Abstract
Data implicate IL-1 in the altered matrix biology that characterizes human intervertebral disc (IVD) degeneration. In the current study we investigated the enzymic mechanism by which IL-1 induces matrix degradation in degeneration of the human IVD, and whether the IL-1 inhibitor IL-1 receptor antagonist (IL-1Ra) will inhibit degradation. A combination of in situ zymography (ISZ) and immunohistochemistry was used to examine the effects of IL-1 and IL-1Ra on matrix degradation and metal-dependent protease (MDP) expression in explants of non-degenerate and degenerate human IVDs. ISZ employed three substrates (gelatin, collagen, casein) and different challenges (IL-1β, IL-1Ra and enzyme inhibitors). Immunohistochemistry was undertaken for MDPs. In addition, IL-1Ra was introduced into degenerate IVD explants using genetically engineered constructs. The novel findings from this study are: IL-1Ra delivered directly onto explants of degenerate IVDs eliminates matrix degradation as assessed by multi-substrate ISZ; there is a direct relationship between matrix degradation assessed by ISZ and MDP expression defined by immunohistochemistry; single injections of IVD cells engineered to over-express IL-1Ra significantly inhibit MDP expression for two weeks. Our findings show that IL-1 is a key cytokine driving matrix degradation in the degenerate IVD. Furthermore, IL-1Ra delivered directly or by gene therapy inhibits IVD matrix degradation. IL-1Ra could be used therapeutically to inhibit degeneration of the IVD.
Highlights
Chronic low back pain either alone or in association with sciatica (CLBP), is a common musculoskeletal disorder causing considerable population morbidity (6% prevalence) and an £11 billion pound annual cost to the UK economy through social and healthcare expenditure and loss of work
The novel findings from this study are: IL-1 receptor antagonist (IL-1Ra) delivered directly onto explants of degenerate intervertebral disc (IVD) eliminates matrix degradation as assessed by multi-substrate in situ zymography (ISZ); there is a direct relationship between matrix degradation assessed by ISZ and metal-dependent protease (MDP) expression defined by immunohistochemistry; single injections of IVD cells engineered to over-express IL-1Ra significantly inhibit MDP expression for two weeks
In the current study we have addressed the lack of direct evidence for this mechanism in human tissue, believing this to be an essential step in translating current laboratory data into clinical applications, by examining the hypothesis 'matrix degradation in degeneration of the intervertebral disc (DIVD) is inhibited by IL-1Ra'
Summary
Chronic low back pain either alone or in association with sciatica (CLBP), is a common musculoskeletal disorder causing considerable population morbidity (6% prevalence) and an £11 billion pound annual cost to the UK economy through social and healthcare expenditure and loss of work. Individual intervertebral discs (IVDs) are part of a complex of interdependent spinal structures known as the 'motion segment', in which IVDs facilitate movement and maintain optimal separation and orientation of other elements This is achieved by a biomechanical balance between the IVD's two main structural elements, the nucleus pulposus (NP) and the annulus fibrosus (AF). Excessive swelling is ADAMTS = a disintegrin and metalloproteinase with thrombospondin motifs; Ad-GFP = adenoviral constructs incorporating green fluorescent protein; AF = annulus fibrosus; ANOVA = Analysis of variables; BSIP = broad spectrum inhibitor of proteinases; CcCl = Cesium chloride; CLBP = chronic low back pain with or without sciatica; DIVD = degeneration of the intervertebral disc; GFP = green fluorescent protein; IHC = immunohistochemistry; IL = interleukin; IL-1Ra = interleukin-1 receptor antagonist; ISZ = in situ zymography; IVD = intervertebral disc; MDP = metal-dependent proteases; MMP = matrix metalloproteinase; MOI = Multiplicity of infection; mRNA = messenger ribose nucleic acid; NP = nucleus pulposus; PMSF = phenyl methyl sulphonyl fluoride; RNA = ribose nucleic acid; T = thymidine
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