Association of facial expression, biochemistry, and microRNAs with lameness in endurance horses

BACKGROUND: Lameness is common in endurance horses. Conventional gait-based assessment may not fully capture its various aspects. Facial expression scales can provide complementary, non-locomotor indicators of lameness-related pain. OBJECTIVE: To evaluate the performance of facial expression scales Horse Grimace Scale (HGS) and Equine Utrecht University Scale for Facial Assessment of Pain (EQUUS-FAP) for assessing pain-related lameness in endurance horses and their associations with the American Association of Equine Practitioners (AAEP) lameness grading system, biochemical, and molecular markers. STUDY DESIGN: Case-control study. METHODS: Forty-four Arabian horses (24 stallions, 20 mares) were enrolled as lame (n = 23), non-lame (n = 11), and control (n = 10). Field-based grouping was determined by veterinary-gate assessment: horses eliminated for irregular gait were classified as lame, and finishers passing veterinary inspection as non-lame. Blinded trot-up videos were reviewed to verify grouping and assign AAEP lameness grades; facial expression videos recorded at the veterinary gate were used to assess facial action units. Blood analyses measured biochemical markers. Real-time PCR quantified serum miR-21 and miR-155. RESULTS: Facial expression scales had high area under the curve values for distinguishing lame from non-lame horses (HGS: 0.947, 95% CI: 0.850-1.000; EQUUS-FAP: 0.917, 95% CI: 0.817-1.000). Facial scores were strongly correlated with AAEP lameness grades (r ≈ 0.78, p ≤ 0.001). Lame horses had higher oxidative stress index, lactate, lactate dehydrogenase, cortisol, aspartate aminotransferase concentrations, and miR-21 expression than other groups (p ≤ 0.015). MAIN LIMITATIONS: The small sample size warrants confirmation of the results in larger populations. CONCLUSIONS: Facial expression scores were associated with AAEP lameness grades and selected biochemical and molecular parameters in endurance horses. Facial expression assessment may provide complementary information alongside established lameness evaluation methods. Further studies are needed to confirm its role in pain-related lameness assessment.