Determination of kinetic changes of aggrecan-hyaluronan interactions in solution from its rheological properties

Abstract

The kinetics of interactions between aggregating cartilage proteoglycan (aggrecan) and hyaluronan was examined through their rheological flow behavior using a cone on plate viscometer. The mixing of the two types of molecules was carried out directly on the plate of the viscometer, and aggregation process was monitored through the changes of the sample's steady-shear viscosity and/or dynamic shear modulus as a function of time. The effect of flow conditions on the aggrecan-hyaluronan interaction rates was examined by subjecting samples to steady-shearing motions at specified shear rates, and to oscillatory shear motions of specified frequencies and amplitudes. The characteristics of the kinetics of interaction between aggrecan and hyaluronan molecules depended not only on the flow conditions under which proteoglycan aggregation took place, but also on the concentration of the components in the solution. At high shear rates (> 10 s −1), viscosity of the mixture solution increased monotonically, starting near the viscosity of the aggrecan solution, and reaching the viscosity of the aggregate solution in approximately 35 min. Surprisingly, under slow shearing motions (< 10 s −1), the viscosities of the mixture solutions exceeded those of control aggregate solutions at identical hyaluronan: aggrecan ratios and concentrations. In addition, the aggregation under oscillatory motions took place near physiologic frequency (10 rad s −1) although the rate of aggregation process was much slower than under steady-shearing motion (> 100 min). However, the high-frequency oscillatory shearing (62.8 rad s −1) tended to impede aggregation resulting in a reduction of dynamic modulus over time. The influence of loading conditions on the rate of aggregation and aggregate size observed in this study seems to suggest a close relationship between proteoglycan structure and content, and degree of physiological stress throughout the joint and frequency of the joint motion.