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A method for the purification of serine proteinases from the bovine intervertebral disc using affinity chromatography on basic pancreatic trypsin inhibitor (BPTI) immobilized to the hydroxyalkyl methacrylate copolymer Separon HEMA 1000 E is reported. Its advantage is the possibility of obtaining serine proteinases without an artificial alteration in relative molecular mass.
Human intervertebral disc (IVD) degeneration often initiated from the human nucleus pulposus (hNP) with aging leading to IVD destruction and extracellular matrix (ECM) depletion. Previously, we have successfully employed transforming growth factor-beta1 (TGF-beta1) to promote chondrogenesis of mesenchymal progenitor cells (MPCs) and immortalized human mesenchymal stem cells. In this study, we examine the role of TGF-beta1 in platelet-rich plasma (PRP) on disc regeneration, including proliferation, redifferentiation, and the reconstitution of tissue-engineered NP. hNP cells were isolated from volunteers with different ages and cultured in the presence of PRP. We found that the most effective concentration for hNP proliferation was 1 ng/ml TGF-beta1 in PRP, which was further applied in the following experiments. hNP cell proliferation in all age groups were increased time-dependently by PRP and cell morphologies showed aggregation. The mRNA of Sox9, type II collagen, and aggrecan were all significantly upregulated by PRP through RT-PCR. Glycosaminoglycan (GAG) accumulation reached the highest value at day 7 and continued to day 9 culture. PRP promoted NP regeneration via the Smad pathway was also determined and highly activated p-Smad2/3 at 30 min and continuously sustained to 120 min. Immunostaining of type II collagen indicates that PRP participates in chondrogenesis of tissue-engineered NP with collagen scaffolds. We concluded that growth factors in PRP can effectively react as a growth factor cocktail to induce hNP proliferation and differentiation, and also promote tissue-engineered NP formation. These findings are the first to demonstrate that PRP might be a therapeutic candidate for prevention of disc degeneration.
A simple finite element model of the L5-S1 intervertebral disc body has been constructed; it is circular and symmetrical about the sagittal plane. The annulus fibrosus of the model was idealized as an inhomogeneous composite of an isotropic ground substance, reinforced by helically oriented collagen fibres so that the model has six different structural components namely: cortical bone, cancellous bone, cartilaginous endplates, nucleus pulposus, ground substance and collagen fibres. A sensitivity analysis of the material properties of each structural component was carried out by varying those properties for one structural component at a time and evaluating the changes in the biomechanical response to compressive displacements. Experimentally available relations between the applied compressive force and the vertical displacements, the nucleus pulposus pressure increase and the disc lateral bulge were used to evaluate the biomechanical responses for each set of material properties. Results showed that both the Poisson's ratio and the Young's modulus of the ground substance play an important role in the prediction of the biomechanical response.
Low back pain (LBP) is a common, debilitating and economically important disorder. Current evidence implicates loss of intervertebral disc (IVD) matrix consequent upon 'degeneration' as a major cause of LBP. Degeneration of the IVD involves increases in degradative enzymes and decreases in the extracellular matrix (ECM) component in a process that is controlled by a range of cytokines and growth factors. Studies have suggested using anabolic growth factors to regenerate the normal matrix of the IVD, hence restoring disc height and reversing degenerative disc disease. However, for such therapies to be successful it is vital that the target cells (i.e. the disc cells) express the appropriate receptors. This immunohistochemical study has for the first time investigated the expression and localization of four potentially beneficial growth factor receptors (i.e. TGFbetaRII, BMPRII, FGFR3 and IGFRI) in non-degenerate and degenerate human IVDs. Receptor expression was quantified across regions of the normal and degenerate disc and showed that cells of the nucleus pulposus (NP) and inner annulus fibrosus (IAF) expressed significantly higher levels of the four growth factor receptors investigated. There were no significant differences between the four growth factor expression in non-degenerate and degenerate biopsies. However, expression of TGFbetaRII, FGFR3 and IGFRI, but not BMP RII, were observed in the ingrowing blood vessels that characterize part of the disease aetiology. In conclusion, this study has demonstrated the expression of the four growth factor receptors at similar levels in the chondrocyte-like cells of the NP and IAF in both non-degenerate and degenerate discs, implicating a role in normal disc homeostasis and suggesting that the application of these growth factors to the degenerate human IVD would stimulate matrix production. However, the expression of some of the growth factor receptors on ingrowing blood vessels might be problematic in a therapeutic approach.
