Alteration of viscoelastic properties is associated with a change in cytoskeleton components of ageing chondrocytes from rabbit knee articular cartilage

The cytoskeleton network is believed to play an important role in the biomechanical properties of the chondrocyte. Ours and other laboratories have demonstrated that chondrocytes exhibit a viscoelastic solid creep behavior in vitro and that viscoelastic properties decrease in osteoarthritic chondrocytes. In this study, we aimed to understand whether the alteration of viscoelastic properties is associated with changes in cytoskeleton components of ageing chondrocytes from rabbit knee articular cartilage. Three age groups were used for this study: young (2-months-old, N=23), adult (8-months-old, N=23), and old (31-months-old, N=23) rabbit groups. Cartilage structure and proteoglycan and type II collagen content were determined by H&E and Toluidine Blue staining, and type II collagen antibody. The detailed structure of the chondrocytes in all groups was visualized using transmission electron microscopy (TEM). Chondrocytes were isolated from full-thickness knee cartilage of rabbits from all groups and their viscoelastic properties were quantified within 2 hours of isolation using a micropipette aspiration technique combined with a standard linear viscoelastic solid model. The components and network of the cytoskeleton within the cells were analyzed by laser scanning confocal microscopy (LSCM) with immunofluorescence staining as well as real time PCR and western blotting. With ageing, articular cartilage contained less chondrocytes and less proteoglycans and type II collagen. TEM observations showed that the cell membranes were not clearly defined, organelles were fewer and the nuclei were deformed or shrunk in the old cells compared with the young and adult cells. In suspension, chondrocytes from all three age groups showed significant viscoelastic creep behavior, but the deformation rate and amplitude of old chondrocytes were increased under the same negative pressure when compared to young and adult chondrocytes. Viscoelastic properties of the old cells, including equilibrium modulus (E infinity), instantaneous modulus (E0) and apparent viscosity (mu) were significantly lower than that those of the young and adult ones (P < 0.001). No significant differences were detected between young and adult chondrocytes (P > 0.05). Moreover, we found that the cytoskeletal networks of old cells were sparser, and that the contents of the various components of the intracellular networks were reduced in old cells, compared with adult and young cells. Aged chondrocytes had a different response to mechanical stimulation when compared to young and adult chondrocytes due to alteration of their viscoelastic properties, which was in turn associated with changes in cell structure and cytoskeleton composition.