Stephen M. Eisenstein

Donor site pain from the ilium. A complication of lumbar spine fusion

Chronic pain at the donor site was reported by 25% of 290 patients who had undergone anterior lumbar spine fusion for low back pain. Donor site pain has characteristic clinical features, may be severely disabling and is stubbornly resistant to treatment. The highest prevalence was in patients who had a tricortical full thickness graft taken through a separate incision overlying the iliac crest. Patients with a clinically unsatisfactory result from the spine fusion also had a significantly higher prevalence of donor site pain.

Matrix Metalloproteinases And Aggrecanase : Their Role in Disorders of the Human Intervertebral Disc

Study Design. A comprehensive immunohistochemical study of matrix metalloproteinase activity in discs from patients with different disc diseases.— Objectives. To identify individual matrix metalloproteinase enzymes that could contribute to the degeneration of the matrix of the intervertebral disc, to identify the cells that produce matrix metalloproteinases (for example, the endogenous disc cells or invading cells associated with vascularisation), and to determine if “aggrecanase” contributes to degradation of proteoglycans in disc disorders. Summary of Background Data. Matrix disorganization and loss of substance are the most common findings in degenerate discs, and proteinase enzyme activity is one means of causing these changes. Methods. Forty-nine discs from 46 patients with degenerative disc disease, posterior anular tears, spondylolisthesis, or disc herniation were studied immunohistochemically to determine the presence of matrix metalloproteinases 1, 2, 3, 7, 8, 9 and 13, tissue metalloproteinases 1 and 2, and proteoglycan degradation products generated by either matrix metalloproteinases or aggrecanase activity. In addition, in situ zymography was used to confirm matrix metalloproteinase activity. Results. The most extensive staining was seen for matrix metalloproteinases 1, 2, 3, and 9, with 91%, 71%, 65%, and 72% of samples having some immunopositivity for the respective antibodies. In contrast, staining for matrix metalloproteinases 7 and 8 was much less (38% for both). Tissue inhibitor of metalloproteinases 1 and 2 were expressed in 34% and 79% of specimens, respectively. Matrix metalloproteinases were found particularly in cell clusters and blood vessels of degenerate discs, with staining correlating positively with macroscopic degenerative grade. For all of the enzymes, there was most staining in the herniation specimens and least in the autopsy samples. The opposite was true of staining for the matrix metalloproteinases inhibitor, tissue inhibitor of metalloproteinases 2, with most found in the autopsy specimens. Enzyme activity was confirmed by in situ zymography and staining for matrix metalloproteinase degradation products of proteoglycans. In addition, there was staining with antibodies demonstrating aggrecanase degradation products. Conclusions. Matrix metalloproteinase activity is more prevalent in herniated discs than in other disc disorders studied, although matrix metalloproteinases may have been more common earlier in the disease progression. Matrix metalloproteinases can be produced by invading blood vessels and associated cells, as well as by indigenous disc cells. Aggrecanase activity, although present in some samples, was not as obvious as that of matrix metalloproteinases. In addition to altered matrix metalloproteinase production, there appears to be a change in the balance between enzymes and endogenous inhibitors, tissue inhibitors of metalloproteinases. This study highlights specific matrix metalloproteinases that might be most efficient to target in developing therapeutics for minimizing degradation of the extracellular matrix of the disc.

Are animal models useful for studying human disc disorders / degeneration?

Intervertebral disc (IVD) degeneration is an often investigated pathophysiological condition because of its implication in causing low back pain. As human material for such studies is difficult to obtain because of ethical and government regulatory restriction, animal tissue, organs and in vivo models have often been used for this purpose. However, there are many differences in cell population, tissue composition, disc and spine anatomy, development, physiology and mechanical properties, between animal species and human. Both naturally occurring and induced degenerative changes may differ significantly from those seen in humans. This paper reviews the many animal models developed for the study of IVD degeneration aetiopathogenesis and treatments thereof. In particular, the limitations and relevance of these models to the human condition are examined, and some general consensus guidelines are presented. Although animal models are invaluable to increase our understanding of disc biology, because of the differences between species, care must be taken when used to study human disc degeneration and much more effort is needed to facilitate research on human disc material.

