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PubMed2005

Advanced glycation end products in intervertebral discs and hip joint capsules: correlation with senile amyloid?

Advanced glycation end-products (AGEs) may be involved in either amyloidogenesis or complications related to amyloid. The incidence of AGE increases with age as does the prevalence of amyloid affecting the hip joint capsule and intervertebral discs. We hypothesized that AGEs may be involved in the pathology of these amyloidoses and investigated the spatial and temporal relationship between AGEs and amyloid of intervertebral discs and hip joint capsules. Using immunohistochemistry, AGEs were found in all 71 intervertebral discs and all 87 hip joint capsules. Amyloid was present in 59 (83%) intervertebral discs and 65 (75%) hip joint capsules. AGEs were found in the immediate vicinity of amyloid deposits in 104 of 253 (41%) investigated amyloid deposits of the intervertebral discs and 159 of 311 (51%) investigated amyloid deposits of the hip joint capsules. However, only rarely were AGEs demonstrated within amyloid deposits. No linear correlation was found between the amount of AGEs and the amount of amyloid deposited. As AGEs are more common than amyloid in intervertebral discs and hip joint capsules, it is conceivable to suggest that AGEs might be involved in the pathogenesis of amyloid at these anatomical sites. However, the amyloid proteins appear not to be modified and AGE modification is not a general characteristic of senile amyloidoses.

Adult↗
PubMed1995

The development of fibrocartilage in the rat intervertebral disc.

The development of fibrocartilage in rat lumbar intervertebral discs has been correlated with an immunohistochemical analysis of the changing distribution of extracellular matrix components. Disc anlagen were first recognised by embryonic day 14 as segmental cell condensations. By E16, the notochord formed a series of bulges, each representing a future nucleus pulposus, and the annulus fibrosus had differentiated in the disc anlagen. The inner part of the annulus was composed of cartilage which linked that of adjacent vertebral bodies. The outer part was fibroblastic, with layers of parallel fibroblasts. The long axes of the cells in successive layers lay at an angle of approximately 90 degrees to each other. This criss-cross orientation of cells preceded the oriented deposition of collagen fibres to form the lamellae. Disc anlagen were immunolabelled weakly for types I and III collagen, chondroitin 6-sulphate and dermatan sulphate. Later tissue differentiation was marked by the appearance of type II collagen, chondroitin 4-sulphate and keratan sulphate in the inner annulus. These components also appeared in the outer annulus, but only in adult animals, and indicated metaplastic change in the lamellar fibroblasts. Fibrocartilage in the nucleus pulposus was only seen in old animals, and the origin of the tissue was less clear. However, the fibrocartilage cells appeared to be derived from the cartilage end plate and/or from the inner annulus. We conclude that fibrocartilage in the intervertebral disc is derived from several sources and that the radial distribution patterns of extracellular matrix components in the adult disc are explained by the embryonic origins of its parts.

Animals↗
PubMed1997

Aggrecan degradation in human intervertebral disc and articular cartilage.

Aggrecan degradation in human intervertebral disc and articular cartilage has been studied by using anti-neoepitope antibodies specific for the N-terminal degradation products generated by cleavage within the interglobular domain at the metalloproteinase and aggrecanase sites. Immunoblot analysis of extracts of annulus fibrosus, nucleus pulposus and articular cartilage demonstrated age-related patterns in the abundance of both degradation products. In all three tissues the metalloproteinase-generated fragment was present at very low levels in young individuals but increased in abundance with age. In the disc tissues, the abundance of this degradation product levelled off in the juvenile; for cartilage this occurred in early adulthood. Despite these temporal differences, the levels attained in adults were comparable for the three tissues. In contrast, the aggrecanase-generated degradation product exhibited tissue-specific differences in the variation of its abundance with age. Whereas this degradation product increased with age in annulus fibrosus and articular cartilage and had levelled off by adulthood, in nucleus pulposus it was present in greatest abundance in young individuals and decreased to very low levels with age. Examination of discs exhibiting various degrees of degeneration did not reveal any differences in the levels of the metalloproteinase and aggrecanase-generated cleavage products that could not be accounted for by differences in age. In adults the product of aggrecanase action was much more abundant in articular cartilage than in either of the disc tissues, despite the age-related increase also observed for annulus fibrosus. Analysis of tissue extracts with an antibody recognizing the G1 domain of aggrecan identified two major degradation products whose abundance and size were correlated with the fragments detected by the anti-neoepitope antibodies. Taken together, these results indicate that cleavage at the metalloproteinase and aggrecanase sites are quantitatively important events in aggrecan catabolism in both articular cartilage and intervertebral disc in vivo. Moreover the two enzyme systems act independently and exhibit differences in the degree to which they contribute to aggrecan degradation in these tissues.

