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Background: The modified Meek technique is not commonly used in equine wound management, despite the consistent reliable and superior results compared with other grafting techniques. Major drawbacks are the need for specialised, expensive equipment and general anaesthesia. Objectives: To describe adjustments of the modified Meek technique enabling use in the standing horse without the need for the full equipment. This implied the use of full‐thickness skin grafts manually harvested from the pectoral area and manually cut into micrografts. Graft acceptance; healing progress; and final functional and cosmetic result were outcome parameters. Study design: Descriptive case series. Methods: Eight horses with traumatic wounds at the dorsal side of the carpus or tarsus, healing by second intention, were treated. Original wound areas and areas of graft acceptance and rejection were determined from post‐processing of digital photographs and percentage acceptance, wound contraction and epithelialisation were calculated. Results: The initial mean wound area was 55.4 cm2. Graft acceptance was 95.3 ± 2.5%. Wound closure was due to 46.0 ± 25.6% wound contraction and 54.0 ± 25.6% epithelialisation and resulted in 96.8 ± 1.9% reduction of the initial wound area 28.0 ± 8.5 days after grafting. The scar was flat, flexible and functional, usually with thin and regular hair growth. The adapted procedure was fast and efficient, with a learning curve for the increased manual work. Main limitations: Small study population. Conclusions: This adapted modified Meek technique can successfully be performed in the standing horse and obviates the need for the full expensive equipment and general anaesthesia. The acceptance of the full‐thickness grafts is excellent resulting in fast and satisfactory healing. [ABSTRACT FROM AUTHOR]

Background: Data on equine lumbar pathology hardly exist in breeds other than Thoroughbreds. Objectives: To describe pathological changes of the osseous lumbar vertebral column in Warmblood horses, Shetland ponies and Konik horses. Study design: Descriptive post‐mortem study. Methods: The lumbar vertebral columns of 34 Warmblood horses, 28 Shetland ponies, and 18 Konik horses were examined by computed tomography (CT). Osteoarthritis (OA) of articular processes (APJs), OA of intertransverse joints (ITJs), intervertebral disc (IVD) mineralisation, impingement of spinous (SPs) and transverse (TPs) processes and spondylosis were scored. Breed differences in prevalence and severity of pathologies were analysed by linear regression analysis and by calculating Spearman's rank correlation coefficients (rs). Results: In Warmblood horses, the prevalence of OA of APJs, impingement of SPs and TPs was respectively 90%, 36%, and 35%, significantly higher than in the other breeds (p < 0.001). In Konik horses, IVD mineralisation (40%) and spondylosis (10%) were more frequent than in Warmbloods and Shetland ponies (p = 0.03). Severity score for OA of ITJs was highest in Shetland ponies (p < 0.001). For impingement of SPs, severity score was highest in Warmbloods (p = 0.03), and of TPs lowest in Shetland ponies (p = 0.003). For all parameters, except for spondylosis in Shetland ponies, there was a positive correlation between percentage of vertebrae affected and age, with IVD mineralisation scores increasing faster in Konik horses (p < 0.001). In all breeds, there was also a positive relation between scores of severity and age for OA of APJs and ITJs and for IVD mineralisation, with severity scores increasing faster in Shetland ponies (p = 0.04). Strong left/right correlations of the severity scores were seen for OA of the APJ, ITJ, impingement of TPs, and paramedian spondylosis (rs = 0.74–0.86, all p < 0.001). Main limitations: Clinical histories were not available. Conclusions: There are distinct breed differences between prevalence and severity of osseous pathologies of the lumbar spine. Warmblood horses have higher scores for most pathologies with IVD mineralisation being more important in Konik horses and OA of ITJs in Shetland ponies. [ABSTRACT FROM AUTHOR]

In articular cartilage (AC), the collagen arcades provide the tissue with its extraordinary mechanical properties. As these structures cannot be restored once damaged, functional restoration of AC defects remains a major challenge. We report that the use of a converged bioprinted, osteochondral implant, based on a gelatin methacryloyl cartilage phase, reinforced with precisely patterned melt electrowritten polycaprolactone micrometer‐scale fibers in a zonal fashion, inspired by native collagen architecture, can provide long‐term mechanically stable neo‐tissue in an orthotopic large animal model. The design of this novel implant was achieved via state‐of‐the‐art converging of extrusion‐based ceramic printing, melt electrowriting, and extrusion‐based bioprinting. Interestingly, the cell‐free implants, used as a control in this study, showed abundant cell ingrowth and similar favorable results as the cell‐containing implants. Our findings underscore the hypothesis that mechanical stability is more determining for the successful survival of the implant than the presence of cells and pre‐cultured extracellular matrix. This observation is of great translational importance and highlights the aptness of advanced 3D (bio)fabrication technologies for functional tissue restoration in the harsh articular joint mechanical environment. [ABSTRACT FROM AUTHOR]

