Dawn M. Elliott

Nanofibrous biologic laminates replicate the form and function of the annulus fibrosus

Successful engineering of load-bearing tissues requires recapitulation of their complex mechanical functions. Given the intimate relationship between function and form, biomimetic materials that replicate anatomic form are of great interest for tissue engineering applications. However, for complex tissues such as the annulus fibrosus, scaffolds have failed to capture their multi-scale structural hierarchy. Consequently, engineered tissues have yet to reach functional equivalence with their native counterparts. Here we present a novel strategy for annulus fibrosus tissue engineering that replicates this hierarchy with anisotropic nanofibrous laminates seeded with mesenchymal stem cells. These scaffolds directed the deposition of organized, collagen-rich extracellular matrix that mimicked the angle-ply, multi-lamellar architecture and achieved mechanical parity with native tissue after 10 weeks of in vitro culture. Further, we identified a novel role for inter-lamellar shearing in reinforcing the tensile response of biologic laminates, a mechanism that has not previously been considered for these tissues.

Anisotropic and inhomogeneous tensile behavior of the human anulus fibrosus: experimental measurement and material model predictions.

The anulus fibrosus (AF) of the intervertebral disc exhibits spatial variations in structure and composition that give rise to both anisotropy and inhomogeneity in its material behaviors in tension. In this study, the tensile moduli and Poissons ratios were measured in samples of human AF along circumferential, axial, and radial directions at inner and outer sites. There was evidence of significant inhomogeneity in the linear-region circumferential tensile modulus (17.4+/-14.3 MPa versus 5.6+/-4.7 MPa, outer versus inner sites) and the Poissons ratio v21 (0.67+/-0.22 versus 1.6+/-0.7, outer versus inner), but not in the axial modulus (0.8+/-0.9 MPa) or the Poissons ratios V12 (1.8+/-1.4) or v13 (0.6+/-0.7). These properties were implemented in a linear an isotropic material model of the AF to determine a complete set of model properties and to predict material behaviors for the AF under idealized kinematic states. These predictions demonstrate that interactions between fiber populations in the multilamellae AF significantly contribute to the material behavior, suggesting that a model for th

Assessment of Human Disc Degeneration and Proteoglycan Content Using T1ρ-weighted Magnetic Resonance Imaging

Study Design. T1&rgr; relaxation was quantified and correlated with intervertebral disc degeneration and proteoglycan content in cadaveric human lumbar spine tissue. Objective. To show the use of T1&rgr;-weighted magnetic resonance imaging (MRI) for the assessment of degeneration and proteoglycan content in the human intervertebral disc. Summary of Background Data. Loss of proteoglycan in the nucleus pulposus occurs during early degeneration. Conventional MRI techniques cannot detect these early changes in the extracellular matrix content of the disc. T1&rgr; MRI is sensitive to changes in proteoglycan content of articular cartilage and may, therefore, be sensitive to proteoglycan content in the intervertebral disc. Methods. Intact human cadaveric lumbar spines were imaged on a clinical MR scanner. Average T1&rgr; in the nucleus pulposus was calculated from quantitative T1&rgr; maps. After MRI, the spines were dissected, and proteoglycan content of the nucleus pulposus was measured. Finally, the stage of degeneration was graded using conventional T2 images. Results. T1&rgr; decreased linearly with increasing degeneration (r = −0.76, P < 0.01) and age (r = −0.76, P < 0.01). Biochemical analysis revealed a strong linear correlation between T1&rgr; and sulfated-glycosaminoglycan content. T1&rgr; was moderately correlated with water content. Conclusions. Results from this study suggest that T1&rgr; may provide a tool for the diagnosis of early degenerative changes in the disc. T1&rgr;-weighted MRI is a noninvasive technique that may provide higher dynamic range than T2 and does not require a high static field or exogenous contrast agents.

Effect of fiber orientation and strain rate on the nonlinear uniaxial tensile material properties of tendon.

