In situ and ex vivo evaluation of an arthroscopic indentation instrument to estimate the health status of articular cartilage in the equine metacarpophalangeal joint

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.     

Measurement of articular cartilage stiffness of the femoropatellar, tarsocrural, and metatarsophalangeal joints in horses and comparison with biochemical data

OBJECTIVE: To determine normal cartilage stiffness values in different weight-bearing and non-weight-bearing areas of 3 different equine joints, and to evaluate the relationship between cartilage stiffness and glycosaminoglycan (GAG) and collagen content. STUDY DESIGN: Compressive stiffness of the articular cartilage was measured in 8 horse cadaver femoropatellar (FP), tarsocrural (TC), and metatarsophalangeal (MT) joints. Gross evaluation, collagen content, GAG content, and histologic appearance were assessed for each measurement location. ANIMALS: Eight equine cadavers (4 intact females, 4 castrated males; 7 Quarter Horse or Quarter Horse type, 1 Arabian; aged 4-12 years, weighing 400-550 kg). METHODS: The articular surfaces of 8 equine cadaver FP, TC, and MT joints were grossly evaluated for signs of articular cartilage pathology. Stiffness at preselected sites (FP joint-6 sites; TC joint-3 sites; MT joint-4 sites) was determined using an arthroscopic indentation instrument. Biochemical composition (collagen, GAG content) and histologic evaluation (modified Mankin score) were assessed for each measurement site. RESULTS: All cartilage from all sites evaluated was determined to be normal based on macroscopic and histologic assessments. No significant correlation between Mankin scores and cartilage stiffness values was observed. Site differences in cartilage stiffness were measured in all 3 joints (P<.001). GAG or collagen content had a significant positive correlation with stiffness values in 6 of 13 sites (P<.05, r>0.622, r2>0.387). CONCLUSION: Relative cartilage stiffness values measured in healthy equine joints are site dependent and can be measured using an indentation device intended for arthroscopic application. CLINICAL RELEVANCE: An indentation instrument provided an objective means of determining relative compressive stiffness of articular cartilage. Further research needs to be performed to confirm the site and joint differences observed in this study in clinically normal horses and to determine if the tester can be used clinically to predict articular cartilage pathology.     

Functional adaptation of articular cartilage from birth to maturity under the influence of loading: a biomechanical analysis

REASONS FOR PERFORMING STUDY: The concept of functional adapatation of articular cartilage during maturation has emerged from earlier biochemical research. However, articular cartilage has principally a biomechanical function governed by joint loading. OBJECTIVES: To verify whether the concept of functional adaptation can be confirmed by direct measurement of biomechanical properties of cartilage. HYPOTHESIS: Fetuses have homogeneous (i.e. site-independent) cartilage with regard to biomechanical properties. During growth and development to maturity, the biomechanical characteristics adapt according to functional (loading) demands, leading to distinct, site-dependent biomechanical heterogeneity of articular cartilage. METHODS: Osteochondral plugs were drilled out of the surface at 2 differently loaded sites (Site 1: intermittent impact-loading during locomotion, Site 2: low-level constant loading during weightbearing) of the proximal articular cartilage surface of the proximal phalanx in the forelimb from stillborn foals (n = 8), horses of age 5 (n = 9) and 18 months (n = 9) and mature horses (n = 13). Cartilage thickness was measured using ultrasonic, optical and needle-probe techniques. The osteochondral samples were biomechanically tested in indentation geometry. Young's modulus at equilibrium, dynamic modulus at 1 Hz and the ratios of these moduli values between Sites 1 and 2 were calculated. Age and site effects were evaluated statistically using ANOVA tests. The level of significance was set at P<0.05. RESULTS: Fetal cartilage was significantly thicker compared to the other ages with no further age-dependent differences in cartilage thickness from age 5 months onwards. Young's modulus stayed constant at Site 1, whereas at Site 2 there was a gradual, statistically significant increase in modulus during maturation. Values of dynamic modulus at both Sites 1 and 2 were significantly higher in the fetus and decreased after birth. Values for both moduli were significantly different between Sites 1 and 2 from age 18 months onwards. The ratio of values between Sites 1 and 2 for Young's modulus and dynamic modulus showed a gradual decrease from approximately 1.0 at birth to 0.5-0.6 in the mature horse. At age 18 months, all values were comparable to those in the mature horse. CONCLUSIONS: In line with the concept of functional adaptation, the neonate is born with biomechanically 'blank' or homogeneous cartilage. Functional adaptation of biomechanical properties takes place early in life, resulting in cartilage with a distinct heterogeneity in functional characteristics. At age 18 months, functional adaptation, as assessed by the biomechanical characteristics, has progressed to a level comparable to the mature horse and, after this age, no major adaptations seem to occur. POTENTIAL RELEVANCE: Throughout life, different areas of articular cartilage are subjected to different types of loading. Differences in loading can adequately be met only when the tissue is biomechanically adapted to withstand these different loading conditions without injury. This process of functional adaptation starts immediately after birth and is completed well before maturity. This makes the factor of loading at a young age a crucial variable, and emphasises the necessity to optimise joint loading during early life in order to create an optimal biomechanical quality of articular cartilage, which may well turn out to be the best prevention for joint injury later in life.     

