Equine third metacarpal bone assessment by quantitative ultrasound and dual energy X-ray absorptiometry: an ex vivo study

The purpose of this ex vivo study was to analyse two commonly established methods of mechanical bone property assessment for application in horses: Quantitative ultrasound (QUS), which depends on the bone's density and Young's modulus, and dual energy X-ray absorptiometry (DXA), which depends on the areal bone mineral density (BMD). The third metacarpal bone (MC III) of horses was selected as examination region for practical reasons. An interrelationship between QUS- and DXA-values was examined. Both MC III of eleven randomly selected equine cadavers were divided in nine regions of interest (ROI). A multi-site QUS device was used for axial transmission speed of sound (SOS) measurements and a DXA device was used for BMD evaluation. Full cortical thickness BMD (FcBMD), overall aspect BMD and 4 mm cortical border slice BMD (4 mmBMD) were evaluated. In addition, each ROI of one MC III was measured 10 times to determine QUS- and DXA-measurement precision. SOS values and BMD values obtained at different aspects of MC III were different (P < 0.001). FcBMD and overall BMD obtained at different levels were different (P < 0.001). SOS data correlated with FcBMD-, overall BMD- and 4 mmBMD-data at various ROI. FcBMD-, overall BMD- and 4 mmBMD-data were strongly correlated. The intra-operator coefficient of variation was 1.3% for SOS-measurements and ranged between 1.94 and 10.3% for BMD-measurements. Multi-site axial transmission QUS as well as DXA can be used to precisely measure bone characteristics of MC III in horses. However, both techniques do not measure the same bone properties. It is therefore concluded, that QUS and DXA techniques are complementary for application in horses.     

Use of dual energy x-ray absorptiometry for noninvasive body composition measurements in clinically normal dogs

OBJECTIVE: To determine quantitative values for components of body composition in clinically normal dogs of various breeds by use of dual energy x-ray absorptiometry (DEXA) and validate the precision and accuracy of DEXA technology in dogs. ANIMALS: 103 clinically normal sexually intact adult dogs. PROCEDURE: In a cross-sectional study, Beagles, Pembroke Welsh Corgis, Golden Retrievers, Great Danes, Pointers, Rottweilers, and nonpurebred dogs received total body DEXA scans. For the validation portion of the study, the results of DEXA scans of 6 dogs were compared with values obtained by chemical analyses of tissues from euthanatized dogs to determine the accuracy of this modality in dogs. RESULTS: Results (coefficient of variation) of the precision tests ranged from 0.10% for lean tissue to 5.19% for fat tissue, whereas accuracy tests revealed a difference between percentage bone mineral content and ash values. Body composition differed by sex, such as higher lean tissue and bone mineral content in males within some breeds, and among breeds. Regardless of body size or weight, the percentage of body weight that was bone mineral ranged from 3 to 4.0% [corrected]. CONCLUSIONS AND CLINICAL RELEVANCE: Results of this cross-sectional study provide valuable body composition data for clinically normal adult dogs, which may have research and clinical applications.     

Long-term effects of intermittent equine parathyroid hormone fragment (ePTH-1-37) administration on bone metabolism in healthy horses

Intermittent administration of parathyroid hormone (PTH) is an anabolic therapy for osteoporotic conditions in humans. This study evaluated the effects of equine PTH fragment (ePTH-1-37) administration on bone metabolism in 12 healthy horses. Six horses each were treated once daily for 120days with subcutaneous injections of 0.5μg/kg ePTH-1-37 or placebo. Blood was collected to determine ionized calcium (Ca(++)), total Ca (Ca(T)), inorganic phosphorus, serum equine osteocalcin (eOC), carboxy-terminal telopeptide of type I collagen (ICTP), bone-specific alkaline phosphatase, and carboxy-terminal cross-linked telopeptide of type I collagen. Bone mineral density (BMD) was determined with dual X-ray absorptiometry of the metacarpus and calcaneus. Significantly higher blood Ca(++) and plasma Ca(T) concentrations were measured 5h after ePTH-1-37 administration compared to placebo. Higher serum eOC concentrations were found for ePTH-1-37 treatment at days 90 (P<0.05) and 120 (P=0.05). Significantly higher serum ICTP levels were observed with ePTH-1-37 treatment at days 60 and 90. For both study groups, BMD increased significantly in the calcaneus. Long-term use of ePTH-1-37 seemed to have no negative effects on bone metabolism in healthy horses. The absence of undesirable side effects is the premise to ensure safety for further clinical investigations in horses with increased bone resorption processes.     

