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PubMed1987

Surface strain on human intervertebral discs.

The biomechanical functions of the internal components of the intervertebral disc are not well understood. The surface deformation of 17 human cadaveric lumbar intervertebral discs was studied by photogrammetry by adhering small optical targets to the disc surface and thereby recording the length, bulge, and vertical height of lines on the disc surface representing annular fibers. Discs were studied in pure compression, flexion and extension, axial rotation, and shear. Two definitions of a fiber were investigated: first with the end-points of the fiber on the vertebra ("bone-to-bone" definition), second, where the end points of the fiber were just before the disc vertebra junction (the "disc-only" definition). Measurements were compared with a "constant-volume" physical model and with a mathematical model of the intervertebral disc. Fiber strains were 6% or less under physiological conditions. Comparison of results from the two definitions of fiber length showed greater strains for the disc-only definition in compressive loading. Fiber strains were less than in the constant-volume model of comparable dimensions in compressive loading by a factor of about two, thus suggesting fluid loss or end-plate deformations in the physiologic conditions. The mathematical model indicated that the surface strain for intervertebral discs is very sensitive to the disc-height: diameter ratio and to fluid loss from the disc but is less sensitive to the helix angle of the fibers.

Adult↗
PubMed2006

Efficient and stable gene expression in rabbit intervertebral disc cells transduced with a recombinant baculovirus vector.

STUDY DESIGN: A recombinant baculovirus vector expressing GFP (Ac-CMV-GFP) was tested in rabbit intervertebral disc cells cultured in monolayer in vitro. Direct infection of intervertebral disc cells was then assessed in vivo. OBJECTIVES: To test the efficacy of the baculovirus-mediated marked gene transfer to rabbit intervertebral disc cells, and the expression of the transgene in vitro and in vivo. SUMMARY OF BACKGROUND DATA: The potential application of gene therapy for the treatment of degenerative disc disease has been an area of exciting and active research. Many efforts have been made toward the construction of delivery vectors using mammalian viruses, such as adenovirus, adeno-associated virus, retrovirus, and herpesvirus. However, recently, baculovirus-derived vectors have emerged as possible tools for gene transfer into mammalian cells in vitro and in vivo but, to our knowledge, have never been tested in an intervertebral disc cell. METHODS: The intervertebral disc cells cultured in monolayer were treated with 6 different doses of baculovirus carrying the green fluorescence protein gene (Ac-CMV-GFP). Fluorescence microscopy and flow cytometry were used to analyze transgene expression. The Autographa californica nucleopolyhedrovirus/GFP virus was then injected directly into the intervertebral discs of 8 rabbits at 7, 13, 20, and 28 days after injection. The nucleus pulposus tissues of injected discs were evaluated immediately by fluorescence microscopy for GFP expression. RESULTS: A dose of Ac-CMV-GFP at a multiplicity of infection of 200 achieved the highest transduction ratio (approximately 87% of nucleus pulposus cells) and long-term expression without any toxicity to the cells. In vivo assay showed that Ac-CMV-GFP could also mediate GFP expression in rabbit intervertebral disc cells without inducing any symptoms. The GFP expression level at 7 days after transduction was significantly higher than at 21 and 28 days after treatment. CONCLUSIONS: Baculovirus can transfer exogenous genes into rabbit nucleus pulposus cells at a high efficiency, and safely both in vitro and in vivo. The results suggest that baculoviruses might be useful tools as gene therapy vectors for intervertebral disc diseases.

Animals↗
PubMed2001

Magnetic resonance imaging measurement of relaxation and water diffusion in the human lumbar intervertebral disc under compression in vitro.

