Poster AA3 - The dynamic change of cartilige mechanical properties under reputitive shear load

Introduction: Articular cartilage is a complex tissue renowned for its capacity to furnish a low-friction, highly durable, wear-resistant surface[1]. The overuse of articular cartilage, such as repetitive loading of joints, may cause cartilage wear or degradation. Previous research applying both compression and shear loads to cartilage has found that combined loading of articular cartilage can cause damage to the surface region by shear fatigue loading[2].
Additionally, cartilage contains distinct depth dependent zones, each characterized by unique cellular morphology and mechanical properties[1]. Research has explored the effects of shear loading on the depth dependent mechanics of cartilage. Studies indicate that even slight changes in collagen or aggrecan concentrations, typical in the early stages of cartilage disease, cause significant weakening of the shear modulus, potentially leading to tissue collapse[3]. However, depth dependent changes in cartilage mechanics due to repetitive shear loading have yet to be identified.
Our research quantifies the depth dependent shear mechanical properties as repetitive shear load is applied to the surface of articular cartilage. We hypothesize that the mechanical properties of knee cartilage undergo consistent alterations under repetitive shear loading, particularly at the superficial zone. This understanding of the degenerative progression of articular cartilage holds promise for advancing early diagnosis and treatment strategies for osteoarthritis.

Materials and

Methods: Cylindrical explants (3 mm in height and 5 mm in diameter) were excised from the femoral condyles of three-day-old bovine knee joints and bisected to create a flat surface for confocal microscopy. The samples were stained with a 14μg/mL D-TAF solution for 20 minutes and rinsed in PBS to remove excess dye.
Samples were mounted onto a Tissue Deformation Imaging Stage (TDIS), with the deep zone (cartilage adjacent to the bone) secured to the stationary plate using super glue. Shear oscillations were imparted onto the sample's surface via the moving plate. Prior to shear loading, a 15% compressive load was administered to prevent relative motion between the moving plate and the sample. Subsequently, a 1Hz oscillatory shear with a peak 2% shear strain was applied to the sample's surface for a duration of 8 hours.
At intervals of 0 hours, 4 hours, and 8 hours, the samples were imaged using a Zeiss 700 LSM confocal microscope with a 10x objective, employing a 0.0125 Hz oscillatory shear with a peak 2% strain. Five parallel lines were photo-bleached onto the flat surface of the cartilage for automated computer tracking of strain (Fig 1a). Shear forces were measured and used to calculate shear stress. Throughout the mechanical testing process, the samples were immersed in PBS to maintain tissue hydration. Shear stress versus shear strain curves were plotted for the surface region of cartilage. The toe modulus, high strain region modulus, and effective shear modulus were calculated (Fig 2b).

Results, Conclusions, and Discussions:

Results:
Raw shear displacement amplitudes decrease faster in the surface region ( < 150um) of the 8-hour sample than in 0- or 4-hour samples (Fig 1b). This faster decrease causes the middle zone of the cartilage (>150um) to have a lower displacement after 8 hours than the 0-hour sample. By deriving the first derivative of an exponential fit to the displacement versus depth plots, we obtained depth-dependent cartilage strain profiles (Fig 1c). Strain amplitudes are largest at the surface layer ( < 150um) following 8 hours of repetitive loading, while deeper layers (>250μm) show negligible variation across different loading times. Little to no differences in shear stresses on the cartilage boundaries are noted based on loading duration. Since the middle zone of the cartilage showed no differences in the stress or the strain, all further analyses focused on changes in the surface region of the cartilage. From the stress-strain curves of the cartilage surface (Fig 2a), changes in the shape and degree of nonlinearity are observed between 0, 4, and 8 hours of loading. The toe modulus (Fig 2c) and the high strain modulus (Fig 2d) decrease with increased loading durations. The effective modulus does not change with increased loading duration (Fig 2e).

Conclusion &

Discussion:
In contrast to other shear fatigue studies, this study showed changes in the depth-dependent mechanics due to repetitive shear loading, which is common in joint overuse. Specifically, the mechanical properties of the superficial layer of samples were altered, particularly with extended loading durations (8 hours).
Given cartilage's viscoelastic nature, the low loading frequency during imaging and mechanical measurements allows water molecules adequate time to permeate the fiber network. This ensures the observed mechanical response of our cartilage is based mainly on changes in the solid components. Since the surface of the articular cartilage contains mostly collagen, changes in the collagen are most likely occurring during shear fatigue loading.
Our results show that dynamic shear loading of articular cartilage at physiologic levels can cause altered mechanical properties after only 8 hours, providing further evidence to understand fatigue and wear-induced damage mechanisms of articular cartilage and osteoarthritis development.

Acknowledgements (Optional): Reference:
[1] J. Eschweiler et al., “The Biomechanics of Cartilage—An Overview,” Life, vol. 11, no. 4, Art. no. 4, Apr. 2021, doi: 10.3390/life11040302.
[2] S. R. Oungoulian, K. M. Durney, B. K. Jones, C. S. Ahmad, C. T. Hung, and G. A. Ateshian, “Wear and damage of articular cartilage with friction against orthopedic implant materials,” J. Biomech., vol. 48, no. 10, pp. 1957–1964, Jul. 2015, doi: 10.1016/j.jbiomech.2015.04.008.
[3] T. Wyse Jackson et al., “Structural origins of cartilage shear mechanics,” Sci. Adv., vol. 8, no. 6, p. eabk2805, Feb. 2022, doi: 10.1126/sciadv.abk2805.