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OBJECTIVE To verify the existence of intertransverse joints (ITJs) in young foals. ANIMALS 11 warmblood foals. PROCEDURES Postmortem examination of the lumbar area in foals < 200 days old using CT, MRI, dissection, and histomorphology. Data were analyzed with descriptive statistics. RESULTS Age of foals varied between 1 and 200 days (median, 11 days). Ten foals had 6 lumbar (L) vertebrae, and 1 foal had 5. All 11 foals, irrespective of age, had ITJs between the first sacral and last lumbar vertebrae and between the last and second-to-last lumbar vertebrae. In 6 foals (all with 6 L vertebrae), ITJs also existed between the fourth and fifth L vertebra. One foal, also with 6 L vertebrae, additionally had a unilateral (right) ITJ between the transverse processes of the third and fourth L vertebra. Based on CT, width of ITJs was seemingly greater in young (< 1 month old) foals because of the incomplete ossification of the transverse processes. The ITJs were confirmed and further characterized by MRI, dissection, and histomorphology. CLINICAL RELEVANCE ITJs already exist in very young warmblood foals and are present at birth. During the first months of life, these juvenile ITJs develop similarly to other synovial joints with increasing ossification and concomitant decrease of thickness of the cartilage layer. Knowledge of the presence of these ITJs in young animals is clinically relevant, as they should be recognized as nonpathologic when for instance a young foal is presented for presumed arthropathy and examined with advanced imaging techniques.

An established lipopolysaccharide (LPS) model previously described in Warmbloods, was inconsistent in Standardbred horses, where lameness was not detected despite the presence of synovitis. The present study aimed to determine the dose of LPS from E. coli O55:B5 required to induce mild to moderate lameness following middle carpal joint injection in Standardbred horses and to quantitate the induced lameness over time, with and without anti-inflammatory pre-treatment. In a baseline trial, eight healthy, clinically sound Standardbred horses were used in a rule-based dose-escalation design trial, starting at a dose of 10 endotoxin units (EU). Lameness at trot was evaluated visually and quantitatively (using an inertial-sensor system and pressure plate analysis). Synovial fluid aspirates were analysed for total nucleated cell counts, total protein and prostaglandin E2 (PGE2). Following 2 months wash-out, the effective LPS-dose determined in the baseline trial was used to evaluate the effect of anti-inflammatory treatment. A mixed model for repeated measures with horse as random effect was used for analysis. After injection of 10 EU LPS, the desired degree of lameness was observed in the baseline trial, with maximal lameness at post-injection hour (PIH) 4, followed by a rapid decline and return to baseline by PIH 48. No lameness was observed following pre-treatment with meloxicam. In synovial fluid, PGE2 was significantly higher at PIH 8 and PIH 24 in the baseline trial compared with following meloxicam pre-treatment. In conclusion, injection of the middle carpal joint with 10 EU LPS consistently induces a transient lameness and synovitis in Standardbred horses.