STUDY DESIGN: Twelve lumbar intervertebral disc specimens were imaged with magnetic resonance imaging to estimate relaxation constants, T1 and T2, and tissue water diffusion, before and after applying compression. OBJECTIVES: The objectives of the study were to measure T1, T2, and water diffusion for differences with loading state, region of the disc (anulus fibrosus or nucleus pulposus), and grade of degeneration. SUMMARY OF BACKGROUND DATA: Magnetic resonance imaging can be used qualitatively to estimate water content and degeneration of the intervertebral disc. Beyond structural information of images, the relaxation times T1 and T2 may contain information on the changes occurring with degeneration. A modified spin-echo sequence can be used to estimate tissue water diffusion in cartilage and disc specimens with the ability to measure anisotropy. METHODS: Specimens were imaged in a 1.5-Tesla clinical scanner. T1, T2, and water diffusion were estimated from midsagittal images. Magnetic resonance imaging parameters were calculated before and after axial loading. The measured T1, T2, and D (diffusion coefficient) were compared before and after compression, and for the diffusion data, also by direction to consider anisotropy. RESULTS: For the T1 data, a significant difference was found by region, nucleus > anulus, and loading state, loaded > unloaded. For the T2 values, there was a significant difference by region, nucleus > anulus, and Thompson grade. For diffusion, significant differences were found by region, nucleus > anulus, Thompson grade, direction of diffusion, and state of compression, loaded > unloaded. CONCLUSIONS: This study demonstrated that magnetic resonance imaging can be used to measure significant changes in T1, T2, or diffusion in intervertebral disc specimens by region, loading condition, or Thompson grade.

Adolescent↗
PubMed2004

[Characterization of intervertebral disc--disc cells and pericellular microenvironment].

The intervertebral disc is classified as fibrocartilage, and has distinct functional and mechanical properties. The disc, composed of the nucleus pulposus and annulus fibrosus, varies in density, anatomic features, and in the morphology of their constituent cells. This heterogeneity has hampered detailed investigations of disc cellular metabolism. Focusing on the heterogeneity, we cultured disc cells under various conditions to examine the cellular metabolism, specifically glycosaminoglycan accumulation and pericellular microenvironment, and succeeded in mechanically isolating disc chondrons from annulus fibrosus. We believe that disc chondron also plays an important role in maintaining cartilageous phenotype in terms of cell-matrix interaction of the intervertebral disc.

Cell Culture Techniques↗
PubMed1989

[Acute rupture of lumbar intervertebral disc caused by violent manipulation].

Five cases of acute rupture of lumbar intervertebral disc caused by violent manipulation are reported. After protrusion of the lumbar intervertebral disc were treated by violent manipulation, the lumbo-leg pain were severe suddenly. The operations found that the annulus fibrosus had ruptured and the nerve root or cauda equina was constricted by the nucleus pulposus which had entered into the spinal canal. It must be emphasized that their is in danger of more damaging intervertebral disc degenerated by violent manipulation, then the adhesion of the nerve root will occur gradually. We don't suggest to make violent manipulation for the patient with the nerve root injured. We have acquired good results in treating protrusion of lumbar intervertebral disc by combined therapy and they are introduced in this article.

Adult↗
PubMed2004

[Pathological development of researches on intervertebral disc degeneration].

This article reviews the literature about the etiology and pathology of intervertebral disc degeneration. The degeneration of intervertebral disc is associated with certain biomechanical factor. Stress can directly lead to the rupture of intervertebral disc and, more importantly, change its biological properties. And then, it can trigger the process of degeneration via inflammatory mechanism which may include autoimmunity.

Biomechanical Phenomena↗
PubMed1982

The mechanics of prolapsed intervertebral disc.

Cadaveric lumbar intervertebral joints were subjected to compressive loads while wedged in forward flexion. In some specimens disc lesions were produced and we have been able to identify three different types: postero-lateral radial fissure, annular protrusion and nuclear extrusion. The type of lesion depended on the loading regime but all occurred more readily in slightly degenerated discs.

Adult↗
PubMed2003

Correlation between nucleus zone migration within scoliotic intervertebral discs and mechanical properties distribution within scoliotic vertebrae.

Correlations between intervertebral disc degeneration and bone mass were investigated previously, but never on scoliotic patients. Using MRI measurements of intervertebral discs behavior and vertebral bone tomodensitometry, correlations between nucleus zone displacement within intervertebral discs and mechanical center migration within vertebral bodies were investigated in vivo on scoliotic patients. The protocol, performed on eleven scoliotic girls, was composed of a CT scan acquisition of apical and adjacent vertebrae followed by a MRI acquisition of the thoracolumbar spine. The displacement between the vertebral body centroid and inertia center was computed from the CT images and called the mechanical migration. The displacement between nucleus zones and vertebral body centroids was quantified from MRI and called the nucleus zone migration. For apical vertebrae, a significant correlation was found in the coronal plane (r = 0.766, p < 0.01), but not in the sagittal plane (r = -0.349, p > 0.05). For adjacent vertebrae, significant correlations were found in both coronal (r = -0.633, p < 0.05) and sagittal (r = -0.797, p < 0.01) planes. The nucleus zone migration occurred in the convexity of the curvature whereas the mechanical migration occurred in the concavity.Known secondary mechanical phenomenon of scoliosis was quantified using new parameters describing intervertebral discs and vertebral bodies. Further investigations should be performed to explain the mechanical evolution of scoliosis and to use these parameters in predictive criteria of scoliosis.

