Preliminary study of proton magnetic resonance spectroscopy to assess bone marrow adiposity in the third metacarpus or metatarsus in Thoroughbred racehorses.

BACKGROUND: Magnetic resonance spectroscopy (MRS) has been used to investigate metabolic changes within human bone. It may be possible to use MRS to investigate bone metabolism and fracture risk in the distal third metacarpal/tarsal bone (MC/MTIII) in racehorses. OBJECTIVES: To determine the feasibility of using MRS as a quantitative imaging technique in equine bone by using the 1H spectra for the MC/MTIII to calculate fat content (FC). STUDY DESIGN: Observational cross-sectional study. METHODS: Limbs from Thoroughbred racehorses were collected from horses that died or were subjected to euthanasia on racecourses. Each limb underwent magnetic resonance imaging (MRI) at 3 T followed by single-voxel MRS at three regions of interest (ROI) within MC/MTIII (lateral condyle, medial condyle, proximal bone marrow [PBM]). Percentage FC was calculated at each ROI. Each limb underwent computed tomography (CT) and bone mineral density (BMD) was calculated for the same ROIs. All MR and CT images were graded for sclerosis. Histology slides were graded for sclerosis and proximal marrow space was calculated. Pearson or Spearman correlations were used to assess the relationship between BMD, FC and marrow space. Kruskal-Wallis tests were used to check for differences between sclerosis groups for BMD or FC. RESULTS: Eighteen limbs from 10 horses were included. A negative correlation was identified for mean BMD and FC for the lateral condyle (correlation coefficient = -0.60, p = 0.01) and PBM (correlation coefficient = -0.5, p = 0.04). There was a significant difference between median BMD for different sclerosis grades in the condyles on both MRI and CT. A significant difference in FC was identified between sclerosis groups in the lateral condyle on MRI and CT. MAIN LIMITATIONS: Small sample size. CONCLUSIONS: 1H Proton MRS is feasible in the equine MC/MTIII. Further work is required to evaluate the use of this technique to predict fracture risk in racehorses.

QCT-based computational bone strength assessment updated with MRI-derived 'hidden' microporosity.

Microdamage accumulated through sustained periods of cyclic loading or single overloading events contributes to bone fragility through a reduction in stiffness and strength. Monitoring microdamage in vivo remains unattainable by clinical imaging modalities. As such, there are no established computational methods for clinical fracture risk assessment that account for microdamage that exists in vivo at any specific timepoint. We propose a method that combines multiple clinical imaging modalities to identify an indicative surrogate, which we term 'hidden porosity', that incorporates pre-existing bone microdamage in vivo. To do so, we use the third metacarpal bone of the equine athlete as an exemplary model for fatigue induced microdamage, which coalesces in the subchondral bone. N = 10 metacarpals were scanned by clinical quantitative computed tomography (QCT) and magnetic resonance imaging (MRI). We used a patch-based similarity method to quantify the signal intensity of a fluid sensitive MRI sequence in bone regions where microdamage coalesces. The method generated MRI-derived pseudoCT images which were then used to determine a pre-existing damage (Dpex) variable to quantify the proposed surrogate and which we incorporate into a nonlinear constitutive model for bone tissue. The minimum, median, and maximum detected Dpex of 0.059, 0.209, and 0.353 reduced material stiffness by 5.9%, 20.9%, and 35.3% as well as yield stress by 5.9%, 20.3%, and 35.3%. Limb-specific voxel-based finite element meshes were equipped with the updated material model. Lateral and medial condyles of each metacarpal were loaded to simulate physiological joint loading during gallop. The degree of detected Dpex correlated with a relative reduction in both condylar stiffness (p = 0.001, R2 > 0.74) and strength (p < 0.001, R2 > 0.80). Our results illustrate the complementary value of looking beyond clinical CT, which neglects the inclusion of microdamage due to partial volume effects. As we use clinically available imaging techniques, our results may aid research beyond the equine model on fracture risk assessment in human diseases such as osteoarthritis, bone cancer, or osteoporosis.

