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The objective of this study was to evaluate a non-resorbable implant for the focal repair of chondral defects in eight adult horses with 12-month follow-up. The bi-layered construct composed of a polycarbonate-urethane-urea biomaterial which was printed in 3D fashion onto a bone anchor was implanted into surgically created osteochondral defects into the femoropatellar joints of eight horses. The analysis of post-mortem outcomes were compared to defects treated with microfracture in the same animal on the contralateral femoropatellar jointfemoropatellar joint. The overall macroscopic scoring after 12 months yielded higher scores in the OCI-treated stifles compared to MF treatment (p = 0.09) with better quality and filling of the defect. Histology revealed good anchorage of repair tissue growing into the 3D structure of the implant and histopathology scoring for adjacent native cartilage showed no difference between groups. MRI and micro-CT showed overall less sclerotic reactions in the surrounding bone in the implant group and no foreign body reaction was detected. Biomechanical analysis of the repair tissue revealed a significantly higher peak modulus (p < 0.05) in the implant group (0.74 ± 0.45) compared to the microfracture control group (0.15 ± 0.11). Dynamic loading yielded higher values for the repair tissue overgrowing the implant group (0.23 ± 0.17) compared to the microfracture control (0.06 ± 0.06) (p < 0.05). The bi-layered osteochondral implant provided a safe implant for focal repair of full-thickness osteochondral defects, as no adverse reaction was seen within the joints and the level of degeneration of adjacent cartilage to the repair site was not different compared to that seen in defects treated with microfracture after 12 months., (© 2024 The Author(s). Journal of Orthopaedic Research® published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

The ability of articular cartilage to withstand significant mechanical stresses during activities, such as walking or running, relies on its distinctive structure. Integrating detailed tissue properties into subject-specific biomechanical models is challenging due to the complexity of analyzing these characteristics. This limitation compromises the accuracy of models in replicating cartilage function and impacts predictive capabilities. To address this, methods revealing cartilage function at the constituent-specific level are essential. In this study, we demonstrated that computational modeling derived individual constituent-specific biomechanical properties could be predicted by a novel nanoparticle contrast-enhanced computer tomography (CECT) method. We imaged articular cartilage samples collected from the equine stifle joint (n = 60) using contrast-enhanced micro-computed tomography (µCECT) to determine contrast agents' intake within the samples, and compared those to cartilage functional properties, derived from a fibril-reinforced poroelastic finite element model. Two distinct imaging techniques were investigated: conventional energy-integrating µCECT employing a cationic tantalum oxide nanoparticle (Ta 2 O 5 -cNP) contrast agent and novel photon-counting µCECT utilizing a dual-contrast agent, comprising Ta 2 O 5 -cNP and neutral iodixanol. The results demonstrate the capacity to evaluate fibrillar and non-fibrillar functionality of cartilage, along with permeability-affected fluid flow in cartilage. This finding indicates the feasibility of incorporating these specific functional properties into biomechanical computational models, holding potential for personalized approaches to cartilage diagnostics and treatment., (© 2024. The Author(s).)

Background: Client satisfaction depends on the balance between expectations and service experience. Previous research identified seven aspects of equine veterinary professional conduct that are important for client satisfaction: quality of care, quality of service, horsemanship, transfer of knowledge, financial aspects, interpersonal skills and professionalism., Methods: By employing a cross-sectional study design through a survey-based investigation, horse owners' initial contact preferences and their perceptions of the importance of various aspects of veterinary care in different scenarios were explored. Categories included professional versus amateur and competitive versus non-competitive horse owners. Quantitative data analysis was performed., Results: Data from 1153 participants revealed that horse owners promptly contacted veterinarians for colic (92.7%) but delayed for lameness (51.8%) and pre-purchase examinations (63.0%). Overall, quality of care emerged as the most important aspect of veterinary care for horse owners, with financial aspects considered least important. Competitive and professional horse owners prioritised financial aspects and professionalism, whereas non-professional and non-competitive horse owners prioritised quality of care and interpersonal skills (p < 0.005)., Limitations: Survey distribution relied on a snowball effect, internet access was necessary and the study exclusively represents the Western equine community. Potential bias should be acknowledged., Conclusion: The perceived importance of various aspects of veterinary care varies depending on the nature of the consultation and the horse owner type. Tailoring veterinary services can improve client satisfaction by aligning with diverse expectations., (© 2024 The Author(s). Veterinary Record published by John Wiley & Sons Ltd on behalf of British Veterinary Association.)

Background: Strategies for articular cartilage repair need to take into account topographical differences in tissue composition and architecture to achieve durable functional outcome. These have not yet been investigated in the equine stifle., Objectives: To analyse the biochemical composition and architecture of three differently loaded areas of the equine stifle. We hypothesise that site differences correlate with the biomechanical characteristics of the cartilage., Study Design: Ex vivo study., Methods: Thirty osteochondral plugs per location were harvested from the lateral trochlear ridge (LTR), the distal intertrochlear groove (DITG) and the medial femoral condyle (MFC). These underwent biochemical, biomechanical and structural analysis. A linear mixed model with location as a fixed factor and horse as a random factor was applied, followed by pair-wise comparisons of estimated means with false discovery rate correction, to test for differences between locations. Correlations between biochemical and biomechanical parameters were tested using Spearman's correlation coefficient., Results: Glycosaminoglycan content was different between all sites (estimated mean [95% confidence interval (CI)] for LTR 75.4 [64.5, 88.2], for intercondylar notch (ICN) 37.3 [31.9, 43.6], for MFC 93.7 [80.1109.6] μg/mg dry weight), as were equilibrium modulus (LTR2.20 [1.96, 2.46], ICN0.48 [0.37, 0.6], MFC1.36 [1.17, 1.56] MPa), dynamic modulus (LTR7.33 [6.54, 8.17], ICN4.38 [3.77, 5.03], MFC5.62 [4.93, 6.36] MPa) and viscosity (LTR7.49 [6.76, 8.26], ICN16.99 [15.88, 18.14], MFC8.7 [7.91,9.5]°). The two weightbearing areas (LTR and MCF) and the non-weightbearing area (ICN) differed in collagen content (LTR 139 [127, 152], ICN176[162, 191], MFC 127[115, 139] μg/mg dry weight), parallelism index and angle of collagen fibres. The strongest correlations were between proteoglycan content and equilibrium modulus (r: 0.642; p: 0.001), dynamic modulus (r: 0.554; p < 0.001) and phase shift (r: -0.675; p < 0.001), and between collagen orientation angle and equilibrium modulus (r: -0.612; p < 0.001), dynamic modulus (r: -0.424; p < 0.001) and phase shift (r: 0.609; p < 0.001)., Main Limitations: Only a single sample per location was analysed., Conclusions: There were significant differences in cartilage biochemical composition, biomechanics and architecture between the three differently loaded sites. The biochemical and structural composition correlated with the mechanical characteristics. These differences need to be acknowledged by designing cartilage repair strategies., (© 2023 The Authors. Equine Veterinary Journal published by John Wiley & Sons Ltd on behalf of EVJ Ltd.)

Objective: Cationic tantalum oxide nanoparticles (Ta 2 O 5 -cNPs), as a newly introduced contrast agent for computed tomography of cartilage, offer quantitative evaluation of proteoglycan (PG) content and biomechanical properties. However, knowledge on the depth-wise impact of cartilage constituents on nanoparticle diffusion, particularly the influence of the collagen network, is lacking. In this study, we aim to establish the depth-dependent relationship between Ta 2 O 5 -cNP diffusion and cartilage constituents (PG content, collagen content and network architecture)., Methods: Osteochondral samples (n = 30) were harvested from healthy equine stifle joints (N = 15) and the diffusion of 2.55 nm diameter cationic Ta 2 O 5 -cNPs into the cartilage was followed with micro computed tomography (µCT) imaging for up to 96 hours. The diffusion-related parameters, Ta 2 O 5 -cNP maximum partition (P max ) and diffusion time constant, were compared against biomechanical and depth-wise structural properties. Biomechanics were assessed using stress-relaxation and sinusoidal loading protocols, whereas PG content, collagen content and collagen network architecture were determined using digital densitometry, Fourier-transform infrared spectroscopy and polarized light microscopy, respectively., Results: The P max correlates with the depth-wise distribution of PGs (bulk Spearman's ρ = 0.87, p < 0.001). More open collagen network architecture at the superficial zone enhances intake of Ta 2 O 5 -cNPs, but collagen content overall decreases the intake. The P max values correlate with the equilibrium modulus (ρ = 0.80, p < 0.001) of articular cartilage., Conclusion: This study establishes the feasibility of Ta 2 O 5 -cNPs for the precise and comprehensive identification of biomechanical and structural changes in articular cartilage via contrast-enhanced µCT., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Individuals working in the field of veterinary care are regularly affected by their profession. High levels of responsibility to often provide life-saving health care to animals combined with having to manage owners' expectations and irregular working hours can cause considerable levels of work-related stress among professionals in equine veterinary practice. On the positive side, research also shows that working in the veterinary profession can have a positive impact on personal wellbeing and feelings of fulfillment. A limited number of studies has investigated work satisfaction and engagement among veterinarians across the globe, and none specifically in the equine veterinary work field. The aim of the current study was to identify relevant predictors of employee engagement and work satisfaction in relation to demographic and work environment related factors in the equine veterinary profession., Methods: A cross-sectional study design was used to investigate work satisfaction and employee engagement among equine veterinary professionals from the UK, the US and the Netherlands using an online survey., Results: Results suggest that levels of work engagement and satisfaction in the veterinary profession may be gauged using four factors. These factors encompass Pride and purpose (the extent to which personal core values align with the mission of the employer, i.e., the veterinary practice), Company culture and relationship with management (the manner in which staff members interact with each other and the management), Working conditions and compensation (formal employment conditions relating to responsibilities and rewards and levels of collegiality) and Team culture and learning possibilities (encouragement to pursue personal and professional growth)., Discussion: Findings underline the importance of being particularly mindful of inexperienced colleagues, those with demanding family commitments and, where feasible, of providing employees with a modicum of autonomy in order to ensure a satisfied equine veterinary workforce., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Elte, Acton, Martin, Nielen, van Weeren and Wolframm.)