Simple Summary: Repeated episodes of joint inflammation play a key role in the progression of joint diseases such as osteoarthritis. In order to better understand diseases and develop treatments, animal studies are needed. Most models of joint inflammation cause severe discomfort and irreversible damage to joints which is neither truly reflective of naturally occurring disease processes nor desirable for the welfare of the experimental animals. This study examines a potential model of recurrent lower levels of inflammation. Minute doses of lipopolysaccharide (LPS), a substance that causes inflammation, were injected into the joints of horses three times at two-week intervals, and the effect of these injections on the horses' comfort and welfare and markers of inflammation within the joint fluid were closely monitored. We found that each of these injections produced reliable and comparable levels of inflammation within the joints, with minimal impact on the horses' comfort and welfare. The joints also showed complete recovery when re-examined at a later timepoint. These results suggest that this model has potential as a refined translational model of repeated episodes of joint inflammation that is more representative of natural disease states and can be used to evaluate potential therapeutics over several weeks. This study investigates repeated low-dose lipopolysaccharide (LPS) injections in equine joints as a model for recurrent joint inflammation and its impact on animal welfare. Joint inflammation was induced in eight horses by injecting 0.25 ng of LPS three times at two-week intervals. Welfare scores and clinical parameters were recorded at baseline and over 168 h post-injection. Serial synoviocentesis was performed for the analysis of a panel of synovial fluid biomarkers of inflammation and cartilage turnover. Clinical parameters and a final synoviocentesis were also performed eight weeks after the last sampling point to assess the recovery of normal joint homeostasis. Statistical methods were used to compare the magnitude of response to each of the 3 LPS inductions and to compare the baseline and final measurements. Each LPS injection produced consistent clinical and biomarker responses, with minimal changes in welfare scores. General matrix metalloproteinase (MMP) activity and joint circumference showed greater response to the second LPS induction, but response to the third was comparable to the first. Gylcosaminoglycans (GAG) levels showed a significantly decreased response with each induction, while collagen-cleavage neoepitope of type II collagen (C2C) and carboxypropetide of type II collagen epitope (CPII) showed quicker responses to the second and third inductions. All parameters were comparable to baseline values at the final timepoint. In conclusion, a consistent, reliable intra-articular inflammatory response can be achieved with repeated injections of 0.25 ng LPS, with minimal impact on animal welfare, suggesting potential as a refined translational model of recurrent joint inflammation. [ABSTRACT FROM AUTHOR]

Summary: Background: Post‐traumatic osteoarthritis is a frequent joint disease in the horse. Currently, equine medicine lacks effective methods to diagnose the severity of chondral defects after an injury. Objectives: To investigate the capability of dual‐contrast‐enhanced computed tomography (dual‐CECT) for detection of chondral lesions and evaluation of the severity of articular cartilage degeneration in the equine carpus ex vivo. Study design: Pre‐clinical experimental study. Methods: In nine Shetland ponies, blunt and sharp grooves were randomly created (in vivo) in the cartilage of radiocarpal and middle carpal joints. The contralateral joint served as control. The ponies were subjected to an 8‐week exercise protocol and euthanised 39 weeks after surgery. CECT scanning (ex vivo) of the joints was performed using a micro‐CT scanner 1 hour after an intra‐articular injection of a dual‐contrast agent. The dual‐contrast agent consisted of ioxaglate (negatively charged, q = −1) and bismuth nanoparticles (BiNPs, q = 0, diameter ≈ 0.2 µm). CECT results were compared to histological cartilage proteoglycan content maps acquired using digital densitometry. Results: BiNPs enabled prolonged visual detection of both groove types as they are too large to diffuse into the cartilage. Furthermore, proportional ioxaglate diffusion inside the tissue allowed differentiation between the lesion and ungrooved articular cartilage (3 mm from the lesion and contralateral joint). The mean ioxaglate partition in the lesion was 19 percentage points higher (P < 0.001) when compared with the contralateral joint. The digital densitometry and the dual‐contrast CECT findings showed good subjective visual agreement. Main limitations: Ex vivo study protocol and a low number of investigated joints. Conclusions: The dual‐CECT methodology, used in this study for the first time to image whole equine joints, is capable of effective lesion detection and simultaneous evaluation of the condition of the articular cartilage. [ABSTRACT FROM AUTHOR]

Background: Gait kinematics measured during equine gait analysis are typically evaluated by analysing (asymmetry‐based) discrete variables (eg, peak values) obtained from continuous kinematic signals (eg, timeseries of datapoints). However, when used for the assessment of complex cases of lameness, such as bilateral lameness, discrete variable analysis might overlook relevant functional adaptations. Objectives: The overall aim of this paper is to compare continuous and discrete data analysis techniques to evaluate kinematic gait adaptations to lameness. Study design: Method comparison. Methods: Sixteen healthy Shetland ponies, enrolled in a research programme in which osteochondral defects were created on the medial trochlear ridges of both femurs, were used in this study. Kinematic data were collected at trot on a treadmill before and at 3 and 6 months after surgical intervention. Statistical parametric mapping and linear mixed models were used to compare kinematic variables between and within timepoints. Results: Both continuous and discrete data analyses identified changes in pelvis and forelimb kinematics. Discrete data analyses showed significant changes in hindlimb and back kinematics, where such differences were not found to be significant by continuous data analysis. In contrast, continuous data analysis provided additional information on the timing and duration of the differences found. Main limitations: A limited number of ponies were included. Conclusions: The use of continuous data provides additional information regarding gait adaptations to bilateral lameness that is complementary to the analysis of discrete variables. The main advantage lies in the additional information regarding time dependence and duration of adaptations, which offers the opportunity to identify functional adaptations during all phases of the stride cycle, not just the events related to peak values. [ABSTRACT FROM AUTHOR]

