Bone formation on apatite-coated titanium incorporated with bone morphogenetic protein and heparin

Bone morphogenetic proteins (BMPs) strongly induce osteogenesis and are enhanced by heparin. In this study, a potent osteoinductive material was developed by coating the surface of titanium with apatite incorporated with BMP and heparin.

Titanium samples were treated with a 5 N NaOH solution and heated at 600 degrees C for 24 hours. The treated titanium was soaked in simulated body fluid (SBF) for 4 days and additionally soaked in SBF containing recombinant human BMP-2 (rhBMP-2; 1,000 ng/mL) with or without heparin (30 microg/mL) for 3 days at 37 degrees C. The surfaces of each sample were examined with scanning electron microscopy. The presence of rhBMP-2 on the surface of the apatite-coated titanium was examined using the immunogold method. MC3T3-E1 osteoblast-like cells were cultured on the surface of each sample. The number and morphology of the adherent cells, alkaline phosphatase activity, and osteocalcin mRNA expression were examined.

Apatite was formed on the surface of alkaline heat-treated titanium after soaking in SBF for 7 days. The presence of rhBMP-2 was confirmed by the distribution of BMP-positive immunogold particles. The incorporation of (> or = 3 microg/mL) heparin significantly increased alkaline phosphatase activity and osteocalcin mRNA expression in the cells on apatite-coated titanium containing rhBMP-2.

These findings demonstrate that heparin enhanced BMP-2-induced osteogenesis on apatite-coated titanium without the loss of BMP-2 activity. The combination of BMP-2 and heparin was effective even in the thin apatite layer formed on titanium using the alkaline heat treatment method.