Bone Density↗
PubMed2006

Substance P-saporin down-regulates substance P receptor immunoreactive sensory dorsal root ganglion neurons innervating the lumbar intervertebral discs in rats.

STUDY DESIGN: To examine changes in substance P receptors on dorsal root ganglion cells innervating the rat lumbar intervertebral discs using immunohistochemistry and a retrograde neurotracing method. OBJECTIVE: We evaluated the effects of intradiscal administration of substance P-saporin, a toxin selective for cells expressing substance P receptors. SUMMARY OF BACKGROUND DATA: The rat L5/6 intervertebral disc is multi-segmentally innervated from the L1-L6 dorsal root ganglions. Substance P and the neurokinin-1 receptor contribute to inflammatory pain transmission. Substance P immunoreactive-sensory nerve fibers in human intervertebral discs and immunoreactive-dorsal root ganglion neurons innervating rat intervertebral discs have been reported to be important in the transmission of discogenic low back pain. In the current study, we evaluated the effects of intradiscal administration of substance P-saporin, a toxin selective for cells expressing substance P receptor. METHODS: Sixteen rats were used (control group, n = 8; substance P-saporin group, n = 8). To detect dorsal root ganglion neurons innervating the L5/6 intervertebral disc, neurotracer (fluoro-gold crystals) was placed into the intervertebral disc. Seven days after fluoro-gold application, the L5/6 intervertebral disc was exposed and injected with 175 ng of sterile substance P-saporin (substance P-saporin group, n = 8). Fourteen days after the first operation, each dorsal root ganglion was harvested, sectioned, and processed for neurokinin-1 immunohistochemistry using rabbit antibody to neurokinin-1. The numbers of fluoro-gold labeled neurons, and fluoro-gold labeled and neurokinin-1 immunoreactive neurons were counted in both groups. RESULTS: Neurons innervating the L5/6 intervertebral discs, retrogradely labeled with fluoro-gold, were distributed throughout dorsal root ganglions from L1 to L6 in both groups. Of fluoro-gold labeled neurons, the proportion of neurokinin-1 immunoreactive neurons was 35% in the control group. However, the proportion of neurokinin-1 immunoreactive neurons was 8% after administration of substance P-saporin into the intervertebral discs (substance P-saporin group). Substance P-saporin significantly decreased the ratio of neurokinin-1 immunoreactive neurons. CONCLUSION: Substance P-saporin decreased the ratio of neurokinin-1 immunoreactive neurons innervating the disc related to discogenic low back pain. Substance P-saporin may be a useful tool to investigate the mechanism of discogenic low back pain.

Animals↗
PubMed2006

An in vitro organ culturing system for intervertebral disc explants with vertebral endplates: a feasibility study with ovine caudal discs.

STUDY DESIGN: Whole ovine caudal intervertebral discs with vertebral endplates were cultured under uniaxial diurnal loading for 7 days. OBJECTIVES: To establish and characterize an organ culture system for intervertebral discs, in which disc cells may be "maintained" in their native three-dimensional environment under load. SUMMARY OF BACKGROUND DATA: In vitro culturing of entire discs with preserved biologic and structural integrity would be a useful model to study the effects of nutrition and mechanical loading. METHODS: To maintain endplate permeability, sheep were systemically anticoagulated before death and their caudal vasculature was evacuated with saline postmortem. The first 4 caudal discs were explanted with their adjacent endplates and cultured in bioreactors under uniaxial diurnal loading (0.2 MPa for 8 hours and 0.8 MPa for 16 hours) for 4 or 7 days. Solute transport into the center of the disc was measured after 4 days of culture using a low molecular weight fluorescent marker. Cell viability, glycosaminoglycan synthesis rate, and gene expression profile were measured after 7 days of culture and compared with fresh tissue. RESULTS: Fluorescent images showed that solutes could diffuse into the disc under both static and diurnal loading, but penetration through the endplate increased with diurnal loading. Cell viability and glycosaminoglycan synthesis rates remained unchanged after 7 days of culture. Expression of catabolic genes was significantly up-regulated, whereas anabolic genes tended to be down-regulated after 7 days. CONCLUSIONS: With this novel preparation and culturing technique, endplate permeability could be maintained, which allowed culturing of intact disc explants with endplates for up to 7 days.

