Purpose: Post-traumatic osteoarthritis (PTOA) occurs after physical injury to a joint. To mimic this disease, the most widely published in vivo model is ACL surgical transection. However, this procedure does not simulate the normal development of this disease, as it is an invasive process cutting through the skin and joint capsule, which affects the native biological response to knee injury. Moreover, there is a compelling need to detect PTOA at an early stage due to its progressive pathogenic mechanism. An early sign of cartilage degeneration is swelling which leads to lower mechanical properties of the cartilage. Hence, the aim of this study is first to replicate a non-invasive ACL rupture (Maerz-2015) and secondly to investigate the early early effects of a ligamentous injury on cartilage mechanical properties using 3D indentation mapping.

Objective: To evaluate cross-correlations of ex vivo electromechanical properties with cartilage and subchondral bone plate thickness, as well as their sensitivity and specificity regarding early cartilage degeneration in human tibial plateau. Method: Six pairs of tibial plateaus were assessed ex vivo using an electromechanical probe (Arthro-BST) which measures a quantitative parameter (QP) reflecting articular cartilage compression-induced streaming potentials. Cartilage thickness was then measured with an automated thickness mapping technique using Mach-1 multiaxial mechanical tester. Subsequently, a visual assessment was performed by an experienced orthopedic surgeon using the International Cartilage Repair Society (ICRS) grading system. Each tibial plateau was finally evaluated with mCT scanner to determine the subchondral-bone plate thickness over the entire surface. Results: Cross-correlations between assessments decreased with increasing degeneration level. Moreover, electromechanical QP and subchondral-bone plate thickness increased strongly with ICRS grade (p = 0.86 and p = 0.54 respectively), while cartilage thickness slightly increased (p = 0.27). Sensitivity and specificity analysis revealed that the electromechanical QP is the most performant to distinguish between different early degeneration stages, followed by subchondral-bone plate thickness and then cartilage thickness. Lastly, effect sizes of cartilage and subchondral-bone properties were established to evaluate whether cartilage or bone showed the most noticeable changes between normal (ICRS 0) and each early degenerative stage. Thus, the effect sizes of cartilage electromechanical QP were almost twice those of the subchondral-bone plate thickness, indicating greater sensitivity of electromechanical measurements to detect early osteoarthritis. Conclusion: The potential of electromechanical properties for the diagnosis of early human cartilage degeneration was highlighted and supported by cartilage thickness and mCT assessments. (c) 2017 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Material & Methods: Electromechanical ex vivo mappings of articular surfaces were obtained in distal condyles from 5 sheep (8 – 9 y-o, 9 months post-surgery, bone marrow stimulation model) and 2 control sheep (8 years old) with the Arthro-BST (Biomomentum, Laval) which has a hemispherical indenter (r=3.175mm) to measure electromechanical properties by the quantitative parameter (QP), followed by automated indentation mappings to 50 μm with the Mach-1 (Biomomentum, Laval) with a spherical indenter (r=0.5mm) at the same positions to measure structural stiffness (load/indentation depth). Selected test sites were analyzed histologically and by unconfined compression.