Tendons are exposed to complex loading scenarios that can only be quantified by mathematical models, requiring a full knowledge of tendon mechanical properties. This study measured the anisotropic, nonlinear, elastic material properties of tendon. Previous studies have primarily used constant strain-rate tensile tests to determine elastic modulus in the fiber direction. Data for Poissons ratio aligned with the fiber direction and all material properties transverse to the fiber direction are sparse. Additionally, it is not known whether quasi-static constant strain-rate tests represent equilibrium elastic tissue behavior. Incremental stress-relaxation and constant strain-rate tensile tests were performed on sheep flexor tendon samples aligned with the tendon fiber direction or transverse to the fiber direction to determine the anisotropic properties of toe-region modulus (E0), linear-region modulus (E), and Poissons ratio (v). Among the modulus values calculated, only fiber-aligned linear-region modulus (E1) was found to be strain-rate dependent. The E1 calculated from the constant strain-rate tests were significantly greater than the value calculated from incremental stress-relaxation testing. Fiber-aligned toe-region modulus (E(1)0 = 10.5 +/- 4.7 MPa) and linear-region modulus (E1 = 34.0 +/- 15.5 MPa) were consistently 2 orders of magnitude greater than transverse moduli (E(2)0 = 0.055 +/- 0.044 MPa, E2 = 0.157 +/- 0.154 MPa). Poissons ratio values were not found to be rate-dependent in either the fiber-aligned (v12 = 2.98 +/- 2.59, n = 24) or transverse (v21 = 0.488 +/- 0.653, n = 22) directions, and average Poissons ratio values in the fiber-aligned direction were six times greater than in the transverse direction. The lack of strain-rate dependence of transverse properties demonstrates that slow constant strain-rate tests represent elastic properties in the transverse direction. However, the strain-rate dependence demonstrated by the fiber-aligned linear-region modulus suggests that incremental stress-relaxation tests are necessary to determine the equilibrium elastic properties of tendon, and may be more appropriate for determining the properties to be used in elastic mathematical models.

Effects of degeneration on the biphasic material properties of human nucleus pulposus in confined compression.

Study Design. The biphasic compressive material properties of normal and degenerate human nucleus pulposus tissue were measured in confined compression. Objectives. The objective of this study was to determine the effects of degeneration and age on the mechanical properties of human nucleus pulposus. Summary of Background Data. The nucleus pulposus exhibits swelling behavior in proportion to proteoglycan content. In shear, the nucleus exhibits both fluid-like and solid-like properties, suggesting a biphasic nature. To date, biphasic compressive properties of human nucleus pulpous have not been reported. Methods. Human nucleus pulposus samples were tested in confined compression. Isometric swelling stress and effective aggregate modulus were measured. Linear biphasic theory was used to determine the permeability of the tissue. Mechanical behavior was correlated with proteoglycan and water content. Results. Degeneration produced significant decreases in swelling stress (Psw = 0.138 ± 0.029 MPa nondegenerate, Psw = 0.037 ± 0.038 MPa degenerate) and effective aggregate modulus (HAeff = 1.01 ± 0.43 MPa nondegenerate, HAeff = 0.44 ± 0.19 MPa degenerate). Both properties were inversely correlated with proteoglycan content. Permeability increased with degeneration (ka = 0.9 ± 0.43 × 10−15 m4/N-s nondegenerate, ka = 1.4 ± 0.58 × 10−15 m4/N-s degenerate). Conclusions. Swelling is the primary load-bearing mechanism in both nondegenerate and degenerate nucleus pulposus. Knowledge of the biphasic material properties of the nucleus pulposus will aid the development of new treatment strategies for disc degeneration aimed at restoring mechanical function of the intervertebral disc.

Degeneration and regeneration of the intervertebral disc: lessons from development

Degeneration of the intervertebral discs, a process characterized by a cascade of cellular, biochemical, structural and functional changes, is strongly implicated as a cause of low back pain. Current treatment strategies for disc degeneration typically address the symptoms of low back pain without treating the underlying cause or restoring mechanical function. A more in-depth understanding of disc degeneration, as well as opportunities for therapeutic intervention, can be obtained by considering aspects of intervertebral disc development. Development of the intervertebral disc involves the coalescence of several different cell types through highly orchestrated and complex molecular interactions. The resulting structures must function synergistically in an environment that is subjected to continuous mechanical perturbation throughout the life of an individual. Early postnatal changes, including altered cellularity, vascular regression and altered extracellular matrix composition, might set the disc on a slow course towards symptomatic degeneration. In this Perspective, we review the pathogenesis and treatment of intervertebral disc degeneration in the context of disc development. Within this scope, we examine how model systems have advanced our understanding of embryonic morphogenesis and associated molecular signaling pathways, in addition to the postnatal changes to the cellular, nutritional and mechanical microenvironment. We also discuss the current status of biological therapeutic strategies that promote disc regeneration and repair, and how lessons from development might provide clues for their refinement.