Animals↗
PubMed2001

Intradiscal solid phase displacement as a determinant of the centripetal fluid shift in the loaded intervertebral disc.

STUDY DESIGN: The movement of cross sections of the monofilament nylon threads inserted into the axially loaded intervertebral disc was traced with magnetic resonance imaging (MRI). This technique allowed the observation of the sequential solid phase displacement of the loaded intervertebral disc. OBJECTIVES: To clarify sequential solid phase displacement of the axially loaded intervertebral disc to elucidate the cause of centripetal fluid shift within a disc. SUMMARY OF BACKGROUND DATA: We already have reported that there is a centripetal fluid shift within the axially loaded intervertebral disc during the early phase of loading. We assumed that there should be an elaborate intradiscal matrix displacement that generates a pressure gradient within the disc to cause a centripetal fluid shift. METHODS: Thirteen freshly obtained bovine caudal intervertebral discs were prepared. Three to five monofilament nylon threads were inserted into each disc in the anterior-posterior direction to trace the intradiscal solid phase displacement on the midcoronal MR images. Sequential displacement of the disc matrix was recorded during a 294 N axial loading. RESULTS: Relatively large centrifugal expansion at the inner layer of the anulus fibrosus compared with less centrifugal expansion of the outer anulus fibrosus was observed in accord with gradual creep of the disc thickness. CONCLUSIONS: The uneven displacement of the intradiscal solid phase observed in the present study expels the fluid phase from the inner anulus fibrosus, thus resulting in accumulation of fluid phase in the nucleus pulposus. The present study suggests the presence of a mechanism that retains water within the normal intervertebral disc, in spite of an external load, because it forms a water-abundant nucleus pulposus, which is surrounded by an anulus fibrosus with decreased water permeability caused by fluid loss. A more detailed analysis is required to clarify topographic volumetric changes within the disc.

Animals↗
PubMed2004

Ad/CMV- hTGF-beta1 treats rabbit intervertebral discs degeneration in vivo.

To investigate therapeutic efficiency of Ad/CMV- hTGF-beta1 gene for rabbit intervertebral disc degeneration model. 60 Japanese white rabbits were selected to form the 1.5-L6 Anterior-Lateral-Anulus-Fibrosus-Incision-Induced model in order to simulate human intervertebral disc degeneration. 36 rabbits, whose corresponding intervertebral discs were injected with 20 microl (10 x 10(6) pfu) of Ad/CMV- hTGF-beta1 gene, constituted the therapy group, 12 were injected with 20 microl (10 x 10(6) pfu)of Ad/CMV-LacZ gene as comparison group, while 12 were only injected with equivalent capacity of saline for empty comparison group, 3 weeks after injection, examples were taken for investigation of HE staining, MRI, Western Blotting and immunohistochemical research TGF-beta1. Wide distribution of TGF-beta1 was detected by immunohistochemical research in the degenerated annulus fibrosus after injection. Western Blotting research showed significant increase of TGF-beta1 content in intervertebral discs treated with TGF-beta1 gene than comparison groups. MRI signal transformed from low to comparatively high and that intervertebral disc pathological degree improved. Ad/CMV- hTGF-beta1 gene transfection is a potential method to increase TGF-beta1 content and reverse intervertebral disc degeneration.

Adenoviridae↗
PubMed1986

Secretory cells in the nucleus pulposus of the adult human intervertebral disc. A preliminary report.

