Tai Yi Yuan

EFFECT OF COMPRESSION AND ANISOTROPY ON THE DIFFUSION OF GLUCOSE IN ANNULUS FIBROSUS

Study Design. Investigation of the effect of static compression and anisotropy on the apparent diffusivity of glucose in bovine annulus fibrosus (AF). Objective. To determine the apparent glucose diffusivity in 2 directions (axial and radial) of bovine AF under 3 levels of compressive strain (0%, 10%, and 20%). Summary of Background Data. Knowledge of diffusivity of small molecules is important for understanding nutritional supply in intervertebral discs and the mechanisms of disc degeneration. However, little is known regarding the strain-dependent and anisotropic behavior of glucose diffusivity in intervertebral discs. Methods. Apparent glucose diffusivity measurements were performed on 10 axial and 10 radial AF specimens from bovine coccygeal discs. The dependence of diffusivity on compression was determined using 3 levels of strain (0%, 10%, and 20%). Results. The apparent glucose diffusivity (mean ± standard deviation) of the bovine AF in the axial direction was 1.38 ± 0.015 × 10−6 cm2/s (n = 10) at 0%, 1.00 ± 0.070 × 10−6 cm2/s (n = 10) at 10%, and 7.65 ± 0.552 × 10−7 cm2/s (n = 10) at 20% compression. For radial specimens, the apparent glucose diffusivity was determined to be 9.17 ± 1.12 × 10−7 cm2/s (n = 10), 7.29 ± 0.863 × 10−7 cm2/s (n = 10), and 5.43 ± 1.16 × 10−7 cm2/s (n = 10) for 0%, 10%, and 20% compressions, respectively. A significant decrease in diffusivity with increasing strain was found for both axial and radial specimens [analysis of variance (ANOVA), P < 0.05]. Diffusion in the radial direction was determined to be significantly less than that in the axial direction (ANOVA, P < 0.05). A significant interaction was found between the level of strain and the direction of diffusion (ANOVA, P < 0.05). Conclusion. Diffusion of glucose in bovine AF is dependent on strain and the direction of diffusion.

Mechanical Loading Affects the Energy Metabolism of Intervertebral Disc Cells

Research has shown that mechanical loading affects matrix biosynthesis of intervertebral disc (IVD) cells; however, the pathway(s) to this effect is currently unknown. Cellular matrix biosynthesis is an energy demanding process. The objective of this study was to investigate the effects of static and dynamic compressive loading on energy metabolism of IVD cells. Porcine annulus fibrosus (AF) and nucleus pulposus (NP) cells seeded in 2% agarose were used in this experiment. Experimental groups included 15% static compression and 0.1 and 1 Hz dynamic compression at 15% strain magnitude for 4 h. ATP, lactate, glucose, and nitric oxide (NO) contents in culture media, and ATP content in cell–agarose construct were measured using biochemical assays. While the total ATP content of AF cells was promoted by static and dynamic loading, only 1 Hz dynamic loading increased total ATP content of NP cells. Increases in lactate production and glucose consumption of AF cells suggest that ATP production via glycolysis is promoted by dynamic compression. ATP release and NO production of AF and NP cells were significantly increased by dynamic loading. Thus, this study clearly illustrates that static and dynamic compressive loading affect IVD cell energy production while cellular responses to mechanical loading were both cell type and compression type dependent.

Effects of low glucose concentrations on oxygen consumption rates of intervertebral disc cells.