Background: Quantitative gait analysis offers objective information to support clinical decision-making during lameness workups including advantages in terms of documentation, communication, education, and avoidance of expectation bias. Nevertheless, hardly any data exist comparing outcome of subjective scoring with the output of objective gait analysis systems., Objectives: To investigate between- and within-veterinarian agreement on primary lame limb and lameness grade, and to determine relationships between subjective lameness grade and quantitative data, focusing on differences between (1) veterinarians, (2) live vs video assessment, (3) baseline assessment vs assessment following diagnostic analgesia., Study Design: Clinical observational study., Methods: Kinematic data were compared to subjective lameness assessment by clinicians with ≥8 years of orthopaedic experience. Subjective assessments and kinematic data for baseline trot-ups and response to 48 diagnostic analgesia interventions in 23 cases were included. Between and within-veterinarian agreement was investigated using Cohen's Kappa (κ). Asymmetry parameters for kinematic data ('forelimb lame pattern', 'hindlimb lame pattern', 'overall symmetry', 'vector sum head', 'pelvic sum') were determined, and used as outcome variables in mixed models; explanatory variables were subjective lameness grade and its interaction with (1) veterinarian, (2) live or video evaluation and (3) baseline or diagnostic analgesia assessment., Results: Agreement on lame limb between live and video assessment was 'good' between and within veterinarians (median κ = 0.64 and κ = 0.53). There was a positive correlation between subjective scoring and measured asymmetry. The relationship between lameness grade and objective asymmetry differed slightly between (1) veterinarians (for all combined parameters, p-values between P < .001 and 0.04), (2) between live and video assessments ('forelimb lame pattern', 'overall symmetry', both P ≤ .001), and (3) between baseline and diagnostic analgesia assessment (all combined parameters, between P < .001 and .007)., Main Limitations: Limited number of veterinarians (n = 4) and cases (n = 23), only straight-line soft surface data, different number of subjective assessments live vs from video., Conclusions: Overall, between- and within-veterinarian agreement on lame limb was 'good', whereas agreement on lameness grade was 'acceptable' to 'poor'. Quantitative data and subjective assessments correlated well, with minor though significant differences in the number of millimetres, equivalent to one lameness grade between veterinarians, and between assessment conditions. Differences between baseline assessment vs assessment following diagnostic analgesia suggest that addition of objective data can be beneficial to reduce expectation bias. The small differences between live and video assessments support the use of high-quality videos for documentation, communication, and education, thus, complementing objective gait analysis data., (© 2021 The Authors. Equine Veterinary Journal published by John Wiley & Sons Ltd on behalf of EVJ Ltd.)

Background: High-power laser therapy gained popularity recently as a regenerative treatment for tendinitis and desmitis in the horse. However, studies evaluating the effects of laser therapy on tissue repair at the histological level in large mammals are lacking., Objectives: To evaluate the effects of high-power laser therapy on suspensory desmitis healing, using a model of suspensory ligament branch injury., Study Design: In vivo experiments., Methods: Standardised lesions were surgically induced in all four lateral suspensory branches of 12 healthy Warmblood horses. Laser therapy (class 4, 15W) was applied daily on two of four induced lesions for four consecutive weeks. Horses were randomly assigned to either short-term study (horses were sacrificed after 4 weeks) or long-term study (6 months). Suspensory ligament samples were scored after staining with haematoxylin-eosin and immunostaining for collagen 1- collagen 3- and factor VIII., Results: In the short-term study, significantly better (lower) scores for variation in density (17% above cut-off score in treated lesions vs. 31% above cut-off score in controls, P = .03), shape of nuclei (54% vs 92%, P = .02), fibre alignment (32% vs 75%, P = .003) and fibre structure (38% vs 71%, P = .02) were found in laser-treated lesions when compared to controls. Collagen 3 expression was significantly higher (32% vs 19%, P = .006) in control lesions. In both short- and long-term studies combined, parameters lesion size (44% vs 56%, P = .02) and shape of nuclei (53% vs 84%, P = .05) scored significantly better in treated lesions. Long-term, significantly better (lower) scores were found in the laser-treated group for lesion size (15% vs 45%, P = .008) and a higher percentage above cut-off score for density of the nuclei (27% vs 9%, P = .02), compared to controls., Main Limitations: The model of suspensory branch injury is not an exact representation of clinical overstrain lesions., Conclusions: These results suggest that high-power laser therapy enables better lesion healing than conservative treatment., (© 2022 EVJ Ltd.)

The central regions of galaxy clusters are permeated by magnetic fields and filled with relativistic electrons 1 . When clusters merge, the magnetic fields are amplified and relativistic electrons are re-accelerated by turbulence in the intracluster medium 2,3 . These electrons reach energies of 1-10 GeV and, in the presence of magnetic fields, produce diffuse radio halos 4 that typically cover an area of  around 1 Mpc 2 . Here we report observations of four clusters whose radio halos are embedded in much more extended, diffuse radio emission, filling a volume 30 times larger than that of radio halos. The emissivity in these larger features is about 20 times lower than the emissivity in radio halos. We conclude that relativistic electrons and magnetic fields extend far beyond radio halos, and that the physical conditions in the outer regions of the clusters are quite different from those in the radio halos., (© 2022. The Author(s).)

Background: The increasing popularity of objective gait analysis makes application in prepurchase examinations (PPE) a logical next step. Therefore, there is a need to have more understanding of asymmetry during a PPE in horses described on clinical evaluation as subtly lame., Objectives: The objective of this study is to objectively compare asymmetry in horses raising minor vet concerns in a PPE and in horses raising major vet concerns with that found in horses presented with subtle single-limb lameness, and to investigate the effect of age/discipline on the clinicians' interpretation of asymmetry on the classification of minor vet concerns in a PPE., Study Design: Clinical case-series., Methods: Horses presented for PPE (n = 98) or subjectively evaluated as single limb low-grade (1-2/5) lame (n = 24, 13 forelimb lame, 11 hindlimb lame), from the patient population of a single clinic, were enrolled in the study provided that owners were willing to participate. Horses undergoing PPE were assigned a classification of having minor vet concerns (n = 84) or major vet concerns (n = 14) based on findings during the dynamic-orthopaedic part of the PPE. Lame horses were only included if pain-related lameness was confirmed by an objective improvement after diagnostic analgesia exceeding daily variation determined for equine symmetry parameters using optical motion capture. Clinical evaluation was performed by six different clinicians, each with ≥8 years of equine orthopaedic experience. Vertical movement symmetry was measured using optical motion capture, simultaneously with the orthopaedic examination. Data were analysed using previously described parameters and mixed model analysis and least squares means were used to calculate differences between groups., Results: There was no effect of age or discipline on the levels of asymmetry within PPE horses raising minor vet concerns. MinDiff and RUD of the head discriminated between forelimb lame and PPE horses raising minor vet concerns; MinDiff, MaxDiff, RUD of the Pelvis, HHDswing and HHDstance did so for hindlimb lameness. Two lameness patterns differentiated both forelimb and hindlimb lame from PPE horses with minor vet concerns: RUD Poll + MinDiff Withers - RUD Pelvis and RUD Pelvis + RUD Poll - MinDiff Withers. Correcting for vertical range of motion enabled differentiation of PPE horses with minor vet concerns from PPE horses with major vet concerns., Main Limitations: Objective data only based on trot on soft surface, limited number of PPE horses with major vet concerns., Conclusions: Combinations of kinematic parameters discriminate between PPE horses with minor vet concerns and subtly lame horses, though overlap exists., (© 2021 The Authors. Equine Veterinary Journal published by John Wiley & Sons Ltd on behalf of EVJ Ltd.)

