Local shear properties of rabbit articular cartilage capture surface region mechanics of human, equine, and bovine tissue

Abstract

New Zealand white rabbits are a prevalent model species used to study preclinical articular cartilage repair therapies. The composition and structure of rabbit articular cartilage have been extensively characterized, yet the local shear properties of the tissue are unknown. Characterizing the local shear properties is essential for understanding the structure-function relationship in the tissue and relating the rabbit preclinical model to human disease. Therefore, the objectives of this study were to (1) characterize the local shear properties of articular cartilage from the femoral condyles of New Zealand white rabbits, (2) determine if local protein content or matrix structure correlated with local shear properties, and (3) compare microscale shear moduli values of rabbit cartilage to those previously reported for human, equine, and bovine tissues. Local shear strains and moduli varied with rabbit cartilage tissue depth; shear modulus was highest ∼ 50 µm below the tissue surface and decreased to plateau values around 150 µm, mirroring the trend with shear strains. Local shear strains showed significant correlations with local protein content but not matrix organization. Rabbit cartilage shear properties followed similar spatial trends as bovine, equine, and human tissue in the first ∼ 100 um of the tissue depth. However, rabbit tissue then differentiated from the larger animals as shear modulus values plateaued and did not increase by an order of magnitude like that seen in the larger species. Local shear properties of rabbit articular cartilage capture the surface properties of human, equine, and bovine cartilage but mechanically lack the deep zone region.

Keywords: Arthritis; Mechanics; Microscale; Osteoarthritis; Preclinical model; Strain; Zone.

Fig. 1.. Microscale Shear Properties Vary with Tissue Depth.

Photobleached lines were created in the rabbit cartilage prior to shear loading to track tissue displacements (A). Photobleached lines in the sample were tracked to obtain tissue displacements and calculate shear strain (B) and shear modulus (C) as a function of tissue depth. Shapes denote individual animals and the black line denotes the geometric average (n = 4).

Fig. 2.. DTAF Staining of Protein Captures Variation in Fibrillar Collagen with Depth.

Cartilage tissue was histologically assessed for GAGs (A) and fibrillar collagen (BC). GAG concentration is highest in the middle of the tissue and notably absent at the surface and deep/calcified region of the tissue (A). Fibrillar collagen staining is enriched where GAGs are absent in the surface and deep zones of the tissue (BC). DTAF staining for protein shows the same depth-dependent staining intensity as Picrosirius Red where staining is highest at the surface and deep zones of the tissue (D). The DTAF sample is thinner compared to the histology images as it has been subjected to ~ 12 % axial compression.

Fig. 3.. Quantification of Protein Concentration and Matrix Organization with Depth through DTAF Staining.

From confocal images of tissue stained with DTAF, FFTs were calculated along the depth of the tissue to characterize matrix organization (A). Protein intensity increased ~ 50 μm from the surface and then decreased to plateau values at depths greater than 150 μm (B). The orientation index calculated from FFTs were highest at the surface and then decreased through the depth of the tissue (C). The orientation angle decreased at the surface until a depth 75 μm and then increased to plateau values for the remaining depth of the sample (D). Shapes denote individual animals. Black lines denote the geometric average for DTAF intensity and orientation index and the arithmetic mean for orientation angle due to the zero values (n = 4).

Fig. 4.. Microscale Shear Mechanics Correlate with Matrix Composition but Not Matrix Organization Measures.

Microscale shear modulus and shear strains had significant correlations with matrix composition (A,D), but not matrix organization measures (B,C,E,F). Shear strains decreased with increased protein concentration (DTAF intensity, A). Shear modulus measures showed similar trends to the shear strain data, but these correlations did not have slopes that were significantly different than zero (D-F). Shape and color denote individual animals (n = 4). R² and slope (m) values are from linear correlations and are bolded if statistically significant (p < 0.05).

Fig. 5.. Rabbit Tissue Captures Surface and Initial Middle Zone Shear Properties of Adult Human, Adult Equine, and Neonatal Bovine Cartilage.

Average shear modulus for rabbit cartilage was plotted along with microscale shear modulus measurements from adult human, adult equine, and neonatal bovine cartilage reported in the literature. Color denotes species.