Study Design. Investigation of the effects of low glucose concentrations on the oxygen consumption rates of intervertebral disc cells. Objectives. To determine the oxygen consumption rate of porcine anulus fibrosus (AF) cells at different glucose concentrations and to examine the differences in the oxygen consumption rate between AF and nucleus pulposus (NP) cells at different glucose levels. Summary of Background Data. Poor nutrient supply has been suggested as a potential mechanism for degeneration of the intervertebral disc (IVD). Distribution of nutrients in the IVD is strongly dependent on transport properties of the tissue and cellular metabolic rates. Previous studies have shown dependence of oxygen consumption rate of IVD cells on oxygen tension, pH levels, and glucose levels outside the physiologic range. However, the oxygen consumption rate of AF cells at in vivo glucose levels has not been investigated. Methods. IVD cells were isolated from the outer AF and NP of 4- to 5-month-old porcine lumbar discs. The changes in oxygen tension were recorded when cells were cultured in sealed metabolism chamber. The oxygen consumption rate of cells was determined by theoretical curve fitting using the Michaelis-Menten equation. Results. The outer AF cells cultured in high glucose medium (25 mmol/L) exhibited the lowest oxygen consumption rate, whereas no significant differences in oxygen consumption rates were found among outer AF cells cultured at physiologic glucose levels (i.e., 1 mmol/L, 2.5 mmol/L, 5 mmol/L). The oxygen consumption rate of NP cells was significantly greater than that of outer AF cells. Conclusion. Since the oxygen consumption rates determined in this study are comparable to the findings in the literature, this study has developed a new alternative method for determining oxygen consumption rate. The oxygen consumption rates of IVD cells reported in this study will be valuable for theoretically predicting local oxygen concentrations in IVD, which can provide a better understanding of transport of oxygen in the discs.

Strain Dependent Oxygen Diffusivity in Bovine Annulus Fibrosus

Intervertebral disc (IVD) is the largest avascular structure in the human body and nutrition supply into IVD is mainly through diffusion from the peripheral blood vessels. Poor nutrition supply to the disc is believed to be one of the causes for disc degeneration. While many studies have aimed at studying and analyzing the effect of mechanical loading on water content, chemical composition, and nutritional levels in IVD [1–3], no study has been reported to investigate the effect of mechanical compression on oxygen diffusion in the IVD tissue. The objective of this study was to determine oxygen diffusivity in annulus fibrosus (AF) samples under different levels of compression.Copyright

Verification of Measurements of Cartilaginous Tissue Constructs in the Online Characterizing Bioreactor System

In cartilage tissue engineering the self-repair of articular cartilage requires monitoring of the cells deposition of extracellular matrix, but current methods of observing destroy the sample. Sample destruction does not allow for long-term, continuous monitoring. The objective of the study was to test the bioreactor system ability to non-destructively measure properties of the tissue-engineered constructs. Porcine articular cartilage was decellularized and proteoglycans were removed. Porcine chondrocytes were seeded into the scaffolds using centrifugation within a cell seeding device and the scaffolds were put into culture plates for up to 21 days in culture medium. At different intervals, the scaffolds were analyzed using the bioreactor system, DMMB quantification, and histological techniques. The bioreactor measurements and DMMB quantification both confirmed there was a consistent increase in proteoglycan content within the scaffolds up to the 21 day endpoint, but the proteoglycan content of the constructs was only at 30% of the measured average content in native tissue. The bioreactor was able to measure the proteoglycan content accurately when compared to the measurements taken by DMMB quantification. The results of the study showed the bioreactor is useful for monitoring tissue growth in cartilage tissue engineering constructs non-destructively.

Mechanical Compression Affects Nutritional Transport in Human Intervertebral Disc

Although the exact cause is not clear, low back pain has been attributed to degeneration of the intervertebral disc (IVD) of the spine, which has been linked to poor nutritional supply to the disc. Because the IVD is the largest avascular structure in the human body, disc cells must rely on diffusional transport for delivery of important nutrients, such as oxygen and glucose, from the surrounding vasculature. Thus, understanding factors affecting nutritional supply to the cells is important in elucidating the etiology of disc degeneration and related back pain. While knowledge of how mechanical strain affects nutritional transport is important in understanding these phenomena, little can be found in the literature regarding strain-dependent diffusion of nutrients in human IVD.Copyright

A two-point conductivity approach to measuring fixed charge density in intervertebral disc tissue

Low back pain, a major socio-economic concern in the United States, is believed to result from degeneration of the intervertebral disc (IVD) of the spine [1]. The IVD is characterized as a charged, hydrated soft tissue made up of a central nucleus pulposus (NP) surrounded by the layered annulus fibrosus (AF). The negatively-charged nature of the disc derives from the charged groups attached to the glycosaminoglycan (GAG) molecules found on proteoglycans (PG) in the extracellular matrix of the disc. The fixed charge density (FCD) is a measure of the number of negative charges attached to the disc matrix per unit volume. FCD is important to disc function, both mechanically (i.e., swelling pressure) and in terms of transport through the disc [2].Copyright