Background: The pressure on equine veterinarians to provide services that meet with a client's demands, that is, realizing client satisfaction, is considerable., Aim: The aim of this paper is to analyze existing literature, with a view to distill the most relevant components for client satisfaction in equine veterinary practice., Materials and Methods: A structured literature search was conducted. Included papers were systematically organised and analysed using an inductive approach., Results: Seven components relevant to client satisfaction in equine veterinary practice were identified: quality of care, quality of service, horsemanship of the veterinarian, costs of service, interpersonal skills, professional attitude and transfer of knowledge., Discussion: The limited amount and variable quality of available records regarding client satisfaction in equine veterinary practice are the main limitations of this study., Conclusion: The seven categories identified are likely to play a fundamental role in achieving client satisfaction in equine veterinary practice. Additional research is required to validate these categories and relate them to different types of clients as well as map their needs and expectations, so that they can be used to assist equine veterinary professionals in tailoring customer experience to the individual client., (© 2021 The Authors. Veterinary Record published by John Wiley & Sons Ltd on behalf of British Veterinary Association.)

Human societies and natural ecosystems are under threat by growing populations, overexploitation of natural resources and climate change. This calls for more sustainable utilization of resources based on past experiences and insights from many different disciplines. Interdisciplinary approaches to studies of historical commons have potential to identify drivers of change and keys to success in the past, and offer advice about the management and use of shared resources in contemporary and future systems. We address these issues by applying an ecological perspective to historical data on social-ecological systems. We perform comparisons and time series analyses for nine successful Dutch commons for which high-resolution data on the regulatory activities and use of shared resources is available for on average 380 years (range 236 to 568) during the period 1300 to 1972. Within commons, institutional developments were oscillating, with periods of intense regulatory activity being separated by periods of low activity, and with the dynamics of regulations being largely independent across commons. Ecological theory posits that species that occupy similar niches should show correlated responses to environmental challenges; however, commons using more similar resources did not have more parallel or similar institutional developments. One notable exception was that sanctioning was more frequent in commons that directed more regulatory activities towards non-renewable subsoil resources, whereas there was no association between sanctioning and the use of renewable resources. This might indicate that commoners were aware of potential resource depletion and attempted to influence freeriding by actively trying to solve the underlying social dilemmas. Sanctioning regulations were more frequent during the first than during the second part of a common's life, indicating that while sanctioning might have been important for the establishment of commons it was not key to the long-term persistence of historical commons., Competing Interests: The authors have declared that no competing interests exist.

Background: Left-right movement symmetry is a highly desirable characteristic in sport horses., Objectives: This study compared movement symmetry in well-trained dressage horses in unridden and unrestrained position and ridden in a dressage frame, and investigated possible associations between gaits., Study Design: Experimental study., Methods: Seven sound, high-level dressage horses were measured at walk and sitting trot on a treadmill at several speeds under two conditions: with and without rider. Left-right differences in stance duration, stance protraction and retraction based on longitudinal hoof positions, ipsilateral limb tracking, minimum and maximum vertical positions of the dorsal spinous processes of the sixth thoracic (T6), third sacral vertebrae (S3) and wing of atlas, and vertical ground reaction forces were calculated and analysed in mixed models., Results: At walk, five body variables indicated increased asymmetry in the ridden condition compared with unridden condition: forelimb stance duration (unridden/ridden left-right differences 9 vs 13 ms; P = .008), forelimb stance protraction (P = .004), stance retraction (P = .001) and first force peak (P = .003), and hindlimb stance retraction (P = .01). At trot, six body variables were more asymmetrical in the ridden condition: forelimb stance duration (2.5 vs 3.8 ms, P = .004); hindlimb stance protraction (P < .0001) and retraction (P = .01), T6 minimum (4 vs 6 mm, P = .001), T6 maximum (9 vs 11 mm, P = .01) and S3 maximum (6 vs 12 mm, P < .001). Five variables had significant associations between asymmetries at walk and trot, but only three demonstrated a positive slope., Main Limitations: A limited number of horses and riders were studied. Measurements were performed on a treadmill., Conclusions: High-level horses moved slightly more asymmetrically when ridden in a dressage frame than in the unridden condition., (© 2020 The Authors. Equine Veterinary Journal published by John Wiley & Sons Ltd on behalf of EVJ Ltd.)

One of the important challenges in bone tissue engineering is the development of biodegradable bone substitutes with appropriate mechanical and biological properties for the treatment of larger defects and those with complex shapes. Recently, magnesium phosphate (MgP) doped with biologically active ions like strontium (Sr 2+ ) have shown to significantly enhance bone formation when compared with the standard calcium phosphate-based ceramics. However, such materials can hardly be shaped into large and complex geometries and more importantly lack the adequate mechanical properties for the treatment of load-bearing bone defects. In this study, we have fabricated bone implants through extrusion assisted three-dimensional (3D) printing of MgP ceramics modified with Sr 2+ ions (MgPSr) and a medical-grade polycaprolactone (PCL) polymer phase. MgPSr with 30 wt% PCL (MgPSr-PCL30) allowed the printability of relevant size structures (>780 mm 3 ) at room temperature with an interconnected macroporosity of approximately 40%. The printing resulted in implants with a compressive strength of 4.3 MPa, which were able to support up to 50 cycles of loading without plastic deformation. Notably, MgPSr-PCL30 scaffolds were able to promote in vitro bone formation in medium without the supplementation with osteo-inducing components. In addition, long-term in vivo performance of the 3D printed scaffolds was investigated in an equine tuber coxae model over 6 months. The micro-CT and histological analysis showed that implantation of MgPSr-PCL30 induced bone regeneration, while no bone formation was observed in the empty defects. Overall, the novel polymer-modified MgP ceramic material and extrusion-based 3D printing process presented here greatly improved the shape ability and load-bearing properties of MgP-based ceramics with simultaneous induction of new bone formation., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

T2* mapping is promising for the evaluation of articular cartilage collagen. In this work, a groove model in a large animal is used as a model for posttraumatic arthritis. We hypothesized that T2* mapping could be employed to differentiate between healthy and (subtly) damaged cartilage. Eight carpal joints were obtained from four adult Shetland ponies that had been included in the groove study. In this model, grooves were surgically created on the proximal articular surface of the intermediate carpal bone (radiocarpal joint) and the radial facet of the third carpal bone (middle carpal joint) by either coarse disruption or sharp incision. After 9 months, T2* mapping of the entire carpal joint was carried out on a 7.0-T whole-body magnetic resonance imaging (MRI) scanner by means of a gradient echo multi-echo sequence. Afterwards, assessment of collagen orientation was carried out based on Picrosirius Red-stained histological sections, visualized by polarized light microscopy (PLM). The average T2* relaxation time in grooved samples was lower than in contralateral control sites. Opposite to the grooved areas, the "kissing sites" had a higher average T2* relaxation time than the grooved sites. PLM showed mild changes in orientation of the collagen fibers, particularly around blunt grooves. This work shows that T2* relaxation times are different in healthy cartilage vs (early) damaged cartilage, as induced by the equine groove model. Additionally, the average T2* relaxation times are different in kissing lesions vs the grooved sites., (© 2020 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

Bioactive hydrogels based on naturally-derived polymers are of great interest for regenerative medicine applications. Among naturally-derived polymers, silk fibroin has been extensively explored as a biomaterial for tissue engineering due to its unique mechanical properties. Here, we demonstrate the rapid gelation of cell-laden silk fibroin hydrogels by visible light-induced crosslinking using riboflavin as a photo-initiator, in presence of an electron acceptor. The gelation kinetics were monitored by in situ photo-rheometry. Gelation was achieved in minutes and could be tuned owing to its direct proportionality to the electron acceptor concentration. The concentration of the electron acceptor did not affect the elastic modulus of the hydrogels, which could be altered by varying the polymer content. Further, the biocompatible riboflavin photo-initiator combined with sodium persulfate allowed for the encapsulation of cells within silk fibroin hydrogels. To confirm the cytocompatibility of the silk fibroin formulations, three cell types (articular cartilage-derived progenitor cells, mesenchymal stem cells and dental-pulp-derived stem cells) were encapsulated within the hydrogels, which associated with a viability >80% for all cell types. These results demonstrated that fast gelation of silk fibroin can be achieved by combining it with riboflavin and electron acceptors, which results in a hydrogel that can be used in tissue engineering and cell delivery applications.

