[In vivo osteogenic functions of injectable biological bone cement]

To evaluate the osteogenic functions of injectable biological bone cement during repairing inclusive bone defects in rabbit femoral condyles.

Encapsulated rhBMP-2 gene modified rabbit bone mesenchymal cells (BMSCs) were seeded into calcium phosphate cement (CPC) to prepare injectable biological bone cement. The live/dead cell ratio was detected by calcein-AM/ethidium homodimer staining and cement microstructure examined by electron microscope. The anti-compression strength of CPC were tested. Inclusive bone defects were created in the bilateral femoral condyles of 6 rabbits. The biological CPC was implanted randomly at one side and pure CPC at the other side. The radiological films were taken immediately, at Weeks 6 & 12 post-operation. The animals were sacrificed at Week 12 post-operation and both femoral condyles were retrieved to prepare decalcified slides. The morphometry parameters of bone tissue, such as cement area (CA), calcified trabecular area (TBA), osteoblast index (OBI) and osteoclast (OCI) were measured. Immunohistochemical staining was performed to detect the expressions of BMP (bone morphogenetic protein)-2, TGF (transforming growth factor)-β1 and VEGF (vascular endothelial growth factor).

The calcein-AM/ethidium homodimer staining showed that the live rate of encapsulated cells was over 90%. It was found under electron microscope that there was the formation of hydroxyapatite crystals and the presence of connective microporous structure. The anti-compression strength of biological cement was (20.19 ± 1.75) MPa. And it was much less than that of pure CPC. Radiological study showed that both types of CPC could fill bone defects completely without gap. The biological CPC degraded a little faster. As demonstrated by decalcified slides, the cements were absorbed into numerous small blocks around newborn trabeculae without insertion of fibrous tissues. CA decreased to 9.68% in biological CPC group and it was much less than 17.47% for pure CPC group. TBA was 58.75% in biological CPC group and it was much greater than 34.34% for pure CPC group. There was no significant difference in OBT and OCT between two groups. Immunohistochemical staining showed there were a large number of oval-shaped nuclear mast cells in biological CPC group. They were strongly positive for the expression of BMP-2. A moderate number of cells had a positive expression of TGF-β1 and VEGF. More positive cells were present in biological CPC group.

The introduction of BMP-2 into CPC renders CPC capable of bone induction. Thus CPC induce both the proliferation of BMSCs and the expressions of TGF-β1 and VEGF. The synergistic effects of these three factors accelerate the degradation of CPC.