Prognostic value of power doppler ultrasonography for equine superficial digital flexor tendon injury in thoroughbred racehorses.

The potential value of hypervascularity detected with power Doppler ultrasonography (PDU) within equine superficial digital flexor tendon (SDFT) as a prognostic factor of SDFT injury is not clear. The purpose of this study was to test the hypothesis that hypervascularity within SDFT is one of the risk factors for subsequent severe SDFT injury and to evaluate the prognostic value. A prospective cohort study of 97 Thoroughbred racehorses without any clinical signs of SDFT injury was conducted. Six variables of age, body weight, sex, the cross-sectional area of SDFT, PDU signal within SDFT and experience of steeplechase were assessed for the possibility of risk factors of subsequent SDFT injury in follow-up period of 1 year. Multivariable logistic regression analyses were used for assessment of the odds ratios (ORs) and 95 % confidence intervals (CIs) of SDFT injury. Multivariable logistic regression analysis revealed that the PDU signal within SDFT was a risk factor for the development of SDFT injury in follow-up period (P = 0.017). The adjusted OR of SDFT injury was significantly higher in PDU positive group than in PDU negative group (OR 3.17, 95 % CIs 1.20-8.35). Although further studies are required, these results would be useful for early detection and/or prevention of development for clinical severe SDFT injury.

A retrospective cohort study investigating risk factors for the failure of Thoroughbred racehorses to return to racing after superficial digital flexor tendon injury.

A retrospective cohort study was conducted to investigate risk factors for the failure of Thoroughbred racehorses to return to racing after an injury of the superficial digital flexor tendon (SDFT). Successful return was defined as the completion of five or more races after SDFT injury. The official Japan Racing Association (JRA) medical records of racehorses with a core-type SDFT injury were reviewed for clinical variables related to the characteristics of the horse and the severity of SDFT injuries at the time of diagnosis. Data on racing outcomes were obtained from the official JRA racing database. Risk factors were screened using univariable logistic regression and subsequent multivariable model building. Forty-nine of 346 (14.2%) horses successfully returned to racing after SDFT injuries. Multivariable model building revealed that an increase in the total number of injured zones (defined as the total number of zones in which the injured hypoechoic area was observed at the time of ultrasonographic diagnosis of SDFT injury) was associated with an increased risk of failure to return to racing after SDFT injury. Horse characteristics, such as age, body mass and sex, were not associated with a successful return to racing. In the rehabilitation of cases with larger (longer) lesions, more effective and careful medical management may be needed for an improvement in the athletic outcomes.

The use of sonoelastography to assess the recovery of stiffness after equine superficial digital flexor tendon injuries: A preliminary prospective longitudinal study of the healing process.

BACKGROUND: The objective assessment of the mechanical properties of the superficial digital flexor tendon (SDFT) could provide useful information for the rehabilitation of horses with SDFT injuries. Assessment of strain ratio (the strain of a standard reference divided by that of lesions) is a quantitative method in sonoelastography for evaluating tissue stiffness in vivo. As yet, no longitudinal studies have used strain ratio to evaluate the progression of stiffness in SDFT injuries. OBJECTIVES: To test the hypothesis that strain ratio can evaluate the recovery of stiffness during the healing of SDFT injuries. STUDY DESIGN: Prospective and longitudinal study with observer-blinded evaluation. METHODS: Ultrasonography, including sonoelastography, was performed in seven Thoroughbred horses with naturally occurring SDFT injuries at five time points: within 20 days of the injury, and at 2, 3, 6 and 9 months after the injury. Blinded sonoelastographic images were independently evaluated by two veterinarians to assess interobserver agreement. The recovery of stiffness and echogenicity in lesions were evaluated using the strain ratio and grey-scale ratio (echogenicity of lesions divided by that of the surrounding area), respectively. RESULTS: Interobserver agreement was assessed as 'almost perfect'. Strain ratios were significantly higher at 9 months after injury than at the other time points (all P<0.05). Strain ratios at 6 months after injury were significantly higher than those at earlier time points (P<0.05). Grey-scale ratios within 20 days of injury were significantly lower than those at the other time points (all P<0.05). MAIN LIMITATIONS: Validations of SDFT status were evaluated only by recovery of the echogenicity in lesions and not by histopathological examination. CONCLUSIONS: Although further studies are needed to validate the relationships between injured SDFT status and sonoelastographic findings, this preliminary study shows that strain ratio may provide a means to monitor the recovery of stiffness in lesions during rehabilitation, even when the grey-scale ratio remains unchanged from a few months after SDFT injury. The Summary is available in Chinese - see Supporting Information.