This study aimed to quantify the long‐term progression of blunt and sharp cartilage defects and their effect on joint homeostasis and function of the equine carpus. In nine adult Shetland ponies, the cartilage in the radiocarpal and middle carpal joint of one front limb was grooved (blunt or sharp randomized). The ponies were subjected to an 8‐week exercise protocol and euthanized at 39 weeks. Structural and compositional alterations in joint tissues were evaluated in vivo using serial radiographs, synovial biopsies, and synovial fluid samples. Joint function was monitored by quantitative gait analysis. Macroscopic, microscopic, and biomechanical evaluation of the cartilage and assessment of subchondral bone parameters were performed ex vivo. Grooved cartilage showed higher OARSI microscopy scores than the contra‐lateral sham‐operated controls (p < 0.0001). Blunt‐grooved cartilage scored higher than sharp‐grooved cartilage (p = 0.007) and fixed charge density around these grooves was lower (p = 0.006). Equilibrium and instantaneous moduli trended lower in grooved cartilage than their controls (significant for radiocarpal joints). Changes in other tissues included a threefold to sevenfold change in interleukin‐6 expression in synovium from grooved joints at week 23 (p = 0.042) and an increased CPII/C2C ratio in synovial fluid extracted from blunt‐grooved joints at week 35 (p = 0.010). Gait analysis outcome revealed mild, gradually increasing lameness. In conclusion, blunt and, to a lesser extent, sharp grooves in combination with a period of moderate exercise, lead to mild degeneration in equine carpal cartilage over a 9‐month period, but the effect on overall joint health remains limited. [ABSTRACT FROM AUTHOR]

Microfracture is the current standard in treatment of focal full-thickness cartilage lesions in horses, but clinical outcome may vary. Nanofracture is a novel technique that uses a commercially developed device to yield smaller diameter perforations with deeper penetration into the subchondral bone. Experimentally, in rabbits and sheep, nanofracture has been shown to result in superior repair compared to microfracture. The objective was to study the feasibility and preliminary outcome of nanofracture using a commercial device for treatment of cartilage defects in horses. Nanofracture was tested ex vivo in n = 2 cadaveric equine stifle joints and in vivo in n = 8 horses with experimental partial thickness cartilage defects in the medial femoral trochlear ridge. These were treated with an experimental biomaterial or nanofracture, and repair tissue was studied macroscopically (ICRS-I score) and microscopically (histological ICRS-II score and micro-CT) after 7 months. Both in cadaveric equine stifle joints and in vivo, the nanofracture device could readily be applied and allowed easy penetration of the subchondral bone. Repair tissue after 7 months was graded 'near-normal' macroscopically, while histologically, the abundant repair tissue proved mainly fibrocartilaginous in nature. Micro-CT revealed near-full restoration of mid-lesion cartilage layer thickness but altered subchondral bone microarchitecture. The in vivo study did not include a control group treated with conventional microfracture for comparison. To our knowledge, this is the first report on bone marrow stimulation using nanofracture as a potential method to enhance chondral defect repair in horses. In the in vivo study, no clinical adverse effects were observed, and promising good defect filling with fibrocartilaginous tissue was seen 7 months after treatment. [ABSTRACT FROM AUTHOR]

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

The use of extracellular vesicles (EVs) as a potential therapy is currently explored for different disease areas. When it comes to the treatment of joint diseases this approach is still in its infancy. As in joint diseases both inflammation and the associated articular tissue destruction are important factors, both the immune-suppressive and the regenerative properties of EVs are potentially advantageous characteristics for future therapy. There is, however, only limited knowledge on the basic features, such as numerical profile and function, of EVs in joint articular tissues in general and their linking medium, the synovial fluid, in particular. Further insight is urgently needed in order to appreciate the full potential of EVs and to exploit these in EV-mediated therapies. Physiologic joint homeostasis is a prerequisite for proper functioning of joints and we postulate that EVs play a key role in the regulation of joint homeostasis and hence can have an important function in re-establishing disturbed joint homeostasis, and, in parallel, in the regeneration of articular tissues. In this mini-review EVs in the joint are explained from a historical perspective in both health and disease, including the potential niche for EVs in articular tissue regeneration. Furthermore, the translational potential of equine models for human joint biology is discussed. Finally, the use of MSC-derived EVs that is recently gaining ground is highlighted and recommendations are given for further EV research in this field. [ABSTRACT FROM AUTHOR]