Thoroughbred Racehorses in Hong Kong Require Vitamin D Supplementation to Mitigate the Risk of Low Vitamin D Status.

There is a paucity of data relating to the vitamin D status of racehorses. We hypothesised that the management of racehorses in Hong Kong (HK) predisposes to low vitamin D status unless they receive dietary supplementation. Serum concentrations of 25-hydroxyvitamin D2 (25OHD2), 25-hydroxyvitamin D3 (25OHD3) and total 25-hydroxyvitamin D (total 25OHD) for 79 non-grazing HK racehorses were compared with those for 22 racehorses training in the United Kingdom (UK) that grazed for ≥1 h/d, and for which published data exists. A nested group of 41 HK horses was sampled twice to determine the effect of the duration in HK on vitamin D status. The HK horses had significantly lower serum concentrations of total 25OHD and 25OHD2 than the UK horses; 25OHD2 was undetectable in 15/79 HK sera and serum concentrations of 25OHD2 declined with the duration in HK. The main determinants of vitamin D status were assessed using linear regression; the retained variables were the 25OHD3 concentration and the duration in HK. The inverse relationship between the serum concentrations of 25OHD2 and 25OHD3, previously identified in humans, was observed for the first time in horses. In conclusion, HK racehorses have low serum 25OHD2 and total 25OHD concentrations and rely on D3 supplementation to maintain adequate vitamin D status. Further study is required to determine the optimal form of dietary vitamin D supplementation for Thoroughbred racehorses.

The effect of season, management and endocrinopathies on vitamin D status in horses.

BACKGROUND: Vitamin D deficiency is common in humans and is increasingly linked to the pathogenesis of a multitude of diseases including obesity and metabolic syndrome. The biology of vitamin D in horses is poorly described; the relative contribution of the diet and skin synthesis to circulating concentrations is unclear and associations with the endocrine disease have not been explored. OBJECTIVES: To determine the relationship between management, season and endocrine disease and vitamin D status in horses. STUDY DESIGN: Cross-sectional cohort study. METHODS: Plasma concentrations of 25-hydroxyvitamin D2 (25(OH)D2 ) and D3 (25(OH)D3 ) were measured by liquid chromatography-tandem mass spectrometry in 34 healthy unsupplemented grazing ponies and 22 stabled Thoroughbreds receiving supplementary vitamin D3 in feed. A nested group of 18 grazing ponies were sampled on long and short days (>12 and <12 h of light/day) to determine the effect of sunlight exposure. In addition, the relationships between age, sex, adiposity, serum insulin, adrenocorticotropic hormone and vitamin D status were assessed in a mixed group of 107 horses using a linear regression model. RESULTS: All animals had a measurable level of 25(OH)D2 (median 10.7 nmol/L) whilst 25(OH)D3 was only detected in Thoroughbreds receiving D3 supplementation. Thoroughbreds had lower concentrations of 25(OH)D2 than ponies (7.4 vs. 12.6 nmol/L, p < 0.01). In grazing ponies, 25(OH)D2 concentrations were significantly higher on long days compared to short days (14.4 vs. 8.7 nmol/L, p < 0.01), whilst 25(OH)D3 was undetectable. Measures of increased adiposity, but not basal insulin, were associated with higher 25(OH)D2 concentrations, conversely to humans. Increasing ACTH was associated with lower 25(OH)D2 (p < 0.01). MAIN LIMITATIONS: Vitamin D2 concentrations were not measured in grass or forage. CONCLUSIONS: In horses 25(OH)D2 is the predominant vitamin D metabolite, and there is an apparent lack of endogenous vitamin D3 production. The relationship between vitamin D and endocrine disorders in horses does not reflect that of other species and warrants further investigation.

Training associated alterations in equine respiratory immunity using a multiomics comparative approach.