Application of sonoelastography for evaluating the stiffness of equine superficial digital flexor tendon during healing.

Sonoelastography can assess the inner stiffness of tissues. Sonoelastographic evaluation of injured equine superficial digital flexor tendons (SDFTs) is considered to be useful for assessing the stiffness of a lesion even during late-stage rehabilitation. The purpose of this study was to investigate and compare the sonoelastographic appearance of injured SDFTs over time from the onset of the injury. Eighteen horses were classified into three groups according to the length of time from injury onset: group A, within two weeks after injury; group B, approximately five months after injury; and group C, approximately nine months after injury. Longitudinal and transverse images of all injured SDFTs were obtained using grey-scale ultrasonography and sonoelastography. Grey-scale and sonoelastographic images were evaluated by two observers using echogenicity-grading and colour-grading systems, respectively. The authors evaluated the interobserver agreement and compared the grades among the three groups. The results indicated almost perfect interobserver agreement. Significant differences were found in the sonoelastography among the three groups, whereas no significant difference was found in the grey-scale ultrasonography between groups B and C. Sonoelastography is a feasible and useful modality to evaluate the equine injured SDFTs in vivo and to distinguish between them among the different phases even during the chronic phase.

Feasibility study of monitoring of plasma etching chamber conditions using superimposed high-frequency signals on rf power transmission line.

An in situ monitoring system that can detect changes in the conditions of a plasma etching chamber has been developed. In the system, low-intensity high-frequency signals are superimposed on the rf power transmission line used for generating plasma. The system measures reflected high-frequency signals and detects the change in their frequency characteristics. The results indicate that the system detects the changes in the conditions in etching chambers caused by the changes in the electrode gap and the inner wall condition and demonstrate the effectiveness of the system. The system can easily be retrofitted to mass-production equipment and it can be used with or without plasma discharge. Therefore, our system is suitable for in situ monitoring of mass-production plasma etching chambers. The system is expected to contribute to development of predictive maintenance, which monitors films deposited on the inner wall of the chamber and prevents equipment faults caused by misalignment of chamber parts in mass-production equipment.

Note: practical monitoring system using characteristic impedance measurement during plasma processing.

A method for characteristic impedance monitoring (CIM) is developed for anomaly detection during plasma processing. Advantages of the method include high-sensitivity and real-time monitoring, as well as structural simplicity to install the measurement system in the mass production equipment. To obtain real-time monitoring without time delay, our CIM system consists of a directional coupler and a newly developed vector processing system instead of the conventional high voltage and current probes. The system can measure the time variations of characteristic and load impedances even while the plasma is generated.

Cell-based therapies for tendon and ligament injuries.

Tendon and ligament injuries have proved difficult to treat effectively. Cell-based therapies offer the potential to harness the complex protein synthetic machinery of the cell to induce a regenerative response rather than fibrous scarring. This article reviews the current state of play with respect to the clinically used cell preparations for the treatment of tendon and ligaments overstrain injuries.

Optimisation of bone marrow aspiration from the equine sternum for the safe recovery of mesenchymal stem cells.