The quantitative assessment of photodensity of the third carpal bone in the horse

AIM: To determine whether variation in x-ray-beam angle significantly affected photodensity of the third carpal bone (C3) in the horse using the tangential radiographic view, and indirectly determine whether radioabsorptiometry (RA) could be used to assess differences in bone mineral density (BMD) of C3 between exercised and non-exercised horses. METHODS: The left distal carpal row was isolated post-mortem from 2-year-old Thoroughbred horses that had been either exercised (n=7) for up to 14 weeks using a standard training regimen for racehorses, or not exercised (n=7). The effect of variation in x-ray-beam angle on photodensity was determined on these isolated carpal bones in vitro. Image analysis was used to assess photodensity (compared to a known thickness of aluminium) of four regions of interest (ROI) in C3 and one ROI in the fourth carpal bone (C4) of 14 isolated distal rows of carpal bones of the horse. The isolated carpal bones were placed fl at on a x-ray cassette and radiographed at 90 degrees (i.e. with the x-ray beam perpendicular to the cassette). The x-ray-beam angle was varied in the dorsal sagittal plane by 5 degrees increments to a total of 15 degrees from 90 degrees and from a base angle of 60 degrees (the x-ray-beam angle at which the tangential view is taken in clinical cases). RESULTS: Variation in beam angle of <10 degrees from 90 degrees significantly affected photodensity, and photodensity was significantly affected when the angle was varied <5 degrees from 60 degrees . When taken at an x-ray-beam angle of 60 degrees , the abaxial aspect of the radial facet of C3 had a consistently higher photodensity than the rest of C3 and C4. The photodensity of the third and fourth carpal bones was higher in exercised horses than in non-exercised horses. CONCLUSION: As variation in x-ray-beam angle significantly affected photodensity, RA using the tangential view is not considered clinically applicable for assessing BMD of C3 and the accuracy of subjective assessment of BMD of C3 using the tangential view in horses is questionable.     

Use of multisite quantitative ultrasonography for noninvasive assessment of bone in horses

OBJECTIVE: To evaluate the usefulness of multisite quantitative ultrasonography for noninvasive assessment of bone in horses. SAMPLE POPULATION: 12 healthy horses and both forelimbs from 8 clinically normal horses. PROCEDURE: For in vivo measurements, various regions of interest (ROI) were examined on the third metacarpal bone, radius, and tibia. Precision error for speed of sound (SOS) measurements was obtained by measuring each ROI of 4 horses 10 times with probe repositioning. Additionally, 3 operators measured each aspect of the third metacarpal bone of 6 horses 5 times each. For ex vivo measurements, third metacarpal bones were examined at 9 ROI, and SOS measurements were performed before and after soft tissue removal. One ROI of a single forelimb was subjected to 96 ex vivo measurements with 3 different contact media. RESULTS: The lateral aspect of the third metacarpal bone had significantly higher SOS values than the dorsal and medial aspect of the third metacarpal bone. No difference was obtained between SOS values of the lateral and medial aspect of the radius. The tibia had significantly higher SOS values than the lateral aspect of the radius and the dorsal and medial aspect of the third metacarpal bone. Intraoperator coefficients of variation ranged from 0.62 to 3.15%, and interoperator coefficients of variation ranged from 0.78 to 2.70%. Values of SOS were highest when silicone oil was used as the contact medium. CONCLUSIONS AND CLINICAL RELEVANCE: Speed of sound measurements obtained by quantitative ultrasonography in axial transmission mode can be used to precisely measure superficial cortical bone properties of third metacarpal bone, radius, and tibia in horses.     

Effects of immobilization followed by remobilization on mineral density, histomorphometric features, and formation of the bones of the metacarpophalangeal joint in horses