Neutrophilic airway inflammation is highly prevalent in racehorses in training, with the term mild to moderate equine asthma (MMEA) being applied to the majority of such cases. Our proposed study is largely derived from the strong association between MMEA in racehorses and their entry into a race training program. The objectives of this study are to characterise the effect of training on the local pulmonary immune system by defining the gene and protein expression of tracheal wash (TW) derived samples from Thoroughbred racehorses prior to and following commencement of race training. Multiomics analysis detected 2138 differentially expressed genes and 260 proteins during the training period. Gene and protein sets were enriched for biological processes related to acute phase response, oxidative stress, haemopoietic processes, as well as to immune response and inflammation. This study demonstrated TW samples to represent a rich source of airway cells, protein and RNA to study airway immunity in the horse and highlighted the benefits of a multiomics methodological approach to studying the dynamics of equine airway immunity. Findings likely reflect the known associations between race-training and both airway inflammation and bleeding, offering further insight into the potential mechanisms which underpin training associated airway inflammation.

Application across species of a one health approach to liquid sample handling for respiratory based -omics analysis.

Airway inflammation is highly prevalent in horses, with the majority of non-infectious cases being defined as equine asthma. Currently, cytological analysis of airway derived samples is the principal method of assessing lower airway inflammation. Samples can be obtained by tracheal wash (TW) or by lavage of the lower respiratory tract (bronchoalveolar lavage (BAL) fluid; BALF). Although BALF cytology carries significant diagnostic advantages over TW cytology for the diagnosis of equine asthma, sample acquisition is invasive, making it prohibitive for routine and sequential screening of airway health. However, recent technological advances in sample collection and processing have made it possible to determine whether a wider range of analyses might be applied to TW samples. Considering that TW samples are relatively simple to collect, minimally invasive and readily available in the horse, it was considered appropriate to investigate whether, equine tracheal secretions represent a rich source of cells and both transcriptomic and proteomic data. Similar approaches have already been applied to a comparable sample set in humans; namely, induced sputum. Sputum represents a readily available source of airway biofluids enriched in proteins, changes in the expression of which may reveal novel mechanisms in the pathogenesis of respiratory diseases, such as asthma and chronic obstructive pulmonary disease. The aim of this study was to establish a robust protocol to isolate macrophages, protein and RNA for molecular characterization of TW samples and demonstrate the applicability of sample handling to rodent and human pediatric bronchoalveolar lavage fluid isolates. TW samples provided a good quality and yield of both RNA and protein for downstream transcriptomic/proteomic analyses. The sample handling methodologies were successfully applicable to BALF for rodent and human research. TW samples represent a rich source of airway cells, and molecular analysis to facilitate and study airway inflammation, based on both transcriptomic and proteomic analysis. This study provides a necessary methodological platform for future transcriptomic and/or proteomic studies on equine lower respiratory tract secretions and BALF samples from humans and mice.

Ex Vivo Comparison of the Accuracy of a Clamped with a Hand-Held Drill Guide for Drilling Distal Third Metacarpals and Metatarsals in Equids.

OBJECTIVES: The aim of this study was to determine whether a clamped drill guide can be used effectively to drill across equine distal third metacarpals and metatarsals and to compare accuracy and speed of a drilling with a hand-held drill guide. STUDY DESIGN: Eighty equine cadaver limbs were divided between four surgeons with varying experience. The limbs were randomly allocated to clamped or hand-held drill guides and placed in a stand in an upright/standing position on a metal table. Drilling times from cis- to trans cortices were measured. Post-drilling computed tomography images were used to assess drilling deviations in the dorsopalmar and proximodistal directions. RESULTS: Mean proximodistal deviation of the drill hole was distad for both guides and significantly (p = 0.01) less for the clamped drill guide +0.35° (range: -5.42°, +6.61°, standard deviation [SD] ± 3.18°) than the hand-held drill guide +2.05° (range: -5.24°, +9.81°, SD ± 3.69°). Mean dorsopalmar/plantar deviation was non-significantly (p = 0.17) less for the clamped drill guide -0.27° (range: -9.72°, +6.58°, SD ± 3.57°) than the hand-held drill guide +0.60° (range: -9.74°, + 12.33°, SD ± 4.51°). Mean time taken to drill from cis to trans cortex was significantly (p < 0.001) shorter with the clamped drill guide (41.2s) than the hand-held guide (56.6s). CONCLUSION: The clamped drill guide could be used successfully on the equine distal third metacarpals and metatarsals and resulted in some improved accuracy and reduced drilling time compared with the hand-held guide. The use of the clamped drill guide in vivo could be supported. This may be of benefit for certain fracture repairs in equids.