Background: In today's porcine industry, lameness has a major welfare and economic impact, and is often caused by osteochondrosis (OC). The etiological factors of the disease have been studied in depth, however, to this day, little is known about the natural course of the disorder and how it can be detected at an early stage in pigs. The aim of this pilot study was to assess the potential of three non-invasive techniques for the detection and monitoring of early OC processes in piglets. A group of weaned piglets (n = 19) were examined longitudinally using radiographs, a visual lameness scoring scheme and a quantitative pressure-mat based locomotion analysis system to detect OC in the humeroradial, femoropatellar and tarsocrural joints. At several time points, a selection of animals was euthanized for post-mortem examinations, including histology, which was the gold standard. Results: In this study, clear signs of subclinical signs of OC were observed, however, we were unsuccessful in producing clinical OC. Lesions were observed to be commonly bilaterally symmetric in the joints examined in 80 % of cases. The radiographic examinations showed a clear correlation with the gold standard, particularly when subclinical lesions were of a high histological score. Moreover, radiography was also able to detect the early repair processes, which appeared to take place at least until 14 weeks of age. Both visual scoring and pressure mat analyses showed good intra-assay reproducibility, with the pressure mat showing intra-class correlation values between 0.44 and 0.6 and the inter-observer agreement of visual scoring method was between 88 and 96 %, however their correlation to OC lesions detected by histology was very weak, with only 2 out of 12 traits for the visual scoring method showing significant and biologically logical relations to a specific joint having histological OC lesions. For the pressure mat, only a maximum of 5 associations for specific joints with histological OC lesions were found out of a possible 8. Conclusion: All tested in-vivo methods showed good reproducibility. Radiography was the most reliable technique to detect and monitor longitudinally the earliest signs of OC in these piglets. It also demonstrated that the "Point of No Return" (PNR) of the disease, when repair processes end, might be later than anticipated, after 13 weeks of age. All in all, our study shows that the timing of the use of these in-vivo methods is critical to detect and monitor OC, especially in the early phases of the disease. It also shows the difficulty in producing OC regardless of the optimization of the experimental settings in relation to the etiological factors known to induce OC. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) in synovial fluid (SF) are gaining increased recognition as important factors in joint homeostasis, joint regeneration, and as biomarkers of joint disease. A limited number of studies have investigated EVs in SF samples of patients with joint disease, but knowledge on the role of EVs in healthy joints is lacking. In addition, no standardized protocol is available for isolation of EVs from SF. Based on the high viscosity of SF caused by high concentrations of hyaluronic acid (HA) - a prominent extracellular matrix component - it was hypothesized that EV recovery could be optimized by pretreatment with hyaluronidase (HYase). Therefore, the efficiency of EV isolation from healthy equine SF samples was tested by performing sequential ultracentrifugation steps (10,000g, 100,000g and 200,000g) in the presence or absence of HYase. Quantitative EV analysis using high-resolution flow cytometry showed an efficient recovery of EVs after 100,000g ultracentrifugation, with an increased yield of CD44+ EVs when SF samples were pretreated with HYase. Morphological analysis of SF-derived EVs with cryo-transmission-electron microscopy did not indicate damage by high-speed ultracentrifugation and revealed that most EVs are spherical with a diameter of 20-200 nm. Further protein characterization by Western blotting revealed that healthy SF-derived EVs contain CD9, Annexin-1, and CD90/Thy1.1. Taken together, these data suggest that EV isolation protocols for body fluids that contain relatively high amounts of HA, such as SF, could benefit from treatment of the fluid with HYase prior to ultracentrifugation. This method facilitates recovery and detection of CD44+EVs within the HA-rich extracellular matrix. Furthermore, based on the findings presented here, it is recommended to sediment SF-derived EVs with at least 100,000g for optimal EV recovery. [ABSTRACT FROM AUTHOR]

The role of intrinsic and extrinsic healing in injured tendons is still debated. In this study, we characterized cell plasticity, proliferative capacity, and migration characteristics as proxy measures of healing potential in cells derived from the peritenon (extrinsic healing) and compared these to cells from the tendon core (intrinsic healing). Both cell populations were extracted from horse superficial digital flexor tendon and characterized for tenogenic and matrix remodeling markers as well as for rates of migration and replication. Furthermore, colony-forming unit assays, multipotency assays, and real-time quantitative polymerase chain reaction analyses of markers of osteogenic and adipogenic differentiation after culture in induction media were performed. Finally, cellular capacity for differentiation towards a myofibroblastic phenotype was assessed. Our results demonstrate that both tendon- and peritenon-derived cell populations are capable of adipogenic and osteogenic differentiation, with higher expression of progenitor cell markers in peritenon cells. Cells from the peritenon also migrated faster, replicate more quickly, and show higher differentiation potential toward a myofibroblastic phenotype when compared to cells from the tendon core. Based on these data, we suggest that cells from the peritenon have substantial potential to influence tendon-healing outcome, warranting further scrutiny of their role. [ABSTRACT FROM AUTHOR]

