Decellularisation processes for the intervertebral disc

Intervertebral disc (IVD) degeneration is a major cause of back pain, particularly in the lumbar region. Current surgical interventions, such as spinal fusion and total disc replacements, have limitations. An alternative solution could be to replace the degenerated IVD with a decellularised IVD from a natural source. Removal of the cellular components from natural IVDs should render them non immunogenic upon implantation into the host. The aim of this study was to develop a decellularisation protocol using large animal IVDs with vertebral bone attachments and to translate the protocol to human tissue. Seven different protocols were investigated based on hypotonic low concentration sodium dodecyl sulphate with proteinase inhibitors, freeze/thaw cycles and nuclease treatments. It was found that reduction of the bony tissue in bovine samples and the incorporation of sonication increased protocol efficacy. Cells and DNA were removed to acceptable levels and the glycosaminoglycans (GAGs), important for IVD compressive function, were largely retained. Cyclic compression testing showed sufficient sensitivity to detect a significant increase in stiffness of the decellularised tissue, when compared to the native state. Encouragingly, this change in biomechanical properties was within natural tissue variation. The protocol was then applied to human thoracic IVDs. Results showed that total DNA levels in all the decellularised tissue regions investigated were below 50 ng.mg-1 dry tissue weight and the tissue retained high levels of GAGs. Application of the developed decellularisation protocol has the potential to successfully remove cells and DNA from both bovine and human bone disc bone tissue, while retaining functional molecules, such as GAGs, and tissue biomechanical properties. The retention of bone in the IVD samples should allow incorporation of the tissue into the recipient spine. Although there is a need for further studies to investigate repeatability, biocompatibility and functional performance, the potential of a decellularised IVD was demonstrated in this study.