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Normal age-related viscoelastic properties of chondrons and chondrocytes isolated from rabbit knee.

BACKGROUND
The mechanical microenvironment of the chondrocytes plays an important role in cartilage homeostasis and in the health of the joint. The pericellular matrix, cellular membrane of the chondrocytes, and their cytoskeletal structures are key elements in the mechanical environment. The aims of this study are to measure the viscoelastic properties of isolated chondrons and chondrocytes from rabbit knee cartilage using micropipette aspiration and to determine the effect of aging on these properties.


METHODS
Three age groups of rabbit knees were evaluated: (1) young (2 months, n = 10); (2) adult (8 months, n = 10); and (3) old (31 months, n = 10). Chondrocytes were isolated from the right knee cartilage and chondrons were isolated from left knees using enzymatic methods. Micropipette aspiration combined with a standard linear viscoelastic solid model was used to quantify changes in the viscoelastic properties of chondrons and chondrocytes within 2 hours of isolation. The morphology and structure of isolated chondrons were evaluated by optical microscope using hematoxylin and eosin staining and collagen-6 immunofluorescence staining.


RESULTS
In response to an applied constant 0.3 - 0.4 kPa of negative pressure, all chondrocytes exhibited standard linear viscoelastic solid properties. Model predictions of the creep data showed that the average equilibrium modulus (E(∞)), instantaneous modulus (E(0)), and apparent viscosity (m) of old chondrocytes was significantly lower than the young and adult chondrocytes (P < 0.001); however, no difference was found between young and adult chondrocytes (P > 0.05). The adult and old chondrons generally possessed a thicker pericellular matrix (PCM) with more enclosed cells. The young and adult chondrons exhibited the same viscoelastic creep behavior under a greater applied pressure (1.0 - 1.1 kPa) without the deformation seen in the old chondrons. The viscoelastic properties (E(∞), E(0), and m) of young and adult chondrons were significantly greater than that observed in young and adult cells, respectively (P < 0.001). The adult chondrons were stiffer than the young chondrons under micropipette aspiration (P < 0.001).


CONCLUSIONS
Our findings provide a theoretical model to measure the viscoelastic properties of the chondrons as a whole unit by micropipette aspiration, and further suggest that the properties of the chondrocytes and PCM have an important influence on the biomechanical microenvironment of the knee joint cartilage degeneration that occurs with aging.