High-power laser therapy is often used as a treatment for human sport injuries but controlled standardized studies on its efficacy are lacking. The technique has also been introduced in the equine field and recently promising results were reported in a retrospective study focusing on 150 sporthorses suffering from tendinopathy and desmopathy of the SDFT, DDFT, suspensory ligament, and suspensory branches. The goal of the present study was to evaluate the effect of high-power laser in a standardized lesion model in horses. Lesions were created in all lateral suspensory branches of 12 warmblood horses. In each horse, 2 of the 4 lesioned branches were treated daily with a multi-frequency high-power laser for 4 weeks. Color Doppler ultrasonography was performed during and after the treatment period. Six horses were euthanized 4 weeks post-surgery (short-term) and 6 were further rehabilitated until 6 months and then euthanized (long-term). High-field MRI evaluation was performed on all cadaver limbs. On ultrasound, transverse size of the lesion was significantly smaller after 2- and 3 months ( p = 0.026 and p = 0.015) in the treated branches. The expected post-surgery enlargement of the lesion circumference and cross-sectional area (CSA) over time, was significantly lower in the short-term laser treated group ( p = 0.016 and p = 0.010). Treated lesions showed a significantly increased Doppler signal during treatment ( p < 0.001) compared with control. On MRI, in the short and long-term group, the CSA of the lesions was significantly smaller ( p = 0.002), and the mean signal significantly lower in the treatment groups ( p = 0.006). This standardized controlled study shows that multi-frequency high-power laser therapy significantly improves healing of a suspensory branch ligament lesion., (Copyright © 2020 Pluim, Martens, Vanderperren, van Weeren, Oosterlinck, Dewulf, Kichouh, Van Thielen, Koene, Luciani, Plancke and Delesalle.)

Historical commons represent self-governed governance regimes that regulate the use and management of natural and man-made shared resources. Despite growing scientific interests, analyses of commons evolution and temporal dynamics are rare and drivers of change (birth, adaptation, dissolution) remain obscure. We apply an interdisciplinary approach and address these issues from an eco-evolutionary perspective. Analyses of > 400 Dutch commons over more than a millennium (between the 9th and the 20th century) uncovered that most commons originated between 1200 and 1700, and that there was a particularly high rate of evolution during 1300-1550, a pattern intermediate to gradualism and punctuated equilibrium in biological evolution. Dissolutions of commons were rare prior to 1800 and peaked around 1850, comparable to a mass extinction in biology. Temporal trends in number, spatial distribution, density, and dispersion of historical commons were distinctive and resembled developments seen at the levels of species and individuals in the growth of biological communities and populations, in that they showed signs of saturation determined by the abundance and distribution of resources. The spatiotemporal dynamics of commons also pointed to important roles of social, economic and political factors, such as new reclamations of resources and pressure on resources due to population growth. Despite internal and external pressures, the self-governing commons studied here were very successful, in the sense that they persisted for on average >350 years. There was a weak positive relationship between the use of multiple resources and the lifespan of commons, resembling associations between diversity and persistence seen in biological systems. It is argued that eco-evolutionary perspectives can further the understanding of the long-term dynamics of commons as institutions for collective action, vitalize future research, improve management of shared goods, and advise about sustainable utilization of finite resources., Competing Interests: The authors have declared that no competing interests exist.

Articular cartilage contains a subpopulation of tissue-specific progenitors that are an ideal cell type for cell therapies and generating neocartilage for tissue engineering applications. However, it is unclear whether the standard chondrogenic medium using transforming growth factor beta (TGFβ) isoforms is optimal to differentiate these cells. We therefore used pellet culture to screen progenitors from immature bovine articular cartilage with a number of chondrogenic factors and discovered that bone morphogenetic protein-9 (BMP9) precociously induces their differentiation. This difference was apparent with toluidine blue staining and confirmed by biochemical and transcriptional analyses with BMP9-treated progenitors exhibiting 11-fold and 5-fold greater aggrecan and collagen type II (COL2A1) gene expression than TGFβ1-treated progenitors. Quantitative gene expression analysis over 14 days highlighted the rapid and phased nature of BMP9-induced chondrogenesis with sequential activation of aggrecan then collagen type II, and negligible collagen type X gene expression. The extracellular matrix of TGFβ1-treated progenitors analyzed using atomic force microscopy was fibrillar and stiff whist BMP9-induced matrix of cells more compliant and correspondingly less fibrillar. Polarized light microscopy revealed an annular pattern of collagen fibril deposition typified by TGFβ1-treated pellets, whereas BMP9-treated pellets displayed a birefringence pattern that was more anisotropic. Remarkably, differentiated immature chondrocytes incubated as high-density cultures in vitro with BMP9 generated a pronounced anisotropic organization of collagen fibrils indistinguishable from mature adult articular cartilage, with cells in deeper zones arranged in columnar manner. This contrasted with cells grown with TGFβ1, where a concentric pattern of collagen fibrils was visualized within tissue pellets. In summary, BMP9 is a potent chondrogenic factor for articular cartilage progenitors and is also capable of inducing morphogenesis of adult-like cartilage, a highly desirable attribute for in vitro tissue-engineered cartilage.

The clinical translation of three-dimensionally printed bioceramic scaffolds with tailored architectures holds great promise toward the regeneration of bone to heal critical-size defects. Herein, the long-term in vivo performance of printed hydrogel-ceramic composites made of methacrylated-oligocaprolactone-poloxamer and low-temperature self-setting calcium-phosphates is assessed in a large animal model. Scaffolds printed with different internal architectures, displaying either a designed porosity gradient or a constant pore distribution, are implanted in equine tuber coxae critical size defects. Bone ingrowth is challenged and facilitated only from one direction via encasing the bioceramic in a polycaprolactone shell. After 7 months, total new bone volume and scaffold degradation are significantly greater in structures with constant porosity. Interestingly, gradient scaffolds show lower extent of remodeling and regeneration even in areas having the same porosity as the constant scaffolds. Low regeneration in distal regions from the interface with native bone impairs ossification in proximal regions of the construct, suggesting that anisotropic architectures modulate the cross-talk between distant cells within critical-size defects. The study provides key information on how engineered architectural patterns impact osteoregeneration in vivo, and also indicates the equine tuber coxae as promising orthotopic model for studying materials stimulating bone formation., (© 2020 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Treating joint diseases remains a significant clinical challenge. Conventional in vitro cultures and animal models have been helpful, but suffer from limited predictive power for the human response. Advanced models are therefore required to mimic the complex biological interactions within the human joint. However, the intricate structure of the joint microenvironment and the complex nature of joint diseases have challenged the development of in vitro models that can faithfully mimic the in vivo physiological and pathological environments. In this review, we discuss the current in vitro models of the joint and the progress achieved in the development of novel and potentially more predictive models, and highlight the application of new technologies to accurately emulate the articular joint., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Galaxy clusters are the most massive gravitationally bound structures in the Universe. They grow by accreting smaller structures in a merging process that produces shocks and turbulence in the intracluster gas. We observed a ridge of radio emission connecting the merging galaxy clusters Abell 0399 and Abell 0401 with the Low-Frequency Array (LOFAR) telescope network at 140 megahertz. This emission requires a population of relativistic electrons and a magnetic field located in a filament between the two galaxy clusters. We performed simulations to show that a volume-filling distribution of weak shocks may reaccelerate a preexisting population of relativistic particles, producing emission at radio wavelengths that illuminates the magnetic ridge., (Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Lightning is a dangerous yet poorly understood natural phenomenon. Lightning forms a network of plasma channels propagating away from the initiation point with both positively and negatively charged ends-called positive and negative leaders 1 . Negative leaders propagate in discrete steps, emitting copious radio pulses in the 30-300-megahertz frequency band 2-8 that can be remotely sensed and imaged with high spatial and temporal resolution 9-11 . Positive leaders propagate more continuously and thus emit very little high-frequency radiation 12 . Radio emission from positive leaders has nevertheless been mapped 13-15 , and exhibits a pattern that is different from that of negative leaders 11-13,16,17 . Furthermore, it has been inferred that positive leaders can become transiently disconnected from negative leaders 9,12,16,18-20 , which may lead to current pulses that both reconnect positive leaders to negative leaders 11,16,17,20-22 and cause multiple cloud-to-ground lightning events 1 . The disconnection process is thought to be due to negative differential resistance 18 , but this does not explain why the disconnections form primarily on positive leaders 22 , or why the current in cloud-to-ground lightning never goes to zero 23 . Indeed, it is still not understood how positive leaders emit radio-frequency radiation or why they behave differently from negative leaders. Here we report three-dimensional radio interferometric observations of lightning over the Netherlands with unprecedented spatiotemporal resolution. We find small plasma structures-which we call 'needles'-that are the dominant source of radio emission from the positive leaders. These structures appear to drain charge from the leader, and are probably the reason why positive leaders disconnect from negative ones, and why cloud-to-ground lightning connects to the ground multiple times.

