Abstract
Purpose and Background: There is increasing evidence that events within the diseased intervertebral disc (IVD) are mediated by locally synthesised cytokines. A prominent histological, imaging and surgical feature of IVD disease is degradation of the cartilaginous discal matrix. Whilst the mechanism by which this is mediated is unknown, in other situations where connective tissues are degraded degradation is the result of production of matrix-degrading enzymes by local connective tissue cells stimulated by cytokines, particularly the beta isoform of interleukin-1 (IL-1β). Included amongst these disorders is osteoarthritis (OA) of diarthrodial joints. OA has many similarities to the discal “degeneration” seen in mechanical back pain syndromes. In the current study, we have used a combination of in-situ techniques to establish if IL-1β is responsible for stimulating matrix degradation in the IVD.
Methods: Using a combination of radioactive in-situ hybridisation (ISH) and competitive in situ zymography (ISZ) we have studied expression of IL-1β and IL-1R – its type 1 receptor (ISH) and matrix degradation (ISZ) in five diseased lumbar IVD taken at spinal fusion surgery and 10 cadaveric IVD (five normal and five diseased). The nucleus pulposus (NP) was separated from the annulus fibrosus and diced into 0.5cm cubes. Half the cubes (typically three) were fixed in formalin and processed into paraffin wax for ISH, and half were used for ISZ. For ISH, 5 μm sections of paraffin-embedded tissue were reacted with cDNA probes radiolabelled with 35S to 580 and 530 base segments of the IL-1β and IL-1R molecules. Hybridisation was disclosed using autoradiography. For ISZ, 50 μm vibratome sections were placed into wells on microscope slides precoated with gelatin. Sections were incubated for 10 days, half in culture medium and half in medium supplemented with human recombinant IL-1 receptor antagonist (IL-1Ra – an inhibitor of IL-1). Sections were photographed at daily intervals to detect evidence of gel degradation.
Results: Chondrocytes within patient and cadaveric diseased but not normal discs expressed mRNA for both IL-1β and IL-1R. By ISZ, the same cells degraded gelatin. Degradation was inhibited by recombinant IL-1Ra.
Conclusion: This study shows that chondrocytes of diseased discs express IL-1 and its receptor. The same cells produced matrix-degrading enzymes by a mechanism that can be inhibited by the IL-1 inhibitor IL-1Ra. IL-1 is a potential therapeutic target for the management of IV disc disease.
The abstracts were prepared by Dr P Dolan. Correspondence should be addressed to him at the British Orthopaedic Association, Royal College of Surgeons, 35-43 Lincoln’s Inn Fields, London WC2A 3PN.
