Chondrogenically differentiated mesenchymal stromal cell pellets stimulate endochondral bone regeneration in critical-sized bone defects

Abstract

Grafting bone defects or atrophic non-unions with mesenchymal stromal cells (MSCs)-based grafts is not yet successful. MSC-based grafts typically use undifferentiated or osteogenically differentiated MSCs and regenerate bone through intramembranous ossification. Endochondral ossification might be more potent but requires chondrogenic differentiation of MSCs. Here, we determined if chondrogenically differentiated MSC (ch-MSC) pellets could induce bone regeneration in an orthotopic environment through endochondral ossification. Undifferentiated MSC pellets (ud-MSC) and ch-MSC pellets were generated from MSCs of human donors cultured on chondrogenic medium for respectively 3 (ud-MSC) and 21 (ch-MSC) days. A 6mm femoral bone defect was made and stabilised with an internal plate in 27 athymic rats. Defects were left empty for 6 weeks to develop an atrophic non-union before they were grafted with ch-MSC pellets or ud-MSC pellets. Micro-CT scans made 4 and 8 weeks after grafting showed that ch-MSC pellets resulted in significantly more bone than ud-MSC pellets. This regenerated bone could completely bridge the defect, but the amount of bone regeneration was donor-dependent. Histology after 7 and 14 days showed slowly mineralising pellets containing hypertrophic chondrocytes, as well as TRAP-positive and CD34-positive cells around the ch-MSC pellets, indicating osteoclastic resorption and vascularisation typical for endochondral ossification. In conclusion, grafting critical femoral bone defects with chondrogenically differentiated MSC pellets led to rapid and pronounced bone regeneration through endochondral ossification and may therefore be a more successful MSC-based graft to repair large bone defects or atrophic non-unions. But, since bone regeneration was donor-depend, the generation of potent chondrogenically differentiated MSC pellets for each single donor needs to be established first.