No abstract

Low back pain is a common and debilitating disorder. Current evidence implicates intervertebral disc (IVD) degeneration and herniation as major causes, although the pathogenesis is poorly understood. While several cytokines have been implicated in the process of IVD degeneration and herniation, investigations have predominately focused on Interleukin 1 (IL-1) and tumor necrosis factor alpha (TNFα). However, to date no studies have investigated the expression of these cytokines simultaneously in IVD degeneration or herniation, or determined which may be the predominant cytokine associated with these disease states. Using quantitative real time PCR and immunohistochemistry we investigated gene and protein expression for IL-1β, TNFα and their receptors in non-degenerate, degenerate and herniated human IVDs. IL-1β gene expression was observed in a greater proportion of IVDs than TNFα (79% versus 59%). Degenerate and herniated IVDs displayed higher levels of both cytokines than non-degenerate IVDs, although in degenerate IVDs higher levels of IL-1β gene expression (1,300 copies/100 ng cDNA)were observed compared to those of TNFα (250 copies of TNFα/100 ng cDNA). Degenerate IVDs showed ten-fold higher IL-1 receptor gene expression compared to non-degenerate IVDs. In addition, 80% of degenerate IVD cells displayed IL-1 receptor immunopositivity compared to only 30% of cells in nondegenerate IVDs. However, no increase in TNF receptor I gene or protein expression was observed in degenerate or herniated IVDs compared to non-degenerate IVDs. We have demonstrated that although both cytokines are produced by human IVD cells, IL-1β is expressed at higher levels and in more IVDs, particularly in more degenerate IVDs (grades 4 to 12). Importantly, this study has highlighted an increase in gene and protein production for the IL-1 receptor type I but not the TNF receptor type I in degenerate IVDs. The data thus suggest that although both cytokines may be involved in the pathogenesis of IVD degeneration, IL-1 may have a more significant role than TNFα, and thus may be a better target for therapeutic intervention.

Current evidence implicates intervertebral disc degeneration as a major cause of low back pain, although its pathogenesis is poorly understood. Numerous characteristic features of disc degeneration mimic those seen during ageing but appear to occur at an accelerated rate. We hypothesised that this is due to accelerated cellular senescence, which causes fundamental changes in the ability of disc cells to maintain the intervertebral disc (IVD) matrix, thus leading to IVD degeneration. Cells isolated from non-degenerate and degenerate human tissue were assessed for mean telomere length, senescenceassociated β-galactosidase (SA-β-gal), and replicative potential. Expression of P16 INK4A (increased in cellular senescence) was also investigated in IVD tissue by means of immunohistochemistry. RNA from tissue and cultured cells was used for real-time polymerase chain reaction analysis for matrix metalloproteinase-13, ADAMTS 5 (a disintegrin and metalloprotease with thrombospondin motifs 5), and P16 INK4A . Mean telomere length decreased with age in cells from nondegenerate tissue and also decreased with progressive stages of degeneration. In non-degenerate discs, there was an agerelated increase in cellular expression of P16 INK4A . Cells from degenerate discs (even from young patients) exhibited increased expression of P16 INK4A , increased SA-β-gal staining, and a decrease in replicative potential. Importantly, there was a positive correlation between P16 INK4A and matrix-degrading enzyme gene expression. Our findings indicate that disc cell senescence occurs in vivo and is accelerated in IVD degeneration. Furthermore, the senescent phenotype is associated with increased catabolism, implicating cellular senescence in the pathogenesis of IVD degeneration.

Following a previous description of nociceptive nerve fibre growth into usually aneural inner parts of painful intervertebral disc (IVD), this study has investigated whether nociceptive nerve ingrowth into painful IVD is stimulated by local production of neurotrophins. Immunohistochemistry and in situ hybridization have been used to investigate expression of the candidate neurotrophin, nerve growth factor (NGF), and its high- and low-affinity receptors trk-A and p75, respectively, in painful IVD excised for the management of low back pain. IVD from patients with back pain were of two types: those that when examined by discography reproduced the patient symptoms (pain level IVD) and those that did not (non-pain level IVD). Microvascular blood vessels accompanied nerve fibres growing into pain level IVD and these expressed NGF. The adjacent nerves expressed the high-affinity NGF receptor trk-A. These vessels entered the normally avascular IVD through the discal end plates. NGF expression was not identified in non-pain level or control IVD. Some non-pain level IVD had vessels within them, which entered through the annulus fibrosus. These did not express NGF nor did nerves accompany them. These findings show that nociceptive nerve ingrowth into painful IVD is causally linked with NGF production by blood vessels growing into the IVD, from adjacent vertebral bodies.

The histological and biochemical changes that occur in the extracellular matrix of the intervertebral disc (IVD) during ageing and degeneration have been investigated extensively. However, the mechanisms behind these changes are not fully understood. A number of studies have suggested the involvement of matrix metalloproteinases (MMPs) and ADAMTS in IVD degeneration, but few have localized the site of production of these enzymes to the cells of the degenerate disc. This study uses immunohistochemical techniques to localize and quantify the production of degrading enzymes (MMPs 1, 3, and 13, and ADAMTS 4) and their inhibitors (TIMPS 1, 2, and 3) within non-degenerate and degenerate discs of varying severity of degeneration. In all discs investigated, the cells that produced the enzymes and their inhibitors were the chondrocyte-like cells of the nucleus pulposus and inner annulus fibrosus (AF), with little immunopositivity in the outer AF. Non-degenerate discs showed low numbers of cells expressing the degradative enzymes MMP 1 and ADAMTS 4, suggesting a role for these enzymes in normal homeostasis. No MMP 3 or MMP 13 immunopositivity was observed in non-degenerate discs. In degenerate discs, the number of cells immunopositive for MMPs 1, 3, 13 and ADAMTS 4 increased with the severity of degeneration. This increase in degrading enzymes was also accompanied by increases in the number of cells immunopositive for TIMPs 1 and 2 but not TIMP 3. This study highlights that although the expression of a number of MMPs increases with degeneration, this is accompanied by an increase in their inhibitors. However, the increase in the number of cells immunoreactive for ADAMTS 4 with increasing degeneration was not paralleled by a rise in its inhibitor TIMP 3. This finding indicates that the aggrecanases, rather then the MMPs, are a possible therapeutic target for the inhibition of disc degeneration.