The walk and trot are inherently symmetrical gaits, making them potentially suitable for the detection of left-right asymmetries. The aims of this study were to describe asymmetrical vertical excursions of the withers at walk in non-lame high-level dressage horses and to seek associations between these asymmetric movements and other kinematic variables and vertical ground reaction forces (vGRFs). Seven dressage horses, judged clinically as being sound, walked unridden and unrestrained on a treadmill with an integrated force measuring system (480 Hz), from which spatiotemporal and vGRF variables were extracted. Markers were tracked by 12 infrared cameras (240 Hz). The vertical position of the sixth thoracic vertebra (T6), limb protraction and retraction distances throughout stance, and global limb lengths were determined. Contralateral trial-mean differences were calculated, including difference in T6 minimum vertical position between contralateral steps (T6minDiff). Mixed models were used to study associations between symmetry parameters. Trial-mean T6minDiff ranged between 0.3-23 mm. Of the seven horses, five consistently dropped the withers more in early left forelimb stance, one was fairly symmetrical, and one dropped the withers more in early right forelimb stance. Comparisons between contralateral limbs showed the following associations. The forelimb that was retracted when T6min was lowest showed greater retraction at toe-off (1 mm increase predicted 0.17 mm T6minDiff increase) and shorter stance duration (1 ms decrease predicted 0.3 mm T6minDiff increase). The hind limb that was in midstance when T6min was lowest showed a greater range of motion during the stance phase (1 mm increase in protraction or retraction predicted 0.2 mm T6minDiff increase). The haunches were displaced away from the side of the forelimb that was protracted when T6min was lowest (1 mm lateral shift predicted 0.07 mm T6minDiff increase). Forelimb and hind limb vGRF parameters were non-significant. Asymmetry of vertical withers movement in horses assessed as being sound at trot was related to a complex pattern of asymmetries in spatiotemporal variables throughout the stride cycle rather than to vertical load redistribution between the forelimbs. This suggests that the asymmetry may be due to inherent laterality rather than weight-bearing lameness., Competing Interests: The authors have declared that no competing interests exist.

Saddle slip, defined as a progressive lateral displacement of the saddle during ridden exercise, has recently been given attention in the scientific press as a potential sign of lameness. The aim of this study was to objectively quantify the normal lateral movement (oscillations) of the saddle relative to the horse in non-lame horses, and associate this movement to the movements of the horse and rider. Data from seven Warmblood dressage horses competing at Grand Prix (n = 6) or FEI Intermediate (n = 1) level, ridden by their usual riders, were used. Simultaneous kinetic, kinematic and saddle pressure measurements were conducted during sitting and rising trot on a force-measuring treadmill. The maximum lateral movement of the caudal part of the saddle relative to the horse's spine (MAX) was determined for each diagonal step. A mixed model was applied, with MAX as outcome, and T6 and S3 vertical position, rigid body rotation angles (roll, pitch, yaw) of the horse's and rider's pelvis, vertical ground reaction forces, saddle force, and rider position (rising in rising trot, sitting in rising trot or sitting in sitting trot) as explanatory variables. The least square means for MAX were 14.3 (SE 4.7) mm and 23.9 (SE 4.7) mm for rising and sitting in rising trot, and 20.3 (SE 4.7) mm for sitting trot. A 10 mm increase in maximum pelvic height at push off increased MAX by 1.4 mm (p<0.0001). One degree increase in rider pelvis roll decreased MAX 1.1 mm, and one degree increase in rider pelvis yaw increased MAX 0.7 mm (both p<0.0001). The linear relationships found between MAX and movements of both horse and rider implies that both horse and rider movement asymmetries are reflected in the lateral movements or oscillations of the saddle in non-lame horses., Competing Interests: The authors have declared that no competing interests exist.

The aim of this study was to develop a formulation with a sustained intra-articular release of the anti-inflammatory drug tacrolimus. Drug release kinetics from the prepared tacrolimus loaded monodisperse biodegradable microspheres based on poly(d-l-lactide-PEG)- b -poly(l-lactide) multiblock copolymers were tunable by changing polymer composition, particularly hydrophobic-hydrophilic block ratio. The monospheres were 30 μm and released the drug, depending on the formulation, in 7 to >42 days. The formulation exhibiting sustained release for 1 month was selected for further in vivo evaluation. Rat knees were injected with three different doses of tacrolimus (10 wt %) loaded monospheres (2.5, 5.0, and 10 mg), contralateral control knees with saline. Micro-CT and histology showed no negative changes on cartilage, indicating good biocompatibility. Minor osteophyte formation was seen in a dose dependent fashion, suggesting local drug release and therapeutic action thereof. To investigate in vivo drug release, tacrolimus monospheres were injected into horse joints, after which multiple blood and synovial fluid samples were taken. Sustained intra-articular release was seen during the entire four-week follow-up, with negligible systemic drug concentrations (<1 ng/mL), confirming the feasibility of local intra-articular drug delivery without provoking systemic effects. Intra-articular injection of unloaded monospheres led to a transient inflammatory reaction, measured by total synovial leucocyte count (72 h). This reaction was significantly lower in joints injected with tacrolimus loaded monospheres, showing not only the successful local tacrolimus delivery but also local anti-inflammatory action. This local anti-inflammatory potential without systemic side-effects can be beneficial in the treatment of inflammatory joint diseases, among which is osteoarthritis.

Diagnosis of sacroiliac dysfunction in horses includes manual motion palpation of the equine ilium and sacrum. Motion of the ilium and sacrum during manual force application to the equine pelvis has been measured previously in vitro. The aim of this study was to measure the amount and direction of motion in vivo, including comparison of bone-fixated and skin-mounted inertial sensors. Sensors were skin-mounted over tuber sacrale (TS) and third sacral spinous process of six Thoroughbred horses and later attached via Steinmann pins inserted into the same bony landmarks. Orientations of each TS and sacrum were recorded by one investigator during six trials of manual force applied to the pelvis, inducing cranial, caudal, and oblique rotations. Mean values were reported in Euler angles for the three orthogonal planes lateral bending, flexion-extension (FE), and axial rotation (AR). Differences between skin- and bone-fixated markers were determined with significance set at P < .05. The largest mean values recorded during rotations applied to the pelvises were for FE, (2.08° ± 0.35°) with bone-fixated sensors. AR gave the largest values recorded with skin mountings (1.70° ± 0.48°). There was a poor correlation between skin-mounted and bone-fixated markers with AR being the orthogonal plane in which results from skin mounting were closest to results from bone-fixated sensors Bony kinematics during external movement applied to the pelvis cannot be predicted from skin-mounted sensors, due to differences between skin- and bone-mounted sensors., (Copyright © 2018 Elsevier Inc. All rights reserved.)

In this paper, we describe and validate the EquiMoves system, which aims to support equine veterinarians in assessing lameness and gait performance in horses. The system works by capturing horse motion from up to eight synchronized wireless inertial measurement units. It can be used in various equine gait modes, and analyzes both upper-body and limb movements. The validation against an optical motion capture system is based on a Bland-Altman analysis that illustrates the agreement between the two systems. The sagittal kinematic results (protraction, retraction, and sagittal range of motion) show limits of agreement of ± 2.3 degrees and an absolute bias of 0.3 degrees in the worst case. The coronal kinematic results (adduction, abduction, and coronal range of motion) show limits of agreement of - 8.8 and 8.1 degrees, and an absolute bias of 0.4 degrees in the worst case. The worse coronal kinematic results are most likely caused by the optical system setup (depth perception difficulty and suboptimal marker placement). The upper-body symmetry results show no significant bias in the agreement between the two systems; in most cases, the agreement is within ±5 mm. On a trial-level basis, the limits of agreement for withers and sacrum are within ±2 mm, meaning that the system can properly quantify motion asymmetry. Overall, the bias for all symmetry-related results is less than 1 mm, which is important for reproducibility and further comparison to other systems., Competing Interests: Mihai Marin-Perianu, Raluca Marin-Perianu and Paul Havinga founded Inertia-Technology B.V. (Enschede, The Netherlands), which sells the inertial sensor system (ProMove-mini) that is used as the basis of the EquiMoves system, which is evaluated in this study. Stephan Bosch and Berend-Jan van der Zwaag are employees of Inertia-Technology B.V.