Adolescent↗
PubMed2002

CTM brace effect on scoliotic intervertebral discs using MRI method.

MRI has been clinically only used for investigation of intervertebral disc disorders. In this study, MR images were used and a new 3D modelling of the intervertebral discs was proposed. MRI examination had been performed on fourteen girls presenting an idiopathic scoliosis and wearing a first CTM brace. Using an in-house image processing software and the pre-post processing software Patran, geometrical models were obtained with and without brace for each patient. These models included the outline of the intervertebral high intensity zone, composed of the nucleus and a part of the annulus. The shift forward between disc high intensity zone centres and body centres was found to be varying from 0 to 8mm. The sagittal and coronal shifts forward appeared in the curvature convexity and were maximum at the curvature apex. The intervertebral disc wedging was found to be varying from -10 degrees to +10 degrees. On these fourteen analysed patients, the CTM brace decreased the coronal shift forward between disc high intensity zone centres and body centres, and increased the sagittal intervertebral wedging. The intervertebral disc informations obtained represented new data in the scoliotic deformation description. But this method was not adapted for a clinical use. The qualitative and quantitative data obtained will help the orthopaedist in the brace design and also the clinician in the scoliosis comprehension.

Adolescent↗
PubMed2001

Intervertebral disc disorganisation and its relationship to age adjusted vertebral body morphometry and vertebral bone architecture.

Vertebral deformity, intervertebral disc disorganisation, and change to vertebral bone architecture are morphological features that are associated with low back pain. The purpose of this study was to examine the influence of the morphological disorganisation of the intervertebral disc on vertebral body shape indices and vertebral cancellous bone architecture. Lumbar spines, T12-S1, were collected from 27 cadavers. The motion segments T12-L1, L2-L3 and L4-L5 were selected for the study. There were 8 females aged 35-94 years and 19 males aged 20-90 years. An intervertebral disc grade signifying the severity of disc disorganisation was assigned to each disc using the macroscopic disc grading criteria of Hansson and Roos (Spine, 1981; 6:147-153.). Vertebral shape indices and vertebral body bone histomorphometric analyses were performed on the vertebral bodies. Where appropriate, data were age adjusted and the influence of morphological disc disorganisation on vertebral body deformity and cancellous bone architecture analysed. Increased vertebral body axial area and the ratio of vertebral body axial area to sagittal area were associated with an increase in vertebral deformity and disc disorganisation. This suggests that vertebral deformity that remains clinically silent in the general population is influenced by intervertebral disc disorganisation. Vertebral cancellous bone architecture undergoes change associated with increased disc disorganisation, consistent with increased vertebral deformity. Vertebral bodies adjacent to degenerate discs (Grade 4) showed increased BV/TV and Tb.Th and decreased BS/BV. This shows that disc disorganisation may modulate vertebral cancellous bone architecture such that it protects against age-related bone changes. In addition, vertebral body wedging and concavity are associated with smaller vertebral body size and vertebral body compression is associated with larger vertebral body size and compromised cancellous bone architecture.

Adult↗
PubMed2004

Conditioned medium differentially regulates matrix protein gene expression in cells of the intervertebral disc.

