Viscosupplementation is a symptomatic treatment of osteoarthritis (OA) intended to restore rheologic homeostasis of the synovial fluid by injecting hyaluronic acid intraarticularly. Despite the long history of this therapy, little is known about its mechanisms of action and differences between commercial preparations. We investigated the rheologic behavior of OA synovial fluid with time, when stored at 4°C, before and after the addition of two hyaluronic acid commercial preparations (linear and cross-linked). Thirteen OA synovial fluids were stored at 4°C and assayed using steric exclusion chromatography, which allows hyaluronic acid to be separated from the remaining pool of proteins and its molecular weight and concentration to be determined without any pretreatment and calibration. The synovial fluid rheology also was studied in vitro, before and after addition of two viscosupplements, over 6 weeks. The non-Newtonian behavior of synovial fluid throughout followup appears to be the result of loose interactions between proteins and hyaluronic acid. When mixed with the linear hyaluronic acid, synovial fluid becomes less non-Newtonian whereas the non-Newtonian behavior was reinforced when mixed with the cross-linked hyaluronic acid. The rheology was nearly unchanged for all synovial fluids over 6 weeks. Our preliminary trial shows it is possible to study synovial fluid, stored at 4°C, over a long time and suggests the enzymatic degradation of hyaluronic acid is negligible under these experimental conditions.

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Different pre-treatments applied to synovial fluids (SF) before their analyses are tested to characterize SF after storage under different conditions and to investigate their evolution along a viscosupplementation treatment. The main techniques proposed involve steric exclusion chromatography with triple detection (SEC) and viscometry; it is the first time that such a study is developed. SEC gives the molecular weight distribution and concentration of hyaluronan (HA) and proteins separately; the steady state viscosity is always non-Newtonian and not directly related to SF composition. Pre-treatment of SF (storage in cold, filtration, centrifugation) allows us to conclude that, in order to store SF, it is best to freeze it, even if in some cases, viscosity is modified but not the composition. All the data obtained (including protease pre-treatment) allow us to conclude that a small fraction of HA-protein complex forms a loose 3D-network and controls the rheology.

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