Between 1986 and 1988 100 patients in whom MRI had been used to diagnose prolapses of lumbar intervertebral discs were admitted to the neurosurgical department of the Munich Technical University. MRI proved to be a valuable diagnostic technique, even if one has to concede certain limitations to the usefulness of the results when compared to conventional methods, particularly in such cases where clinical symptoms are not clear. Typical findings are illustrated and compared with those seen in MRI, whose relative importance is discussed.
A comparison of the higher hydrostatic pressure in the nucleus of the healthy intervertebral disc during daily loading with the relatively lower osmotic swelling pressure in the disc during rest suggests the existence of direction-dependent flow resistance such that all of the fluid exuded from the disc during loading is recovered during rest. In this study, this direction-dependent resistance was demonstrated for flows through the cartilage endplates and the underlying marrow contact channels in the bony endplates. Using an ex vivo sheep endplate model, the resistance ratio (Rout/Rin) for flow through the endplate was 39.0+/-3.8 (mean +/- S.E.). In addition, a path of fluid flow through the marrow contact holes was revealed using fluorescent staining.
An analysis of the bulging of the annulus fibrosus as a function of the height of the intervertebral disc during compression is presented. The theory was compared with the results of experimental tests on rabbit lumbar discs. When viewed in profile the ventral bulge closely resembled an arc of a circle; at each stage of compression it appeared to extend from the cranial to the caudal extremities of the annulus fibrosus. The radius of curvature of the arc was measured as a function of disc height and the results compared with theoretical predictions. Good agreement was obtained if it was assumed that about half the volume change swept out by the approach of the cranial and caudal end-plates was accommodated by the annular bulge. The lengths of the collagenous fibres reinforcing the outer lamella of the ventral annulus were estimated from the dimensions of the ventral bulge; no appreciable length increase appeared to accompany compression.
An epidemiologic case-control study of herniated lumbar intervertebral disc was conducted in Springfield, Massachusetts, New Brunswick, New Jersey, and New York, New York, to evaluate the role of several possible risk factors in the etiology of this disorder. Patients with signs and symptoms of herniated lumbar disc (N = 287) were matched to control subjects without back pain by age, sex, source of care, and geographic area. Of the total case-subject group, 177 were confirmed by surgery, computed tomographic scan, myelogram, or magnetic resonance imaging. This article focuses on non-occupational lifting, an activity not previously reported on. Frequent lifting of objects or children weighing 25 or more pounds with knees straight and back bent was associated with increased risk of herniated lumbar disc. This association was particularly strong among confirmed case subjects (relative risk = 3.95). Positive associations among confirmed case subjects were also seen for frequent lifting with arms extended (relative risk = 1.87) and twisting while lifting (relative risk = 1.90). No associations were found for frequent stretching or carrying. If confirmed in other investigations, these data suggest that instruction in lifting techniques should be extended into the home.
Our objective for this study was to determine the presence and distribution of tenascin in the human intervertebral disc. The tenascins are a family of extracellular matrix proteins with repeated structural domains homologous to epidermal growth factor, fibronectin type III and the fibrinogens. Little is known about the presence of this protein in the disc. Ten normal human discs donated from subjects newborn to 15 years old, 10 control discs from adult donors aged 24-41 years, and 11 surgical disc specimens from patients aged 26-76 years were examined for immunolocalization of tenascin. In young discs, tenascin was localized throughout the annulus; in the nucleus, localization was confined to pericellular matrix. In adult control and degenerating disc specimens, tenascin in the annulus was localized primarily in pericellular matrix regions encircling either single cells or clusters of disc cells; in rare instances localization was more diffuse in the intraterritorial matrix. In young, healthy disc, tenascin was abundant throughout the annulus. In contrast, degenerating discs in adults showed a localization restricted to the pericellular, and rarely, more restricted intraterritorial matrix. These observations indicate that changes in the amount and distribution of tenascin may have a role in disc aging and degeneration, possibly by modulating fibronectin-disc-cell interactions, and causing alterations in the shape of disc cells.