Cellular senescence, that is, the withdrawal from the cell cycle, combined with the acquirement of the senescence associated secretory phenotype has important roles during health and disease and is essential for tissue remodeling during embryonic development. Osteoclasts are multinucleated cells, responsible for bone resorption, and cell cycle arrest during osteoclastogenesis is well recognized. Therefore, the aim of this study was to investigate whether these cells should be considered senescent and to assess the influence of hypoxia on their potential senescence status. Osteoclastogenesis and bone resorption capacity of osteoclasts, cultured from CD14+ monocytes, were evaluated in two oxygen concentrations, normoxia (21% O 2 ) and hypoxia (5% O 2 ). Osteoclasts were profiled by using specific staining for proliferation and senescence markers, qPCR of a number of osteoclast and senescence-related genes and a bone resorption assay. Results show that during in vitro osteoclastogenesis, osteoclasts heterogeneously obtain a senescent phenotype. Furthermore, osteoclastogenesis was delayed at hypoxic compared to normoxic conditions, without negatively affecting the bone resorption capacity. It is concluded that osteoclasts can be considered senescent, although senescence is not uniformly present in the osteoclast population. Hypoxia negatively affects the expression of some senescence markers. Based on the direct relationship between senescence and osteoclastogenesis, it is tempting to hypothesize that contents of the so-called senescence associated secretory phenotype (SASP) not only play a functional role in matrix resorption, but also may regulate osteoclastogenesis., (© 2018 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals Inc.)

Objective: To report on the experiences with the use of commercial and autologous fibrin glue (AFG) and of an alternative method based on a 3D-printed polycaprolactone (PCL) anchor for the fixation of hydrogel-based scaffolds in an equine model for cartilage repair., Methods: In a first study, three different hydrogel-based materials were orthotopically implanted in nine horses for 1-4 weeks in 6 mm diameter full-thickness cartilage defects in the medial femoral trochlear ridge and fixated with commercially available fibrin glue (CFG). One defect was filled with CFG only as a control. In a second study, CFG and AFG were compared in an ectopic equine model. The third study compared the efficacy of AFG and a 3D-printed PCL-based osteal anchor for fixation of PCL-reinforced hydrogels in three horses for 2 weeks, with a 4-week follow-up to evaluate integration of bone with the PCL anchor. Short-term scaffold integration and cell infiltration were evaluated by microcomputed tomography and histology as outcome parameters., Results: The first study showed signs of subchondral bone resorption in all defects, including the controls filled with CFG only, with significant infiltration of neutrophils. Ectopically, CFG induced clear inflammation with strong neutrophil accumulation; AFG was less reactive, showing fibroblast infiltration only. In the third study the fixation potential for PCL-reinforced hydrogels of AFG was inferior to the PCL anchor. PCL reinforcement had disappeared from two defects and showed signs of dislodging in the remaining four. All six constructs fixated with the PCL anchor were still in place after 2 weeks. At 4 weeks, the PCL anchor showed good integration and signs of new bone formation., Conclusions: The use of AFG should be preferred to xenogeneic products in the horse, but AFG is subject to individual variations and laborious to make. The PCL anchor provides the best fixation; however, this technique involves the whole osteochondral unit, which entails a different conceptual approach to cartilage repair.

Cell-laden hydrogels are the primary building blocks for bioprinting, and, also termed bioinks, are the foundations for creating structures that can potentially recapitulate the architecture of articular cartilage. To be functional, hydrogel constructs need to unlock the regenerative capacity of encapsulated cells. The recent identification of multipotent articular cartilage-resident chondroprogenitor cells (ACPCs), which share important traits with adult stem cells, represents a new opportunity for cartilage regeneration. However, little is known about the suitability of ACPCs for tissue engineering, especially in combination with biomaterials. This study aimed to investigate the potential of ACPCs in hydrogels for cartilage regeneration and biofabrication, and to evaluate their ability for zone-specific matrix production. Gelatin methacryloyl (gelMA)-based hydrogels were used to culture ACPCs, bone marrow mesenchymal stromal cells (MSCs) and chondrocytes, and as bioinks for printing. Our data shows ACPCs outperformed chondrocytes in terms of neo-cartilage production and unlike MSCs, ACPCs had the lowest gene expression levels of hypertrophy marker collagen type X, and the highest expression of PRG4, a key factor in joint lubrication. Co-cultures of the cell types in multi-compartment hydrogels allowed generating constructs with a layered distribution of collagens and glycosaminoglycans. By combining ACPC- and MSC-laden bioinks, a bioprinted model of articular cartilage was generated, consisting of defined superficial and deep regions, each with distinct cellular and extracellular matrix composition. Taken together, these results provide important information for the use of ACPC-laden hydrogels in regenerative medicine, and pave the way to the biofabrication of 3D constructs with multiple cell types for cartilage regeneration or in vitro tissue models., Statement of Significance: Despite its limited ability to repair, articular cartilage harbors an endogenous population of progenitor cells (ACPCs), that to date, received limited attention in biomaterials and tissue engineering applications. Harnessing the potential of these cells in 3D hydrogels can open new avenues for biomaterial-based regenerative therapies, especially with advanced biofabrication technologies (e.g. bioprinting). This study highlights the potential of ACPCs to generate neo-cartilage in a gelatin-based hydrogel and bioink. The ACPC-laden hydrogel is a suitable substrate for chondrogenesis and data shows it has a bias in directing cells towards a superficial zone phenotype. For the first time, ACPC-hydrogels are evaluated both as alternative for and in combination with chondrocytes and MSCs, using co-cultures and bioprinting for cartilage regeneration in vitro. This study provides important cues on ACPCs, indicating they represent a promising cell source for the next generation of cartilage constructs with increased biomimicry., (Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Background: Adipose tissue is a promising source of mesenchymal stromal cells (MSCs) for the treatment of tendon disease. The goal of this study was to assess the effect of a single intralesional implantation of adipose tissue-derived mesenchymal stromal cells (AT-MSCs) on artificial lesions in equine superficial digital flexor tendons (SDFTs)., Methods: During this randomized, controlled, blinded experimental study, either autologous cultured AT-MSCs suspended in autologous inactivated serum (AT-MSC-serum) or autologous inactivated serum (serum) were injected intralesionally 2 weeks after surgical creation of centrally located SDFT lesions in both forelimbs of nine horses. Healing was assessed clinically and with ultrasound (standard B-mode and ultrasound tissue characterization) at regular intervals over 24 weeks. After euthanasia of the horses the SDFTs were examined histologically, biochemically and by means of biomechanical testing., Results: AT-MSC implantation did not substantially influence clinical and ultrasonographic parameters. Histology, biochemical and biomechanical characteristics of the repair tissue did not differ significantly between treatment modalities after 24 weeks. Compared with macroscopically normal tendon tissue, the content of the mature collagen crosslink hydroxylysylpyridinoline did not differ after AT-MSC-serum treatment (p = 0.074) while it was significantly lower (p = 0.027) in lesions treated with serum alone. Stress at failure (p = 0.048) and the modulus of elasticity (p = 0.001) were significantly lower after AT-MSC-serum treatment than in normal tendon tissue., Conclusions: The effect of a single intralesional injection of cultured AT-MSCs suspended in autologous inactivated serum was not superior to treatment of surgically created SDFT lesions with autologous inactivated serum alone in a surgical model of tendinopathy over an observation period of 22 weeks. AT-MSC treatment might have a positive influence on collagen crosslinking of remodelling scar tissue. Controlled long-term studies including naturally occurring tendinopathies are necessary to verify the effects of AT-MSCs on tendon disease.

