Injection of recombinant interleukin-1β (reIL-1β) is commonly used in equine joint research to evaluate the clinical and biological outcomes of inflammation, often tracking changes over time to assess the effects of the experimental model. Incorporating functional assessments, such as gait asymmetry analysis, in conjunction with biological assessments, is essential for capturing the real-world impact of joint interventions and ensuring findings translate to meaningful improvements in equine health and locomotion. This study aimed to examine the relationship between synovial fluid biochemical changes and symmetry analysis (inertial measurement unit; IMU) following a 75-ng injection of recombinant interleukin-1β (reIL-1β) into the left intercarpal joint of 11 horses Synovial fluid samples were collected at 0 h (pre-injection; control) and at 6 h and 12 h post-injection and analyzed for prostaglandin E2 (PGE2), glycosaminoglycan (GAG), nitric oxide (NO), chondroitin sulfate epitope 846 (CS846), and resolvin D1 (RvD1). The IMU data were collected at baseline (hour 0) and 6 and 12 h post-injection to assess poll and pelvic movement during in-hand trot trials, with MinDiff (loading asymmetry) and MaxDiff (push-off asymmetry) values calculated. A linear mixed model was used to assess the association between biochemical outcomes and asymmetry measures with significance set at P ≤ 0.05. Increased synovial PGE2 significantly contributed to greater poll MinDiff (estimate = 0.007, SE = 0.003, P = 0.006), whereas elevated CS846 was a key contributor to increased poll MaxDiff (estimate = 0.006, SE = 0.003, P = 0.036). Additionally, higher PGE2 was linked to lower pelvis MaxDiff (estimate = −0.002, SE = 0.001, P = 0.024), whereas increased CS846 was associated with higher pelvis MinDiff (estimate = 0.003, SE = 0.001, P = 0.026). This research emphasizes the value of integrating biochemical and functional data to provide a more comprehensive tool for evaluating interventions for joint health
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