transplanted in vivo were detected in the repair tissue for the entire period of observation with diminishing cell density over time. At one week, the cell density of the transplanted chondrocytes was 100% of the initial density; at two and three weeks, the cell density was 70%; and, after four weeks, the cell density had decreased to 15%. Safranin-O staining of histological sections indicated cartilage-specific matrix production in vitro and in vivo. Integration of transplanted cells into the host repair tissue was not observed. The two-week period of preconditioning in alginate suspension culture had no apparent influence on the temporal fate of the cells or the histological appearance of the repair tissue.

Conclusions and Clinical Relevance: Alginate promotes expression of cartilage-specific genes and allows delivery of chondrocytes into osteochondral defects. Transgenic chondrocytes labeled with enhanced green fluorescent protein are detectable in the defect, but they do not appear to form repair tissue and they decrease in number with time. In view of the clinical application of cell-based cartilage repair, understanding the fate of transplanted cells becomes increasingly relevant. Transgenic chondrocytes are an effective tool to study the role of transplanted chondrocytes in articular cartilage repair.