A light microscopical study was conducted to ascertain the type of cells in the nucleus pulposus of the adult human intervertrebral disc. Three lumbar intervertebral discs were removed from each of 15 male and female adults at autopsy (ages ranged from 19 to 62 years). The tissue was fixed in formalin, decalcified in formic acid, dehydrated in a graded series of ethanol, embedded in paraffin, and serially sectioned at 7-10 micron. Tissue sections were affixed to albuminized glass slides and stained either by hematoxylin and eosin or hematoxylin and Van Gieson's stain. The cells of the bulk of the nucleus pulposus consisted of chondrocytes and a few fibroblasts; however, the subchondral matrix of the nucleus pulposos contained numerous stellate cells with (from 1 to 8) unusually long (up to 80 micron) primary cytoplasmic processes that often branch into secondary processes. The cell processes contained cytoplasmic varicosities at various intervals along their lengths; and their endings often expanded into bulbous, vesicle-filled process terminals. The surrounding extracellular matrix usually contained numerous, vesicle-filled, eosinophil matrix bodies. Morphological similarities of cytoplasmic varicosities, process terminals, and matrix bodies, as well as the apparent budding of process terminals, suggest that these previously unidentified cells are secreting an unknown matrix component into the subchondral matrix of the nucleus pulposus of the adult human.

Adult↗
PubMed1989

Water diffusion pathway, swelling pressure, and biomechanical properties of the intervertebral disc during compression load.

The behavior of water in the intervertebral disc of pig tail and its physiologic and biomechanical properties were investigated in relation to compression load. The water content, chemical composition, and swelling pressure in the intervertebral disc were measured, and the mechanism of the generation of the swelling pressure in relation to compression load stress was studied. The swelling pressure, through regulation of the water content of the disc and the resistance of the external load, differs with the region of the intervertebral disc. In the nucleus pulposus and the inner layer of the anulus fibrosus, the swelling pressure rises in proportion to the load, but few changes occur in the outer layer of the anulus fibrosus, and the constant pressure environment is thus maintained. The tritiated water (3H2O) uptake of the disc under various loads was measured. The molar partition coefficient of tritiated water is almost equal to 1 even under a compression load, which suggests that water is freely exchangeable. The diffusion of 3H2O in the intervertebral disc was traced using two pathway models: the perianular route and the end-plate route. The diffusion of water in the unloaded disc for both uptake and washout was about 2 to 3 times larger in the perianular route than in the end-plate route. Under load, the water diffusion was inhibited in both pathways. The relation between the load and displacement revealed viscoelastic properties indicating creep and stress relaxation. Young's modulus and the stiffness increased with a rise in load speed.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1999

Phenotypic characteristics of rabbit intervertebral disc cells. Comparison with cartilage cells from the same animals.

STUDY DESIGN: Intervertebral disc cells were extracted from the surrounding matrix, and their metabolic activities and phenotypes were studied. OBJECTIVES: To compare the metabolic activities and phenotypes of cell populations extracted from the intervertebral discs of young rabbits with those of articular and growth plate chondrocytes from the same animals. SUMMARY OF BACKGROUND DATA: The phenotype of intervertebral disc cells has been poorly studied and still is debated. METHODS: The intervertebral discs as well as articular and vertebral growth plate cartilage of rabbits were digested enzymatically. The morphology of freshly isolated cells was examined. Their contents of collagen II and X mRNAs were determined by Northern blot analysis, and their sulfation activity by 35S-sulfate incorporation as chondrocytic markers. Cells were cultured at high density or low density and grown in primary culture. The stability of their phenotype was monitored by evaluating the collagen I and II mRNA ratio. The proteoglycans newly synthesized by the cells also were quantified, and their elution profile analyzed on Sepharose 2B columns. RESULTS: The anulus fibrosus cells were morphologically undistinguishable from articular chondrocytes. The nucleus pulposus contained mainly large vacuolated cells and a few smaller cells. All freshly extracted cells expressed different levels of collagen II mRNA. Anulus fibrosus and nucleus pulposus cells contained, respectively, 22% and 8% of collagen II mRNA compared with that found in articular or growth plate chondrocytes from the same animal. Only growth plate chondrocytes expressed collagen X. When anulus fibrosus cells were incubated for 48 hours at high density, they had collagen II mRNA contents similar to those of articular and growth plate chondrocytes, but synthesized five to six times fewer sulfated proteoglycans. When seeded at low density, anulus fibrosus cells divided more slowly than articular chondrocytes and incorporated four times fewer 35S-sulfate into proteoglycans. Their collagen II mRNA content was 2.75-fold lower than that of chondrocytes, and the procollagen alpha 1II/alpha 1I mRNA ratio was 3.1 for anulus fibrosus cells and 7 for chondrocytes. No collagen X mRNA was detected. When incubated for 48 hours at high density, the nucleus pulposus giant cells had four times less collagen II mRNA content than cartilage cells but synthesized the same amounts of sulfated proteoglycans. They did not divide during 21 days in culture and still contained collagen II mRNA but no collagen X mRNA. CONCLUSIONS: Findings showed that intervertebral disc cells all express cartilage-specific matrix proteins with quantitative differences, depending on their anatomic situation. It is suggested that anulus fibrosus cells are chondrocytic cells at a different stage of differentiation than articular and growth plate chondrocytes. The phenotype of nucleus pulposus cells still is unclear. They could be chondrocytic or notochordal. A definitive answer to this important question requires differentiating markers of notochordal cells.

