Engineered biomaterial resorption and growth factor delivery for enhanced bone regeneration

Biomaterials used in bone tissue regeneration have shown limited success in the clinic, with lack of integration and vascularization being the greatest hurdles to overcome. During its tenure at Rutgers University, the New Jersey Center for Biomaterials focused on the development of tyrosine- and tyrosol-based polymers, demonstrating the ability to synthesize large libraries of polymers with tunable chemical, thermal, and mechanical properties. While it was demonstrated that a subset of these polymers performed better than the clinically relevant poly(L-lactic acid) (PLLA) in a bone regeneration setting, a significant drawback to these polymers was their lack of resorption. Biomaterial resorption plays a key role in remodeling and resolution of healing in the bone environment. The aromatic contribution of the tyrosine-based monomers gives the polymer its hydrophobicity and rigidity, but it also decreases the solubility of the degradation products significantly, thus reducing resorption. Replacing the tyrosine-based monomers with analogous tyrosol-based monomers improved resorption slightly, though they still fall short of the polymers used clinically. In an effort to address the limited resorption, a protease-sensitive peptide was co-polymerized with a subset of poly(ester arylates) previously developed in the lab. Protease-sensitive peptides have been used in cross-linked hydrophilic networks to promote cell-specific biomaterial resorption, and the work to translate this into a linear, hydrophobic thermoplastic is described in Chapter 3. Protease sensitivity depends on the accessibility and lability of the target bond, and in this work, it is demonstrated that this sensitivity can be tuned by either altering the chemistry of the polymer backbone, or by incorporating a peptide sequence to provides a more specific mode of degradation. To circumvent the laborious degradation studies that are typically required for this type of analysis, quartz crystal microbalance (QCM-D) was used to assess material resorption in the presence of enzymes. This technique provides a high-throughput way of screening polymers for their surface erosion potential. Incorporation of a proteinase K-sensitive peptide sequence improved resorption of the tyrosol-based poly(ester arylate) in the presence of proteinase K, and it was shown that this resorption is enzyme-specific. Another approach to improve integration and vascularization of biomaterials for bone regeneration is to target the cells that are responsible for physiological bone resorption. Osteoclasts are multi-nucleated giant cells that are capable of digesting collagen type I and dissolving the mineral phase of bone tissue. To assess the ability of osteoclasts to attach and activate on a bone-promoting polymer, a tyrosine-derived polycarbonate with a history of use in bone regeneration (E1001(1k)) was used as a model polymer. In Chapter 4, the functionalization of the polymer and the subsequent culture of osteoclasts is described. The E1001(1k) polymer was functionalized with an RGD peptide to promote cell attachment, functionalized with collagen type I to create a physiologically relevant surface, or blended with -tricalcium phosphate. It was hypothesized that these additional functionalities would be required, and would also promote, osteoclast attachment and activity. Morphological characteristics of osteoclasts were observed on all polymeric substrates, including the unmodified polymer, using transmission light microscopy and confocal microscopy. The transparency of the amorphous E1001(1k) polymer allowed for visualization of the actin ring and ruffled border in high resolution. Active osteoclasts are historically imaged using electron microscopy techniques, but these techniques are destructive and limited in the information they can provide. Observation of an active osteoclast on E1001(1k) provides the basis for studying osteoclast-biomaterial interactions in the future as well as the ability to visualize osteoclast resorption of polymers in real time. While polymeric biomaterials have shown promise in this area of research, their translation into the clinic has also been hindered by inconsistencies between in vitro and in vivo performance. The E1001(1k) polymer has been used as a carrier for bone morphogenetic-2 (BMP-2). While BMP-2 has been used widely as a potent inducer of bone regeneration, its activity assessment in vitro has been presented with conflicting methods in the literature. In Chapter 5, the effect of several assay parameters on measured BMP-2 activity is explored. The model cell line chosen has a significant effect on the concentration range of BMP-2 over which alkaline phosphatase activity is linearly correlated. This information was then used to compare the activity of commercially available BMP-2, and it is also demonstrated that there is a direct correlation between measured activity of BMP-2 and its concentration as measured by ELISA. Together, this work has implications in the design and evaluation of biomaterials used as BMP-2 carriers as they are translated from the in vitro setting to in vivo applications.Ph.D.Includes bibliographical reference