Granulocyte colony-stimulating factor (G-CSF) was shown to promote bone regeneration and mobilization of vascular and osteogenic progenitor cells. In this study, we investigated the effects of a systemic low dose of G-CSF on both bone consolidation and mobilization of hematopoietic stem/progenitor cells (HSPCs), endothelial progenitor cells (EPCs) and mesenchymal stromal cells (MSCs) in a rat model of distraction osteogenesis (DO). Neovascularization and mineralization were longitudinally monitored using positron emission tomography and planar scintigraphy. Histological analysis was performed and the number of circulating HSPCs, EPCs and MSCs was studied by flow cytometry. Contrary to control group, in the early phase of consolidation, a bony bridge with lower osteoclast activity and a trend of an increase in osteoblast activity were observed in the distracted callus in the G-CSF group, whereas, at the late phase of consolidation, a significantly lower neovascularization was observed. While no difference was observed in the number of circulating EPCs between control and G-CSF groups, the number of MSCs was significantly lower at the end of the latency phase and that of HSPCs was significantly higher 4 days after the bone lengthening. Our results indicate that G-CSF accelerates bone regeneration and modulates mobilization of progenitor cells during DO.

This protocol describes the use of a newly developed external fixator for distraction osteogenesis in a rat femoral model. Distraction osteogenesis (DO) is a surgical technique leading to bone regeneration after an osteotomy. The osteotomized extremities are moved away from each other by gradual distraction to reach the desired elongation This procedure is widely used in humans for lower and upper limb lengthening, treatment after a bone nonunion, or the regeneration of a bone defect following surgery for bone tumor excision, as well as in maxillofacial reconstruction. Only a few studies clearly demonstrate the efficiency of their protocol in obtaining a functional regenerated bone, i.e., bone that will support physiological weight-bearing without fracture after removal of the external fixator. Moreover, protocols for DO vary and reproducibility is limited by lack of information, making comparison between studies difficult. The aim of this study was to develop a reproducible protocol comprising an appropriate external fixator design for rat limb lengthening, with a detailed surgical technique that permits physiological weight-bearing by the animal after removal of the external fixator.

Bone marrow contains skeletal progenitor cells (mesenchymal stem cells) which are important actors in skeletal repair. Nevertheless, the therapeutic use of mesenchymal stem cells for articular cartilage repair remains a challenge with a limited efficacy on both clinical outcomes and cartilage regeneration. In rabbit periosteal graft models, we show that such skeletal progenitor cells are recruited and undergo differentiation through different skeletal tissue pathways (bone, cartilage, skeletal muscle) in bone marrow areas that are under periosteal graft influence. Moreover, histological observations show among cartilage differentiated cells two different cartilage pathways, corresponding respectively to articular cartilage formations and to endochondral cartilage formations. This underline the presence in bone marrow of cartilage progenitor cells that escape from the intrinsic endochondral differentiation program resulting in transient rather than permanent cartilage, and follow a specific articular cartilage differentiation pathway. With respect to tissue engineering, and while extremely complex techniques are being developed to artificially generate functionally integrated, stratified articular cartilage like structures, our observations urge the development of protocols that select the proper population of articular cartilage progenitor cells before its use as cartilage building units. Articular cartilage repair; Tissue engineering; Skeletal progenitors; Mesenchymal stem cells; Articular cartilage pathway; Endochondral cartilage pathway; Periosteum