Mammalian articular cartilage serves diverse functions, including shock absorption, force transmission and enabling low-friction joint motion. These challenging requirements are met by the tissue’s thickness combined with its highly specific extracellular matrix, consisting of a glycosaminoglycan-interspersed collagen fiber network that provides a unique combination of resilience and high compressive and shear resistance. It is unknown how this critical tissue deals with the challenges posed by increases in body mass. For this study, osteochondral cores were harvested post-mortem from the central sites of both medial and lateral femoral condyles of 58 different mammalian species ranging from 25 g (mouse) to 4000 kg (African elephant). Joint size and cartilage thickness were measured and biochemical composition (glycosaminoclycan, collagen and DNA content) and collagen cross-links densities were analyzed. Here, we show that cartilage thickness at the femoral condyle in the mammalian species investigated varies between 90 µm and 3000 µm and bears a negative allometric relationship to body mass, unlike the isometric scaling of the skeleton. Cellular density (as determined by DNA content) decreases with increasing body mass, but gross biochemical composition is remarkably constant. This however need not affect life-long performance of the tissue in heavier mammals, due to relatively constant static compressive stresses, the zonal organization of the tissue and additional compensation by joint congruence, posture and activity pattern of larger mammals. These findings provide insight in the scaling of articular cartilage thickness with body weight, as well as in cartilage biochemical composition and cellularity across mammalian species. They underscore the need for the use of appropriate in vivo models in translational research aiming at human applications. [ABSTRACT FROM AUTHOR]

The effectiveness of new therapies to treat tendon injuries is difficult to determine and is often based on semi-quantitative methods, such as grey level analysis of ultrasonographic images or subjective pain scores. The alternatives are costly and long-lasting end-stage studies using experimental animals. In this study, a method of ultrasonographic tissue characterisation (UTC), using mathematical analysis of contiguous transverse ultrasonographic images, was used for intra-vital monitoring of the healing trajectory of standardised tendon lesions treated with platelet rich plasma (PRP) or placebo. Using UTC it was possible to detect significant differences between the groups in the various phases of repair. At end stage, over 80% of pixels showed correct alignment in the PRP group, compared with just over 60% in the placebo group (P < 0.05). UTC also showed significant differences in the course of the healing process between PRP treated and placebo treated animals throughout the experiment. It was concluded that computerised analysis of ultrasonographic images is an excellent tool for objective longitudinal monitoring of the effects of treatments for superficial digital flexor tendon lesions in horses. [ABSTRACT FROM AUTHOR]

Abstract: Pain recognition and management in animals has advanced considerably in the last decade and currently animal welfare is receiving increasing public interest. However, the comprehensive assessment of pain has been inadequately addressed in horses and till recently composite pain scales (CPSs) have hardly been validated for use in patients. We investigated the reliability and clinical applicability of a CPS, originally developed under experimental conditions for assessing pain in horses with various acute soft-tissue and orthopedic conditions before and after general anesthesia and/or (non)elective surgery. These clinical cases (n = 94) were scored by means of the CPS twice daily. Horses without painful conditions and horses admitted for nonpainful diagnostic procedures under general anesthesia were compared with those that were admitted with either acute or chronic surgical and nonsurgical painful conditions of both visceral and somatic origin. Scores of observer 1 were compared with observer 2 to study inter-observer reliability. Composite pain scores showed low baseline values in healthy animals with nonpainful conditions and were not affected when general anesthesia was the only intervention. Inter-observer reliability was very high (n = 23 horses; weighted kappa correlation coefficient, κ = 0.81). Horses with painful conditions responding well to analgesic treatment could be discriminated from horses that had to be euthanized on humane grounds because of painful nonresponsive conditions. We found the CPS to be a promising tool that has the potential to provide a good basis for direct day-to-day assessment of pain status in equine patients with various painful conditions in the future. [ABSTRACT FROM AUTHOR]

Tendon injuries are notorious for their slow and functionally inferior healing. Intratendinous application of platelet-rich plasma (PRP) has been reported to stimulate the repair process of tendon injuries, but there is little conclusive evidence for its effectiveness. A placebo-controlled experimental trial was performed to test the hypothesis that a single intratendinous PRP treatment enhances the quality of tendon repair, as evidenced by improved biochemical, biomechanical, and histological tissue properties. In six horses, tendon lesions were created surgically in the Superficial Digital Flexor Tendons (SDFT) of both front limbs, one of which was treated with PRP and the other with saline. After 24 weeks, the tendons were harvested for biochemical, biomechanical, and histological evaluations. Collagen, glycosaminoglycan, and DNA content (cellularity) was higher in PRP-treated tendons ( p = 0.039, 0.038, and 0.034, respectively). The repair tissue in the PRP group showed a higher strength at failure ( p = 0.021) and Elastic Modulus ( p = 0.019). Histologically, PRP-treated tendons featured better organization of the collagen network ( p = 0.031) and signs of increased metabolic activity ( p = 0.031). It was concluded that PRP increases metabolic activity and seems to advance maturation of repair tissue over nontreated experimentally induced tendon lesions, which suggests that PRP might be beneficial in the treatment of clinical tendon injuries. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:211-217, 2010 [ABSTRACT FROM AUTHOR]

