Influence of microarchitecture on stressed volume and mechanical fatigue behaviour of equine subchondral bone.

Fractures of the equine metacarpophalangeal (MCP) joint are among the most common and fatal injuries experienced by racehorses. These bone injuries are a direct result of repetitive, high intensity loading of the skeleton during racing and training and there is consensus that they represent a mechanical fatigue phenomenon. Existing work has found the fatigue life of bone to be strongly determined by bone microarchitecture and the resulting stressed volume (i.e., the volume of bone stressed above assumed yield). The purpose of this study was to quantify the influence of bone microarchitecture on the mechanical fatigue behaviour of equine subchondral bone from the MCP joint across a wide variety of sample types. Forty-eight subchondral bone samples were prepared from the third metacarpal (MC3) and proximal phalanx (P1) of 8 horses and subsequently imaged using high resolution micro-computed tomography (µCT) to quantify microarchitectural features of interest, including bone volume fraction, tissue mineral density, pore size, pore spacing, and pore number. Samples were cyclically loaded in compression to a stress of 70 MPa, and fatigue life was defined as the number of cycles until failure. Finite element models were created from the µCT images and used to quantify stressed volume. Based on the expected log point-wise predictive density, stressed volume was a strong predictor of fatigue life in both the MC3 and P1. A regional analysis indicated fatigue life was more strongly associated with bone volume fraction in the superficial (r2 = 0.32, p < 0.001) and middle (r2 = 0.70, p < 0.001) regions of the subchondral bone, indicating the prominent role that the cortical plate played in the fatigue resistance of equine subchondral bone. By improving our understanding of the variance in fatigue life measurements, this research helps clarify the underlying mechanisms of the mechanical fatigue process and provides a basic understanding of subchondral bone injuries in the equine fetlock joint.

Comparing Inertial Measurement Units to Markerless Video Analysis for Movement Symmetry in Quarter Horses.

BACKGROUND: With an increasing number of systems for quantifying lameness-related movement asymmetry, between-system comparisons under non-laboratory conditions are important for multi-centre or referral-level studies. This study compares an artificial intelligence video app to a validated inertial measurement unit (IMU) gait analysis system in a specific group of horses. METHODS: Twenty-two reining Quarter horses were equipped with nine body-mounted IMUs while being videoed with a smartphone app. Both systems quantified head and pelvic movement symmetry during in-hand trot (hard/soft ground) and on the lunge (left/right rein, soft ground). Proportional limits of agreement (pLoA) were established. RESULTS: Widths of pLoA were larger for head movement (29% to 50% in-hand; 22% to 38% on lunge) than for pelvic movement (13% to 24% in-hand; 14% to 24% on lunge). CONCLUSION: The between-system pLoAs exceed current "lameness thresholds" aimed at identifying the affected limb(s) in lame horses. They also exceed published limits of agreement for stride-matched data but are similar to repeatability values and "lameness thresholds" from "non-lame" horses. This is encouraging for multi-centre studies and referral-level veterinary practice. The narrower pLoA values for pelvic movement asymmetry are particularly encouraging, given the difficulty of grading hind limb lameness "by eye".

Upper Body Movement Symmetry in Reining Quarter Horses during Trot In-Hand, on the Lunge and during Ridden Exercise.

Veterinary lameness examinations often comprise assessing ridden horses. Quantitative movement symmetry measurements can aid evidence-based decision making. While these are available for 'English' style riding, they are not for 'Western' style riding. This quantitative observational study quantified movement symmetry in reining Quarter Horses (QHs). Movement symmetry of the head, withers and sacrum (differences between minima, maxima and upward amplitudes) were quantified with inertial sensors in N = 30 medium/high level reining QHs during trot in-hand, on the lunge and ridden by one experienced rider (straight-line/circles) on reining-purpose riding surfaces. Mixed linear models for movement symmetry assessed the effects of ridden exercise and movement direction (fixed factors), stride time (covariate) and horse (random factor): single factors and two-way interactions with Bonferroni correction at p < 0.05. Three withers and pelvic parameters showed marginally more symmetrical movement when ridden (p ≤ 0.044; 1−5 mm differences). Three withers, three sacrum and one head parameter were significantly affected by movement direction (all p ≤ 0.026), five showed increased asymmetry on the inside rein, and two, quantifying vertical displacement maximum difference, showed the opposite. Riding QHs in 'Western' style showed small movement symmetry differences. Circular exercise confirmed increases in weight bearing asymmetry on the inside rein and in pushoff asymmetry on the outside rein. This should be further investigated for differentiating between different causes of lameness.

