BMP depletion occurs during prolonged acid demineralization of bone: characterization and implications for graft preparation

Abstract

Demineralization of allograft bone increases the bioavailability of matrix-associated bone morphogenetic proteins (BMPs), rendering these grafts osteoinductive. While osteoinductivity is related to BMP content, little is known about how the demineralization protocol, in particular, extended demineralization times, affects graft BMP levels. We characterized the BMP-7 content of <710 μm bovine bone powder demineralized under various conditions. Using 1 g of bone per 50 ml of 0.125 N, 0.25 N, or 0.5 N HCl, demineralization was performed at room temperature for 5 min to 24 h. Minimum residual calcium levels were obtained within 90 min and were <1 wt % using the 0.25 N and 0.5 N baths and 17 wt % using the 0.125 N bath. Measured peak BMP-7 levels were also obtained within 90 min and were 161-165 ng g(-1) using the 0.25 N and 0.5 N baths and 55.2 ng g(-1) using the 0.125 N bath. This compares to 5.1 ng g(-1) for undemineralized bone. Further acid bath exposure to 24 h resulted in BMP-7 decline to about 50% of the peak value, which was significant. The BMP-7 half-life was estimated to be 26 h. It is likely that the decline was due to diffusion of BMP-7 from the bone matrix into the acid. These results suggest the importance of not over demineralizing bone grafts and should stimulate further research that can be incorporated into the processing methodology followed by tissue banks.

Characterization of Bone and Bone-Based Graft Materials Using FTIR Spectroscopy

“…As reported in a recent study about BMPs depletion in particles of bovine cortical bone under acid exposure (0.25 and 0.5 M HCl) (Pietrzak et al, 2009), the availability of BMP-7 increases as demineralization occurs but, after reaching a maximum in the extraction bath, continuously declines. These results alert for the need to control the demineralization processing conditions.…”

“…In fact, as mineral is removed, the matrix associated BMPs become available rendering DBM grafts osteoinductive (Pietrzak et al, 2009). These grafts can be used either alone (Libin et al, 1975;Morone & Boden, 1998;Pietrzak et al, 2005) or in combination with bone marrow, autogenous bone graft, or other materials (Kim et al, 2002;Kucukkolbasi et al, 2009;Nade & Burwell, 1977).…”

In vitro effect of mineralized and demineralized bone allografts on proliferation and differentiation of MG-63 osteoblast-like cells

“…These observations may be explained by the possibility that differences in type, concentration and release rate of water-soluble osteoinductive factors released from mineralized and demineralized allografts (irrespective of physical contact effect and insoluble osteogenic factors) may affect their biological behavior and osteoinductive properties. This means that the process of demineralization not only decreases the calcium content, but also may damage some important soluble osteoinductive proteins such as BMPs (Zhang et al 1997;Pietrzak et al 2009). In addition to BMPs, presence and solubility of minerals such as calcium/phosphate ions are necessary to achieve osteoinductive properties (Habibovic et al 2007).…”

Influence of hydrochloric acid concentration on the demineralization of cortical bone

“…In the case of using HCl (the most frequently used acid), the major inorganic constituent of bone (hydroxyapatite) reacts to form monocalcium phosphate and calcium chloride (Horneman et al, 2004;Dorozhkin, 1997). Bone samples are usually demineralized in 0.5-0.6 M HCl in order to remove bone mineral efficiently while leaving the graft osteoinductive (Pietrzak et al, 2009). However, and as mentioned before, sometimes the objective of bone demineralization is just to provide a flexible material regardless of its osteoinductive potential, or an "inert" matrix (containing no significant amount of signal molecule content) to be used in some clinical and research applications.…”