Transport properties of the human cartilage endplate in relation to its composition and calcification.

Study Design The transport properties of solutes of different sizes and conformations were studied in cartilage endplates. Objectives The results were correlated with the composition of the cartilage matrix to determine if a relationship existed between this and the movement of molecules within it. Summary of Background Data Solute transport through the hyaline cartilage endplate is important not only for the physiologic and metabolic processes of that tissue, but also for those of the adjacent intervertebral disc. Movement of solutes depends on solute size, shape or charges, and the composition of the matrix itself. Changes in composition of the cartilage endplate, such as those that occur in degeneration or scoliosis, may affect transport. Methods Partition and diffusion coefficients of solutes ranging in molecular weight from 115 to 70,000 d have been measured on cores of cartilage endplate. Transport properties were assessed in relation to core composition. Results The shape and size of the solutes were found to affect their transport through cartilage matrix, with larger molecules being more highly excluded and diffusing more slowly. Long‐chain polymers were able to penetrate the matrix less readily than the more globular molecules. The more hydrated the matrix, the higher the degree of penetration and the more easily solutes could move, in contrast to the inverse relationship between the other components of the matrix and solute transport. With increased proteoglycan, collagen, or calcification in the tissue, there was greater restriction of solute movement. Conclusions The proteoglycans normally found in the endplate regulate movement of solutes into and out of the disc. It has been shown previously that removal of proteoglycans from the endplate accelerates the loss of proteoglycans from the nucleus. Hence, a major function of the cartilage endplate may be to prevent fragments of osmotically active proteoglycans from leaving the disc.

Matrix metalloproteinases in the human intervertebral disc: role in disc degeneration and scoliosis.

Study Design. Biochemical study of human intervertebral discs collected at surgery from patients with low back pain associated with disc degeneration or scoliosis. Matrix metalloproteinases were studied by quantitative zymography. Objective. To determine whether changes in the expression of matrix metalloproteinases will bring about tissue remodelling that contributes to the progressive nature and pathology of these diseases of the intervertebral disc. Summary of Background Data. The diseases of the intervertebral disc, degenerative disc disease and scoliosis, are both characterized by changes in the extracellular matrix components that will affect the mechanical function of the tissue. Matrix metalloproteinases are known to have the capability of degrading all the known extracellular matrix components of the disc. Methods. Matrix metalloproteinases 2 and 9 were detected by gelatin‐gel zymography and quantified by laser scanning densitometry. Both pro and active forms of the enzymes were measured. Thirty‐four discs from patients with low back pain and 29 from patients with scoliosis were investigated. Results. A correlation was found between the increasing levels of matrix metalloproteinases 2 and 9 and the grade of degenerative disc disease. In addition, the levels of these enzymes show a differential expression across the scoliotic disc with the highest levels in samples taken from the convexity of the curve. Conclusions. The difference between the concave and convex side of the scoliotic curve indicates that mechanical loads might influence the expression of these enzymes. The increased expression of these enzymes in both degenerative disc disease and scoliosis strongly suggests that they may affect the progressive nature of these diseases.

Mechanoreceptors in intervertebral discs. Morphology, distribution, and neuropeptides.