The role of synovial fluid constituents in the lubrication of collagen-glycosaminoglycan scaffolds for cartilage repair.

Extracellular matrix (ECM)-derived scaffolds have shown promise as tissue-engineered grafts for promoting cartilage repair. However, there has been a lack of focus on fine-tuning the frictional properties of scaffolds for cartilage tissue engineering as well as understanding their interactions with synovial fluid constituents. Proteoglycan-4 (PRG4) and hyaluronan (HA) are macromolecules within synovial fluid that play key roles as boundary mode lubricants during cartilage surface interactions. The overall objective of this study was to characterize the role PRG4 and HA play in the lubricating function of collagen-glycosaminoglycan (GAG) scaffolds for cartilage repair. As a first step towards this goal, we aimed to develop a suitable in vitro friction test to establish the boundary mode lubrication parameters for collagen-GAG scaffolds articulated against glass in a phosphate buffered saline (PBS) bath. Subsequently, we sought to leverage this system to determine the effect of physiological synovial fluid lubricants, PRG4 and HA, on the frictional properties of collagen-GAG scaffolds, with scaffolds hydrated in PBS and bovine synovial fluid (bSF) serving as negative and positive controls, respectively. At all compressive strains examined (ε = 0.1-0.5), fluid depressurization within hydrated collagen-GAG scaffolds was >99% complete at ½ minute. The coefficient of friction was stable at all compressive strains (ranging from a low 0.103 ± 0.010 at ε = 0.3 up to 0.121 ± 0.015 at ε = 0.4) and indicative of boundary-mode conditions. Immunohistochemistry demonstrated that PRG4 from recombinant human (rh) and bovine sources adsorbed to collagen-GAG scaffolds and the coefficient of friction for scaffolds immersed in rhPRG4 (0.067 ± 0.027) and normal bSF (0.056 ± 0.020) solution decreased compared to PBS (0.118 ± 0.21, both p < 0.05, at ε = 0.2). The ability of the adsorbed rhPRG4 to reduce friction on the scaffolds indicates that its incorporation within collagen-GAG biomaterials may enhance their lubricating ability as potential tissue-engineered cartilage replacements. To conclude, this study reports the development of an in vitro friction test capable of characterizing the coefficient of friction of ECM-derived scaffolds tested in a range of synovial fluid lubricants and demonstrates frictional properties as a potential design parameter for implants and materials for soft tissue replacement.

Proteoglycan-4 and hyaluronan composition in synovial fluid and serum from clinical equine subjects: relationship to cartilage boundary lubrication and viscosity of synovial fluid.

Purpose: In experimental models of equine joint-injury and osteoarthritis synovial fluid (SF) composition (proteoglycan-4, hyaluronan) can vary, along with changes to SF mechanical function (lubrication, viscosity). The study hypotheses were a) clinical equine joint-injury and disease results in altered SF composition and diminished mechanical function, and b) serum composition (proteoglycan-4 or hyaluronan) changes concurrently. The objectives were to characterize composition (proteoglycan-4, hyaluronan), and function of SF and serum from normal horses compared to clinical groups: osteoarthritis, acute-joint-injury, and osteochondrosis.Materials and Methods: Equine samples of SF (from various joints) and blood were collected at the point-of-care. Proteoglycan-4 concentrations were measured by amplified-luminescence-proximity-assay and enzyme-linked-immunosorbent-assay in SF and serum, respectively. Molecular-weight of hyaluronan was characterized by agarose-gel-electrophoresis, and concentrations were measured by enzyme-linked-immunosorbent-assay kit. Biomechanical function of SF was characterized by an in vitro cartilage-on-cartilage friction test, and viscosity test.Results: SF proteoglycan-4 concentration increased in acute-joint-injury (1185 ± 276 versus normal 205 ± 106 µg/mL, µ± SEM, p < 0.01), with increased percentage of lower molecular-weight hyaluronan in acute-joint-injury and osteochondrosis. SF and serum proteoglycan-4 concentrations were correlated in normal horses (r2 = 0.85, p < 0.05), but not in clinical groups. Cartilage-lubricating ability was unchanged, although steady-shear viscosity of acute-joint-injury SF decreased from normal.Conclusion: Composition of SF from cases of equine acute-joint-injury changed; both proteoglycan-4 concentration and hyaluronan molecular-weight were altered, with decreased SF viscosity, but no associated changes to serum. Serum proteoglycan-4 and hyaluronan concentrations alone may not be useful biomarkers for equine joint-injury or disease.