Purpose: The purpose of the current study is to investigate whether electromechanical properties of articular cartilage in human tibial plateau correlated with histological scores and unconfined compression parameters as in human distal femurs (Sim et al., 2014). Methods: Right and left tibial plateaus from six human donors (5 males and 1 female, average age 48 years) were provided by RTI Surgical (Alachua, FL, USA). Ex vivo mappings of electromechanical properties of entire articular surfaces were performed using the Arthro-BST. The device calculates a quantitative parameter (QP) of cartilage electromechanical activity corresponding to the number of microelectrodes in contact with the cartilage when the sum of their streaming potential reaches 100 mV. A high QP indicates weak electromechanical properties and vice-versa. Subsequently, osteochondral cores were harvested from normal and visually abnormal regions on the tibial plateau. A total of 56 cores were tested in unconfined compression to obtain the fibril modulus (Ef), equilibrium modulus (Em), and permeability (k) prior to histoprocessing. Safranin O-Fast Green-stained paraffin sections were scored with the Mankin histological-histochemical grading system. The electromechanical QP corresponding to each cored site was calculated as the average of all QPs measured within 6 mm from the core center location. Results: As the electromechanical QP increases, safranin O/Fast Green stained sections showed that GAG staining in the cartilage matrix and structural integrity decreases (Fig. 1). A strong correlation was found between electromechanical QP and Mankin score (r = 0.50, p = 0.0004). A strong correlation was also found between QP and Ef (r = −0.73, p < 0.0001; Fig. 2A) and QP and permeability (log k) (r = 0.64, p < 0.0001; Fig. 2B). Weak correlations were found between QP and Em (r = −0.30, p = 0.0186; Fig. 2C) and between QP and thickness of the articular cartilage (r = 0.42, p = 0.0006; Fig. 2D). As expected, the electromechanical QP decreases with increasing Ef and Em whereas the electromechanical QP increases with Mankin Score, permeability and thickness.Figure 2(A) Correlation between the electromechanical QP and fibril modulus Ef;. (B) Correlation between the electromechanical QP and permeability logk; (C) Correlation between the electromechanical QP and matrix modulus Em; (D) Correlation between the electromechanical QP and cartilage thickness.View Large Image Figure ViewerDownload Hi-res image Download (PPT) Conclusions: The electromechanical QP measured in human tibial plateau correlated significantly with the histological Mankin score and with unconfined compression mechanical parameters, similar to what was previously seen in human distal femurs. Moreover, a weak correlation was found between the thickness of the cartilage on the tibial plateau and the electromechanical QP. This is in contrast to what was previously observed in the distal femurs but could be related to topographical variations present on the tibial plateau surface, where cartilage is very thin and rigid in regions covered by the meniscus but thicker and softer in regions not covered by meniscus. Considering these results, we believe that the Arthro-BST can provide a reliable tool for articular cartilage assessment on human tibial plateau.

Purpose: Due to their size (∼1mm), mouse models pose significant challenges to map biomechanical properties over their articular surfaces. The purpose of this study was to determine if an automated indentation technique could be used to map the biomechanical properties of the articular surfaces in murine knees and to identify early alterations of the articular cartilage of a mouse strain (STR/ort) that spontaneously develops osteoarthritis (OA) on the medial side of their knees. Methods: The biomechanical measurements were performed ex vivo, on the left femoral condyles and tibial plateaus of five healthy Balb/c males (age of 12-15 weeks) and five age- and sex-matched STR/ort mice, using a 3-axis mechanical tester (Mach-1 v500css, Biomomentum, Laval) equipped with a multiple-axis load cell. Indentation measurements (30-42/surfaces) were performed using a 0.35 mm diameter spherical indenter (30 μm indentation in 1 second with 20 seconds relaxation). Following biomechanical testing, the articular surfaces were fixed in 4% paraformaldehyde for histological assessment. Data reported is the structural stiffness at an indentation depth of 10μm. To compare the structural stiffness between healthy and OA-developing animals each articular surface were divided into 4 regions, exterior medial (I), inner medial (II), inner lateral (III) and exterior lateral (IV). Data is reported as the mean ± SE (n= 5) for each of these regions. Statistical analysis was performed by ANOVA. Results: In healthy animals, mapping of the structural stiffness at an indentation depth of 10 μm showed a spatial distribution similar to that of larger animals (Figure 1A,B, inserts). The structural stiffness of the lateral and inner half of the medial condyles (Figure 1A) was similar in OA and healthy mice. The stiffness of the exterior lateral plateau was also not significantly different OA and healthy mice. In contrast, the stiffness of the exterior half of the medial condyle in OA mice (5.9±0.7 N/mm) was significantly lower than that of the healthy mice (10.2±1.1 N/mm, ANOVA, p<0.05). The structural stiffness of the exterior medial condyle and the inner half of the lateral plateaus in OA mice was inferior to that of healthy mice (ANOVA, p<0.05). Conclusions: This study shows that this automated indentation technique can map the biomechanical properties of murine knee joints. The mapping of mechanical properties shows similar distribution patterns to those previously observed for larger species (human, sheep and rat). The identification of cartilage regions with lower structural stiffness, at sites known to develop OA in the STR/ort strain, suggests this method can be used to identify and characterize OA affected articular surfaces. Studies are ongoing to validate, by histology, the cartilage quality of the affected areas. These results show that indentation mapping could be used in mouse models to test the efficacy of drugs aiming to inhibit cartilage degradation or improving its healing in OA or following a joint injury.