Horses with solar foot penetration, deep digital flexor tendon injury, and absence of concurrent synovial sepsis can have a positive outcome.

Solar foot penetration is one of the causes of deep digital flexor tendon injuries in horses, however, limited information is available on the prognosis for return to soundness in the absence of synovial sepsis. Objectives of this retrospective observational study were to describe low-field magnetic resonance imaging (MRI) findings and long-term outcome for a group of horses with this combination of clinical problems. Horses were included if low-field standing MRI of the foot was performed following puncture wounds, injury of the deep digital flexor tendon was diagnosed, and sepsis was confirmed to be absent in all adjacent synovial structures (distal interphalangeal joint, navicular bursa, and digital flexor tendon sheath). Medical records were reviewed and MRI studies were re-interpreted. Follow-up information was obtained via a telephone questionnaire at a minimum of 6 months post-injury. A total of 11 horses met inclusion criteria. In three horses, the deep digital flexor tendon injury was only visible in the T2 fast spin echo sequence and contrast radiography improved diagnostic certainty. The most commonly affected area was between the distal border of the distal sesamoid bone and the facies flexoria of the distal phalanx (6/11, 55%). Six horses (60%) had an excellent outcome (5, show jumping; 1, general purpose) and returned to full athletic function. Five horses (40%) were sound but had not yet resumed full work at the time of follow-up. Findings indicated that the prognosis for return to soundness can be good for horses with solar penetration, deep digital flexor injury, and absence of synovial sepsis.

Magnetic resonance imaging assisted management in five cases of suspected quittor.

OBJECTIVES: Assessment of the usefulness of magnetic resonance imaging (MRI) in treatment planning in suspected cases of quittor in the horse. METHODS: Five horses with chronic discharging tracts at the level of the foot underwent MRI for treatment planning. RESULTS: The MRI examination revealed variable involvement of soft tissue and osseous structures of the foot in addition to abnormalities of the ungular cartilages in all cases. In two cases, follow-up MRI examination was performed. Four of five horses had a successful outcome, with three of these undergoing only one surgical procedure and one being managed medically. CLINICAL SIGNIFICANCE: We believe that the use of preoperative MRI facilitated accurate determination of the structures involved in cases of quittor, guiding the management, surgical approach and postoperative therapy.

Partial ostectomy of the dens to correct atlantoaxial subluxation in a pony.

OBJECTIVE: To report a surgical technique that allows decompression and anatomic re-alignment for management of subluxation of the atlantoaxial articulation. STUDY DESIGN: Clinical report. ANIMALS: Four-year-old Welsh Section B pony gelding. METHODS: Attempts at closed reduction of subluxation of the atlantoaxial articulation under general anesthesia were unsuccessful, so a ventral surgical approach allowing partial ostectomy of the dens of the axis was used to facilitate reduction. RESULT: Recovery from surgery was uneventful and the pony remained comfortable without neurologic deficits. Both neck stiffness and ataxia resolved. Radiographic examination at 5 days and 6 months postoperatively showed normal anatomic position of the dens of the axis and stabilization of the atlantoaxial joint. CONCLUSION: Partial ostectomy of the dens using a ventral approach can be used to facilitate reduction of atlantoaxial subluxation.