30 High impact loading organ culture model to investigate the post-traumatic disc degenerative condition
Objective: The aim of this study was to analyse the biological and biomechanical behaviour of the intervertebral disc (IVD) in response to a single high-impact traumatic loading.
Methods: Bovine caudal IVDs were cultured under 2 different loading conditions: 1) Control group under physiological sinusoidal dynamic loading; 2) In the model group, a single high impact traumatic damage was induced. After another culture period of 1 or 7 days under physiological loading, the stiffness of the IVD was measured, the loss of matrix molecules in the culture medium was quantified, the tissue structure was assessed by histology, the cell viability was analysed, and the gene expression of catabolic and inflammatory markers was analysed. RNA samples from day 1 were also analysed by RNA-sequencing. The Gene Ontology terms and Kyoto Encyclopaedia of Genes and Genomes analyses were performed.
Results: The model group exposed to single high impact loading had a lower stiffness compared to control group, indicating loss of mechanical properties. Extracellular matrix degrading enzymes and inflammatory genes were up-regulated in the model group compared to the control group. Single high impact loading induced higher release of extracellular matrix components into the culture medium, more severe cell death, and more disorganized tissue structure. RNA sequencing revealed 460 differentially expressed genes (nucleus pulposus(NP):252, annulus fibrosus(AF):208) in IVD tissue one day after the single impact traumatic event. Five dysregulated pathways were also enriched in the degenerated disc tissue.
Conclusion: The single high impact traumatic loading induced a degenerative cascade, as indicated by significant drop of cell viability, altered gene expression and tissue structure, and increased extracellular matrix degradation. This model has a high potential for investigation of the degeneration mechanism in post-traumatic IVD disease.