Influence of rhBMP-2 on Rat Bone Marrow Stromal Cells Cultured on Titanium Fiber Mesh

Abstract

Titanium (Ti) fiber mesh is a candidate scaffold material for the creation of bone graft substitutes (BGS). Two densities (3.54 × 10⁴ cells/cm² [LD or low density] and 3.54 × 10⁵ cells/cm² [HD or high density]) of rat bone marrow stromal cells were seeded on Ti-fiber mesh discs. Cells were cultured for up to 16 days, 7 days of which the cells were in the presence of various concentrations of rhBMP-2 (0, 10, 100, and 1,000 ng/mL) in order to evaluate osteogenic expression. Scanning electron microscopy (SEM), light microscopy (LM), energy dispersive spectroscopy (EDS), DNA and calcium (Ca) content measurements, and x-ray diffraction (XRD) analysis were performed. SEM and EDS evaluation showed that a confluent layer of cells was present on top of the meshes together with collagen bundles and calcified globular accretions. Light microscopical evaluation showed a densely stained layer in the upper part of the mesh. SEM and Ca content measurement showed that calcification starts at 8 days. In addition, it was demonstrated that DNA content peaked at 8 days. LM, SEM, and Ca content evaluation revealed positive effects of increasing the cell seeding density, the rhBMP-2 concentration and the culture time on mineralization. Increasing the cell seeding density also showed a positive effect on DNA content. No effects of rhBMP-2 concentration were seen on DNA content. Finally, XRD revealed that the deposited matrix contained a precipitate of a stable calcium phosphate phase. We conclude that (1) titanium fiber mesh sustains excellent osteogenic expression in vitro, (2) increasing the cell seeding density has a positive effect on osteogenic expression in titanium mesh in vitro, and (3) in high density specimens, rhBMP-2 concentrations of 100 ng/mL and 1,000 ng/mL stimulate extracellular matrix calcification in a dose-responsive manner.