Motion of 6 clinically sound horses trotting at a speed of 4 m/s on a treadmill was captured by video cameras before and 9, 16, and 23 days after amphotericin-induced lameness to determine the quantitative variables of three-dimensional computer-assisted image analysis that objectively describe carpal lameness. Amphotericin-B was used to induce lameness, and phenylbutazone (2.2 mg/kg of body weight, PO, once) and butorphanol tartrate (0.1 mg/kg IM, q 6 h, to effect) were used to control discomfort. Four 60-Hz cameras were symmetrically placed around the treadmill to capture 6 seconds of images from retroreflective spheres taped to the trotting horses. Images were transferred to a video-based digitizer and a computer work station, where 4 files of two-dimensional data were reduced to 1 file of three-dimensional data. The effect of lameness on motion analyzed was assessed by use of two-way ANOVA. Differences between means were assessed, using the Student-Newman-Keul's test (P less than or equal to 0.05). Head and withers excursions, (dorsal vertical displacement of head and withers targets, respectively) during the sound forelimb support phase were increased significantly during all lameness measurement periods. Head excursion, but not withers excursion, during the lame forelimb support phase, was decreased significantly during all lameness measurement periods. Computer determinations of stride length swing phase, stance phase, forelimb abduction, and carpal and fetlock ranges of motion did not consistently characterize the lameness. It was concluded that three-dimensional computer-assisted image analysis could be used for objective lameness evaluation in horses and that head and withers excursions were the most consistent variables for assessing equine carpal lameness.

Tissue variation in microscope slides made for spermatozoon analysis and variation introduced by the subjective techniques used to analyze these slides reduce the statistical power of studies that seek to use spermatozoon morphology to predict fertility. A simple specimen preparation method was developed to standardize stallion spermatozoon morphologic smears, and a new, automated spermatozoa morphometry instrument was used to objectively analyze the efficacy of the specimen preparation technique. The method achieved a standard spermatozoon concentration and reduced field-to-field variation in the number of spermatozoa analyzed. Metric measurements of spermatozoon head dimensions from clinically normal, fertile stallions revealed small, but highly significant, differences between stallions. The variation in metric measurements between replicate slides within stallions was small, indicating that replicate slide analysis probably is not necessary for clinically normal stallions. Coefficients of variation were generally less than 11% for metric measurements between stallions, and were less than 4% within stallions. This study revealed that a high degree of statistical power can be achieved when using these new, standardized specimen preparation and objective analysis techniques. Such power makes possible the detection of subtle differences between clinically normal stallions, and may facilitate accurate detection of abnormal fertility (ie, subfertility) in stallions.

Ultrasonographic cross-sectional area (CSA) measurements of equine superficial digital flexor (SDF) tendon were obtained to determine the feasibility of ultrasonography for CSA measurement of tendon in vivo and in vitro. Ultrasonographic measurements were compared with a more traditional CSA measurement method, ink-blot analysis. In addition, values for ultrasonographic SDF tendon mean echogenicity were obtained in vivo and in vitro. The left forelimb SDF tendons of 23 horses were evaluated ultrasonographically. Cross-sectional images were acquired at 4-cm intervals distal to the base of the accessory carpal bone (DACB) to the level of the proximal sesamoid bones while horses were standing squarely. After euthanasia, the left forelimbs were mounted in a materials testing system (MTS) and loaded under tension to standing load. Ultrasonographic images were again acquired at the same locations. The ultrasonographic images were digitized, and values for ultrasonographic CSA and mean echogenicity were obtained for each level. Immediately after mechanical testing, a 1-cm-thick transverse section of SDF tendon at 12 cm DACB was removed. Three ink blots were prepared from each end of the removed tendon section and digitized. The 6 CSA values were averaged to generate a value for morphologic CSA for each SDF tendon at 12 cm DACB. Standing ultrasonographic tendon CSA at 12 cm DACB was consistently smallest (mean +/- SD CSA = 86 +/- 11 mm2), followed by MTS ultrasonographic CSA (mean, 95 +/- 12 mm2), with ink-blot morphologic CSA being largest (mean, 99 +/- 15 mm2). Comparison of standing and MTS ultrasonographic values at 12 cm DACB revealed a strong positive linear correlation between methods (R2 = 0.74, P = 0.001). Comparison of ink-blot CSA at 12 cm DACB with standing and MTS ultrasonographic CSA revealed strong positive linear correlations (R2 = 0.64, P = 0.001 and R2 = 0.72, P = 0.001, respectively). For ultrasonographic mean echogenicity, standing values insignificantly exceeded MTS values at each level. The authors conclude that ultrasonography is a useful technique for the noninvasive assessment of SDF tendon CSA that can be applied in vivo and in vitro.