Animals↗
PubMed2001

Collagen gene expression and mechanical properties of intervertebral disc cell-alginate cultures.

Cells of the intervertebral disc have a limited capacity for matrix repair that may contribute to the onset and progression of degenerative disc changes. In this study, the biosynthetic capacity of cells isolated from specific regions of the porcine intervertebral disc was evaluated in vitro. Using a competitive reverse transcription-polymerase chain reaction technique, gene expression levels for types I and II collagen were quantified in cells cultured for up to 21 d in a three-dimensional alginate culture system and compared to levels obtained for cells in vivo. The mechanical properties of cell-alginate constructs were measured in compression and shear after periods of culture up to 16 weeks. Cells from the anulus fibrosus expressed the most type I collagen mRNA in vivo and in vitro, while cells from the transition zone expressed the most type II collagen mRNA in vivo and in vitro. Mechanical testing results indicate that a mechanically functional matrix did not form at any time during the culture period; rather, decreases of up to 50% were observed in the compressive and shear moduli of the cell-alginate constructs compared to alginate with no cells. Together with results of prior studies, these results suggest that intervertebral disc cells maintain characteristics of their phenotype when cultured in alginate, but the molecules they synthesize are not able to form a mechanically functional matrix in vitro.

Alginates↗
PubMedDate not supplied

1980 Volvo award in basic science. Proteoglycans in experimental intervertebral disc degeneration.

An animal model of intervertebral disc degeneration induced surgically by ventral nuclear herniation in the rabbit produces morphologic changes of disc degeneration. Histologic characteristics and proteoglycan changes have been studied at various times after herniation. After injury, there was metaplasia into fibrocartilage originating from the cells along the margins of the annular wound, with proliferation of cells changing almost the entire disc space into fibrocartilage. A vertebral osteophyte occurred through an endochondral ossification sequence. Aggregating proteoglycans had two periods of repletion in the early course of degeneration. The water content of the disc was rapidly but only transiently restored in the first two days after herniation, whilst the changes in the total proteoglycan content of the disc paralleled these changes. Hyaluronic acid content decreased rapidly after herniation, but the size of the proteoglycan monomers did not change with degeneration. It is suggested that loss of confined fluid mechanics signals an abortive repair attempt rather than that of biochemical changes in proteoglycans initiate disc degeneration.

Animals↗
PubMed2003

[Growth factor and regeneration of intervertebral disc].

OBJECTIVE: To review research progress of the relation between growth factor and repair of intervertebral disc. METHODS: The recent articles on growth factor and repair of intervertebral disc were extensively reviewed. The expression of growth factor in intervertebral disc and the effect of growth factor on disc cells were investigated. RESULTS: Some growth factors play roles in the development and degeneration of intervertebral disc. Exogenous growth factor can increase proliferation of disc cells and production of proteoglycans and collagens. Gene of growth factor can be transferred to intervertebral disc cell by adenovirus. CONCLUSION: Growth factor plays an important role in the regulation of development and degeneration of interertebral disc. The above results show that the feasibility of usage of growth factor in the treatment of disc degeneration and in repair and reconstruction of disc.

Cell Division↗