STUDY DESIGN: Matrix protein gene expression was determined for cells of the anulus fibrosus (AF) and nucleus pulposus (NP) regions of the intervertebral disc when cultured in AF or NP cell-conditioned medium. OBJECTIVES: To investigate changes in mRNA levels for type I collagen, type II collagen and aggrecan in cells of the AF and NP in response to cell-conditioned medium. SUMMARY OF BACKGROUND DATA: Cells of the intervertebral disc have been shown to respond to exogenous soluble mediators such as the growth factors TGF-beta and IGF-1. Little is known of their biologic response to endogenous factors that may be secreted locally or by cells of neighboring regions. METHODS: Porcine cells were cultured for 48 hours in alginate gel in the presence or absence of conditioned medium. Gene expression for aggrecan and collagens was quantified using real-time reverse transcriptase-polymerase chain reaction. RESULTS.: AF cell gene expression was generally stimulated by the conditioned medium of either AF or NP cells. In contrast, the notochordal cell-containing NP cells showed little change in gene expression with either source of conditioned medium. CONCLUSIONS: Cells of the NP and AF secrete soluble factors in culture at similarly effective doses to stimulate matrix protein gene expression in AF cells of the intervertebral disc. Unlike AF cells, however, NP cell gene expression was not stimulated by any conditioned medium, suggesting that differences exist in the responsiveness of cells of notochordal (NP) and fibrocartilaginous (AF) phenotypes. Understanding these differences between cells of the intervertebral disc may reveal unique stimulatory factors important to repair and regeneration of the degenerated intervertebral disc.

Aggrecans↗
PubMed1997

[Investigation on matrix degrading enzymes of lumbar intervertebral discs].

Changes in the macromolecular matrix of the intervertebral disc may predispose to biomechanical failure of the disc. Such changes would involve extracellular enzymes capable of altering the collagen and proteoglycan of the disc matrix. In this study, tritium-labeled type I collagen was used as a substrate to estimate the activity of collagenase in the discs of 41 cases of lumbar disc protrusion (LDP) patients by surgical intervention. The annulus fibrous (AF) and nucleus pulposus (NP) were measured separately. 34 normal discs harvested by autopsy acted as controls. For estimation of relative neutral proteinase content of 6 normal and 16 degenerated lumbar discs, polyacrylamide gelelectrophoresis (PAGE), heat-denatured collagen as a substrate, and photo-density scanning with peak area autocalculating system were adopted. The results presented that both AF and NP of the normal discs had a similar lower collagenolytic activity and a very limited activity of neutral proteinase, while the degenerated discs showed a higher activity, especially in the degenerated NP. The extruded type of LDP got a higher collagenolytic activity in NP than that of the prolapsed LDP. The fact showed that the matrix degrading enzymes play a very important role in the process of lumbar disc degeneration. The difference of disc degeneration is the biochemical basis of different clinical types of LDP. Matrix degrading enzyme system is a very complexed multienzymatic system. Other neutral proteinases may join this system besides the collagenase.

Adolescent↗
PubMed2002

Classification of age-related changes in lumbar intervertebral discs: 2002 Volvo Award in basic science.

STUDY DESIGN: A histologic study on age-related changes of the human lumbar intervertebral disc was conducted. OBJECTIVES: To investigate comprehensively age-related temporospatial histologic changes in human lumbar intervertebral disc, and to develop a practicable and reliable classification system for age-related histologic disc alteration. SUMMARY OF THE BACKGROUND DATA: No comprehensive microscopic analysis of age-related disc changes is available. There is no conceptual morphologic framework for classifying age-related disc changes as a reference basis for more sophisticated molecular biologic analyses of the causative factors of disc aging or premature aging (degeneration). METHODS: A total of 180 complete sagittal lumbar motion segment slices obtained from 44 deceased individuals (fetal to 88 years of age) were analyzed with regard to 11 histologic variables for the intervertebral disc and endplate, respectively. In addition, 30 surgical specimens (3 regions each) were investigated with regard to five histologic variables. Based on the semiquantitative analyses of 20,250 histologic variable assessments, a classification system was developed and tested in terms of validity, practicability, and reliability. The classification system was applied to cadaveric and surgical disc specimens not included in the development of the classification system, and the scores were assessed by two additional independent raters. RESULTS: A semiquantitative analyses provided clear histologic evidence for the detrimental effect of a diminished blood supply on the endplate, resulting in the tissue breakdown beginning in the nucleus pulposus and starting in the second life decade. Significant temporospatial variations in the presence and abundance of histologic disc alterations were observed across levels, regions, macroscopic degeneration grades, and age groups. A practicable classification system for age-related histologic disc alterations was developed, resulting in moderate to excellent reliability (kappa values, 0.49-0.98) depending on the histologic variable. Application of the classification system to cadaveric and surgical specimens demonstrated a significant correlation with age ( < 0.0001) and macroscopic grade of degeneration ( < 0001). However, substantial data scatter caution against reliance on traditional macroscopic disc grading and favor a histology-based classification system as a reference standard. CONCLUSIONS: Histologic disc alterations can reliably be graded based on the proposed classification system providing a morphologic framework for more sophisticated molecular biologic analyses of factors leading to age-related disc changes. Diminished blood supply to the intervertebral disc in the first half of the second life decade appears to initiate tissue breakdown.