OBJECTIVE: To determine microradiographic appearance, bone histomorphometry, and mineral density of the long bones of the metacarpophalangeal joint in horses after immobilization followed by remobilization. ANIMALS: 5 healthy horses. PROCEDURE: One forelimb of each horse was immobilized in a fiberglass cast for 7 weeks, followed by 8 weeks of increasing exercise. Calcein and oxytetracycline were administered IV during the immobilization and exercise phases, respectively, for bone labeling and analysis after euthanasia. Sagittal sections of metacarpal bones and proximal phalanges were examined via radiography, dual energy x-ray absorptiometry, histomorphometry, and bone label analysis. RESULTS: Radiography revealed loss of bone mineral opacity in the subarticular regions of the immobilized metacarpal bones and phalanges and subchondral lesions in metacarpal bones in 2 horses. In phalanges, a significant decrease in subarticular volumetric bone mineral density was detected. There was significantly less bone volume and calcein-labeled bone surface and more vascular volume and oxytetracycline-labeled bone surface in immobilized phalanges, compared with contralateral phalanges. CONCLUSIONS AND CLINICAL RELEVANCE: Eight weeks of exercise after single-limb immobilization is insufficient for recovery of volumetric bone mineral density. During immobilization and remobilization, the subchondral and trabecular bone appear to be actively remodeling.     

ACCURACY AND PRECISION OF DUAL-ENERGY X-RAY ABSORPTIOMETRY FOR EX VIVO DETERMINATION OF MINERAL CONTENT IN TURKEY POULT BONES

The right humerus was removed from 30 20-week-old male turkey poults for humerus strength analysis using dual-energy x-ray absorptiometry and humerus-breaking strength. Specimens were cleaned and dried before scanning. To determine the most precise and accurate protocol of bone densitometry analysis for avian long bones, we scanned each specimen using five different techniques, all aimed to simulate soft-tissue thickness. Correlation coefficients and linear regression equations between 1) bone mineral content and humerus ash, and 2) bone mineral density and humerus-breaking strength were estimated with each technique and compared. The coefficient of variation values for precision ranged from 0.40% to 1.69% for bone mineral content and from 0% to 4.19% for bone mineral density. The accuracy was determined by comparing the bone mineral content of each humerus with the corresponding ash weight; the correlation coefficients between the two parameters were highly significant (range 0.949-0.963; P < or = 0.01). Significant correlations were also observed between humerus-breaking strength and bone density measurements (range 0.762-0.785; P < or = 0.01). Linear regression coefficients relating both parameters considered were also highly significant. We concluded that dual-energy x-ray absorptiometry is an accurate and precise method with which to determine ex vivo bone mineral content and strength in turkey bones. Further investigations are requested for field applications of this method to study factors affecting bone physiology and strength.     

Evaluation of cutaneous analgesia after non-focused extracorporeal shock wave application over the 3rd metacarpal bone in horses

Extracorporeal shock wave therapy is becoming increasingly used in equine practice to treat musculoskeletal disorders. Although the exact effects of extracorporeal shock waves on tissues still need to be elucidated, transient cutaneous analgesia has been observed within treated areas in humans and horses. The purpose of this study was to determine the onset, magnitude, and duration of cutaneous analgesia after non-focused extracorporeal shock wave (ESW) application by comparing the limb withdrawal reflex latency (LWRL) in treated and untreated areas on the metacarpus of horses, using a focused light source. A single treatment with non-focused ESW was applied to 3 areas over the dorsal aspect of one metacarpus in 12 horses. The LWRL was measured to assess cutaneous sensation in treated and untreated control areas of the treated metacarpus and in an untreated area on the opposite metacarpus, before and at various time points after ESW application. Most treated and control areas revealed a significant decrease in LWRL over time compared with baseline values. Although the results of our study do not suggest cutaneous analgesia after ESW application to the equine metacarpus, we advise cautious use of this treatment modality for horses in training or before competition until further research is done.     

Comparative assessment of bone mineral measurements obtained by use of dual-energy x-ray absorptiometry, peripheral quantitative computed tomography, and chemical-physical analyses in femurs of juvenile and adult dogs