In this study, we investigated the use of microspheres with a narrow particle size distribution ('monospheres') composed of biodegradable poly(DL-lactide)-PEG-poly(DL-lactide)-b-poly(L-lactide) multiblock copolymers that are potentially suitable for local sustained drug release in articular joints. Monospheres with sizes of 5, 15 and 30μm and a narrow particle size distribution were prepared by a micro-sieve membrane emulsification process. During in vitro degradation, less crystallinity, higher swelling and accelerated mass loss during was observed with increasing the PEG content of the polymer. The monospheres were tested in both a small (mice/rat) and large animal model (horse). In vivo imaging after injection with fluorescent dye loaded microspheres in mice knees showed that monospheres of all sizes retained within the joint for at least 90days, while the same dose of free dye redistributed to the whole body within the first day after intra-articular injection. Administration of monospheres in equine carpal joints caused a mild transient inflammatory response without any clinical signs and without degradation of the cartilage, as evidenced by the absence of degradation products of sulfated glycosaminoglycans or collagen type 2 in the synovial fluid. The excellent intra-articular biocompatibility was confirmed in rat knees, where μCT-imaging and histology showed neither changes in cartilage quality nor quantity. Given the good intra-articular retention and the excellent biocompatibility, these novel poly(DL-lactide)-PEG-poly(DL-lactide)-b-poly(L-lactide)-based monospheres can be considered a suitable platform for intra-articular drug delivery., Statement of Significance: This paper demonstrates the great potential in intra-articular drug delivery of monodisperse biodegradable microspheres which were prepared using a new class of biodegradable multi-block copolymers and a unique membrane emulsification process allowing the preparation of microspheres with a narrow particle size distribution (monospheres) leading to multiple advantages like better injectability, enhanced reproducibility and predictability of the in vivo release kinetics. We report not only on the synthesis and preparation, but also in vitro characterization, followed by in vivo testing of intra-articular biocompatibility of the monospheres in both a small and a large animal model. The favourable intra-articular biocompatibility combined with the prolonged intra-articular retention (>90days) makes these monospheres an interesting drug delivery platform. What should also be highlighted is the use of horses; a very accurate translational model for the human situation, making the results not only relevant for equine healthcare, but also for the development of novel human OA therapies., (Copyright © 2016. Published by Elsevier Ltd.)

Chondral and osteochondral lesions due to injury or other pathology are highly prevalent conditions in horses (and humans) and commonly result in the development of osteoarthritis and progression of joint deterioration. Regenerative medicine of articular cartilage is an emerging clinical treatment option for patients with articular cartilage injury or disease. Functional articular cartilage restoration, however, remains a major challenge, but the field is progressing rapidly and there is an increasing body of supportive clinical and scientific evidence. This review gives an overview of the established and emerging surgical techniques employed for cartilage repair in horses. Through a growing insight in surgical cartilage repair possibilities, surgeons might be more stimulated to explore novel techniques in a clinical setting., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Cosmic rays are the highest-energy particles found in nature. Measurements of the mass composition of cosmic rays with energies of 10(17)-10(18) electronvolts are essential to understanding whether they have galactic or extragalactic sources. It has also been proposed that the astrophysical neutrino signal comes from accelerators capable of producing cosmic rays of these energies. Cosmic rays initiate air showers--cascades of secondary particles in the atmosphere-and their masses can be inferred from measurements of the atmospheric depth of the shower maximum (Xmax; the depth of the air shower when it contains the most particles) or of the composition of shower particles reaching the ground. Current measurements have either high uncertainty, or a low duty cycle and a high energy threshold. Radio detection of cosmic rays is a rapidly developing technique for determining Xmax (refs 10, 11) with a duty cycle of, in principle, nearly 100 per cent. The radiation is generated by the separation of relativistic electrons and positrons in the geomagnetic field and a negative charge excess in the shower front. Here we report radio measurements of Xmax with a mean uncertainty of 16 grams per square centimetre for air showers initiated by cosmic rays with energies of 10(17)-10(17.5) electronvolts. This high resolution in Xmax enables us to determine the mass spectrum of the cosmic rays: we find a mixed composition, with a light-mass fraction (protons and helium nuclei) of about 80 per cent. Unless, contrary to current expectations, the extragalactic component of cosmic rays contributes substantially to the total flux below 10(17.5) electronvolts, our measurements indicate the existence of an additional galactic component, to account for the light composition that we measured in the 10(17)-10(17.5) electronvolt range.

Background: Adipose tissue-derived mesenchymal stromal cells (AT-MSCs) are frequently used to treat equine tendinopathies. Up to now, knowledge about the fate of autologous AT-MSCs after intralesional injection into equine superficial digital flexor tendons (SDFTs) is very limited. The purpose of this study was to monitor the presence of intralesionally injected autologous AT-MSCs labelled with superparamagnetic iron oxide (SPIO) nanoparticles and green fluorescent protein (GFP) over a staggered period of 3 to 9 weeks with standing magnetic resonance imaging (MRI) and histology., Methods: Four adult warmblood horses received a unilateral injection of 10 × 10(6) autologous AT-MSCs into surgically created front-limb SDFT lesions. Administered AT-MSCs expressed lentivirally transduced reporter genes for GFP and were co-labelled with SPIO particles in three horses. The presence of AT-MSCs in SDFTs was evaluated by repeated examinations with standing low-field MRI in two horses and post-mortem in all horses with Prussian blue staining, fluorescence microscopy and with immunofluorescence and immunohistochemistry using anti-GFP antibodies at 3, 5, 7 and 9 weeks after treatment., Results: AT-MSCs labelled with SPIO particles were detectable in treated SDFTs during each MRI in T2*- and T1-weighted sequences until the end of the observation period. Post-mortem examinations revealed that all treated tendons contained high numbers of SPIO- and GFP-labelled cells., Conclusions: Standing low-field MRI has the potential to track SPIO-labelled AT-MSCs successfully. Histology, fluorescence microscopy, immunofluorescence and immunohistochemistry are efficient tools to detect labelled AT-MSCs after intralesional injection into surgically created equine SDFT lesions. Intralesional injection of 10 × 10(6) AT-MSCs leads to the presence of high numbers of AT-MSCs in and around surgically created tendon lesions for up to 9 weeks. Integration of injected AT-MSCs into healing tendon tissue is an essential pathway after intralesional administration. Injection techniques have to be chosen deliberately to avoid reflux of the cell substrate injected. In vivo low-field MRI may be used as a non-invasive tool to monitor homing and engraftment of AT-MSCs in horses with tendinopathy of the SDFT.

Background: The use of tack (equipment such as saddles and reins) and especially of bits because of rein tension resulting in pressure in the mouth is questioned because of welfare concerns. We hypothesised that rein tension patterns in walk and trot reflect general gait kinematics, but are also determined by individual horse and rider effects. Six professional riders rode three familiar horses in walk and trot. Horses were equipped with rein tension meters logged by inertial measurement unit technique. Left and right rein tension data were synchronized with the gait., Results: Stride split data (0-100 %) were analysed using mixed models technique to elucidate the left/right rein and stride percentage interaction, in relation to the exercises performed. In walk, rein tension was highest at hindlimb stance. Rein tension was highest in the suspension phase at trot, and lowest during the stance phase. In rising trot there was a significant difference between the two midstance phases, but not in sitting trot. When turning in trot there was a significant statistical association with the gait pattern with the tension being highest in the inside rein when the horse was on the outer fore-inner hindlimb diagonal., Conclusions: Substantial between-rider variation was demonstrated in walk and trot and between-horse variation in walk. Biphasic rein tensions patterns during the stride were found mainly in trot.

Reasons for Performing Study: Articular cartilage regeneration is the focus and goal of considerable research effort. Since articular chondrocytes descend from a distinct cohort of progenitor cells located in embryonic nascent joints (interzones), establishing the timing of equine interzone formation is an essential first step towards understanding equine joint and articular cartilage development., Objectives: To establish the time frame during which the equine femorotibial interzone forms., Study Design: Descriptive anatomical study., Methods: Equine embryos were harvested at 37 (E37), 40, 42, 45, 50 and 65 days' gestation. The femorotibial interzone was examined using high-resolution episcopic microscopy of E37, E42, E45, E50 and E65. Additional histology and collagen-II-immunohistochemistry were performed on E42., Results: At E37, the femorotibial interzone is first visible as a uniform layer, while at E42 the interzone is fully formed and consists of 3 morphologically distinct layers. The first evidence of cavitation was seen at E45. At E50, the cruciate ligaments were well formed and by E65, joint formation appeared complete., Conclusions: The embryogenesis of the equine femorotibial joint is similar to the developmental timeline of stage-matched human and murine embryos. Further studies looking at interzone formation on a cellular and molecular level may further our understanding of the intricate developmental patterns and pathways of articular cartilage development., (© 2014 EVJ Ltd.)

We present measurements of radio emission from cosmic ray air showers that took place during thunderstorms. The intensity and polarization patterns of these air showers are radically different from those measured during fair-weather conditions. With the use of a simple two-layer model for the atmospheric electric field, these patterns can be well reproduced by state-of-the-art simulation codes. This in turn provides a novel way to study atmospheric electric fields.