Study Design The present study investigated the occurrence and morphology of mechanoreceptors in human and bovine intervertebral discs and longitudinal ligaments. Objective To determine the type and frequency of mechanoreceptors present in intervertebral discs and anterior longitudinal ligaments in two patient groups, those with low back pain and those with scoliosis. Bovine coccygeal discs were examined. Summary of Background Data Nerves have been described in intervertebral tissues, but there is little information on the endings of these nerves and their receptors, stimulation of which can cause a nerve impulse. Methods The presence of mechanoreceptors were investigated by immunolocalization of nerves and neuropeptides.By examining sequential sections, the frequency of receptors was assessed. Results Immunoreactivity to neural antigens showed mechanoreceptors in the anulus fibrosus and longitudinal ligaments of bovine and human specimens. Their morphology resembled Pacinian corpuscles, Ruffni endings, and, most frequently, Golgi tendon organs. They were found in 50% of discs investigated from patients with low back pain and in 15% of those with scollosis. Conclusions Mechanoreceptors were found in the outer 2–3 lamellae of the human intervertebral disc and anterior longitudinal ligament. Physiologic studies in other tissues indicate that these provide the individual with sensation of posture and movement, and in the case of Golgi tendon organs, of nociception. In addition to providing proprioception, mechanoreceptors are thought to have roles in maintaining muscle tone and reflexes. Their presence in the intervertebral disc and longitudinal ligament can have physiologic and clinical implications.

Cell Cluster Formation in Degenerate Lumbar Intervertebral Discs is Associated with Increased Disc Cell Proliferation

Healthy human intervertebral discs contain relatively few cells and these are sparsely distributed. A characteristic feature of disc degeneration, however, is the appearance of cell clusters, particularly in damaged areas. How these clusters form is currently unknown. We have examined excised pathological human discs for evidence of cell proliferation. Disc sections were immunostained for the proliferating cell nuclear antigen (PCNA) and the proliferation-associated Ki-67 antigen. PCNA immunopositive cells were observed within degenerate discs, commonly though not exclusively, in cell clusters. Cells immunopositive for the Ki-67 antigen were less prevalent than those for PCNA, but similarly were observed frequently within clusters in degenerate discs. In contrast, immunopositivity for these markers was not common in less degenerate discs or in areas of the disc where cell clusters were not observed. These observations suggest that disc cell proliferation is associated with disc degeneration and is the likely cause of cell cluster formation.

Changes in collagen cross-linking in degenerative disc disease and scoliosis

Study Design. Biochemical study of the changes in the collagen cross‐link profile of human intervertebral discs collected at surgery from patients with either low back pain associated with disc degeneration or scoliosis. Objective. To determine whether changes occur in the collagen cross‐link profile in the disc of patients with either low back pain associated with disc degeneration or scoliosis, which may well influence matrix integrity. Such changes in the cross‐link profile of a tissue indicates increased matrix turnover and tissue remodeling and may have implications for the progression of these disorders. Summary of Background Data. The diseases of the intervertebral disc, degenerative disc disease and scoliosis, are both characterized by changes in the extracellular matrix components that will affect the mechanical function of the tissue. The stability of the collagenous components and hence the mechanical integrity of connective tissues such as the disc is dependent on the degree and type of cross‐links between the collagen molecules. This article reports results on the distribution of the different cross‐links in the disc and the changes that occur with age, degenerative disc disease, and scoliosis. Methods. Thirty‐three discs were obtained from patients with degenerative disc disease and 29 discs from patients with scoliosis. Samples were acid hydrolyzed and the collagen cross‐links analyzed by either fractionation on an amino acid analyzer configured for cross‐link analysis using ninhydrin postcolumn detection or fractionation by high‐pressure liquid chromatography with fluorescence detection. Results. The reducible cross‐links and the mature cross‐link all increased from the outer anulus fibrosus through into the nucleus pulposus. The highest levels of the mature cross‐link were found in the cartilage endplate. The nonenzymic derived cross‐link, pentosidine, in contrast, showed little difference across the disc, but did show the expected age‐related increase. In degenerative disc disease, no change in the levels of the reducible or mature cross‐links was found, but a decrease was observed in the levels of the age‐related cross‐link pentosidine in the more severe disease samples. In scoliosis, significantly higher levels of the reducible cross‐links were found on the convex than on the concave side of the scoliotic disc. Conclusions. These changes in the cross‐link profile of the intervertebral disc in degenerative disc disease and scoliosis are indicative of increased matrix turnover and tissue remodeling and likely to have implications for the progression of these disorders.