OBJECTIVE: To compare bone mineral measurements obtained by use of dual-energy x-ray absorptiometry (DEXA), peripheral quantitative computed tomography (pQCT), and chemical-physical analyses and determine effects of age and femur size on values obtained for the various techniques. SAMPLE POPULATION: Femurs obtained from 15 juvenile and 15 adult large-breed dogs. PROCEDURE: n each femur, 7 regions of interest were examined by use of DEXA to measure the bone mineral content (BMC) and bone mineral density (BMD), and 5 were examined by use of pQCT to measure BMD. Among these, 1 region was examined by both noninvasive methods and an invasive method. Volume of the femur was determined by water displacement. Volumetric bone density (VBD) was calculated. Calcium (Ca), phosphorus (P), total Ca, and total P contents were determined. RESULTS: DEXA- and pQCT-derived results revealed that all values increased with age in juvenile dogs. In adults, VBD and pQCT-derived BMD decreased significantly and DEXA-derived BMD increased with increasing femur length.The pQCT-derived BMD correlated well with VBD and Ca content, whereas DEXA-derived BMC was strongly correlated with Ca content. In juveniles, values correlated regardless of the technique used, whereas in adult dogs, DEXA-derived BMD did not correlate with pQCT-derived BMD, Ca concentration, or VBD unless data were adjusted on the basis of femur length. CONCLUSIONS AND CLINICAL RELEVANCE: DEXA-derived BMD adjusted for femur length yields approximately the same percentage variability in VBD as for pQCT-derived BMD. However, pQCT-derived BMD is still more sensitive for determining variability BMD in Ca concentration, compared with DEXA-derived BMD adjusted for femur length.     

Effect of focused and radial extracorporeal shock wave therapy on equine bone microdamage

OBJECTIVES : To determine whether bone microcracks are altered after application of focused and radial extracorporeal shock wave therapy (ESWT) to the equine distal limb. STUDY DESIGN : An ex vivo experimental model. SAMPLE POPULATION : A contralateral limb specimen was obtained from 11 Thoroughbred racehorses with a unilateral catastrophic injury. Distal limb specimens were also obtained from 5 non-racing horses. METHODS : Three separate skin-covered bone segments were obtained from the mid-diaphysis of the metacarpus (MC3) or metatarsus (MT3). Focused (9,000 shockwaves, 0.15 mJ/mm2, 4 Hz) and radial (9,000 shockwaves, 0.175 mJ/mm2, 4 Hz) ESWT treatments were randomized to the proximal and distal segments and the middle segment was used as a treatment control for pre-existing microcracks. After treatment, bone specimens were bulk-stained with basic fuchsin and microcracks were quantified in transverse calcified bone sections. RESULTS : ESWT had small but significant effects on microcracks. Microcrack density (Cr.Dn) and microcrack surface density (Cr.S.Dn) were increased after focused ESWT, whereas Cr.Le was increased after radial ESWT. In racing Thoroughbreds, Cr.Le increased with increased number of races undertaken. Cr.Dn and Cr.S.Dn were not significantly influenced by the number of races undertaken. CONCLUSION : ESWT has small but significant effects on bone microcracking ex vivo. CLINICAL RELEVANCE : These preliminary data suggest that ESWT has the potential to increase bone microcracking in equine distal limb bone in vivo. Such effects may be more pronounced in Thoroughbreds that are actively being raced, because in vivo microcracking increases with increased number of races undertaken.     

Autologous osteochondral grafting (mosaic arthroplasty) for treatment of subchondral cystic lesions in the equine stifle and fetlock joints

OBJECTIVE: To describe treatment of equine subchondral bone cysts (SBCs) by reconstruction of the articular surface with osteochondral grafts. STUDY DESIGN: Case series of horses with SBCs unresponsive to conservative therapy. ANIMALS: Eleven horses (1-12 years). METHODS: SBCs were identified in 4 locations: medial femoral condyle (5 horses), lateral femoral condyle (1), distal epiphysis of the metacarpus (4), or metatarsus (1). Osteochondral autograft transplantation (mosaic arthroplasty) was performed, taking grafts from the abaxial border of the medial femoral trochlea of the unaffected limb. Graft implantation was achieved through a small arthrotomy or by arthroscopy depending on SBC location. RESULTS: All horses improved postoperatively; 10 horses had successful outcomes with radiographic evidence of successful graft incorporation and 7 returned to a previous or higher activity level. On follow-up arthroscopy (5 horses) there was successful reconstitution of a functional gliding surface. One horse had delayed incorporation of a graft because of a technical error but became sound. One horse had recurrence after 4 years of work and soundness. One stallion was used for breeding and light riding because of medial meniscal injuries on the same limb. CONCLUSIONS: Implantation of osteochondral grafts should be considered for SBC when conservative management has not improved lameness and there is a risk of further joint injury and degeneration. CLINICAL RELEVANCE: Mosaic arthroplasty should be considered for treatment of subchondral bone cysts of the femoral condyle and distal articular surface of the metacarpus/tarsus in horses that are refractory to non-surgical management.     