Adolescent↗
PubMed2006

The involvement of aggrecan polymorphism in degeneration of human intervertebral disc and articular cartilage.

The functions of the intervertebral disc and of articular cartilage are intimately related to their aggrecan content. Aggrecan is a proteoglycan that interacts with hyaluronan to form large aggregates, which are responsible for the ability of the tissues to resist compressive loads. This function is related to the structure of aggrecan, and in particular to the large number of chondroitin sulphate chains present on its core protein. The chondroitin sulphate chains are present in two adjacent regions of the aggrecan core protein, termed the CS1 and CS2 domains. In the human, the region of the aggrecan gene encoding the CS1 domain exhibits size polymorphism, which can result in variation in the degree of chondroitin sulphate substitution of aggrecan in different individuals. This raises the possibility that the functional properties of aggrecan may vary between individuals, and that those individuals with an inferior aggrecan structure may be more susceptible to premature intervertebral disc or articular cartilage degeneration. Several studies have been performed to demonstrate such an association, but the results have been ambiguous. This review explains the relationship between aggrecan structure and function, describes the technique used to assess aggrecan polymorphism and the conclusions and limitations of the data obtained to date, and discusses the implications for tissue degeneration and clinical practice.

Aggrecans↗
PubMed1999

[Relationship between smoking and drinking and herniation of intervertebral disc].

To study whether there is relationship between smoking and drinking and herniation of intervertebral disc, we investigated 214 patients who had been proved with herniation of intervertebral disk during 1986-1996. The results were that in the younger (< or = 35 year old), no matter whether they drank or not, the number of smoker in the patients with herniation of intervertebral disc was more than that in the control group; no matter whether they smoked or not, the number of drinker in the patients with herniation of intervertebral disc was more than that in the control group.

Adolescent↗
PubMed1994

The dose-related effect of intradiscal chymopapain on rabbit intervertebral discs.

STUDY DESIGN: This study analyzed the histological and biochemical responses of intervertebral disc tissue to intradiscal injection of varying amounts of chymopapain. OBJECTIVE: To determine the appropriate amount of chymopapain needed to accomplish effective degradation of proteoglycans (PG) in the nucleus pulposus of intervertebral discs. SUMMARY OF BACKGROUND DATA: Chymopapain is an accepted treatment alternative for patients with disc herniations. The recommended clinical dose of 2,000-4,000 pKats per injection is derived from early animal studies and empirical results in man. A lower effective dose could reduce the complication rate while providing similar clinical results. METHODS: Twenty to 4,000 pKat of chymopapain was injected into rabbit discs, and the level of keratan sulfate (KS) epitope in serum was measured at different times after the injection. The animals were killed after 6 days and the injected and two neighboring discs were examined histologically. RESULTS: The serum KS level did not change appreciably after injection of 20 pKat, rose moderately at 100 and 200 pKat, and rose strongly at 500 pKat. Doses greater than 500 pKat did not result in further increase in the KS level. CONCLUSION: Degradation of the disc proteoglycans is dose dependent and reaches a maximum at 500 pKat. Higher doses appear not to cause further loss of aggrecan molecules, and injection of more than 1,000 pKat produces significant annular destruction.

Animals↗
PubMed1979

[Relationships between the height of intervertebral discs and the degree of lordosis of the cervical spine in children. A roentgenometric study (author's transl)].

On lateral roentgenograms of the cervical spine of 120 normal children between 3 and 14 years of age the ventral and dorsal height of all 5 intervertebral discs were measured and the "index of the intervertebral disc" was calculated. In addition, the "degree of lordosis" of the cervical spine was determined according to ISHIHARA (1964). Evaluation was in 3 age groups: 3 to 6, 7 to 10, 11 to 14 years; and the arithmetic mean and standard deviation were calculated for each group. All intervertebral discs are more or less "cuneiform" (ventrally higher than dorsally); they narrow from cranial to caudal. In children the degree of lordosis of the cervical spine shows a continuous decrease with advancing years. In all age groups girls show a lower degree of lordosis than boys. Statistically the differences of average values of the intervertebral-disc-index among the various shapes of the cervical spine in neutral position (straight, sublordotic, lordotic and hyperlordotic) are highly significant. By means of roentgenometric methods our examinations demonstrate that the lordotic curvature of the healthy cervical spine primarily depends on the cuneiform shape of intervertebral discs.