Purpose: An important measure of articular cartilage function in health and disease is its biomechanical properties. While much research has mouse models of osteoarthritis, the assessment of biomechanical properties in these small joints is quite challenging. We have previously developed novel and easily implemented indentation technique on sheep stifle joints and rat knee joints. The purpose of this study was to determine if this novel technique could be used to map the biomechanical properties of entire mice articular surfaces in indentation. Methods: Wild-type mice of different age were used for this study. The left and right femoral condyles and tibial plateaus of an 8-week-old C57BL/6 male mouse were first biomechanically mapped in vitro. The mechanical tester used was a 3-axis Mach-1 v500css from Biomomentum Inc. A spherical indenter (Fig. 1) of 0.35 mm diameter (tolerance of ± 2.5 μm and sphericity of 0.625 μm) was use for the indentation measurement with a multiple-axis load cell of 7.0 kg range and 350 mg z-resolution. A 26G intradermal bevel needle was used for the thickness measurement. Following biomechanical testing, the articular surface was fixed in 4% PFA for histological assessment. Results: The normal force versus time curve (Fig. 2) shows the typical behavior of articular cartilage in a stress relaxation configuration. High-resolution maps of the load at 10μm indentation depth were obtained for the tibial plateau and femoral condyles (Fig. 3). The average load at 10μm of indentation depth (mean ± SD) for the tibial plateau was 5.7 ± 2.7 g and 7.8 ± 4.1 g for the femoral condyles. It takes approximately 1 minute for each indentation measurement and 30 seconds for each thickness measurement (≈25 positions for each condyles and ≈ 40 positions for each tibial plateau). Conclusions: The mapping of biomechanical properties of cartilage using indentation is applicable to articular cartilage as thin as mice with any type of articular surface curvature, such as the tibial plateau or femoral condyle. We are currently comparing the thickness found by this novel method with the thickness on histological slides to confirm cartilage thickness maps and cartilage stiffness maps calculated using an elastic model developed by Hayes in 1972. Ongoing studies include examination of the effect of age and gene modifications (knock-out models) on indentation properties of articular cartilage. This novel indentation method will provide comparative biomechanical properties of articular cartilage in different OA models by reliably measuring the biomechanical properties of entire articular surfaces.Figure 2Typical stress relaxation (Normal force vs. Time) curve of articular cartilage obtained with a 20 μm indentation depthView Large Image Figure ViewerDownload Hi-res image Download (PPT)Figure 3Load at 10 μm of indentation depth mappings obtained on an 8-week-old wild-type mouse (right and left joints has been averaged)View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Objective: The hand-held Arthro-BST (TM) device is used to map electromechanical properties of articular cartilage. The purpose of the study was to evaluate correlation of electromechanical properties with histological, biochemical and biomechanical properties of cartilage.Method: Electromechanical properties (quantitative parameter (QP)) of eight human distal femurs were mapped manually ex vivo using the Arthro-BST (1 measure/site, 5 s/measure, 3209 sites). Osteochondral cores were then harvested from different areas on the femurs and assessed with the Mankin histological score. Prior to histoprocessing, cores were tested in unconfined compression. A subset of the cores was analyzed with polarized light microscopy (PLM) to assess collagen structure. Biochemical assays were done on additional cores to obtain water content and glycosaminoglycan (GAG) content. The QP corresponding to each core was calculated by averaging all QPs collected within 6 mm of the core center.Results: The electromechanical QP correlated strongly with both the Mankin score and the PLM score (r = 0.73, P < 0.0001 and r = -0.70, P < 0.0001 respectively) thus accurately reflecting tissue quality and collagen architecture. Electromechanical QP also correlated strongly with biomechanical properties including fibril modulus (r = -0.76, P < 0.0001), matrix modulus (r = -0.69, P < 0.0001), and log of permeability (r = 0.72, P < 0.0001). The QP correlated weakly with GAG per wet weight and with water content (r = -0.50, P < 0.0003 and r = 0.39, P < 0.006 respectively).Conclusion: Non-destructive electromechanical QP measurements correlate strongly with histological scores and biomechanical parameters providing a rapid and reliable assessment of articular cartilage quality. (C) 2014 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.