Functional consequences of cartilage degeneration in the equine metacarpophalangeal joint: quantitative assessment of cartilage stiffness

REASONS FOR PERFORMING STUDY: No quantitative data currently exist on the relationship of the occurrence of cartilage degeneration and changes in site-specific biomechanical properties in the metacarpophalangeal (MCP) joint in the horse. OBJECTIVES: To gain insight into the biomechanical consequences of cartilage deterioration at 2 differently loaded sites on the proximal articular surface of the proximal phalanx (P1). HYPOTHESIS: Static and dynamic stiffness of articular cartilage decreases significantly in degenerated cartilage. METHODS: Cartilage degeneration index (CDI) values were measured at the lateral dorsal margin (Site 1), lateral central fovea (Site 2) and entire joint surface of P1 (CDIP1) in 30 horses. Group 1 contained joints without (CDIP1 values <25 %, n = 22) and Group 2 joints with (CDIP1 values >25 %, n = 8) signs of cartilage degeneration. Cartilage thickness at Sites 1 and 2 was measured using ultrasonic and needle-probe techniques. Osteochondral plugs were drilled out from Sites 1 and 2 and subsequently tested biomechanically in indentation geometry. Young's modulus at equilibrium and dynamic modulus were determined. RESULTS: Cartilage thickness values were not significantly different between the 2 groups and sites. Young's modulus at Site 1 was significantly higher in Group 1 than in Group 2; at Site 2, the difference was not significant. Dynamic modulus values were significantly higher in Group 1 than in Group 2 at both sites. CONCLUSIONS: Degenerative cartilage changes are clearly related to loss of stiffness of the tissue. Absolute changes in cartilage integrity in terms of CDI are greatest at the joint margin, but concomitant changes are also present at the centre, with a comparable decrease of the biomechanical moduli at the 2 sites. Therefore, significant cartilage degradation at the joint margin not only reflects local deterioration of biomechanical properties, but is also indicative of the functional quality in the centre. POTENTIAL RELEVANCE: These findings may be important for improving prognostication and developing preventative measures.     

Influence of site and age on biochemical characteristics of the collagen network of equine articular cartilage

OBJECTIVE: To determine variations in biochemical characteristics of equine articular cartilage in relation to age and the degree of predisposition for osteochondral disease at a specific site. SAMPLE POPULATION: Articular cartilage specimens from 53 horses 4 to 30 years old. PROCEDURE: Healthy specimens were obtained from 2 locations on the proximal articular surface of the first phalanx that had different disease prevalences (site 1 at the mediodorsal margin and site 2 at the center of the medial cavity). Water, total collagen, and hydroxylysine contents and enzymatic (hydroxylysylpyridinoline [HP]) and nonenzymatic (pentosidine) crosslinking were determined at both sites. Differences between sites were analyzed by ANOVA (factors, site, and age), and age correlation was tested by Pearson's product-moment correlation analysis. Significance was set at P< 0.01. RESULTS: Correlation with age was not found for water, collagen, hydroxylysine contents, and enzymatic cross-linking. Nonenzymatic crosslinking was higher in older horses and was linearly related to age (r = 0.94). Water and collagen contents and HP and pentosidine crosslinks were significantly higher at site 1. Hydroxylysine content was significantly lower at site 1. CONCLUSIONS: Except for nonenzymatic glycation, the composition of articular cartilage collagen does not change significantly in adult horses. A significant topographic variation exists in biochemical characteristics of the articular cartilage collagen network in equine metacarpophalangeal joints. These differences may influence local biomechanical properties and, hence, susceptibility to osteochondral disease, as will greater pentosidine crosslinks in older horses that are likely to cause stiffer and more brittle cartilage.     