REASONS FOR PERFORMING STUDY: Mesenchymal stem cell (MSC) therapy for orthopaedic disease is being used with increasing frequency; there is a need to define a safe, reliable and effective technique for the recovery of MSCs from the sternum of the horse. OBJECTIVES: To describe an optimised safe technique for obtaining bone marrow-derived MSCs from the sternum of the Thoroughbred horse. METHODS: The anatomical relationship of the sternum with the heart and internal anatomy was demonstrated in cadavers. Sternal anatomy was evaluated ultrasonographically and after midline sectioning. Sternebrae were examined histologically after aspiration to determine the effect of needle insertion. The quality of the aspirate was evaluated as the number of colony-forming units from sequential and separately aspirated 5 ml aliquots and assessed for their multipotency using trilineage differentiation. RESULTS: The optimal safe location for the needle was the 5th sternebra because it had a safe dorsoventral thickness and was cranial to the apex of the heart. This sternebra could be reliably identified ultrasonographically. Aspirates could also be obtained from the 4th and 6th sternebrae, although the former is between the front limbs and the latter closer to the heart. Minimal disruption of the internal bony architecture was seen after needle insertion through the thin outer cortex and the first 5 ml aliquot contained the greatest number of colony-forming units of mesenchymal stem cells with trilineage capabilities. CONCLUSIONS: Accurate placement of a Jamshidi needle into the medullary cavity of the 4th-6th individual sternebrae is facilitated by the use of ultrasonography and enables aspiration of bone marrow reliably with minimal damage to the sternum and risk to the horse. POTENTIAL CLINICAL RELEVANCE: Sternal marrow aspiration as described is a safe and reliable technique to obtain MSCs for orthopaedic cell-based therapies.

Can exercise modulate the maturation of functionally different immature tendons in the horse?

Tendons can be considered in two functional groups, those contributing to energetics of locomotion and those acting solely to position the limb. The energy-storing tendons in both human and equine athletes have a high frequency of injury with similar pathophysiology. In previous studies, high-intensity exercise appears to induce a disruption of the matrix rather than functional adaptation in adults. Here we explore the hypothesis that the introduction of controlled exercise during growth would result in an adaptive response without deleterious effects. Young horses were given a controlled exercise program similar to that previously shown to induce matrix changes in energy-storing tendons of skeletally mature animals. The tendons were assessed in relation to mechanical properties, molecular composition, and morphology. Results showed a significant increase in cartilage oligomeric matrix protein (COMP) in the positional tendon but not in the energy-storing tendon. Other matrix properties and mechanical properties were not significantly changed. While the imposition of high-strain-rate exercise in immature horses failed to augment the development of the energy-storing tendon over and above that induced by normal pasture exercise, it did not induce deleterious changes, supporting an earlier introduction of athletic training in horses.

Estimation of heritability for superficial digital flexor tendon injury by Gibbs sampling in the Thoroughbred racehorse.

The superficial digital flexor tendon (SDFT) injury causes considerable wastage of racing Thoroughbreds. In this study, the heritability of SDFT injury of the forelimbs was estimated in the Thoroughbred racehorses based on the Bayesian analysis with Gibbs sampling using threshold animal models. New horse patients of SDFT injury at the age of 2-5 years accommodated in the training centers in 2005 were used for the analyses. Based on clinical data of 8198 horses, heritability of SDFT was estimated to be the order of 0.17 (posterior mode) to 0.19 +/- 0.05 (posterior mean +/- posterior SD).

Effect of uphill exercise on equine superficial digital flexor tendon forces at trot and canter.

REASONS FOR PERFORMING STUDY: One cause of overstrain injury to the superficial digital flexor tendon (SDFT) in horses is the force loaded on the SDFT during repeated running. Therefore, decreasing this force may reduce SDFT injury. It has been reported that strain on the SDFT decreases with a toe-wedge shoe. Uphill courses are used for training of racehorses, and the angle of hoof-sole to the horizon during uphill running is similar to that of the toe-wedge shoe. OBJECTIVES: To determine the effects of uphill exercise on the force on the SDFT during trotting and cantering. METHODS: Arthroscopically implantable force probes (AIFP) were implanted into the SDFT of the left or right forelimb of 7 Thoroughbred horses and AIFP output recorded during trotting and cantering on a treadmill inclined at slopes of 0, 3 or 8%, and then 0% again. Superficial digital flexor tendon force was calculated as a relative value, with the amplitude of AIFP output voltage at initial 0% slope equal to 100. RESULTS: Out of 14 sets of experiments, AIFP data were analysed successfully in 9 at the trot, in 3 at the canter in the trailing forelimb on a slope of 3 and 8%, and in 2 at the canter in the leading forelimb on a slope of 3%. Increasing the incline from 0-8% tended to decrease peak force in the SDFT at the trot, and in the trailing forelimb at the canter. However, force in the SDFT was unchanged in the leading forelimb at the canter on the 3% incline. CONCLUSIONS: The force in the SDFT trotting or cantering uphill is unchanged or lower than that loaded at the same speed on a flat surface. Because at similar speeds the workload for uphill exercise is greater than on the flat, uphill running increases exercise intensity without increasing force in the SDFT. POTENTIAL RELEVANCE: Uphill exercise may reduce the risk of SDFT injury as both running speed and SDFT force are decreased on an incline as compared to the flat, even when exercise intensity is the same. Further study is needed to confirm these findings at canter in a larger population of horses.