We investigated the effects of exercise-induced loading on the collagen network of equine articular cartilage. Collagen fibril architecture at a site (1) subjected to intermittent high-intensity loading was compared with that of an adjacent site (2) sustaining continuous low-level load. From horses exposed to forced exercise (CONDEX group) or not (PASTEX group), the spatial parallelism of fibrils and the orientation angle between fibrils and the surface at depths 9 µm apart through cartilage from surface to tidemark were determined using polarized light microscopy, and expressed as parallelism index (PI) and orientation index (OI). PI was significantly higher in site 2 than 1 in CONDEX and PASTEX groups. PI was significantly higher in forced exercised horses at site 2 but not site 1. OI was significantly greater (more perpendicular to the surface) in the superficial and deep cartilage of site 2 than 1 in both CONDEX and PASTEX groups. Superficial zone OI was higher in exercised horses at site 1 but not at site 2. Exercise increased collagen parallelism and affected orientation. The site differences in OI indicate that Benninghoff's classic predominantly perpendicular arcades appear not to be a consistent architectural feature, but adapt to local forces sustained. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res [ABSTRACT FROM AUTHOR]

Abstract: During biomineralization the organism controls the nature, orientation, size and shape of the mineral phase. The aim of this study was to investigate whether proteins or vesicles that are constitutively released by growing ATDC5 cells have the ability to affect the formation of the calcium phosphate crystal. Therefore, subconfluent cultured ATDC5 cells were incubated for 1 h in medium without serum. Subsequently, medium was harvested and incubated for 24 h in the presence of additional Pi. This resulted in the formation of flat mineralizing structures (FMS), consisting of complex irregularly shaped flat crystals, which occasionally contained fiber-like structures (∼40 μm in size). Without pre-incubation of medium with cells, only small punctate (dot like) calcium phosphate precipitates were observed. The formation of FMS was shown to be caused by soluble factors released by subconfluent ATDC5 cells. Proteomic analysis by mass spectrometry showed that FMS contained a specific set intracellular proteins, serum proteins, and extracellular matrix proteins. Bulk cytosolic proteins derived from homogenized cells or serum proteins did, however, not induce the formation of FMS. Conditioned medium from HeLa, CHO K1, RAW 264.7 and MDCK cells was also capable to form FMS under our experimental conditions. Therefore the formation of FMS seems to be caused by specific soluble factors constitutively released by ADTC5 and other cells. This in vitro model system can be used as a tool to identify factors that affect the shape of the biomineral phase. [Copyright &y& Elsevier]

The purpose of this study was to determine whether individual limb forces could be calculated accurately from kinematics of trotting and walking horses. We collected kinematic data and measured vertical ground reaction forces on the individual limbs of seven Warmblood dressage horses, trotting at 3.4 m s-1 and walking at 1.6 m s-1 on a treadmill. First, using a segmental model, we calculated from kinematics the total ground reaction force vector and its moment arm relative to each of the hoofs. Second, for phases in which the body was supported by only two limbs, we calculated the individual reaction forces on these limbs. Third, we assumed that the distal limbs operated as linear springs, and determined their force-length relationships using calculated individual limb forces at trot. Finally, we calculated individual limb force-time histories from distal limb lengths. A good correspondence was obtained between calculated and measured individual limb forces. At trot, the average peak vertical reaction force on the forelimb was calculated to be 115±0.9 N kg-1 and measured to be 11.7±0.9 N kg-1, and for the hindlimb these values were 9.8±0.7 N kg-1 and 10.0±0.6 N kg-1, respectively. At walk, the average peak vertical reaction force on the forelimb was calculated to be 6.9±0.5 N kg-1 and measured to be 7.1±03 N kg-1, and for the hindlimb these values were 4.8±0.5 N kg-1 and 4.7±03 N kg-1, respectively. It was concluded that the proposed method of calculating individual limb reaction forces is sufficiently accurate to detect changes in loading reported in the literature for mild to moderate lameness at trot. [ABSTRACT FROM AUTHOR]

Abstract: Sodium nitroprusside (SNP) is a nitric oxide (NO) donor drug, which is therapeutically used as a vasodilating drug in heart transplantations. In our previous study it was found that SNP at a concentration of 100 μM inhibited mineralization in a cell culture system, indicating that the beneficial effects of this drug may also include inhibition of vascular calcification. The aim of this study was to investigate which bioactive compounds generated from SNP inhibit mineralization. ATDC5 cells were grown for 14 days and mineralization was induced by addition of 5 mM phosphate for 24 h. Mineralization was determined by staining precipitated calcium with an alizarin red stain. It was found that the NO donors S-nitrosogluthatione and S-nitroso-N-acetylpenicillamine were not able to inhibit mineralization and NO scavengers could not antagonize the inhibiting effect of SNP on mineralization. The iron chelator deferoxamine (200 μM) antagonized the inhibiting effect on mineralization mediated by SNP and ammonium iron sulfate inhibited mineralization in a dose-dependent manner (10–100 μM). Furthermore, iron ions (30 μM) were detected to be released from SNP in the cell culture. These data show that the iron moiety of sodium nitroprusside, rather than nitric oxide inhibits mineralization. [Copyright &y& Elsevier]

