Comparison of intra-articular injectable polydactyly-derived chondrocyte sheet-minis with direct transplantation of intact polydactyly-derived chondrocyte sheets in a rat full-thickness cartilage defect model.

Polydactyly-derived chondrocyte (PD) sheets are being developed as an allogeneic cell sheet therapy for cartilage defects associated with knee osteoarthritis. Although intact PD sheets can promote hyaline cartilage repair, transplantation requires invasive open procedures. Injectable fragments of PD sheets (PD sheet-minis) may reduce invasiveness. However, their in vivo repair capacity after intra-articular administration remains unclear. PD cells were isolated from discarded cartilage tissue obtained during polydactyly surgery. PD sheets and PD sheet-minis were fabricated for 14 days using temperature-responsive culture inserts and RepCell plates, respectively. PD cells, PD sheets, and PD sheet-minis were compared by cell counting, viability assays, flow cytometry, soluble factor assays, and quantitative polymerase chain reaction. A full-thickness cartilage defect was created in the femoral trochlea of male athymic nude rats. The animals were randomized by body weight into vehicle, PD cell suspension, PD sheet-mini suspension, or intact PD sheet transplantation groups (n = 6 each). Hindlimb weight-bearing was assessed after surgery, and cartilage repair was evaluated histologically 4 weeks after transplantation. The viable cell numbers administered per knee were comparable between PD cells, PD sheet-minis, and intact PD sheets. The PD sheet-minis and PD sheets showed similar surface marker profiles, whereas PD cells showed significantly lower CD26 and higher CD146, CD49a, and CD166 expression. Compared to PD cells, PD sheet-minis secreted more melanoma inhibitory activity and dickkopf-related protein 1 and less transforming growth factor-β, monocyte chemoattractant protein-1, and matrix metalloproteinase-3. Gene expression analysis revealed lower COL1A1 and RUNX2 expression and higher MMP3 expression in PD sheet-minis than in PD cells; COL2A1 and COL10A1 were not detected. Weight-bearing showed no sustained intergroup difference. Histologically, intact PD sheets produced consistent Safranin O-positive, type II collagen-positive, and human vimentin-positive repair tissue, whereas PD cells and PD sheet-minis mainly produced fibrous repair tissue. The mean Osteoarthritis Research Society International (OARSI) scores were 11.4 ± 5.4, 10.7 ± 6.2, 9.3 ± 7.1, and 4.4 ± 3.2 in the vehicle, PD cell, PD sheet-mini and PD sheet groups, respectively. PD sheet-minis retained several sheet-like in vitro characteristics; however, single intra-articular administration did not reproduce the cartilage repair achieved with intact PD sheets. These findings suggest that lesion-specific retention and persistence, rather than injectability alone, may be important determinants of structural cartilage repair by PD cell-sheet products.