Identification of concentric lamellae in the articular calcified cartilage of osteoarthritic mice

Abstract

Purpose: The structure, ultrastructure and function of hyaline articular cartilage (HAC) and subchondral bone (SCB) have been the subject of much investigation and their potential involvement in the pathogenesis of osteoarthritis (OA) has been widely recognised. However much less attention has been focussed on the intervening tissue, articular calcified cartilage (ACC). The ultrastructure and cell biology of ACC is poorly described and it is not known to what extent it is involved as in the initiation and progression of OA. Examination of ACC by high quality light microscopy revealed the presence of distinct morphological features around the chondrocytes in the calcified cartilage of mice. This prompted us to undertake a transmission electron microscopy (TEM) study of ACC in wild type (WT) mice and in mice with genetic or environmentally-induced OA to identify changes which may further our understanding of the role played by ACC in the early stages of OA. Methods: Aged WT and OA mice were sacrificed, their knee’s fixed in either PBFS or glutaraldehyde for 24 hours, and decalcified for 7 days in EDTA. Samples underwent routine TEM processing, sectioning with an ultramicrotome, and post-staining with uranyl acetate and lead citrate. Results: In both WT and OA mice, we identified the appearance of concentric lamellae surrounding chondrocytes in ACC. The lamellae appeared to be laid down in association with the advancing tidemark indicating that they are formed during the calcification of cartilage matrix. These lamellar structures were present around hypertrophic chondrocytes in the deeper zones of ACC but were also observed in association with viable chondrocytes towards the calcification front, where some chondrons appeared to be partially engulfed by lamellae. The number of lamellae per chondron varied between 5 and 20. Quantitative analysis revealed that there was an increase of lamellae with age and that there were more in degenerating joints. The composition of the lamellae has not been elucidated but collagen fibres could be detected. Conclusions: We describe the presence of concentric lamellae in ACC. These novel structures appear to play a role in mineralisation and they may be related to the lamellae detected using SEM by Hirotani et al in 1974. Those authors proposed the existence of a lamellar system around chondrocytes in the deep matrix of the articular cartilage in patients with secondary OA, but other than that this is a novel finding in ACC. The association with mineralisation and the advancement of the tidemark, and their greater abundance in OA indicate that the formation of these lamellae might be important in the pathogenesis of OA since thinning of the ACC due to advancing mineralisation is reported to be a characteristic of joints undergoing osteoarthritic degeneration. Studies are ongoing to determine the periodicity of the lamellae. Identification of the mechanism by how the lamellae are formed should provide a better understanding of the function and regulation of ACC, and its role in the initiation and progression of OA.