Tissue engineered scaffold for lamellar bone regeneration
Critical sized bone defects (CSD) account for 82% of all extremities injuries from blast injuries, and are commonly repaired using autologous bone grafts. However, many problems are associated with this technique, including infections and donor site morbidity. Bone tissue engineers have developed several approaches to promote bony in-growth, yet mechanical stability remains an inadequacy of these implants even though it is crucial to the success of the implant. With the goal to investigate osteon modeling through collagen organization, calcium phosphate (CaP) scaffolds (Hydroxyapatite---HAp, beta-Tricalcium Phosphate---beta-TCP, biphasic CaP---BCaP) were developed. First cross sectional microchannels mimicking osteonal structure were created in hydroxyapatite disks with diameters ranging from 50 to 500mum. The change in extra cellular matrix (ECM) secretion and cell attachment/orientation was investigated as a function of microchannel diameter. A 3D scaffold was then built with longitudinal microchannels spanning the entire length of the construct resembling naturally occurring osteons in cortical bone. The scaffold was evaluated for the ability to support osteoblast precursor proliferation, differentiation and ECM secretion within the 3D osteonal microchannels. Results showed a difference in the time required for cells to create active orientation within the microchannels as a function of channel diameter, as well as a specific shift in orientation with time for the entire cell layer. Overall in this study it was demonstrated it is possible to replicate osteoid secretion in vitro. Moreover, we determined that the BCaP surface is favored over the HAp because of both the higher degradation profile and also because it may increase the availability of calcium to osteoblasts during osteoid formation for mineralization, which in the long run improves the hardness of the ECM. In addition to affecting the quality of the ECM secreted, material properties were not found to have any other effects on osteoid secretion. However, it was determined that multiple cell seedings, as well as time, have a significant role in increasing the size of the osteoid secretion close to the surface. These observations indicate that the microchannel architecture demonstrates both the necessary biomechanical properties as well as the appropriate cell response to serve as a suitable load bearing graft substitute in pre-clinical testing. This observed mechanism can be translated into functional load bearing collagen-I matrices that could withstand physiological loads in clinical reconstructions. It was determined that substrate curvature affects cell orientation, the time required for initial response, and shift in orientation with time. These findings not only demonstrate that bone cells actively respond to curved substrate attachment, but also prove the ability to fabricate a scaffold that recreates the architecture of cortical bone and that can be used to repair bone with lamellar, mechanically competent organization.