Two techniques for evaluating argyrophilic nucleolar organizer regions (AgNOR) were compared on 74 canine mammary tumors to discriminate between benign and malignant lesions. For each lesion, direct counting of AgNOR on at least 100 cell nuclei was compared with area, perimeter, and integrated optical density AgNOR dot values determined by image analysis. Significant differences between benign and malignant tumors were observed with both methods; however, lesions determined as aggressive or proliferative by histologic evaluation were only singled out by image analysis measurements. Image analysis, in our hands, was a reliable, precise, and convenient technique to characterize malignancy in canine mammary tumors.

The relationship of muscle and bone structure to limb weakness was examined in 60 Duroc pigs from 3 lines divergently selected for thoracic limb weakness. The lines were designated high, control, or low, with the low line having inferior thoracic limb structure. At approximately 100 kg, 10 pigs of each line and gender were scored for thoracic limb structure and movement. Right and left thoracic limbs were collected at slaughter. A computerized morphometric image analysis system was used to determine cross-sectional areas of muscles, bones, and soft tissues at levels through the brachium, antebrachium, metacarpus, and digits. The statistical model that was used to analyze the data included the effects of line, sire, gender, and side (left vs right), with weight as a covariate. Total bone area was similar for all 3 lines of pigs at all cross-sectional levels, but significant differences in muscle and other soft tissue areas were observed, including significantly greater extensor area for the antebrachium (P less than 0.001) in low-line pigs than in control- and high-line pigs, smaller total area (P less than 0.05) of the metacarpus in low-line pigs than in control and high-line pigs, and less total area of the medial digit (P less than 0.01) in low-line pigs than in control- or high-line pigs. Total area of bone and soft tissue for each cross-sectional region was significantly greater (P less than 0.05) in boars than in gilts. Side differences also were observed in total cross-sectional areas of bone and soft tissue of the antebrachium, metacarpus, and digits. Pigs selected for inferior thoracic limb structure had less total soft tissue cross-sectional areas in distal limb regions than did control- or high-line pigs. Limb weakness may be related to altered distribution of soft tissue supporting structures of the thoracic limb.

Eight untrained 2-year-old Thoroughbred horses were used in a study of the remodeling response of the proximal sesamoid bone (PSB) to training-related stimuli. Two horses each were assigned to 1 of 4 groups: group 1, untrained, pasture turnout (control); group 2, modified-classically trained, dirt track; group 3, classically trained, dirt track; and group 4, classically trained, wood chip track. Horses were given fluorochromic bone labels every 28 days during training. All horses were euthanatized after 5 months of training, and the proximal sesamoid bones (PSB) were removed. A midsagittal section of bone 85- to 95-micromole thick was prepared for histomorphometric analysis by use of computerized image analysis and epifluorescent microscopy. Porosity (percent), trabecular width (micrometer), extent of anisotropy (percent), mineralizing surface (percent), fractional mineralizing surface (percent), and mineral apposition rate (micrometers per day) were determined at 5 circular regions of each specimen. Region 1 was located within the apex of the PSB, regions 2, 3, and 4 were subjacent to the subchondral plate, and region 5 was within the basilar articular margin. Data were pooled to allow comparison by training group and by region. The PSB from horses trained on dirt tracks (groups 2 and 3) had significantly (P < 0.05) lower porosities and greater trabecular width, compared with the control group. The PSB from all training group specimens had significantly larger mineralizing surfaces than control group specimens. The fractional mineralizing surface revealed a rapid and vigorous response of the endosteal surface of the PSB in horses trained on dirt tracks. When group data were pooled, region 2 was found to have the lowest porosity and greatest trabecular width of any region. Region 1 was found to have the highest porosity and lowest structural anisotropy of any region. Structural anisotropy of the cancellous bone was greatest at regions 2, 3, and 4. The results of this study demonstrate a substantial stress adaptive remodeling response of the PSB to training-related stimuli. Regional morphologic variations were found that presumably reflect the load history of the PSB in vivo. Adaptive changes may allow the PSB to withstand without failure large stresses generated during maximal exercise.