Application Understanding muscular adaptations could inform objective lameness-detection for early diagnosis/treatment,\ud ultimately serving to detect sub-clinical issues in supposed healthy horses and to reduce pain/ incapacity in lame horses.\ud Introduction The prevalence and impact of lameness on equine welfare has led to extensive research, which has\ud biomechanically analysed lameness-related alterations in movement. Despite this, limited information is available about\ud adaptive muscle activity that facilitates movement during lameness. Surface electromyography (sEMG) is a non-invasive\ud method for quantifying muscle activity. However, no equine studies have employed sEMG to compare inherent and\ud adaptive activity during non-lame and standardised lameness conditions, respectively. The aim of this preliminary study\ud was to compare Triceps Brachii (TB) muscle activity in horses before and after induced forelimb (FL) lameness, using\ud sEMG data.\ud Material and methods Six clinically non-lame horses (5 mares, 1 stallion, age: 7.0±3.7 years, height: 162.3±4.0 cm, body\ud mass: 572.7±45.8 kg) were used. sEMG sensors (Delsys Trigno, Delsys Inc.) were attached bilaterally to locations above\ud TB (long head), that were prepared by removing all hair and cleaning with isopropyl alcohol. Retro-reflective markers were\ud attached to anatomical landmarks for quantitative lameness evaluation (QHorse, Qualisys AB) and gait event detection.\ud sEMG (2000 Hz) and 3D kinematic (200 Hz) data were synchronously collected from horses during in-hand trot trials,\ud conducted on a straight, hard surfaced runway before (baseline) and after FL lameness induction. Baseline data were\ud initially collected, then temporary, mild FL lameness (2-3/5 AAEP Lameness Scale) was induced using mechanical bolt\ud pressure, applied to the tip of the frog and monitored by qualified veterinarians (T.S., F.S.B.) using a modified horseshoe\ud (Merkens and Schamhardt, 1988). Left and right FL lameness induction were randomised. Following data collection, the\ud bolt/ sole pressure was removed and no horses showed adverse reactions to lameness inductions, or residual lameness. For\ud stride segmentation, gait events were detected using kinematic data that were low-pass filtered (Butterworth 4th order, 10\ud Hz cut-off) and analysed in accordance with the methods described by Holt et al. (2017). To quantify lameness, MinDiff\ud was calculated using poll vertical displacement data, where healthy horses exhibit MinDiff between -6 – 6mm and left and\ud right FL lameness are exhibited as more positive and negative values, respectively (Rhodin et al., 2016). Raw sEMG\ud signals were DC-offset removed, high-pass filtered (Butterworth 4th order, 40 Hz cut-off) (St. George et al., 2018), and fullwave\ud rectified. Integrated EMG (iEMG) and average rectified value (ARV) were calculated using stride duration as\ud temporal domain. To reduce inter-subject variability, iEMG and ARV from each horse were normalised to the maximum\ud value observed for each limb (left/ right FL) across all strides from the baseline condition. Data from the “lame” and “nonlame”\ud limb were grouped, according to the limb where lameness was induced. A 2x2 repeated measures ANOVA was used\ud to compare muscle activity between limb (lame, non-lame) and condition (baseline, induced FL lameness). Post-hoc\ud analyses using Bonferroni correction were performed where significant main effects were found.\ud Results Mean ± sd MinDiff were baseline:-1.8 ± 8.7 mm, left FL lameness induction:-55.3 ± 34.1 mm, right FL lameness:\ud 56.8 ± 17.9 mm. Significant interactions between limb and condition were found for iEMG (p < 0.05, n2=0.74) and ARV (p\ud < 0.05, n2=0.75). Post hoc analyses of iEMG and ARV data revealed muscle activity was significantly higher in the lame\ud limb (p < 0.05) and significantly lower in the non-lame limb (p < 0.01) during the induced FL lameness condition.\ud Conclusion Preliminary findings reveal neuromuscular adaptations in TB during induced FL lameness. Significant\ud increases in stance duration have been reported during FL lameness (Weishaupt et al., 2006). Therefore, significant\ud increases in lame limb muscle activity may be due to prolonged stabilisation of the shoulder and elbow joints, as a\ud compensatory mechanism of gait adaptation to lameness. Further investigations of additional muscles and chronic lameness\ud cases are required to determine whether sEMG can provide a complimentary tool for objective lameness detection.

Extracellular vesicle (EV) concentration, characteristics and function in equine synovial fluid (SF) during normal growth and development has not previously been studied. Isolation of EVs was performed in SF from three healthy foals and two adult horses by differential ultracentrifugation (10,000 g and 200,000 g); EVs were purified by sucrose density gradient floatation and analysed by high-resolution flow cytometry (FCM), buoyant density and western blotting. Additionally, repeated biomarker analysis of sulphated glycosaminoglycans (GAG), matrix metalloproteinase (MMP), C-terminal crosslinked telopeptide type II collagen (CTX-II), collagenase cleaved neopeptide type II collagen (C2C) was performed in SF from 10 foals and six adult horses. In contrast with the quantitative EV profile, the biomarker profile in SF from juvenile joints was substantially different from that in SF from adult animals. However, there were qualitative differences in the high-resolution FCM scatter plots. Future in-depth functional analyses may reveal differences between juvenile and mature EVs in SF.

Objective: To investigate the potential of quantitative susceptibility mapping (QSM) and T2* relaxation time mapping to determine mechanical and structural properties of articular cartilage via univariate and multivariate analysis. Methods: Samples were obtained from a cartilage repair study, in which surgically induced full-thickness chondral defects in the stifle joints of seven Shetland ponies caused post-traumatic osteoarthritis (14 samples). Control samples were collected from non-operated joints of three animals (6 samples). Magnetic resonance imaging (MRI) was performed at 9.4 T, using a 3-D multi-echo gradient echo sequence. Biomechanical testing, digital densitometry (DD) and polarized light microscopy (PLM) were utilized as reference methods. To compare MRI parameters with reference parameters (equilibrium and dynamic moduli, proteoglycan content, collagen fiber angle and -anisotropy), depth-wise profiles of MRI parameters were acquired at the biomechanical testing locations. Partial least squares regression (PLSR) and Spearman’s rank correlation were utilized in data analysis. Results: PLSR indicated a moderate-to-strong correlation (ρ = 0.49–0.66) and a moderate correlation (ρ = 0.41–0.55) between the reference values and T2* relaxation time and QSM profiles, respectively (excluding superficial-only results). PLSR correlations were noticeably higher than direct correlations between bulk MRI and reference parameters. 3-D parametric surface maps revealed spatial variations in the MRI parameters between experimental and control groups. Conclusion: Quantitative parameters from 3-D multi-echo gradient echo MRI can be utilized to predict the properties of articular cartilage. With PLSR, especially the T2* relaxation time profile appeared to correlate with the properties of cartilage. Furthermore, the results suggest that degeneration affects the QSM-contrast in the cartilage. However, this change in contrast is not easy to quantify.