Effect of fiber distribution and realignment on the nonlinear and inhomogeneous mechanical properties of human supraspinatus tendon under longitudinal tensile loading

Tendon exhibits nonlinear stress–strain behavior that may be partly due to movement of collagen fibers through the extracellular matrix. While a few techniques have been developed to evaluate the fiber architecture of other soft tissues, the organizational behavior of tendon under load has not been determined. The supraspinatus tendon (SST) of the rotator cuff is of particular interest for investigation due to its complex mechanical environment and corresponding inhomogeneity. In addition, SST injury occurs frequently with limited success in treatment strategies, illustrating the need for a better understanding of SST properties. Therefore, the objective of this study was to quantitatively evaluate the inhomogeneous tensile mechanical properties, fiber organization, and fiber realignment under load of human SST utilizing a novel polarized light technique. Fiber distributions were found to become more aligned under load, particularly during the low stiffness toe‐region, suggesting that fiber realignment may be partly responsible for observed nonlinear behavior. Fiber alignment was found to correlate significantly with mechanical parameters, providing evidence for strong structure–function relationships in tendon. Human SST exhibits complex, inhomogeneous mechanical properties and fiber distributions, perhaps due to its complex loading environment. Surprisingly, histological grade of degeneration did not correlate with mechanical properties.

Effect of altered matrix proteins on quasilinear viscoelastic properties in transgenic mouse tail tendons

AbstractTendons have complex mechanical behaviors that are viscoelastic, nonlinear, and anisotropic. It is widely held that these behaviors are provided for by the tissues composition and structure. However, little data are available to quantify such structure–function relationships. This study quantified tendon mechanical behaviors, including viscoelasticity and nonlinearity, for groups of mice that were genetically engineered for altered extracellular matrix proteins. Uniaxial tensile stress-relaxation experiments were performed on tail tendon fascicles from the following groups: eight week old decorin knockout, eight week old reduced type I collagen, three week old control, and eight week old control. Data were fit using Fungs quasilinear viscoelastic model, where the model parameters represent the linear viscoelastic and nonlinear elastic response. The viscoelastic properties demonstrated a larger and faster stress relaxation for the decorin knockout and a smaller and slower stress relaxation for the three week control. The elastic parameter, A, in the eight week control group was significantly greater than in the collagen reduction and three week control groups. This study provides quantitative evidence for structure–function relationships in tendon, including the role of proteoglycan in viscoelasticity. Future studies should directly correlate composition and structure with tendon mechanics for the design and evaluation of tissue-engineered constructs or tendon repairs.

Comparison of animals used in disc research to human lumbar disc geometry.

Study Design. Measurement and normalization of disc geometry parameters for several animal models used in disc research. Objectives. To compare normalized values of disc geometry to the human disc geometry to aid in the selection and interpretation of animal model studies. Summary of Background Data. Animal models are widely used to study intervertebral disc degeneration and to evaluate disc treatment methods because of the availability of the tissue, the decreased variability between subjects compared with humans, and the feasibility to perform in vivo experiments. There is a general lack of comparative data with respect to the human disc analog for animal models. Methods. The disc height, lateral width, AP width, area, and the nucleus pulposus lateral width, AP width, area, and centroid offset were all measured and normalized by 2 scaling factors, lateral width and disc area, for comparison to human. Results. The species studied were ranked according to the average percent deviation of the normalized disc height, AP width and nucleus pulposus area from human geometry as: mouse lumbar (12%), rat lumbar (15%), mouse tail (18%), baboon (19%), bovine tail (22%), rabbit (26%), sheep (31%), and rat tail (46%). Conclusions. This paper provides a reference to compare disc geometries of experimental animal models to the human lumbar disc, to aid both in interpretation of and in planning for experimental disc research, and to provide normalized disc geometry parameters for computational models.

Comparison of animal discs used in disc research to human lumbar disc: axial compression mechanics and glycosaminoglycan content.

Study Design. Experimental measurement and normalization of in vitro disc axial compression mechanics and glycosaminoglycan and water content for several animal species used in intervertebral disc research. Objective. To compare normalized axial mechanical properties and glycosaminoglycan and water content from other species to those of the human disc to aid in selection and interpretation of results in animal disc studies. Summary of Background Data. There is a lack of mechanical and biochemical comparative data from animal intervertebral discs with respect to the human disc. Methods. Intervertebral disc axial mechanical properties, glycosaminoglycan, and water content were evaluated for 9 disc types in 7 mammalian species: the calf, pig, baboon, sheep, rabbit, rat and mouse lumbar, and the cow and rat tail. Disc area and height were used for calculation of the normalized mechanical parameters. Glycosaminoglycan content was normalized by dry weight. Results. Many directly measured mechanical parameters varied by orders of magnitude. However, these parameters became comparable and often did not show significant differences after geometric normalization. Both glycosaminoglycan and water content revealed similarity across species. Conclusion. Disc axial mechanics are very similar across animal species when normalizing by the geometric parameters of disc height and area. This suggests that the disc tissue material properties are largely conserved across animal species. These results provide a reference to compare disc axial mechanics and glycosaminoglycan and water composition of experimental animal models to the human lumbar disc, to aid in both selection and interpretation of experimental disc research.