Effects of extracorporeal shock wave therapy on bone

Extracorporeal shock waves have been used for 30 years to fragment uroliths for nonsurgical treatment for urolithiasis in humans. Applied to bone, shock waves delivered at the appropriate energy and pulse number, can stimulate osteogenesis. In Europe, shock waves are routinely used to treat nonunions in humans despite poor understanding of the mechanism of action. Shock wave therapy has also been used clinically in horses. Preliminary experimental studies indicate that shock wave therapy does not damage soft tissue in the distal aspect of the equine limb and can stimulate osteogenesis throughout the depth of the near cortex of the metacarpus and metatarsus.     

Pathophysiology and diagnosis of third carpal bone disease in horses: a review

Third carpal bone (C3) disease is a significant cause of lameness in Standardbred and Thoroughbred horses. The bone density of C3 increases as a result of exercise, reducing the compliance of the bone and predisposing it to injury. Currently, the most widely used method of diagnosis is subjective radiography using the tangential view. Radiographically, increases in bone mineral density (BMD) appear as sclerosis but it is not known at what point increases in sclerosis indicate the onset of disease or increased risk of C3 fracture. A quantitative assessment of the BMD of C3 in horses would improve understanding of the changes that occur within this bone and guide athletic management, as it is thought that BMD changes precede articular cartilage damage. Methods of non-invasive bone-mineral analysis used for the detection of osteoporosis in humans include single photon absorptiometry (SPA), dual x-ray absorptiometry (DXA), computed tomography (CT), radioabsorptiometry (RA), quantitative ultrasonography (QU) and magnetic resonance imaging (MRI). To date, DXA and RA are the most commonly used methods of quantitative non-invasive bone-mineral analysis in horses. The cost of equipment and difficulties in performing DXA in live animals preclude the routine use of this technique for diagnostic purposes. RA may become clinically applicable to C3 analysis in horses, but small variations in x-ray beam angle when taking the tangential view significantly affect results, making this technique clinically inapplicable at this time. Currently, methods of quantitative non-invasive bone-mineral analysis of C3 in horses are not suited to clinical application.     

Characterization of age- and location-associated variations in the composition of articular cartilage from the equine metacarpophalangeal joint

Objective: To characterize the impact of age, gender, location and individual animal variation on the composition of articular cartilage from the metacarpophalangeal joint of horses. Design: Cartilage specimens were obtained from the metacarpophalangeal joints of 28 male, female and castrated male horses ranging in age from one day to 27 years of age. Cartilage samples from the distal metacarpus, proximal first phalanx and proximal sesamoids were analyzed separately. Chondrocyte number, DNA content, proteoglycan concentration and total collagen content were determined for each animal and joint location. Results: Age and joint location had a significant effect on chondrocyte number and DNA content with higher cell counts and DNA content detected in cartilage from the youngest age groups and in cartilage from the metacarpus and proximal sesamoids. The influence of age on chondrocyte numbers was not significant in horses over two years of age. Both age and joint location also influenced total proteoglycan and collagen content. Lower proteoglycan and collagen concentrations were detected in younger horses, and cartilage from the metacarpus had lower proteoglycan and collagen concentrations than that from other joint locations. Gender did not appear to influence chondrocyte number or matrix content of equine articular cartilage. However, there was significant residual variation in cellularity, proteoglycan levels and collagen content between individual animals that could not be explained by the signalment factors considered in this study. Conclusions: Future studies examining equine articular cartilage should avoid direct morphologic comparisons between animals of different ages, and any comparisons made between individuals should be interpreted cautiously. In addition, in vitro tissue culture models should avoid the use of cartilage pooled from different animals or from different locations within the same joint.     

Monitoring response to aminobisphosphonate therapy in canine osteosarcoma using dual energy x‐ray absorptiometry (dexa)