Aim: The horse joint, due to its similarity with the human joint, is the ultimate model for translational articular cartilage repair studies. This study was designed to determine the critical size of cartilage defects in the equine carpus and serve as a benchmark for the evaluation of new cartilage treatment options. Material and methods: Circular full-thickness cartilage defects with a diameter of 2, 4, and 8 mm were created in the left middle carpal joint and similar osteochondral (3.5 mm in depth) defects in the right middle carpal joint of 5 horses. Spontaneously formed repair tissue was examined macroscopically, with MR and µCT imaging, polarized light microscopy, standard histology, and immunohistochemistry at 12 months. Results: Filling of 2 mm chondral defects was good (77.8 ± 8.5%), but proteoglycan depletion was evident in Safranin-O staining and gadolinium-enhanced MRI (T1Gd). Larger chondral defects showed poor filling (50.6 ± 2.7% in 4 mm and 31.9 ± 7.3% in 8 mm defects). Lesion filling in 2, 4, and 8 mm osteochondral defects was 82.3 ± 3.0%, 68.0 ± 4.6% and 70.8 ± 15.4%, respectively. Type II collagen staining was seen in 9/15 osteochondral defects but only in 1/15 chondral defects. Subchondral bone pathologies were evident in 14/15 osteochondral samples but only in 5/15 chondral samples. Although osteochondral lesions showed better neotissue quality than chondral lesions, the overall repair was deemed unsatisfactory because of the subchondral bone pathologies. Conclusion: We recommend classifying 4 mm as critical osteochondral lesion size and 2 mm as critical chondral lesion size for cartilage repair research in the equine carpal joint model.

Since Galileo’s days the effect of size on the anatomical characteristics of the structural elements of the body has been a subject of interest. However, the effects of scaling at tissue level have received little interest and virtually no data exist on the subject with respect to the osteochondral unit in the joint, despite this being one of the most lesion-prone and clinically relevant parts of the musculoskeletal system. Imaging techniques, including Fourier transform infrared imaging, polarized light microscopy and micro computed tomography, were combined to study the response to increasing body mass of the osteochondral unit. We analyzed the effect of scaling on structural characteristics of articular cartilage, subchondral plate and the supporting trabecular bone, across a wide range of mammals at microscopic level. We demonstrated that, while total cartilage thickness scales to body mass in a negative allometric fashion, thickness of different cartilage layers did not. Cartilage tissue layers were found to adapt to increasing loads principally in the deep zone with the superficial layers becoming relatively thinner. Subchondral plate thickness was found to have no correlation to body mass, nor did bone volume fraction. The underlying trabecular bone was found to have thicker trabeculae (r = 0.75, p < 0.001), as expected since this structure carries most loads and plays a role in force mitigation. The results of this study suggest that the osteochondral tissue structure has remained remarkably preserved across mammalian species during evolution, and that in particular, the trabecular bone carries the adaptation to the increasing body mass.

Horses are precocial animals and able to stand and walk within hours after birth. To cope with associated loading, intrauterine bone development has shown to be anticipative. This study provides further insight into the post‐natal development of structurally important features of trabecular and subchondral bone of the talus and sagittal ridge of the tibia of warm‐blooded horses. In all areas studied, the average bone volume fraction showed a gradual increase over time, which was the result of a significant increase in trabecular thickness, without significant changes in the degree of anisotropy. Similar to the mineralised part of the bone, collagen content, measured as average retardation using polarised light microscopy, increased significantly, but the degree of anisotropy of the collagen type I network did not. At birth, the subchondral bone layer had a more trabecular aspect, gradually changing to an even surface with only a few vascular canals at an age of 2 months. Presented results indicate the necessity for a stronger structure, but not for a different structural design after birth, providing further evidence for anticipatory bone development in the horse. More knowledge about the strategies used to cope with mechanical loading after birth might be helpful in understanding the developmental bone and joint diseases.