Objective--To describe spontaneous locomotion activity of foals kept under various management conditions and assess the suitability of global positioning system (GPS) technology for recording foal activity. Animals--59 foals. Procedures--During the foals' first 4 months of life, 921 observation periods (15 minutes each) were collected and analyzed for locomotion activities. The GPS system was evaluated by simultaneously carrying out field observations with a handheld computer. Results--Foals spent 0.5% of total observed time cantering, 0.2% trotting, 10.7% walking, 32.0% grazing, 34.8% standing, and 21.6% lying down. Total observed daytime workload (velocity X distance) in the first month was approximately twice that in the following months. Locomotion activity decreased with increasing age. Colts had more activity than fil-lies in certain periods, and foals that were stabled for some portion of the day had compensatory locomotion activity, which was probably insufficient to reach the level of foals kept continually outside. The GPS recordings and handheld-computer observations were strongly correlated for canter, trot, and walk and moderately correlated for standing and lying. Correlation for grazing was low. Conclusions and Clinical Relevance--Results indicated that domestically managed foals, when kept 24 h/d at pasture, will exercise at a level comparable with feral foals. High workload during the first month of life might be important for conditioning the muscuIoskeletal system. The GPS technique accurately quantified canter, trot, and walk activities; less accurately indexed resting; and was unsuitable for grazing because of the wide array of velocities used while foraging. [ABSTRACT FROM AUTHOR]

Objective--To determine the mechanism that enables horses to partially counteract the shift of the center of pressure under the hoof induced by changes in hoof morphology attributable to growth and wear during a shoeing interval. Animals--18 clinically sound Warmblood horses. Procedures--Horses were evaluated 2 days and 8 weeks after shoeing during trotting on a track containing pressure-force measuring plates and by use of a synchronous infrared gait analysis system set at a frequency of 240 Hz. All feet were trimmed toward straight alignment of the proximal, middle, and distal phalanges and shod with standard flat shoes. ResuIts--Temporal characteristics such as stance time and the time between heel lift and toe off (ie, breakover duration) did not change significantly as a result of shoeing interval. Protraction and retraction angles of the limbs did not change. Compensation was achieved through an increase in the dorsal angle of the metacarpohalangeal or metarsophalangeal (fetlock) joint and a concomitant decrease of the dorsal angle of the hoof wall and fetlock. There was an additional compensatory mechanism in the hind limbs during the landing phase. Conclusions and Clinical Relevance--Horses compensate for changes in hoof morphology that develop during an 8-week shoeing interval such that they are able to maintain their neuromuscular pattern of movement. The compensation consists of slight alterations in the angles between the distal segments of the limb. Insight into natural compensation mechanisms for hoof imbalance will aid in the understanding and treatment of pathologic conditions in horses. [ABSTRACT FROM AUTHOR]

Objective To evaluate an arthroscopic indentation instrument (Artscan 200) for assessment of the health status of equine articular cartilage. Study Design In vitro experiment using equine isolated proximal phalanx (P1) specimens. Sample Population P1 specimens from 39 horses (aged 1.5–22 years). Methods Reproducibility was tested by determination of the coefficient of variation (CV). Dynamic modulus and cartilage degeneration index (CDI) values were measured at 2 predefined sites (site 1, joint margin; site 2, joint center) to assess the accuracy and to evaluate the relation with surface integrity. Results CV was 9.0%. A significant decrease in indenter force was identified when dynamic modulus values decreased to <2.5 MPa (range of tested samples 0.9–8.1 MPa) and when CDI values at site 1 increased to >50% (range 5.4–72.8%). Conclusions Technique reproducibility was adequate but accuracy was limited. The device identified degeneration-associated decreases in cartilage stiffness only when the mechanical properties of the cartilage were considerably changed. Clinical Relevance Usefulness of this indentation instrument during arthroscopic surgery would be limited in the initial phase of OA-like cartilage degeneration, but may yield important information in more advanced OA. [ABSTRACT FROM AUTHOR]

An overview is given of the direct and long-term effects of exercise on the biochemical characteristics of cartilage and subchondral bone, and on the metabolic activity of chondrocytes in the juvenile horse. In the experimental setup 43 foals were reared until weaning at 5 months of age under similar conditions, except for the type and amount of exercise. Fifteen foals remained at pasture (Pasture group and also control group), 14 foals were kept in box stalls (Box group), and 14 foals were kept in the same box stalls but were subjected daily to an increasing number of gallop sprints (Training group). After weaning 8 foals from each group were euthanised. All remaining 19 animals were housed together in a loose box with access to a small paddock to study a possible reversibility of exercise-induced effects. Post mortem subchondral bone and cartilage samples were collected and analysed for bone morphogenic enzymes, matrix composition, chondrocyte metabolic activity, and bone mineral density. It resulted that lack of exercise leads to a retardation of the normal development of the joint. This is largely compensated for when afterwards a more normal exercise regimen is followed. Most parameters in the Training group approximated those of the pastured foals at age 5 months. However, at age 11 months there were indications for a reduced performance of the investigated tissues in this group. It is concluded that regular, sub-maximal loading, as occurred in the Pasture group, seems best for an optimal development of the musculoskeletal tissues. The combination of short bouts of heavy exercise superimposed on a basic box rest regimen appears to have adverse effects on long-term viability of the tissues and may hence lead to an impaired resistance to injury. [ABSTRACT FROM AUTHOR]

