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Orthopedic and Rehabilitation Engineering

Orthopedic and Rehabilitation Engineering - Poster Session E

Poster AA3 - Hyaluronic Acid Modulates the Lubricating Ability of Recombinant Equine Lubricin

Saturday, October 26, 2024
10:00 AM – 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Articular cartilage withstands millions of loading cycles over its lifetime while maintaining low wear and very low friction coefficients. The synovial fluid (SF) that is present in the joint lubricates the cartilage and contributes to its remarkably low friction coefficients. At the molecular level in SF, lubrication of cartilage can be attributed to a synergistic interaction between lubricin and hyaluronic acid (HA). Using the Stribeck framework to characterize cartilage lubrication [4], lubricin acts as a boundary lubricant in SF, whereas HA acts as a viscous lubricant. Together, in healthy joints, lubricin and HA combine synergistically to lower the friction coefficient at the articular surface.

Previously, much work has been done characterizing the friction of HA and its derivatives, as they are currently used clinically in viscosupplementation [1]. A variety of lubricin formulations have also been evaluated, including purified and recombinant forms of varying size [6] [7] [8]. Furthermore, there have been studies that show HA and different lubricin forms work synergistically together [3]. Recently, a recombinant lubricin, eLub, based on the equine PRG4 gene, was shown to lubricate cartilage in a dose-dependent manner [2]. While other forms of lubricin have been shown to interact with HA, this phenomenon has not yet been shown for eLub. The objective of this study is to characterize the interactions of eLub and HA, using the Stribeck framework to evaluate the boundary and viscous friction behavior at different doses.

Materials and

Methods:  Lubricant Formulation: A 20 mg/ml solution of eLub [5] was serially diluted in half logs to obtain concentrations ranging from 0 mg/ml to 5 mg/ml in phosphate-buffered saline (PBS). The HA mixture was formulated by solubilizing HA (2.5 MDa) directly into the diluted eLub solutions, to produce final HA concentrations ranging from 0 to 4 mg/ml. Tribological testing: Friction characterization was performed using a previously described [4], cartilage on glass tribometer. Briefly, femoral condyles were dissected from neonatal bovine joints. Full-thickness cartilage plugs (6 mm diameter) were biopsy-punched from the condyles and trimmed to a thickness of 2 mm. These plugs were then glued onto a brass pivot, the articular surface mated against a polished glass surface in the lubricant bath, compressed to 30% strain, allowed to stress relax for one hour, and then reciprocally slid at speeds ranging from 0.1 to 10 mm/s. For each speed, the friction coefficient was determined by calculating the shear-to-normal load ratio, measured by a biaxial load cell. Curve Fitting: The RMS errors between the data and the 4-parameter model curves were minimized in both Fig 1A and 1B. Fig. 1A curve fits are a function of the Sommerfeld number, which is a dimensionless number that includes the lubricant’s viscosity, sliding speed, contact width, and normal load. Fig. 1B curves are fitted to an antagonist dose-response model as a function of the eLub concentration.

Results, Conclusions, and Discussions: In the absence of eLub, a classical elastoviscous Stribeck model (R-squared = .77) described changes in the friction coefficient: friction was approximately 0.25 in the boundary region (low Sommerfeld numbers), and then decreased to 0.12 in the viscous region (high Sommerfeld numbers) (Fig. 1A). At high concentrations of eLub, there was an inversion of the classical Stribeck curve: boundary friction was approximately 0.04 and viscous friction was higher than the boundary value, equal to 0.10 (Fig. 1A). As the eLub concentration increased from 0 to 5 mg/ml, there was a strong dose-response (R-squared = 0.99) drop in the boundary friction from 0.25 to 0.04 (Fig. 1B). Viscous friction did not display a strong dose-response drop as eLub concentration increased; it only decreased from 0.12 to 0.10 (R-squared = .31).

Utilizing the Stribeck framework to characterize the tribological behavior of combinations of HA and eLub, both expected and unexpected behaviors were revealed. Expectedly, increasing the concentration of eLub decreased the boundary friction (Fig. 1B). Also, as expected, the viscous mode friction coefficient remained relatively constant with an increase in eLub concentration (Fig. 1B).

However, unexpectedly, Figure 1A revealed that as eLub concentration increased, there was a deviation from classical Stribeck behavior. Instead of starting at a high value of friction in the boundary region and decreasing to a lower value in the viscous region, the boundary friction was initially low and as the Sommerfeld number increased, the friction increased and plateaued into the viscous region. This was also seen in Figure 1B, when the viscous and boundary lines cross each other, the Stribeck curve becomes inverted.

The observation that friction was lower at low Sommerfeld numbers than at high Sommerfeld when lubricin concentrations were high was not expected. Given the tendency of recombinant lubricin to self-aggregate, it is possible that highly viscous HA solutions promote this aggregation, which may affect the interactions of eLub with cartilage. Such phenomena may be mediated through molecular design, buffer composition, or lubricant processing.

Acknowledgements (Optional):  LEAP HI NSF CMMI 2245367

References: [1] Bonnevie+ PloS One, 2019; [2] Vishwanath+ JOR, (in review); [3] Bonnevie+ PloS One, 2015; [4] Gleghorn+ JOR, 2008; [5] Colville+ BioRxiv, 2024; [6] Schmidt+ Arthritis Rheum, 2007; [7] Flannery+ Arthritis Rheum, 2009; [8] Gleghorn+ JOR, 2009