Prevalence of superficial digital flexor tendonitis and suspensory desmitis in Japanese Thoroughbred flat racehorses in 1999.

REASONS FOR PERFORMING STUDY: Overstrain injuries to the superficial digital flexor tendon (SDFT) and suspensory ligament (SI) are among the most common musculoskeletal injuries which contribute to the considerable wastage of racing Thoroughbreds. Many epidemiological studies have demonstrated the prevalence of and risk factors for tendon injury when racing but have not included those injuries sustained during training. However, since tendon injury during training is seen commonly in clinical practice, it is appropriate to determine the overall prevalence of tendon injury sustained during both training and racing. OBJECTIVE: To determine the prevalence of overstrain injury to the SDFT and SL during training and racing among Thoroughbred flat racehorses in Japan in 1999. METHODS: A retrospective study was performed using a sample population of 10,262 Thoroughbred racehorses. The medical information database of Thoroughbred racehorses registered by the Japan Racing Association (JRA) in 1999 was analysed for SDFT and SL overstrain injury diagnosed by a veterinarian employed by JRA during training and racing. Jump racehorses were excluded from this study. RESULTS: The prevalence of forelimb SDFT tendonitis and SL desmitis was 11.1% (1130 cases) and 3.61% (370 cases) of the population, respectively. In the hindlimb, there were 0.06% (6 cases) and 0.14% (14 cases), respectively. Risks of SDF tendonitis in the forelimb in 3-year-olds or older horses were significantly higher than in 2-year-olds. In contrast, the risk of SL desmitis in the forelimb at age 3 and 4 years was 2.23 and 2.11 times higher, respectively, than in 2-year-olds, but this increased to 5.07 times in those age > or = 5 years. Entire males were at greater risk in comparison to females and geldings. CONCLUSIONS: The results suggest that the prevalence of SDF tendonitis and SL desmitis in the forelimb was associated with the horse's age and sex. The prevalence of SL desmitis increased further with age compared with SDF tendonitis, possibly reflecting a more rapid accumulation of degeneration in this structure. POTENTIAL RELEVANCE: The age-related risk demonstrated in this study provides further support that overstrain injuries are associated with accumulated degeneration. These data provide a valuable resource for further research into the aetiology of tendon injury in the racehorse.

Effect of controlled exercise on middle gluteal muscle fibre composition in Thoroughbred foals.

REASONS FOR PERFORMING STUDY: Most racehorses are trained regularly from about age 18 months; therefore, little information is available on the effect of training in Thoroughbred foals. HYPOTHESIS: Well-controlled exercise could improve muscle potential ability for endurance running. METHODS: Thoroughbred foals at age 2 months were separated into control and training (treadmill exercise) groups and samples obtained from the middle gluteal muscle at 2 and 12 months post partum. Muscle fibre compositions were determined by histochemical and electrophoretical techniques and succinic dehydrogenase (SDH) activity was analysed in each fibre type. RESULTS: All fibre types were hypertrophied with growth and type I and IIA fibres were significantly larger in the training than the control group at age 12 months. A significant increase of SDH activity was found in type IIX muscle fibres in the training group. CONCLUSIONS: Training in young Thoroughbred horses can facilitate muscle fibre hypertrophy and increase the oxidative capacity of type IIX fibres, which could potentially enhance stamina at high speeds. POTENTIAL RELEVANCE: To apply this result to practical training, further studies are needed to determine more effective and safe intensities of controlled exercise.