Immunohistochemical detection of Schwann cells in innervated and vascularized human intervertebral discs

Study Design. The ingrowth of nerves, blood vessels, and Schwann cells into human intervertebral discs was examined using immunohistochemistry for cell-type-specific markers. Objectives. To determine whether Schwann cells may contribute to disc innervation, and to assess the relation between disc innervation and vascularization. Summary of Background Data. Intervertebral disc degeneration was associated previously with ingrowth of blood vessels and nerves. Schwann cells are known to play an important role in regulating nerve growth and survival in other tissues, but they have not been examined in human pathologic intervertebral discs. Methods. Serial sections of human intervertebral discs were immunostained for the neuronal markers (neurofilament 200, peripherin, protein gene product 9.5), for the Schwann cell marker (glial fibrillary acidic protein), and for the endothelial cell marker (CD34). Results. Glial fibrillary acidic protein–immunopositive cells colocalized with nerves in degenerate discs, but were absent or rarely observed in nondegenerate, aneural discs. These also were seen in the disc matrix, independently of nerves. Much of the nerve and Schwann cell ingrowth was found in vascularized areas of disc tissue, where the lamellar structure of the anulus fibrosus was disrupted. Blood vessels were observed deeper into the discs than nerves or Schwann cells. Conclusions. The appearance of glial fibrillary acidic protein–immunopositive cells in diseased intervertebral discs was closely associated with nerve ingrowth. This novel finding suggests that Schwann cells have a role to play in regulating disc innervation and nerve function in the disc. Because blood vessels were observed furthermost into the disc, it is possible that degenerate disc vascularization occurs before innervation.

Human intervertebral disc aggrecan inhibits endothelial cell adhesion and cell migration in vitro.

Study Design. The effect of human intervertebral disc aggrecan on endothelial cell growth was examined using cell culture assays. Objective. To determine the response of endothelial cells to human intervertebral disc aggrecan, and whether the amount and type of aggrecan present in the intervertebral disc may be implicated in disc vascularization. Summary of Background Data. Intervertebral disc degeneration has been associated with a loss of proteoglycan, and the ingrowth of blood vessels and nerves. Neovascularization is a common feature also of disc herniation. Intervertebral disc aggrecan is inhibitory to sensory nerve growth, but the effects of disc aggrecan on endothelial cell growth are not known. Methods. Aggrecan monomers were isolated separately from the anulus fibrosus and nucleus pulposus of human lumbar intervertebral discs, and characterized to determine the amount and type of sulfated glycosaminoglycan side chains present. The effects of these aggrecan isolates on the cellular adhesion and migration of the human endothelial cell lines, HMEC-1 and EAhy-926, were examined in vitro. Results. Homogenous substrata of disc aggrecan inhibited endothelial cell adhesion and cell spreading in a concentration dependent manner. In substrata choice assays, endothelial cells seeded onto collagen type I migrated over the collagen until they encountered substrata of disc aggrecan, where they either stopped migrating, retreated onto the collagen, or, more commonly, changed direction to align along the collagen-aggrecan border. The inhibitory effect of aggrecan on endothelial cell migration was concentration dependent, and reduced by enzymatic treatment of the aggrecan monomers with a combination of chondroitinase ABC and keratinase/keratinase II. Anulus fibrosus aggrecan was more inhibitory to endothelial cell adhesion than nucleus pulposus aggrecan. However, this difference did not relate to the extent to which the different aggrecan isolates were charged, as determined by colorimetric assay with 1,9-dimethylmethylene blue, or to marked differences in the distribution of chondroitin sulfated and keratan sulfated side chains. Conclusions. Human intervertebral disc aggrecan is inhibitory to endothelial cell migration, and this inhibitory effect appears to depend, in part, on the presence of glycosaminoglycan side chains on the aggrecan monomer.