Adolescent↗
PubMed1994

Tissue distribution of antibiotics in the intervertebral disc.

STUDY DESIGN: This study analyzed the distribution of antibiotics within the intervertebral disc of rabbits. Specimens were tested with specific antibodies against antibiotics using an immunofluorescent technique. OBJECTIVES: The results were correlated to provide a rationale for perioperative prophylaxis of infection. SUMMARY OF BACKGROUND DATA: Several groups of investigators and the recent data from our laboratory showed quantitative changes in penetration of antibiotics into intervertebral disc. No previous study has assessed antibiotic distribution in anulus fibrosus and nucleus pulposus. METHODS: Discs were obtained from rabbits after intravenous injection of penicillin or gentamicin. Antibiotics were localized in tissue sections using specific antibodies with a immunofluorescent method. RESULTS: Penicillin (negatively charged) and gentamicin (positively charged) penetrated the neutrally charged anulus fibrosus, but penicillin had less ability than gentamicin to penetrate into the negatively charged nucleus pulposus. CONCLUSION: Our data suggest that penetration and distribution of antibiotics into avascular intervertebral disc is significantly influenced by the charge of antibiotics.

Animals↗
PubMed2002

Human intervertebral disc aggrecan inhibits nerve growth in vitro.

OBJECTIVE: To assess the effects of human intervertebral disc aggrecan on nerve growth and guidance, using in vitro techniques. METHODS: Aggrecan extracted from human lumbar intervertebral discs was incorporated into tissue culture substrata for the culture of the human neuronal cell line, SH-SY5Y, or explants of chick dorsal root ganglia. The effects on nerve growth of different concentrations of aggrecan extracted from the anulus fibrosus and nucleus pulposus, and of these aggrecan preparations following enzymic deglycosylation, were compared. RESULTS: Disc aggrecan inhibited the growth of neurites from SH-SY5Y cells and induced growth cone turning of chick sensory neurites in a concentration-dependent manner. Aggrecan isolated from the anulus fibrosus was more inhibitory than that isolated from the nucleus pulposus, but enzymic pretreatments to reduce the glycosylation of both types of disc aggrecan partially abrogated their inhibitory effects. CONCLUSION: Nerve growth into degenerate intervertebral discs has been linked with the development of low back pain, but little is known about factors affecting disc innervation. The finding that disc aggrecan inhibits nerve growth in vitro, and that this inhibitory activity depends on aggrecan glycosylation, has important implications for our understanding of mechanisms that may regulate disc innervation in health and disease.

Aggrecans↗
PubMed2006

Effects of growth differentiation factor-5 on the intervertebral disc--in vitro bovine study and in vivo rabbit disc degeneration model study.

STUDY DESIGN: In vitro studies on the effects of recombinant human growth and differentiation factor-5 (rhGDF-5) on matrix metabolism of bovine intervertebral disc cells and an in vivo study on the effect of rhGDF-5 in the rabbit anular puncture model. OBJECTIVE: To determine the reparative capacity of rhGDF-5 on the intervertebral disc. SUMMARY OF BACKGROUND DATA: The in vitro and in vivo effects of rhGDF-5, a crucial protein in the developing musculoskeletal system, on repair of the degenerated intervertebral disc remain unidentified. METHODS: In vitro, bovine nucleus pulposus and anulus fibrosus cells were cultured with or without rhGDF-5 (100 or 200 ng/mL). On days 7, 14, and 21, the contents of deoxyribonucleic acid and proteoglycan, and the synthesis of proteoglycan and collagen were assessed. In vivo, 16 adolescent New Zealand white rabbits received anular punctures in 2 lumbar discs. Four weeks later, phosphate buffered saline or rhGDF-5 (10 ng, 1 and 100 mug) was injected into the nucleus pulposus. The rabbits were followed up for 16 weeks for disc height, magnetic resonance imaging, and histologic grading. RESULTS: In vitro, rhGDF-5 increased the deoxyribonucleic acid and proteoglycan contents of both cell types significantly after day 14. rhGDF-5 at 200 ng/mL significantly stimulated proteoglycan synthesis (nucleus pulposus: +138%, anulus fibrosus: +24%) and collagen synthesis (nucleus pulposus: +95%, anulus fibrosus: +23%) at day 21. In vivo, the injection of rhGDF-5 resulted in a restoration of disc height, improvement of magnetic resonance imaging scores, and histologic grading scores with statistical significance (P < 0.05-0.001). CONCLUSION: A single injection of rhGDF-5 has a reparative capacity on intervertebral discs, presumably based on its effects to enhance extracellular matrix production in vitro.

