Effects of Oxygen Concentration and Culture Time on Porcine Nucleus Pulposus Cell Metabolism: An in vitro Study.

Low back pain is a common ailment that affects millions of individuals each year and is linked to degeneration of the intervertebral discs in the spine. Intervertebral disc degeneration is known to result from an imbalance in anabolic and catabolic activity by disc cells. Due to the avascular nature of the intervertebral disc, oxygen deficiency may occur in the central nucleus pulposus (NP). The resulting hypoxia affects matrix regulation and energy metabolism of disc cells, although the mechanisms are not fully understood. This study investigates in vitro glucose consumption and gene expression by NP cells over time under varying oxygen tensions. Notochordal porcine NP cells were cultured in agarose discs at 21, 5, or 1% oxygen tension for 1, 5, or 10 days. The expression of 10 key matrix genes, as well as Brachyury (T), by NP cells was analyzed using RT-PCR. Glucose consumption was measured using a two-point method. Results show that culture time and oxygen tension significantly affect glucose consumption rates by porcine NP cells. There were also significant changes in T expression based on oxygen level and culture time. The 1% oxygen tension had a significantly higher T expression on day 10 than the other two groups, which may indicate a better maintenance of the notochordal phenotype. MMP 1 and 13 expression increased over time for all groups, while only the 5% group showed an increase over time for MMP 3. TIMP expression followed the direction of MMPs but to a lesser magnitude. Five percent and twenty-one percent oxygen tensions led to decreases in anabolic gene expression while 1% led to increases. Oxygen concentration and culture time significantly impacted glucose consumption rate and the gene expression of matrix regulatory genes with hypoxic conditions most accurately maintaining the proper NP phenotype. This information is valuable not only for understanding disc pathophysiology, but also for harnessing the potential of notochordal NP cells in therapeutic applications.

Hydraulic permeability of meniscus fibrocartilage measured via direct permeation: Effects of tissue anisotropy, water volume content, and compressive strain.

Hydraulic permeability is an important material property of cartilaginous tissues, governing the rate of fluid flow, which is crucial to tissue biomechanics and cellular nutrition. The effects of strain, anisotropy, and region on the hydraulic permeability in meniscus tissue have not been fully elucidated. Using a one-dimensional direct permeation test, we measured the hydraulic permeability within statically compressed porcine meniscus specimens, prepared such that the explants were in either the axial or circumferential direction of either the central or horn (axial direction only) region of the medial and lateral menisci. A constant flow was applied and the pressure difference was measured using pressure transducers. Specimens were tested under 10-20% compressive strain. Permeability values were in the range of 1.53-1.87 × 10-15 m4/Ns, which is comparable to values found in the literature. Permeability was significantly anisotropic, being higher in the circumferential direction than in the axial direction. Additionally, there was a significant negative correlation between strain level and permeability for all groups. Lastly, no statistically significant difference was found between permeability coefficients from different regional locations. This study provides important information regarding structure-function relationships in meniscal tissues that helps to elucidate biomechanics and transport in the tissue, and can aid in the understanding of the tissue's role in the function of the knee joint and onset of osteoarthritis.

Effect of Strain, Region, and Tissue Composition on Glucose Partitioning in Meniscus Fibrocartilage.

A nearly avascular tissue, the knee meniscus relies on diffusive transport for nutritional supply to cells. Nutrient transport depends on solute partitioning in the tissue, which governs the amount of nutrients that can enter a tissue. The purpose of the present study was to investigate the effects of mechanical strain, tissue region, and tissue composition on the partition coefficient of glucose in meniscus fibrocartilage. A simple partitioning experiment was employed to measure glucose partitioning in porcine meniscus tissues from two regions (horn and central), from both meniscal components (medial and lateral), and at three levels of compression (0%, 10%, and 20%). Partition coefficient values were correlated to strain level, water volume fraction, and glycosaminoglycan (GAG) content of tissue specimens. Partition coefficient values ranged from 0.47 to 0.91 (n = 48). Results show that glucose partition coefficient is significantly (p < 0.001) affected by compression, decreasing with increasing strain. Furthermore, we did not find a statistically significant effect of tissue when comparing medial versus lateral (p = 0.181) or when comparing central and horn regions (p = 0.837). There were significant positive correlations between tissue water volume fraction and glucose partitioning for all groups. However, the correlation between GAG content and partitioning was only significant in the lateral horn group. Determining how glucose partitioning is affected by tissue composition and loading is necessary for understanding nutrient availability and related tissue health and/or degeneration. Therefore, this study is important for better understanding the transport and nutrition-related mechanisms of meniscal degeneration.

Electrical conductivity and ion diffusion in porcine meniscus: effects of strain, anisotropy, and tissue region.

The purpose of the present study was to investigate the effects of mechanical strain, anisotropy, and tissue region on electrical conductivity and ion diffusivity in meniscus fibrocartilage. A one-dimensional, 4-wire conductivity experiment was employed to measure the electrical conductivity in porcine meniscus tissues from two tissue regions (horn and central), for two tissue orientations (axial and circumferential), and for three levels of compressive strain (0%, 10%, and 20%). Conductivity values were then used to estimate the relative ion diffusivity in meniscus. The water volume fraction of tissue specimens was determined using a buoyancy method. A total of 135 meniscus samples were measured; electrical conductivity values ranged from 2.47mS/cm to 4.84mS/cm, while relative ion diffusivity was in the range of 0.235 to 0.409. Results show that electrical conductivity and ion diffusion are significantly anisotropic (p<0.001), both being higher in the circumferential direction than in the axial direction. Additionally, the findings show that compression significantly affects the electrical conductivity with decreasing conductivity levels corresponding to increased compressive strain (p<0.001). Furthermore, there was no statistically significant effect of tissue region when comparing axial conductivity in the central and horn regions of the tissue (p=0.999). There was a positive correlation between tissue water volume fraction and both electrical conductivity and relative ion diffusivity for all groups investigated. This study provides important insight into the electromechanical and transport properties in meniscus fibrocartilage, which is essential in developing new strategies to treat and/or prevent tissue degeneration.

Effect of Static Compressive Strain, Anisotropy, and Tissue Region on the Diffusion of Glucose in Meniscus Fibrocartilage.

Osteoarthritis (OA) is a significant socio-economic concern, affecting millions of individuals each year. Degeneration of the meniscus of the knee is often associated with OA, yet the relationship between the two is not well understood. As a nearly avascular tissue, the meniscus must rely on diffusive transport for nutritional supply to cells. Therefore, quantifying structure-function relations for transport properties in meniscus fibrocartilage is an important task. The purpose of the present study was to determine how mechanical loading, tissue anisotropy, and tissue region affect glucose diffusion in meniscus fibrocartilage. A one-dimensional (1D) diffusion experiment was used to measure the diffusion coefficient of glucose in porcine meniscus tissues. Results show that glucose diffusion is strain-dependent, decreasing significantly with increased levels of compression. It was also determined that glucose diffusion in meniscus tissues is anisotropic, with the diffusion coefficient in the circumferential direction being significantly higher than that in the axial direction. Finally, the effect of tissue region was not statistically significant, comparing axial diffusion in the central and horn regions of the tissue. This study is important for better understanding the transport and nutrition-related mechanisms of meniscal degeneration and related OA in the knee.