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We report here the use of novel self-assembling collagen-hyaluronic acid (HyA) membranes to deliver bone morphogenetic protein-2 (BMP-2) for orthopedic applications. Prior work has demonstrated that collagen-HyA membranes are formed initially through electrostatic interactions between the oppositely charged collagen and HyA molecules, and that membrane growth is driven by osmotic pressure imbalances between the collagen and HyA solutions. The purpose of this study was to investigate the potential of incorporating charged growth factors such as BMP-2 within the membrane for regenerative medicine applications. Membrane material properties, protein mass loss, and release kinetics of BMP-2, as well as biocompatibility and osteogenic potential in vitro and in vivo using a subcutaneous mouse model were assessed. Scanning electron microscopy and mechanical testing confirmed no loss of structural or mechanical integrity upon BMP-2 incorporation into the membranes. Slow and steady release of the growth factor was demonstrated with 17% of total loaded BMP-2 released over the course of 49 days. To test biocompatibility and osteogenic potential in vitro, human mesenchymal stem cells were cultured on collagen-HyA membranes and showed greater proliferation rates (for up to 28 days) on membranes without BMP-2, but a greater alkaline phosphatase activity and osteocalcin production on membranes releasing BMP-2. In vivo subcutaneous implantation of the membranes showed a minimal immune response with osteoblasts and mineral deposits present in the ectopic site for BMP-2-releasing membranes, further demonstrating the potential of the BMP-2-releasing membranes to induce osteogenic differentiation. This study presents a novel strategy to create self-assembled membranes using two biocompatible molecules that can deliver bioactive agents in a sustained manner to induce a local regenerative response.

Osteogenic Potential of BMP-2-Releasing Self-Assembled Membranes