Study Design. Study of regional variations in composition in a sample of 9 mildly to moderately degenerated human intervertebral discs.Objective. The aim of this study was to obtain proteoglycan distribution in human lumbar discs with high position resolution in the: 1) sagittal, 2) coronal, and 3) axial directions.Summary of Background Data. Regional variation in disc proteoglycan content has only been reported in coronal sections in a small number of discs and with low spatial resolution in the sagittal direction, and has not been reported in the axial direction.Methods. Each of 9 human L2-L3 or L3-L4 lumbar discs (age, 53-56 years) were dissected into 36 to 41 specimens using a rectangular cutting die, measured for water content and analyzed for glycosaminoglycan content using the dimethylmethylene blue dye binding assay.Results. The intervertebral discs were mildly to moderately degenerated. They had glycosaminoglycan content ranging ϳ40 to 600 g/mg dry tissue, with largest values in the nucleus and lowest in the outer anulus. In general, posterior regions had greater glycosaminoglycan content than anterior regions, although values were not as high as in the nucleus. Small variations in glycosaminoglycan content in the axial direction were observed with the largest values in the center, although this variation was small compared with radial variations. Water content results followed similar trends as glycosaminoglycan content with average values ranging from ϳ66% to 86%.Conclusions. A refined map of proteoglycan content is presented with 3 important findings. First, sagittal variations were distinct from coronal variations. Second, the proteoglycan content was not uniform across the nucleus regions. Third, some specimens had localized variations in proteoglycan and water contents suggestive of focal damage and degeneration.Key words: proteoglycans, aggrecan, glycosaminoglycan content, fixed charge density, intervertebral disc, degeneration. Spine 2007;32: [1493][1494][1495][1496][1497] The distribution of proteoglycan content in the intervertebral disc is functionally important in defining the swelling behaviors, streaming potential, and compressive properties of the tissue.1-4 A loss of proteoglycans in the central nucleus pulposus region is a clear sign of early degeneration. 5,6 The tissue mechanical properties depend on proteoglycan content; therefore, this compositional information is required for mechanical modeling of the disc 1,7-10 and offers promise for improved understanding of disc function and remodeling in healthy and diseased states.Previous reports of proteoglycan content distribution in the disc have provided relatively coarse information on the distribution in the sagittal (front to back) direction on a large number of specimens using sulfated glycosaminoglycan (GAG) measurements 6 or with a higher spatial distribution from a relatively small number of discs using radioactive ion tracers to obtain direct measurements of fixed charge density. 11 The proteoglycan distributions in the coronal ...

This study evaluated how dynamic compression induced changes in gene expression, tissue composition, and structural properties of the intervertebral disc using a rat tail model. We hypothesized that daily exposure to dynamic compression for short durations would result in anabolic remodeling with increased matrix protein expression and proteoglycan content, and that increased daily load exposure time and experiment duration would retain these changes but also accumulate changes representative of mild degeneration. Sprague-Dawley rats (n ¼ 100) were instrumented with an Ilizarov-type device and divided into three dynamic compression (2 week-1.5 h/day, 2 week-8 h/day, 8 week-8 h/day at 1 MPa and 1 Hz) and two sham (2 week, 8 week) groups. Dynamic compression resulted in anabolic remodeling with increased matrix mRNA expression, minimal changes in catabolic genes or disc structure and stiffness, and increased glysosaminoglycans (GAG) content in the nucleus pulposus. Some accumulation of mild degeneration with 8 week-8 h included loss of annulus fibrosus GAG and disc height although 8-week shams also had loss of disc height, water content, and minor structural alterations. We conclude that dynamic compression is consistent with a notion of ''healthy'' loading that is able to maintain or promote matrix biosynthesis without substantially disrupting disc structural integrity. A slow accumulation of changes similar to human disc degeneration occurred when dynamic compression was applied for excessive durations, but this degenerative shift was mild when compared to static compression, bending, or other interventions that create greater structural disruption. Keywords: intervertebral disc; disc degeneration; mechanobiology; animal model; biomechanics Substantial socioeconomic problems that result from low back pain are often associated with intervertebral disc (IVD) degeneration.1 The etiology of disc degeneration is complex and multifactorial with heredity, mechanical loading, and nutrition all playing significant roles.2-7 These contributing factors are interactive and strongly affected by aging. For example, degradation of the molecular structure of the disc during aging can also render it more susceptible to mechanical injuries. 4 Loading type, magnitude, duration, and frequency all influence cell metabolic responses and matrix remodeling. 5,8,9 Mechanical loading may induce remodeling directly via tissue stresses that may predispose the matrix to damage or through alterations in the biosynthetic response due to mechanically altered biosynthesis of proteins and enzymes.8 IVD degeneration is manifested morphologically through a loss in disc height, decreased nucleus volume, and in a loss of distinction between nucleus pulposus (NP) and annulus fibrosus (AF). In more severe degeneration, a more extensive loss in IVD structural organization has been noted, with formation of clefts and tears in the AF.2 Degenerative changes on the biochemical level are noted first in the NP, with a loss of glysosaminoglycans (GA...

Relatively minor disruption in the disc from needle puncture injury had immediate and progressive mechanical and biologic consequences with important implications for the use of discography, and repair-regeneration techniques. Results also suggest diagnostic techniques sensitive to mechanical changes in the disc may be important for early detection of degenerative changes in response to anulus injury.