Introduction: Palliative therapy is essential to improve the quality of life of dogs with osteosarcoma (OSA), when definitive therapy is not considered a valid option. Bisphosphonates, a novel class of antiosteoclastic drugs, are widely used in humans for several painful osteolytic conditions. Dual energy x‐ray absorptiometry (DEXA) is recognized as a reliable tool to measure bone mineral density (BMD), and to monitor treatment response to bisphosphonates in humans. A prospective evaluation of pamidronate, an injectable aminobisphosphonate, is ongoing in dogs with appendicular OSA. The potential value of DEXA for objective evaluation of BMD variations with palliative therapies is concurrently being assessed. Materials and Methods: Dogs with naturally occurring appendicular OSA treated with pamidronate constitute the patient population. A DEXA scan (QDR‐4500 W, Hologic, Bedford, MA) is performed on day 0 (baseline) and on every treatment day with pamidronate thereafter (every 28 days). For each dog, a whole body scan is performed, followed by a scan of the tumor, and contralateral normal bone. Three regions of interest are subsequently analyzed for BMD changes in tumor and normal bone. Statistical analysis was performed using Student t‐test and paired t‐tests, with significance being set at p < 0.05. Results: Nineteen dogs have been enrolled to date. Seven responders and 6 non‐responders have suitable data for analysis. A significant difference is observed (p = 0.04) between tumor BMD variations of responders and non‐responders at day 28 (mean variations +18.0% and ?4.6% respectively). The changes at day 28 are significant only in the responders (p = 0.038 vs p = 0.05 in non‐responders). When BMD of tumor and normal bone at day 84 is compared to day 0 in six responders, only tumor had a significant increase (p = 0.017 vs p = 0.279, respectively). Conclusions: Objective measurements of response to therapy are essential in pain palliation studies. Increased tumor bone BMD, as obtained by DEXA analysis, may correlate with subjective clinical improvement in pamidronate‐treated dogs with appendicular OSA.     

Core decompression of the equine navicular bone: an in vitro biomechanical study

Objective: To determine the effect of core decompression surgery and bone mineral density (BMD) on the mechanical properties of equine navicular bones. Study Design: Experimental, in vitro study. Sample Population: Fore limb navicular bones (n=36 pairs) from sound 2–5‐year‐old horses with no radiographic abnormalities of the distal aspect of the forelimbs. Methods: Navicular BMD was measured using dual energy X‐ray absorptiometry. One randomly assigned navicular bone from each pair served as control. The contralateral test specimen was allocated to 1 of 6 treatment groups defined by drill bit size (3.2 versus 2.5 mm diameter) and by the number of drill channels (1, 2, or 3) created in the proximal border of the bone. Bones were then tested until failure in 3‐point bending. Data were statistically analyzed using ANOVA and regression analysis. Results: There were significant (P<.001) positive correlations between BMD and biomechanical data. A significant (P<.001) reduction in breaking strength was noted between intact and drilled bone pairs; however, the diameter and number of decompression channels did not significantly (P>.05) influence the extent of the reduction in mechanical strength. Conclusion: In vitro core decompression significantly decreases the breaking strength of the equine navicular bone.     

Radiographic assessment of carpal conformation in the horse: Technique development and validation of the consistency of measurements

Carpal conformation is often considered as a contributory factor to performance and lameness in the horse; however, few attempts have been made to objectively measure radiographic variations of carpal conformation in horses due to insufficient measurable carpal parameters. This pilot study used carpal radiographic images acquired from 10 cadaveric equine forelimbs transected at the antebrachial midshaft from 7 adult horses (7.2 ± 2.6 years), positioned at ‘zero lateromedial’ (ZLM) and ‘zero dorsopalmar’ (ZDP) views, to investigate the anatomy of the equine carpus and develop parameters that could be objectively used to assess carpal conformation in horses. Dorsal carpal angle (DCA: 176.61 ± 0.66º), distal radial slope carpal angle (DRSCA: 145.59 ± 2.19º), intermediate carpal bone proximal tuberosity-radial angle (CiPxTRA: 115.69 ± 3.15º) and third carpal bone palmar facet angle (C3PalFCA: 84.43 ± 1.13º) were all developed from the ZLM view while medial carpal angle (MCA: 183.34 ± 1.02º), disto-dorsal slope angle of the third carpal bone (C3DDSA: 8.27 ± 0.92º) and width ratio of distal radius to proximal metacarpus (WDR:WPM = 1.13±0.03) were 3 of the 10 parameters developed from the ZDP view. Easy to identify and measurable parameters will help to provide quantitative assessment of carpal conformation in the horse with potential of eliminating subjective observational variation errors between clinicians. These newly developed parameters will be useful in further studies to measure variations in the conformation of the equine carpus in live horses and comparison between subjective visual assessment and objective radiographic evaluation methods.     

