Purpose: High levels of the chitinase 3-like protein HC-gp39 (human cartilage glycoprotein 39) have been found in the synovial fluid and sera of patients with arthritis. Although the function of this protein is unclear, in chondrocytes at least it appears to limit catabolic responses to cytokines such as Il-1b. Here we have investigated secretion of this protein by intervertebral disc cells and determined if its production is influenced by extracellular osmolarity.

Methods: Cells were isolated from bovine caudal discs by enzyme digestion and cultured in DMEM in alginate beads for 6 days. Medium osmolarity was increased in the physiological range by sodium/potassium addition. Supernatants were collected every 2 days and replaced with fresh media. At the end of experiment the supernatants were used for lactate determination and for detection of GP-39 by western blotting. Beads were assayed for glycosaminoglycans, cell viability and cell density.

Results: GP-39 was a major protein secreted by disc cells. It was evident on day 2 at low osmolarities. By day 4 concentrations in the medium had increased significantly and the protein was present mainly in fragmented form, particularly at high osmolarities. Osmolarity had no effect on cell density or viability. Rates of lactate production and GAG accumulation were greatest at high osmolarities.

Discussion: Changes in osmolarity, equivalent to those experienced by disc cells during the diurnal loss and regain of fluid content, had significant effects on cell metabolism and influenced production of GP-39. Osmotic changes might thus influence responses of disc cells to inflammatory signals.

Background and Purpose: Intervertebral discs are the largest avascular tissue sources in the human body. The transport of vital nutrients and oxygen into and metabolic waste products out of the disc, relies mainly on the diffusion through the disc matrix. The health or degree of degeneration of the disc is thought to be directly related to the transport properties of the disc. The diffusivity of nutrients and metabolites varies with matrix composition and especially with matrix hydration. The hydration of the disc varies by approximately 25% in the normal 24hour loading cycle of human beings. This work addresses the question of the effect of hydration of the disc tissue on the solute diffusivity.

Methods: Measurements of the diffusion of solutes were performed in ca. 2year bovine caudal discs. Diffusivity of dissolved oxygen and nitrous oxide was monitored electrochemically. Diffusivity of 0.05 to 70kDa solute species was determined by measuring concentration gradients using either fluorescent or radiotracers. Hydration was controlled by either mechanical static load or by osmotic equilibration.

Results: Diffusion rates varied with solute molecular weight (MW), decreasing steeply with an increase in MW. For small solutes, the diffusivity was greater in the nucleus than the outer annulus, but this difference was insignificant for the larger solutes. Diffusivity changed by a significant amount with hydration changes, which were significantly affected by loading. Application of a 0.2MPa mechanical load led to a drop in hydration of the outer annulus and nucleus of 33.3% and 42.1% and corresponding falls in diffusivity of glucose of 34.0% and 81.3% respectively.

Conclusions: The large changes in hydration experienced during normal loading of the spine have a marked effect on nutrient and metabolite diffusivity. This effect has not been considered previously but could significantly influence supply of nutrients to the disc cells.

The aim of this study was to measure diffusion coefficients of solutes through the disc in relation to molecular weight.

The intervertebral disc is avascular thus nutrients and other factors from the blood supply are transported into the intervertebral disc by diffusive and convective flow. For small solutes such as lactate and glucose and oxygen, diffusion appears to predominate however convection may aid transport of larger molecules such as growth factors.

At present there however, there is virtually no information on diffusion of solutes of different molecular weights through the disc; this information is necessary for assessing and modelling transport pathways.

Diffusion coefficients were measured in nucleus and annulus sections of bovine intervertebral discs by a novel method which prevented tissue swelling and proteoglycan loss. Briefly strips of fluorescent or radiolabelled solute-saturated filter-paper were placed adjacent to the disc and the resulting concentration gradients measured at appropriate times. Solute sizes from 0.01 to 70 kDa were investigated. All results are reported as mean + s.e.m (n=6).

Diffusion coefficients (D) fell steeply with increase in molecular weight following a log-log relationship as predicted by theory. For small solutes (lactate) D for the outer annulus was 3.4 ± 1.1.10⁻⁶ cm²/sec while for 70 kDa dextran, D was 1.4 ± 0.6.10⁻⁷. There was no significant difference between values of D for nucleus and outer annulus for any solute.

Diffusion coefficients through the disc follow relationships seen in other cartilages and are dependant on tissue properties and molecular weight. The similarities between values for nucleus and outer annulus demonstrate the conflicting roles of proteoglycan and water contents in governing diffusion through the matrix with D decreasing both with increase in proteoglycan and decrease in water content.