Acute joint pathology and synovial inflammation is associated with increased intra-articular fracture severity in the mouse knee

Abstract

Objective: Post-traumatic arthritis is a frequent cause of disability and occurs most commonly and predictably after articular fracture. The objective of this investigation was to examine the effect of fracture severity on acute joint pathology in a novel murine model of intra-articular fracture.

Design: Low and high energy articular fractures (n=25 per group) of the tibial plateau were created in adult male C57BL/6 mice. The acute effect of articular fracture severity on synovial inflammation, bone morphology, liberated fracture area, cartilage pathology, chondrocyte viability, and systemic cytokines and biomarkers levels was assessed at 0, 1, 3, 5, and 7 days post-fracture.

Results: Increasing intra-articular fracture severity was associated with greater acute pathology in the synovium and bone compared to control limbs, including increased global synovitis and reduced periarticular bone density and thickness. Applied fracture energy was significantly correlated with degree of liberated cortical bone surface area, indicating greater comminution. Serum concentrations of hyaluronic acid (HA) were significantly increased 1 day post-fracture. While articular fracture significantly reduced chondrocyte viability, there was no relationship between fracture severity and chondrocyte viability, cartilage degeneration, or systemic levels of cytokines and biomarkers.

Conclusions: This study demonstrates that articular fracture is associated with a loss of chondrocyte viability and increased levels of systemic biomarkers, and that increased intra-articular fracture severity is associated with increased acute joint pathology in a variety of joint tissues, including synovial inflammation, cortical comminution, and bone morphology. Further characterization of the early events following articular fracture could aid in the treatment of post-traumatic arthritis.

Figure 1

Histopathological assessment of synovitis (modified from Krenn et al. Histopathology 2006). Enlargement of the synovial lining cell layer and density of the cells in the synovial stroma were individually assessed from H&E histology sections of mouse knee joints (magnification 100-200×).

Figure 2

Low energy (A) and high energy (B) fractures of the mouse tibial plateau. Radiograph (left), micro computed tomography (microCT) three-dimensional rendered image (middle, scale bar = 1mm), and microCT cross-section (right, scale bar = 1mm) demonstrating joint morphology of fractures at t=0 day post-fracture.

Figure 3

Chondrocyte cell death as assessed by NBT nuclear stain (n=3 per group per timepoint). Vertical bars denote ± 95% CI. A. The total joint score (maximum possible total joint score = 20) of cell death was greater in the fractured experimental limb than the contralateral control limb in both the anterior and central regions of the knee joint (*p=0.014 for anterior region, p=0.016 for central region). B. Cell death by location in the joint (maximum possible site score = 5) was greater in the fractured experimental limb than the contralateral control limb (* p=0.003 for lateral femur, p=0.04 for medial femur, p=0.001 for lateral tibia), and cell death was significantly greater in the lateral tibial than the medial tibia in the fractured experimental limb (p=0.0002) for the anterior region of the knee. For all measures of chondrocyte cell death there was no significant difference between low energy and high energy fractures.

Figure 4

A. MicroCT images were segmented using a semi-automated custom software in both the intact contralateral control limb (left) and fractured experimental limb (right). These models were then registered to each other using an iterative closest point technique, and were used to calculate liberated surface area. B. Fracture severity, as measured from the liberated surface area, was well correlated to the energy of fracture, as calculated from the load-displacement data (R²=0.70).

Figure 5

Assessment of synovitis in low and high energy fractures (n=5 per group per timepoint). A. Synovitis scores at 0 and 7 days post-fracture. Vertical bars denote ± 95% CI. B. Representative histologic sections of the synovial insertion (Sy) at the medial tibia (MT) and lateral tibia (LT) in contralateral control and fractured experimental limbs at 7 days post-fracture (H&E stain; 400× magnification).