It was found that the ageing process is accompanied by significant changes of elastin content in human intervertebral discs. In the case of annuli fibrosi only a slight age-related increase during the first four decades of life followed by a decrease in later periods of life were observed. In the case of nuclei pulposi the changes were much more apparent. A systematic decrease in elastin: collagen ratio was observed, especially in the case of nuclei pulposi. An increase in elastin: GAGs ratio was observed during the first 4 decades of life. After that period no further changes of the ratio were found.
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Cadaveric lumbar discs were injected with chymopapain and subjected to a series of mechanical tests over a period of up to 19 hours. Discs from the same spine injected with saline were used as controls. The results showed that chymopapain had no measurable effect on the mechanical properties of the disc apart from the increased height and stiffening caused by fluid injection. Another series of tests on isolated pieces of disc material showed that chymopapain could reduce the size of prolapsed nuclear material by 24% in one hour and by 80% in 48 hours. It is concluded that, in the short-term, chymopapain has a negligible effect on the mechanics of a disc but it can reduce the size of any prolapsed nuclear material with which it comes in contact.
Recognition of disc degeneration in vivo is important in the investigation of the pathophysiology of intervertebral disc disease as well as the assessment of patients. The purpose of this study was to compare low-field magnetic resonance imaging and histopathologic findings of disc degeneration in the canine caudal lumbar spine. A simple four-stage classification system for disc degeneration is proposed. Most common signs of disc degeneration in magnetic resonance imaging included nuclear clefts, decreased signal intensity of the nucleus pulposus and tears of the annulus fibrosus, or disc herniations. The association between magnetic resonance images and histopathologic findings was highly significant. A sensitivity of 100% and specificity of 79% for magnetic resonance imaging was calculated using histopathology as the gold standard.
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A survey of 37 young patients, 10 to 18 years of age, with lumbar intervertebral disc herniation is presented. Local physical findings and limitation in straight-leg raising are often more marked in the young than in adults, whereas pain may be slight or even absent. Neurological findings were normal in 40 per cent of the cases, and when deficits were found, they were usually of a minor nature. Herniated disc syndrome in children and adolescents is characteristic and should not be difficult to recognize. With surgical treatment good results were obtained in nearly all cases.
BACKGROUND: Epidural steroid injections are commonly used for the treatment of radicular symptoms associated with symptomatic lumbar intervertebral disc herniations. Transforaminal epidural injections are believed to produce better clinical outcomes than interlaminar epidural injections. OBJECTIVE: To determine a difference in short-term pain improvement and longterm surgical rates between interlaminar and transforaminal injection techniques. DESIGN: Case Control Study. METHODS: For each technique, 20 patients were retrospectively identified who received their first fluoroscopically guided epidural steroid injection for radicular symptoms caused by a lumbar intravertebral disc herniation over an 18 months interval. All patients had corresponding MRI findings and failed previous non-invasive therapies. The Verbal Numerical Rating Scale (VNRS, 0-10 scale) before the treatment, within one hour after the treatment and upon follow-up (average 17.1 days) were analyzed, along with the need for repeat injections and surgical interventions over a 1-year follow-up interval. The patient groups were matched for symptom duration, MRI findings and pre-injection VNRS scores. RESULTS: In the transforaminal group, there was a statistically significant improvement in the VNRS scores from before the injection (VNRS mean 5.9) to immediately after the injection (VNRS mean 2.9, p<0.01), and upon follow-up (VNRS mean 3.2, p<0.01, mean 18.7 days). Nine patients (45%) required 1 or 2 repeated injections, 2 patients (10%) underwent surgery. In the interlaminar group, there was a statistically significant improvement in the VNRS scores from before the injection (VNRS mean 7.3) to immediately after the injection (VNRS mean 3.1, p<0.01), and upon follow-up (VNRS mean 5.9, p<0.01, mean 15.6 days). Eight patients (40%) required 1 or 2 repeated injection, 5 patients (25%) underwent surgery. Fourteen patients (70%) had an improvement of 2 points or more on the VNRS scale in the transforaminal group, compared to 9 (45%) in the interlaminar group. CONCLUSIONS: In the current study, transforaminal epidural steroid injections for the treatment of symptomatic lumbar disc herniation resulted in better short-term pain improvement and fewer long-term surgical interventions than interlaminar epidural steroid injection.
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