There is an increasing interest in controlled release systems for local therapy in the treatment of human and equine joint diseases, aiming for optimal intra-articular concentrations with no systemic side effects. In this study, the intra-articular tolerability and suitability for local and sustained release of tacrolimus (FK506) from monospheres composed of [PDLA-PEG1000]-b-PLLA multiblock copolymers were investigated. Unloaded and tacrolimus-loaded (18.4 mg tacrolimus/joint) monospheres were injected into the joints of six healthy horses, with saline and hyaluronic acid (HA) in the contralateral joints as controls. Blood and synovial fluid were analysed for the tacrolimus concentration and biomarkers for inflammation and cartilage metabolism. After an initial burst release, sustained intra-articular tacrolimus concentrations (>20 ng/mL) were observed during the 42 days follow-up. Whole-blood tacrolimus levels were below the detectable level (<0.5 ng/mL). A transient inflammatory reaction was observed for all substances, evidenced by increases of the synovial fluid white blood cell count and total protein. Prostaglandin and glycosaminoglycan release were increased in joints injected with unloaded monospheres, which was mitigated by tacrolimus. Both tacrolimus-loaded monospheres and HA transiently increased the concentration of collagen II cleavage products (C2C). A histologic evaluation of the joints at the endpoint showed no pathological changes in any of the conditions. Together, these results indicate the good biocompatibility of intra-articular applied tacrolimus-loaded monospheres combined with prolonged local drug release while minimising the risk of systemic side effects. Further evaluation in a clinical setting is needed to determine if tacrolimus-loaded monospheres can be beneficial in the treatment of inflammatory joint diseases in humans and animals. [ABSTRACT FROM AUTHOR]

Meloxicam, a preferential COX‐2 inhibitor, is a commonly used NSAID in pigs. Besides having potential side effects on the gastrointestinal tract, this type of drug might potentially affect osteogenesis and chondrogenesis, processes relevant to growing pigs. Therefore, the effects of long‐term meloxicam treatment on growing pigs were studied. Twelve piglets (n=6 receiving daily meloxicam 0.4 mg/kg orally from 48 until 110 days of age; n=6 receiving only applesauce (vehicle control)) were subjected to visual and objective gait analysis by pressure plate measurements at several time points. Following euthanasia a complete postmortem examination was performed and samples of the talus and distal tibia, including the distal physis, were collected. Trabecular bone microarchitecture was analysed by microCT scanning, bone stiffness by compression testing and growth plate morphology using light microscopy. Animals were not lame and gait patterns did not differ between the groups. Pathological examination revealed no lesions compatible with known side effects of NSAIDs. Trabecular bone microarchitecture and growth plate morphology did not differ between the two groups. The findings of this in vivo study reduce concerns regarding the long‐term use of meloxicam in young, growing piglets. [ABSTRACT FROM AUTHOR]

Background: If chondrocytes from the superficial, middle, and deep zones of articular cartilage could maintain or regain their characteristic properties during in vitro culture, it would be feasible to create constructs comprising these distinctive zones.Hypothesis: Zone-specific characteristics of zonal cell populations will disappear during 2-dimensional expansion but will reappear after 3-dimensional redifferentiation, independent of the culture technique used (alginate beads versus pellet culture).Study Design: Controlled laboratory study.Methods: Equine articular chondrocytes from the 3 zones were expanded in monolayer culture (8 donors) and subsequently redifferentiated in pellet and alginate bead cultures for up to 4 weeks. Glycosaminoglycans and DNA were quantified, along with immunohistochemical assessment of the expression of various zonal markers, including cartilage oligomeric protein (marking cells from the deeper zones) and clusterin (specifically expressed by superficial chondrocytes).Results: Cell yield varied between zones, but proliferation rates did not show significant differences. Expression of all evaluated zonal markers was lost during expansion. Compared to the alginate bead cultures, pellet cultures showed a higher amount of glycosaminoglycans produced per DNA after redifferentiation. In contrast to cells in pellet cultures, cells in alginate beads regained zonal differences, as evidenced by zone-specific reappearance of cartilage oligomeric protein and clusterin, as well as significantly higher glycosaminoglycans production by cells from the deep zone compared to the superficial zone.Conclusion: Chondrocytes isolated from the 3 zones of equine cartilage can restore their zone-specific matrix expression when cultured in alginate after in vitro expansion.Clinical Relevance: Appreciation of the zonal differences can lead to important advances in cartilage tissue engineering. Findings support the use of hydrogels such as alginate for engineering zonal cartilage constructs. [ABSTRACT FROM PUBLISHER]