Transplantation of Fetal Instead of Adult Fibroblasts Reduces the Probability of Ectopic Ossification During Tendon Repair

Although cell transplantation therapy can effectively promote functional tendon repair, occasional ectopic ossification during tendon regeneration undermines its efficacy. The effect of transplanted cell types on ectopic ossification has not yet been systematically evaluated. This study compared the rate of ectopic ossification during tendon repair upon transplantation with mouse fetal fibroblasts (FFs) and their adult counterparts (adult fibroblasts [AFs]). Alkaline phosphatase (ALP) staining, immunofluorescence, and gene expression analysis were used to compare the spontaneous osteogenic differentiation of FFs and AFs in vitro. X-ray, histology, and gene expression analysis were used to investigate the ectopic ossification in a mouse Achilles tendon repair model in vivo. ALP staining and immunofluorescence data in vitro showed that FFs had less spontaneous osteogenic differentiation capacity, and lower expression of runt-related transcription factor 2 (runx2). For the in vivo study, the FFs transplant group displayed reduced ectopic ossification (2/7 vs. 7/7, Mann-Whitney test p < 0.01) at 14 weeks post-transplantation and enhanced tendon repair (general histological score at week 6, 7.53 vs. 10.56, p < 0.05). More chondrocytes formed at 6 weeks, and all mice developed bone marrow at 14 weeks post-transplantation in the AFs transplant group. Gene expression analysis of the regenerated tissue showed significantly higher expression levels of transforming growth factor beta1 (TGF-beta 1) and transforming growth factor beta3 (TGF-beta 3) in the AFs group during the early stages of tendon repair. Our study demonstrates that transplantation of fetal instead of AFs is more promising for tendon repair, underscoring the importance of the origin of seed cells for tendon repair.