Biomechanics of Wound Healing in an Equine Limb Model: Effect of Location and Treatment with a Peptide-Modified Collagen-Chitosan Hydrogel.

The equine distal limb wound healing model, characterized by delayed re-epithelialization and a fibroproliferative response to wounding similar to that observed in humans, is a valuable tool for the study of biomaterials poised for translation into both the veterinary and human medical markets. In the current study, we developed a novel method of biaxial biomechanical testing to assess the functional outcomes of healed wounds in a modified equine model and discovered significant functional and structural differences in both unwounded and injured skin at different locations on the distal limb that must be considered when using this model in future work. Namely, the medial skin was thicker and displayed earlier collagen engagement, medial wounds experienced a greater proportion of wound contraction during closure, and proximal wounds produced significantly more exuberant granulation tissue. Using this new knowledge of the equine model of aberrant wound healing, we then investigated the effect of a peptide-modified collagen-chitosan hydrogel on wound healing. Here, we found that a single treatment with the QHREDGS (glutamine-histidine-arginine-glutamic acid-aspartic acid-glycine-serine) peptide-modified hydrogel (Q-peptide hydrogel) resulted in a higher rate of wound closure and was able to modulate the biomechanical function toward a more compliant healed tissue without observable negative effects. Thus, we conclude that the use of a Q-peptide hydrogel provides a safe and effective means of improving the rate and quality of wound healing in a large animal model.

The Effect of Intense Exercise on Equine Serum Proteoglycan-4/Lubricin.

Objective: Local biological and biomechanical-stimuli modulate proteoglycan-4 secretion within synovial joints. For the horse, changes to proteoglycan-4 concentration and function are notable in acute joint injury and osteoarthritis. Proteoglycan-4 (also known as Lubricin) is present in the blood, however the effect of exercise on equine serum levels is unknown. The overall objective of this study was, therefore, to investigate the effect of intense exercise on serum proteoglycan-4 in thoroughbred horses. Methods: Samples of blood were taken from thoroughbreds (n = 12) during a chuckwagon racing event (Alberta, Canada). The chuckwagon race is a sprint racing event where teams of horses pull a combined 1,325 lbs (601 kg) of wagon and driver around a 5/8th mile (1 km) of dirt track, racing at full gallop to the finish. Blood samples were collected 30-min before the race start, and several timepoints post-race: 5-min, 90-min, 3-h, 12-h, and 23-h. Proteoglycan-4 concentrations in serum were quantified by enzyme-linked-immunosorbent-assay using recombinant-human proteoglycan-4 standards and anti-proteoglycan-4 mAb 9G3. The molecular weight of immunoreactive proteoglycan-4 in serum was assessed by western blot. Results: Proteoglyan-4 in serum demonstrated the expected high MW immunoreactivity to mAb 9G3, consistent with that of full length PRG4. Serum proteoglycan-4 decreased five-minutes post-race from baseline concentration (0.815 ± 0.175 to 0.466 ± 0.090 µg/mL, µ ± SEM, p < 0.01). Conclusions: The concentration of serum proteoglycan-4 in horses decreased significantly five min post-exercise. A potential explanation for this finding could be increased proteoglycan-4 clearance from the circulation. Further investigations could extend to complete the detailed characterization of proteoglycan-4 structure and its potential function within the blood as it relates to joint health and exercise.

Effects of delayed serum separation and long-term storage on the measurement of thyroid hormones in equine blood samples.

Studies were conducted to determine the effects of delaying the separation of serum from the clot and of long-term storage of serum samples on the measurement of thyroid hormones in blood from horses using a fluorescence polarization immunoassay. The measured concentrations of T3 and T4 were not affected by leaving serum on the clot for as long as 24 hours at room temperatures. Storage of serum for 19 to 22 months at -20 degrees C resulted in significant increases of measured T4, but not T3. These studies support previous work demonstrating that thyroid hormones are resistant to degradation, immunologically stable, and reasonably insensitive to potential problems of routine specimen handling when measured with an immunoassay.