THE NEUROVASCULAR AND INFLAMMATORY SIGNATURES OF THE DEGENERATING INTERVERTEBRAL DISC

Objective: Inflammatory cytokine production and de novo neurovascularization have been identified in painful, degenerated interver-tebral discs (IVDs). However, the progression of these key pathoanatomical features, including the cascade of inflammatory cytokines and the infiltration of vessels and neurites remain undefined. The objective of this study is to define the progression of cytokine production and neurovascular invasion during the IVD degeneration. Methods: A needle-puncture injury model was applied to the caudal IVDs of 3-month-old C57BL6/J female mice. The animals were euthanized after the IVDs were allowed to degenerate in vivo for 2-, 4-, and 12-weeks following the injury. A set of IVDs were extracted for immunostaining of infiltrating vessels (endomucin) and nerves (protein-gene-product 9.5). Another set of IVDs were placed in organ culture and forty-nine IVD-secreted cytokines and matrix metal-loproteinases (MMPs) were measured from the media. Results: IVD injury resulted in severe degeneration by 2-weeks that remained consistent up to 12-weeks post puncture. Injury increased the secretion of interleukin (IL)-6, tumor necrosis factor (TNF)α, C-C motif ligand (CCL)4, CCL12, CCL17, CCL20, CCL21, CCL22, C-X-C motif ligand (CXCL)2 and MMP-2 proteins. Although the control IVDs did not exhibit degeneration, they produced TNFα and CCL22. The centrality and structure of inflammatory networks in IVDs evolves over the 12-week time course following injury, highlighting distinct responses between the acute and chronic phases. Neurites propagates rapidly within 2-weeks post-injury and remains constant until 12-weeks. Vascular length peaks at 4-weeks post-injury and re-gresses by 12-weeks. These findings identify the temporal flux of inflammatory cytokines and pain-associated pathoanatomy in a model of IVD degeneration of the mouse caudal spine. Conclusions: Caudal puncture provides a non-invasive alternative to lumbar puncture allowing for injury to multiple IVDs providing a platform for more extensive studies while minimizing the number of mice needed.