Animals↗
PubMed1998

Relative increase of biglycan and decorin and altered chondroitin sulfate epitopes in the degenerating human intervertebral disc.

OBJECTIVE: Proteoglycans are major components of the extracellular matrix of the intervertebral disc. They are vital for the biomechanical properties of the tissue, and are subject to changes in disc degeneration. We aimed to further define these changes and their relationship to normal aging. METHODS: Normal discs (age 13-53 years, n = 6) were analyzed from 5 different sites across the sagittal anterior-posterior direction. Degenerated anterior annulus fibrosus was collected from 7 patients aged 39-46 years. Extracted proteoglycans were separated using agarose and polyacrylamide gel electrophoresis and detected with toluidine blue staining and Western blotting. RESULTS: The center of the disc showed the highest level of total proteoglycans, but lowest levels of decorin and biglycan. Western blots displayed reduced signal for both glycanated and nonglycanated biglycan and decorin after adolescence, while an increased signal of biglycan was observed in degenerated annuli. The 7D4(-) and 3B3(-) epitopes on native chondroitin sulfate chains were present in the large proteoglycans of intervertebral discs, but their signal intensity had no correlation to degeneration. Chondroitinase ABC digestion of the blots brought up 7D4(+) signal in the small proteoglycans of degenerated, but not in healthy tissue. Decrease or total loss of 2B6(+) epitope (indicating 4-sulfated stubs of chondroitin sulfate chains) were found in the large proteoglycans of all degenerated annuli. CONCLUSION: Human intervertebral disc degeneration involves the accumulation of decorin and biglycan relative to other uronic acid containing proteoglycans, the disappearance of 4-sulfated core region in aggrecan-like large proteoglycans, and the emergence of a core structure in the chains of small proteoglycans reacting with the 7D4 antibody; these findings indicate a fundamental alteration in matrix properties that may contribute to the pathogenesis of the disease.

Adolescent↗
PubMed2001

Effect of frozen storage on the creep behavior of human intervertebral discs.

STUDY DESIGN: A biomechanical study of the compressive creep behavior of the human intervertebral disc before and after frozen storage. OBJECTIVES: To determine whether frozen storage alters the time-dependent response of the intact human intervertebral disc. SUMMARY OF BACKGROUND DATA: The biomechanical properties of the intervertebral disc are generally determined using specimens that have been previously frozen. Although it is well established that freezing does not alter the elastic response of the disc, recent data demonstrate that freezing permanently alters the time-dependent mechanical behavior of porcine discs. METHODS: Twenty lumbar motion segments from 10 human spines were harvested between 12 and 36 hours postmortem. The specimens were randomly assigned to one of two groups: Group 1 was tested promptly, stored frozen for 3 weeks, then thawed, and tested a second time; Group 2 was stored frozen for 3 weeks, thawed, and then tested. Each specimen was subjected to 5 cycles of compressive creep under 1 MPa for 20 minutes, followed by a 40-minute recovery under no load. After testing each specimen was graded on a degeneration scale. A fluid transport model was used to parameterize the creep data. RESULTS: There was no statistically significant effect of freezing on the elastic or creep response of the discs. The degree of pre-existing degeneration had a significant effect on the creep response, with the more degenerated discs appearing more permeable. CONCLUSIONS: Frozen storage for a reasonable time with a typical method does not significantly alter the creep response of human lumbar discs. Freezing may produce subtle effects, but these potential artifacts do not appear to alter the discs' time-dependent behavior in any consequential way. These results may not apply to tissue kept frozen for long durations and with poor packaging.

Artifacts↗