A comprehensive understanding of multiscale and multiphasic intervertebral disc mechanics is crucial for designing advanced tissue engineered structures aiming to recapitulate native tissue behavior. The bovine caudal disc is a commonly used human disc analog due to its availability, large disc height and area, and similarities in biochemical and mechanical properties to the human disc. Because of challenges in directly measuring subtissue-level mechanics, such as in situ fiber mechanics, finite element models have been widely employed in spinal biomechanics research. However, many previous models use homogenization theory and describe each model element as a homogenized combination of fibers and the extrafibrillar matrix while ignoring the role of water content or osmotic behavior. Thus, these models are limited in their ability in investigating subtissue-level mechanics and stress-bearing mechanisms through fluid pressure. The objective of this study was to develop and validate a structure-based bovine caudal disc model, and to evaluate multiscale and multiphasic intervertebral disc mechanics under different loading conditions and with degeneration. The structure-based model was developed based on native disc structure, where fibers and matrix in the annulus fibrosus were described as distinct materials occupying separate volumes. Model parameters were directly obtained from experimental studies without calibration. Under the multiscale validation framework, the model was validated across the joint-, tissue-, and subtissue-levels. Our model accurately predicted multiscale disc responses for 15 of 16 cases, emphasizing the accuracy of the model, as well as the effectiveness and robustness of the multiscale structure-based modeling-validation framework. The model also demonstrated the rim as a weak link for disc failure, highlighting the importance of keeping the cartilage endplate intact when evaluating disc failure mechanisms in vitro . Importantly, results from this study elucidated important fluid-based load-bearing mechanisms and fiber-matrix interactions that are important for understanding disease progression and regeneration in intervertebral discs. In conclusion, the methods presented in this study can be used in conjunction with experimental work to simultaneously investigate disc joint-, tissue-, and subtissue-level mechanics with degeneration, disease, and injury.
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Radial variation in biochemical composition of the bovine caudal intervertebral disc
Bovine caudal discs have been widely used in spine research due to their increased availability, large size, and mechanical and biochemical properties that are comparable to healthy human discs. However, despite their extensive use, the radial variations in bovine disc composition have not yet been rigorously quantified with high spatial resolution. Previous studies were limited to qualitative analyses or provided limited spatial resolution in biochemical properties. Thus, the main objective of this study was to provide quantitative measurements of biochemical composition with higher spatial resolution than previous studies that employed traditional biochemical techniques. Specifically, traditional biochemical analyses were used to measure water, sulfated glycosaminoglycan, collagen, and DNA contents. Gravimetric water content was compared to data obtained through Raman spectroscopy and differential scanning calorimetry. Additionally, spatial distribution of lipids in the disc's collagen network was visualized and quantified, for the first time, using multi‐modal second harmonic generation (SHG) and Coherent anti‐Stokes Raman (CARS) microscopy. Some heterogeneity was observed in the nucleus pulposus, where the water content and water‐to‐protein ratio of the inner nucleus were greater than the outer nucleus. In contrast, the bovine annulus fibrosus exhibited a more heterogeneous distribution of biochemical properties. Comparable results between orthohydroxyproline assay and SHG imaging highlight the potential benefit of using SHG microscopy as a less destructive method for measuring collagen content, particularly when relative changes are of interest. CARS images showed that lipid deposits were distributed equally throughout the disc and appeared either as individual droplets or as clusters of small droplets. In conclusion, this study provided a more comprehensive assessment of spatial variations in biochemical composition of the bovine caudal disc.
more » « less- Award ID(s):
- 1751212
- NSF-PAR ID:
- 10460600
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- JOR SPINE
- Volume:
- 2
- Issue:
- 3
- ISSN:
- 2572-1143
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Background Water content is a key parameter for simulating tissue swelling and nutrient diffusion. Accurately measuring water content throughout the intervertebral disc (NP = nucleus pulposus; AF = annulus fibrosus) is important for developing patient‐specific models. Water content is measured using destructive techniques, Quantitative MRI has been used to estimate water content and detect early degeneration, but it is dependent on scan parameters, concentration of free water molecules, and fiber architecture.
Purpose To directly measure disc‐tissue water content using quantitative MRI and compare MRI‐based measurements with biochemical assays, and to quantify changes in disc geometry due to compression.
Study Type Basic science, controlled.
Specimen Twenty bone‐disc‐bone motion segments from skeletally mature bovines.
Field Strength/Sequence 7T/3D fast low angle shot (FLASH) pulse sequence and a T2rapid imaging with refocused echoes (RARE) sequence.
Assessment Disc volumes, NP and AF volumetric water content, and T2relaxation times were measured through MRI; NP and AF tissue gravimetric water content, mass density, and glycosaminoglycan content were measured through a biochemical assay.
Statistical Tests Correlations between MRI‐based measurement and biochemical composition were evaluated using Pearson's linear regression.
Results Mechanical dehydration resulted in a decrease in disc volume by up to 20% and a decrease in disc height by up to 35%. Direct water content measurements for the NP was achieved by normalizing MRI‐based spin density by NP mass density (1.10 ± 0.03 g/cm3). However, the same approach underestimated water content in the AF by ~10%, which may be due to a higher concentration of collagen fibers and bound water molecules.
Data Conclusion Spin density or spin density normalized by mass density to estimate NP and AF water content was more accurate than correlations between water content and relaxation times. Mechanical dehydration decreased disc volume and disc height, and increased maximum cross‐sectional area.
Level of Evidence Technical Efficacy Stage J. Magn. Reson. Imaging 2020;52:1152–1162.
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Abstract Extracellular-matrix composition impacts mechanical performance in native and engineered tissues. Previous studies showed collagen type I-agarose blends increased cell-matrix interactions and extracellular matrix production. However, long-term impacts on protein production and mechanical properties of engineered cartilage are unknown. Our objective was to characterize the effect of collagen type I on the matrix production of chondrocytes embedded in agarose hydrogels. We hypothesized that the addition of collagen would improve long-term mechanical properties and matrix production (e.g. collagen and glycosaminoglycans) through increased bioactivity. Agarose hydrogels (2% w/v) were mixed with varying concentrations of collagen type I (0, 2 and 5 mg/ml). Juvenile bovine chondrocytes were added to the hydrogels to assess matrix production over 4 weeks through biochemical assays, and mechanical properties were assessed through unconfined compression. We observed a dose-dependent effect on cell bioactivity, where 2 mg/ml of collagen improved bioactivity, but 5 mg/ml had a negative impact on bioactivity. This resulted in a higher modulus for scaffolds supplemented with lower collagen concentration as compared to the higher collagen concentration, but not when compared to the control. In conclusion, the addition of collagen to agarose constructs provided a dose-dependent impact on improving glycosaminoglycan production but did not improve collagen production or compressive mechanics.