Regular exercise testing in Warmblood sport horses may, as in racing, potentially help to characterise fitness indices in different disciplines and at various competition levels and assist in understanding when a horse is 'fit to compete'. In this review an overview is given of the current state of the art of exercise testing in the Olympic disciplines of eventing, show jumping and dressage, and areas for further development are defined. In event horses, a simple four-step incremental exercise test measuring heart rate (HR), lactate concentration (LA) and velocity (V) is most often used. In dressage and riding horses, a wide variety of exercise tests have been developed, including incremental exercise tests, indoor riding tests and lunging tests. In show jumping, the use of a five-step incremental exercise test and exercise tests evaluating technical skills and fatigue of the horse has been reported. The velocity at a plasma LA of 4 mmol/L (VLA4) and HR recovery during submaximal exercise intensity have been shown to be the best parameters in event horses for predicting performance and impending injuries. In riding horses, the fitness level of horses is also an important determinant of injuries. Implementation of regular exercise testing and monitoring of training sessions may have important added value in the assessment of performance ability and potential future injuries in Warmblood sport horses. However, there is an urgent need to standardise methodologies and outcome parameters in order to make results comparable., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Nascent embryonic joints, interzones, contain a distinct cohort of progenitor cells responsible for the formation of the majority of articular tissues. However, to date the interzone has largely been studied using in situ analysis for candidate genes in the context of the embryo rather than using an unbiased genome-wide expression analysis on isolated interzone cells, leaving significant controversy regarding the exact role of the intermediate and outer interzone layers in joint formation. Therefore, in this study, using laser capture microdissection (three biological replicates), we selectively harvested the intermediate and outer interzones of mouse embryos at gestational age 15.5 days, just prior to cavitation, when the differences between the layers should be most profound. Microarray analysis (Agilent Whole Mouse Genome Oligo Microarrays) was performed and the differential gene expression between the intermediate interzone cells and outer interzone cells was examined by performing a two-sided paired Student's t-test and pathway analysis. One hundred ninety-seven genes were differentially expressed (≥ 2-fold) between the intermediate interzone and the outer interzone with a P-value ≤ 0.01. Of these, 91 genes showed higher expression levels in the intermediate interzone and 106 were expressed higher in the outer interzone. Pathway analysis of differentially expressed genes suggests an important role for inflammatory processes in the interzone layers, especially in the intermediate interzone, and hence in joint and articular cartilage development. The high representation of genes relevant to chondrocyte hypertrophy and endochondral ossification in the outer interzone suggests that it undergoes endochondral ossification.

The crosslinking agent genipin is increasingly invoked for the mechanical augmentation of collagen tissues and implants, and has previously been demonstrated to arrest mechanical damage accumulation in various tissues. This study established an in vitro dose-response baseline for the effects of genipin treatment on tendon cells and their matrix, with a view to in vivo application to the repair of partial tendon tears. Regression models based on a broad range of experimental data were used to delineate the range of concentrations that are likely to achieve functionally effective crosslinking, and predict the corresponding degree of cell loss and diminished metabolic activity that can be expected. On these data, it was concluded that rapid mechanical augmentation of tissue properties can only be achieved by accepting some degree of cytotoxicity, yet that post-treatment cell survival may be adequate to eventually repopulate and stabilize the tissue. On this basis, development of delivery strategies and subsequent in vivo study seems warranted., (Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Reasons for Performing Study: Training regimens of showjumping horses under field conditions are largely undocumented., Objectives: The aims of this study were to quantify and compare training regimens used in professional-level showjumping yards, with respect to time exercised and type of activity., Study Design: Prospective cohort study., Methods: A prospective 6-month cohort study of showjumping horses in 4 European countries (The Netherlands, Sweden, Switzerland, Great Britain) was designed to analyse training and health data, in yards with several horses in training and riders competing at professional level. Riders documented the daily frequency and duration of all physical activities of the horses. Variation in training routines were compared between riders, location and time. Mixed-models analysis was used to examine factors associated with total time exercised and time spent in flatwork., Results: In 4 countries, the 31 participating riders trained 263 European Warmbloods. The total days at risk (e.g. days in which the horses were considered fit for exercise) was 39,262. Mean time spent in daily exercise, including ridden work, lungeing and treadmill exercise, varied between riders from 19-52 min/day at risk. There was considerable variation in activities and level of heavy work and light exercise, i.e. turnout. Total time exercised and time spent in flatwork differed with month, country and proportion of days lost to training. Low variation of activities was associated with decreased total time trained and increased time spent in flatwork., Conclusions: Riders at this elite professional level of showjumping used training regimens that vary substantially in time spent training and other physical activities and showjumping horses are challenged differently during training despite competing at the same level. Whether all training regimens prepare the horses equally for the demands of competition remains to be determined., (© 2013 EVJ Ltd.)

Aims: To investigate whether autologous adipose tissue-derived mesenchymal stem cells (AT-MSCs) treatment of tendon lesions increases neovascularization during tendon healing., Materials & Methods: A standardized surgical model was used to create lesions in both front limb superficial digital flexor tendons (SDFTs) of nine horses. Either AT-MSCs or control substance was injected intralesionally 2 weeks post-surgery. Color Doppler ultrasonography of SDFTs was performed at regular intervals. Horses were euthanized 22 weeks post-treatment and SDFTs were harvested for histology., Results: The color Doppler ultrasonography signal was significantly more extensive at 2 weeks post-treatment and the number of vessels counted on histologic slides was significantly higher at 22 weeks post-treatment in AT-MSC-treated SDFTs., Conclusion: Our findings indicate that AT-MSC treatment has a beneficial effect on neovascularization of healing tendons.

Purpose: To evaluate the status of articular cartilage and bone in an equine model of spontaneous repair by using the sweep imaging with Fourier transform (SWIFT) magnetic resonance (MR) imaging technique., Materials and Methods: Experiments were approved by the Utrecht University Animal Ethics Committee. Six-millimeter-diameter chondral (n = 5) and osteochondral (n = 5, 3-4 mm deep into subchondral bone) defects were created in the intercarpal joints of seven 2-year-old horses and examined with SWIFT at 9.4 T after spontaneous healing for 12 months. Conventional T2 maps and gradient-echo images were obtained for comparison, and histologic assessment of cartilage and micro-computed tomography (CT) of bone were performed for reference. Signal-to-noise ratio (SNR) analysis was performed, and a radiologist evaluated the MR images. Structural bone parameters were derived from SWIFT and micro-CT datasets. Significance of differences was investigated with the Wilcoxon signed rank test and Pearson correlation analysis., Results: SWIFT was able to depict the different outcomes of spontaneous healing of focal chondral versus osteochondral defects. SWIFT produced constant signal intensity throughout cartilage, whereas T2 mapping showed elevated T2 values (P = .06) in repair tissue (mean T2 in superficial region of interest in an osteochondral lesion = 50.0 msec ± 10.2) in comparison to adjacent intact cartilage (mean T2 = 32.7 msec ± 4.2). The relative SNR in the subchondral plate with SWIFT (0.91) was more than four times higher than that with conventional fast spin-echo (0.12) and gradient-echo (0.19) MR imaging. The correlation between bone volume-to-tissue volume fractions determined with SWIFT and micro-CT was significant (r = 0.83, P < .01)., Conclusion: SWIFT enabled assessment of spontaneous osteochondral repair in an equine model., (© RSNA, 2013.)

The aim of this study was to monitor the fitness level of young Friesian horses and to assess whether fitness data are predictive for final performance score and whether familial aggregation of response to training could be detected. Sixty-six young Friesian horses, the offspring of six different stallions (A, B, C, D, E and F), underwent a 7-week performance test. The horses were given a performance score for their ability for dressage (in weeks 5 and 7; 0-110 points) and were evaluated for fitness using standardised exercise tests (SETs) at the beginning (week 2, SET-I) and the end (week 6, SET-II) of the period. Heart rate (HR, beats/min) was measured in both SETs, and plasma lactate concentration (LA, mmol/L) was measured only in SET-II. Fitness of the horses improved moderately but significantly between SET-I and SET-II (P=0.015). There was a large heterogeneity in responsiveness to training; some horses were high responders, whereas others were non- or low responders. There was a familial aggregation of HR canter-1 response to training (P=0.039), while the HR of stallion C's offspring decreased significantly more than those of stallions A (P=0.09), D (P=0.013) and F (P=0.009). Horses that were reluctant to exercise did not differ in HR or LA concentrations compared to those that completed the SET, which may have been a sign of overreaching. HR had no predictive value for the performance score, but horses that did not reach the anaerobic threshold in SET-II scored significantly better (73.8 ± 5.6 points) than horses that did not (69.9 ± 5.9 points, P=0.025). The findings demonstrate for the first time in the horse a familial aggregation of HR response to training, as has been reported previously in humans. Familial aggregation suggests a genetic influence on the effect of training on fitness in horses. HR could not predict final performance score, but LA concentrations during SET-II were predictive., (Copyright © 2013 Elsevier Ltd. All rights reserved.)