Exercise-induced tendon hypertrophy: cross-sectional area changes during growth are influenced by exercise.

Most skeletal tissues are thought to adapt to the mechanical environment they experience. While this has been demonstrated for muscle and bone, previous studies in the mature horse have failed to demonstrate adaptation in the superficial digital flexor tendon (SDFT), which suffers a high frequency of injury. This study tested the hypothesis that imposed exercise during growth would result in an increase in SDFT cross-sectional area (CSA). Fourteen Thoroughbred foals were divided into 2 sex-matched groups. A control group received 4 h pasture exercise and an exercise group had the same amount of pasture exercise with an additional short period of treadmill exercise daily from age 2-15 months. Activity at pasture was assessed objectively using a visual system. There was no significant difference in pasture activity between groups, although males were more active than females. The exercise programme resulted in a significantly larger tendon CSA in the exercise group at several, but not all, timepoints, which may be attributed to levels of variance. However, there was a significantly greater rate of increase in tendon CSA as a function of time in the exercised compared to the control group. This is the first evidence to suggest that tendon development can be modulated by exercise during growth in the horse, potentially increasing the ability of tendon to withstand the rigours of later athletic activity.

Horse lumbrical muscle: possible structural and functional reorganization in regressive muscle.

An anatomical study of horse lumbrical muscle (Lm) was carried out by light and electron microscopy in combination with immunochemical and cytochemical methods. Paraffin sections were subjected to haematoxylin and eosin (H & E) and Masson's trichrome staining for morphometric analysis. Paraffin sections were also used for immunostaining by anti-PGP 9.5 for reaction with nerve-protein associated-structures, anti-heat-shock protein 70 (hsp 70) for detection of gene expression changes, anti-fast myosin for the determination of muscle fibre types, and for detection of apoptotic gene expression of muscle fibres by the TUNEL method. The distribution of muscle fibre types on frozen sections was also examined by assaying ATPase (pH 4.2). We found that the proximal end of the tendon of the unipennate-shaped Lm binds to the deep digital flexor tendon, and the distal end of the Lm tendon connects to the medial surface of the palmar annular ligament. The Lm was not always present, but when found it varied in length greatly, up to 8 cm (muscle part alone), and weighed less than 1 g. The Lm was white, pale, or reddish in colour depending on the ratio of muscle fibre and connective tissue contents. The semi-tendinized regressive Lm was composed of rich vasculature, peripheral nerves, and nerve-like organs similar to the neuromuscular spindle (NMS). The extrafusal muscle fibres (e-lm) that surround the NMS were replaced with a thick outer capsule of connective tissues (CT) in the Lm nerve-like organ, which we named the neurotendinous capsule (NTC) organ. NTC organs exist alone or as multiple structures (up to eight) surrounded by a common outer capsule at the outermost CT ring. The NTC possesses several intrafusal muscle fibres (ifm) just as the NMS does. That the ifm was associated with nerve endings was confirmed by anti-PGP 9.5 and electron microscopic observation. Some muscle fibres in ifm and e-lm reacted with anti-fast twitch myosin and with anti-hsp 70. The e-lm exhibited at least two fibre types, determined by ATPase (pH 4.2) assay. The ifm exhibited mainly type I (slow twitch) fibres. No apoptotic gene expression was detected in either ifm or e-lm, suggesting the Lm is a vital organ. The degenerating fibres observed in ifm and e-lm indicate that the turnover rate of cytoplasmic components is accelerated. We attribute this phenomenon to the necessity for adaptation to new environmental demands. The surprising finding of tubular aggregates (TAs) in ifm of the NTC organ suggests that the Lm is continuously adapting. Some results related to variation in diameter of the collagen fibrils, isolation of the NTC organ and the myofibrillar protein constituents are also discussed. In conclusion, the so-called regressive Lm has rich vasculature, many peripheral nerves, and newly described NTC organs. The induction of heat-shock protein, lack of apoptotic gene expression in ifm and e-lm fibres, and TA formation in ifm suggest that horse Lm responds to environmental stress through reorganization and/or remodelling of cell constituents. We hypothesize that the horse Lm has lost its original role as a contractile element and changed to another function, likely as a vital nerve organ.