Histomorphologic evaluation of extracorporeal shock wave therapy of the fourth metatarsal bone and the origin of the suspensory ligament in horses without lameness

OBJECTIVE: To determine via histologic examination and scintigraphy the effect of focused extracorporeal shock wave therapy (ESWT) on normal bone and the bone-ligament interface in horses. ANIMALS: 6 horses without lameness. PROCEDURE: Origins of the suspensory ligament at the metacarpus (35-mm probe depth) and fourth metatarsal bone (5-mm probe depth) were treated twice (days 0 and 16) with 2,000 shocks (energy flux density, 0.15 mJ/mm2). One forelimb and 1 hind limb were randomly treated, and the contralateral limbs served as nontreated controls. Bone scans were performed on days -1 (before ESWT), 3, 16, and 19. Histomorphologic studies of control and treated tissues were performed on day 30. RESULTS: ESWT significantly increased the number of osteoblasts but caused no damage to associated soft tissue structures and did not induce cortical microfractures. A significant correlation between osteoblast numbers and radiopharmaceutical uptake was noticed on lateral views of the hind limb on days 3 and 16 and on caudal views of the forelimb on day 3. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that ESWT has the potential to increase osteoblast numbers in horses. The correlation between increased osteoblast numbers and radio-pharmaceutical uptake 3 days and 16 days after the first ESWT suggested that stimulation of osteogenesis occurred soon after ESWT. No damage to bone or the bone-ligament interface should occur at the settings used in this study, and ESWT can therefore be administered safely in horses.     

Evaluation of subchondral bone mineral density associated with articular cartilage structure and integrity in healthy equine joints with different functional demands

OBJECTIVE: To determine and correlate subchondral bone mineral density and overlying cartilage structure and tensile integrity in mature healthy equine stifle (low magnitude loading) and metacarpophalangeal (high magnitude loading) joints. ANIMALS: 8 healthy horses, 2 to 3 years of age. PROCEDURE: Osteochondral samples were acquired from the medial femoral condyle (FC) and medial trochlear ridge (TR) of the stifle joint and from the dorsal (MC3D) and palmar (MC3P) aspects of the distal medial third metacarpal condyles of the metacarpophalangeal joint. Articular cartilage surface fibrillation (evaluated via India ink staining) and tensile biomechanical properties were determined. The volumetric bone mineral density (vBMD) of the underlying subchondral plate was assessed via dual-energy x-ray absorptiometry. RESULTS: Cartilage staining (fibrillation), tensile moduli, tensile strength, and vBMD were greater in the MC3D and MC3P locations, compared with the FC and TR locations, whereas tensile strain at failure was less in MC3D and MC3P locations than FC and TR locations. Cartilage tensile moduli correlated positively with vBMD, whereas cartilage staining and tensile strain at failure correlated negatively with vBMD. CONCLUSIONS AND CLINICAL RELEVANCE: In areas of high joint loading, the subchondral bone had high vBMD and the articular cartilage surface layer had high tensile stiffness but signs of structural wear (fibrillation and low failure strain). The site-dependent variations and relationships in this study support the concept that articular cartilage and subchondral bone normally adapt to physiologic loading in a coordinated way.     

Isolation and characterization of bone marrow-derived equine mesenchymal stem cells

OBJECTIVE: To isolate and characterize bone marrow-derived equine mesenchymal stem cells (MSCs) for possible future therapeutic applications in horses. SAMPLE POPULATION: Equine MSCs were isolated from bone marrow aspirates obtained from the sternum of 30 donor horses. PROCEDURES: Cells were cultured in medium (alpha-minimum essential medium) with a fetal calf serum content of 20%. Equine MSC features were analyzed to determine selfrenewing and differentiation capacity. For potential therapeutic applications, the migratory potential of equine MSCs was determined. An adenoviral vector was used to determine the transduction rate of equine MSCs. RESULTS: Equine MSCs can be culture-expanded. Equine MSCs undergo cryopreservation in liquid nitrogen without altering morphologic characteristics. Furthermore, equine MSCs maintain their ability to proliferate and differentiate after thawing. Immunocytochemically, the expression of the stem cell marker CD90 can be detected on equine MSCs. The multilineage differentiation potential of equine MSCs was revealed by their ability to undergo adipogenic, osteogenic, and chondrogenic differentiation. CONCLUSIONS AND CLINICAL RELEVANCE: Our data indicate that bone marrow-derived stromal cells of horses can be characterized as MSCs. Equine MSCs have a high transduction rate and migratory potential and adapt to scaffold material in culture. As an autologous cell population, equine MSCs can be regarded as a promising cell population for tissue engineering in lesions of the musculoskeletal system in horses.