The overall goal of this work is to define more clearly which mechanical loading conditions are associated with accelerated disc degeneration. This article briefly reviews recent studies describing the effects of mechanical loading on the metabolism of intervertebral disc cells and defines hypothetical models that provide a framework for quantitative relationships between mechanical loading and disc-cell metabolism. Disc cells respond to mechanical loading in a manner that depends on loading magnitude, frequency, and duration. On the basis of the current data, four models have been proposed to describe the effects of continuous loading on cellular metabolism: (1) on/off response, in which messenger ribonucleic acid (mRNA) transcription remains altered for the duration of loading; (2) maintenance, characterized by an initial change in mRNA levels with return to baseline levels; (3) adaptation, in which mRNA transcription is altered and remains at a new steady state; and (4) no response. In addition, five hypothetical mechanisms that describe the long-term consequences of these metabolic changes on disc-remodeling are presented. The transient nature of gene expression along with enzyme activation/inhibition is associated with changes at the protein level. The hypothetical models presented provide a framework for obtaining quantitative relationships between mechanical loading, gene expression, and changes at the compositional level; however, additional factors, such as regulatory mechanisms, must also be considered when describing disc-remodeling. A more quantitative relationship between mechanical loading effects and the metabolic response of the disc will contribute to injury prevention, facilitate more effective rehabilitation treatments, and help realize the potential of biologic and tissue engineering approaches toward disc repair.

Damage of the annulus fibrosus is implicated in common spinal pathologies. The objective of this study was to obtain a quantitative relationship between both the number of cycles and the magnitude of tensile strain resulting in damage to the annulus fibrosus. Four rectangular tensile specimens oriented in the circumferential direction were harvested from the outer annulus of 8 bovine caudal discs (n = 32) and subjected to one of four tensile testing protocols: (i) ultimate tensile strain (UTS) test; (ii) baseline cyclic test with 4 series of 400 cycles of baseline cyclic loading (peak strain = 20% UTS); (iii & iv) acute and fatigue damage cyclic tests consisting of 4 x 400 cycles of baseline cyclic loading with intermittent loading to 1 and 100 cycles, respectively, with peak tensile strain of 40%, 60%, and 80% UTS. Normalized peak stress for all mechanically loaded specimens was reduced from 0.89 to 0.11 of the baseline control levels, and depended on the magnitude of damaging strain and number of cycles at that damaging strain. Baseline, acute, and fatigue protocols resulted in permanent deformation of 3.5%, 6.7% and 9.6% elongation, respectively. Damage to the laminate structure of the annulus in the absence of biochemical activity in this study was assessed using histology, transmission electron microscopy, and biochemical measurements and was most likely a result of separation of annulus layers (i.e., delamination). Permanent elongation and stress reduction in the annulus may manifest in the motion segment as sub-catastrophic damage including increased neutral zone, disc bulging, and loss of nucleus pulposus pressure. The preparation of rectangular tensile strip specimens required cutting of collagen fibers and may influence absolute values of results, however, it is not expected to affect the comparisons between loading groups or dose-response reported.

Intervertebral disc organ culture has the capacity to control mechanical and chemical boundary conditions while keeping the tissue largely intact, and allowing interventions that would be impossible or unethical on animal studies. Recent studies on ex vivo organ culture has mostly involved small animals, or been limited to development and validation studies. In this study, bovine caudal discs were used. The large animal model design ensures that sufficient tissue is available for measurement of multiple dependent variables on the same disc, and a similar aspect ratio, diffusion distance, composition and rate of proteoglycan synthesis to human lumbar discs. The first goal of this study was to refine a set of dependent variables capable of characterizing the response of the intervertebral disc to culturing and to develop a technique to measure cell viability in all three regions of the disc. The second goal was to use these variables to compare static and diurnal loading as a method of maintaining intervertebral disc structure, composition, and cell metabolism similar to the in vivo state. Static (0.2 MPa) and diurnal loading (0.1 and 0.3 MPa alternating at 12 h intervals) were applied and intervertebral discs were examined after 4 or 8 days with dependent variables including changes in geometry (disc height and diameter), composition (tissue water content, tissue proteoglycan content and proteoglycan content lost to the culture media), cell viability and metabolism (proteoglycan synthesis). Results indicate that there was a decrease in disc height and water content after culture regardless of culture duration or loading condition. Cell viability significantly decreased with culture duration in the inner annulus and nucleus; however, a significant reduction in cell viability for the diurnal versus static loading condition was only observed after 8 days in the nucleus region. No significant differences were seen in viability of the outer annulus region with time, or in any loading groups. We conclude that our system is capable of keeping bovine caudal discs alive for at least 8 days without significant changes in GAG content, or cell metabolism, and that static loading was slightly better able to maintain cell viability than diurnal loading. This system offers promise for the future studies on large intervertebral discs requiring measurements of multiple mechanical and biological dependent variables on the same tissue.

The purpose of this study was to gain an improved understanding of the mechanical behavior of the intervertebral disc in the presence and absence of the vertebral endplates. Mechanical behaviors of rat caudal motion segments, vertebrae and isolated disc explants under two different permeability conditions were investigated and viscoelastic behaviors were evaluated using a stretched-exponential function to describe creep and recovery behaviors. The results demonstrated that both vertebrae and discs underwent significant deformations in the motion segment even under relatively low-loading conditions. Secondly, disruption of the collagenous network had minimal impact on equilibrium deformations of disc explants as compared to disc deformations occurring in the motion segments provided that vertebral deformations were accounted for; however, differences in endplate permeability conditions had a significant effect on viscoelastic behaviors. Creep occurred more quickly than recovery for motion segment and explant specimens. In addition, disc explants and motion segments both exhibited non-recoverable deformations under axial compression under lowand high-loading conditions. Results have important implications for interpreting the role of vertebral endplates in contributing to disc mechanical behaviors and direct application to mechanobiology studies involving external loading to rodent tail intervertebral discs.

No abstract