X-ray imaging of articular cartilage using anionic contrast agents has been introduced for quantification of tissue glycosaminoglycan (GAG) concentration. In this in vitro study we investigated diffusion and equilibrium distribution of an anionic contrast agent in human articular cartilage and related the results to tissue composition and integrity.
Osteochondral cylinders (d=4.0mm, n=24) were prepared from femoral medial condyles (FMCs, cartilage thickness 2.13+/-0.54 mm, mean+/-standard deviation [SD]), and tibial medial plateaus ([TMPs]1.99+/-0.38 mm) of human cadaver knees. Samples were immersed for 24h at room temperature in 21 mM concentration of anionic contrast agent Hexabrix. The X-ray absorption maps and profiles were measured before immersion, and after every 2h of immersion using clinical peripheral quantitative computed tomography (pQCT).
An increase in X-ray attenuation along cartilage depth, indicating a characteristic density profile increasing from superficial to deep tissue, could be seen in pQCT images acquired without contrast agent. The complete diffusion of the contrast agent into cartilage took more than 12h. However, the uronic acid concentration correlated with the contrast agent concentration in femoral cartilage (r=-0.76, n=12, P=0.004) as early as after 2h of immersion, and the linear correlation was virtually unchanged during the remaining 22 h. Similarly, the histological tissue integrity (Mankin score) correlated positively with the contrast agent concentration in tibial cartilage (r=+0.75, P=0.005) after 2h of immersion. The X-ray absorption profiles before immersion, i.e., without the contrast agent, and after 24h of immersion were significantly correlated (r=-0.76+/-0.34, mean+/-SD).
Although the complete contrast agent diffusion into human articular cartilage in vitro took more than 12h, significant apparent correlations were revealed between the spatial proteoglycan (PG) and contrast agent distributions already after 2h of immersion. At the stage of incomplete penetration, however, the spatial contrast agent concentration distribution cannot directly reflect the true PG distribution as the Donnan equilibrium has not been reached. However, in degenerated cartilage the diffusion rate increases. Obviously, this can lead to the reported correlation between the bulk PG content and the bulk contrast agent concentration already at the early stages of diffusion.