A semi-immobilization of a partial auricle induces hypertrophy and ultrastructural alteration of cardiomyocytes.

Semi-immobilization of a partial area of the ventral edge, lateral epicardium of the left auricle (ventrolateral of left auricle), by using quick adhesion glue induces moderate hypertrophy of myocytes with an average increase of 34% in cross-sectional area. Intercellular connective tissues increased, and cellular sizes varied markedly. The ultrastructure of immobilized (semi-immobilized) myocytes commonly exhibited degenerating features in myofibrils, various cytoplasmic organelles including mitochondrial cristae and sarcoplasmic reticulum (SR) were disrupted, and T-tubules disappeared. Z-line streaming and widening (hypertrophic Z-line, rod bodies) and increase of metabolic particle deposition are typical phenomena in addition to intercalated disc (Id) disorganization. The results suggest that semi-immobilization of the auricle induces hypertrophy of myocytes in association with degeneration and disruption of myofibrils and other cytoplasmic organelles, and an increase of intercellular connective tissues, rather than increase of myofibril mass. This is the first study to immobilize only a part of the heart rather than the whole animal. Our results using artificial immobilization of cardiac myocytes were extremely significant since the structural alterations obtained were similar to that observed in cardiomyopathies. This suggests that myocytes progressing to heart failure are also subjected to inhibition of movement. Therefore, this experiment may prove very useful as a model for studying the functional effect of heart failure observed in cardiomyopathy.

Tubular aggregates observed in spindle muscle fiber of horse lumbrical muscle.

Tubular aggregates (TAs) originate from the sarcoplasmic reticulum (SR) and form polymorphic double (or single) -walled structures in cross section. TAs are involved in various human skeletal muscle disorders including periodic paralysis, congenital myasthenic syndromes, inflammatory myopathies, and malignant hyperthermias. Horse lumbrical muscle (LM) is a slender fusiform muscle that shows varying degrees of regression due to its limited activity in the limb. Double-walled TAs were found in degenerating spindle fibers and with a range of 80-116 nm (average 92 nm, n=135) for outer layer and 50-78 nm (average 59 nm, n=135) for the inner layer. TAs exhibit degradation of myofibrillar proteins, disruption of mitochondria with cristae lost, glycogen accumulation, electron-dense metabolic products, blebbing appearance of sarcolemma, and presence of various vacuoles. LM fibers also show a similarly degenerative state. The disassembly of the SR structure probably produces a large accumulation of SR proteins which remain as molecules without being further degraded and which could aggregate to form the orderly structure of TAs. We believe that TA formation may be an adaptation to store unbalanced extra proteins by forming ordered aggregates in degeneration caused by stress in cells.

Effects of tri-, di- and monobutyltin on synaptic parameters of the cholinergic system in the cerebral cortex of mice.

Triorganotin compounds like tributyltin have been reported to be biodegraded to diorganotin, monoorganotin and then inorganic tin in animals after they have been ingested. Effects of tributyltin, dibutyltin and monobutyltin on various cholinergic parameters that are involved in synaptic transmission in the mouse cerebral cortex were investigated in vitro. Tributyltin and dibutyltin, but not monobutyltin, inhibited the activity of choline acetyltransferase, both the high-affinity and low-affinity uptakes of choline into synaptosomes, and the binding of [3H]quinuclidinyl benzilate to muscarinic acetylcholine receptors. Tributyltin and dibutyltin, but not monobutyltin, had a slightly suppressive effect on the K(+)-induced release and synthesis of acetylcholine in slices of the cortex. All three butyltins at concentrations from 10(-6) to 10(-4) M had no effect on the activity of acetylcholinesterase. The extent of the inhibitory effects on the cholinergic parameters, apart from the activity of acetylcholinesterase, was slightly greater in the case of tributyltin than dibutyltin, in particularly at the highest concentration (10(-4) M) tested. Therefore, it is concluded that tributyltin metabolites inhibit various parameters of cholinergic activity with a potency ranking of tributyltin > dibutyltin > monobutyltin.