Quantification and age-related distribution of articular cartilage degeneration in the equine fetlock joint

REASONS FOR PERFORMING STUDY: The equine fetlock joint has the largest number of traumatic and degenerative lesions of all joints of the appendicular skeleton. OBJECTIVE: To gain insight into the distribution of cartilage degeneration across the articular surface in relation to age in order better to understand the dynamic nature and progression of osteoarthritis (OA). HYPOTHESIS: That there would be a specific age-related distribution pattern of cartilage degeneration in the equine metacarpophalangeal joint. METHODS: The proximal articular cartilage surfaces of the first phalanges (P1) of 73 slaughter horses (age range 0.4-23 years) with different stages of osteoarthritis were scored semiquantitatively on a 0 to 5 scale and also assessed quantitatively using the cartilage degeneration index (CDI(P1)), which ranges from 0 to 100%. Furthermore, CDI values were determined for special areas of interest; medial dorsal surface (CDI(mds)), lateral dorsal surface (CDI(lds)), medial central fovea (CDI(mcf)) and lateral central fovea (CDI(lcf)). Correlations were calculated for CDI(P1) values and CDI values at the specific areas of interest with macroscopic scores and with age. RESULTS: There was a high correlation between the semiquantitative macroscopic score and the quantitative CDI(P1) values (r = 0.92; P < 0.001). A macroscopic score of 0 (i.e. no obvious cartilage degeneration) corresponded with a CDI(P1) mean +/- s.e. value of 25 +/- 2.8% and a macroscopic score of 5 (i.e. severe cartilage degeneration in localised areas) with a mean +/- s.e. value of 38.1 +/- 7.9%. There was a moderate but highly significant correlation between the CDI(P1) value and the age of the horses (r = 0.41; P < 0.001). Highest CDI values were calculated for the medial dorsal surface (from 10.6 +/- 2.8% at macroscopic Grade 0 to 63.1 +/- 8.4% at Grade 5). At the lateral dorsal surface, these values were 5.9 +/- 1.4% and 47.2 +/- 10.4%, respectively. The CDI(mcf) and CDI(lcf) were significantly lower (P < 0.05) than the CDI(mds) and CDI(lds) at all grades. The CDI(mcf) ranged from 1.0 +/- 2.9% at Grade 0 to 43.7 +/- 9.1% at Grade 5; laterally, these values were 1.5 +/- 2.6% and 15.2 +/- 6.2%, respectively. CONCLUSIONS: CDI grading increased from lateral to medial and from central to dorsal. This specific distribution pattern confirms the heterogeneous nature of the OA process and strongly supports an important role for biomechanical loading, superimposed on age-related changes, in the spread of the disorder over the joint. POTENTIAL RELEVANCE: Knowledge of the development of OA across the articular surface is essential for understanding the dynamic nature and progression of the disease and can form a basis for improvements in diagnostic and therapeutic approaches to degenerative joint disease.     

CORRELATION OF ARTICULAR CARTILAGE THICKNESS MEASUREMENTS MADE WITH MAGNETIC RESONANCE IMAGING,MAGNETIC RESONANCE ARTHROGRAPHY,AND COMPUTED TOMOGRAPHIC ARTHROGRAPHY WITH GROSS ARTICULAR CARTILAGE THICKNESS IN THE EQUINE METACARPOPHALANGEAL JOINT

Osteoarthritis of the metacarpophalangeal joint is common cause of lameness in equine athletes, and is hallmarked by articular cartilage damage. An accurate, noninvasive method for measuring cartilage thickness would be beneficial to screen for cartilage injury and allow for prompt initiation of interventional therapy. The objective of this methods comparison study was to compare computed tomographic arthrography (CTA), magnetic resonance imaging (MRI), and magnetic resonance arthrography (MRA) measurements of articular cartilage thickness with gross measurements in the metacarpophalangeal joint of Thoroughbred horses. Fourteen cadaveric, equine thoracic limbs were included. Limbs were excluded from the study if pathology of the metacarpophalangeal articular cartilage was observed with any imaging modality. Articular cartilage thickness was measured in nine regions of the third metacarpal bone and proximal phalanx on sagittal plane MRI sequences. After intra‐articular contrast administration, the measurements were repeated on sagittal plane MRA and sagittal CTA reformations. In an effort to increase cartilage conspicuity, the volume of intra‐articular contrast was increased from 14.5 ml, to maximal distention for the second set of seven limbs. Mean and standard deviation values were calculated, and linear regression analysis was used to determine correlations between gross and imaging measurements of cartilage thickness. This study failed to identify one imaging test that consistently yielded measurements correlating with gross cartilage thickness. Even with the use of intra‐articular contrast, cartilage surfaces were difficult to differentiate in regions where the cartilage surfaces of the proximal phalanx and third metacarpal bone were in close contact with each other.     

New approach for quantitative assessment of articular cartilage degeneration in horses with osteoarthritis

OBJECTIVE: To evaluate a modified digital imaging technique for quantitative assessment of the grade of osteoarthritis across the proximal articular surface of the first phalanx in horses. SAMPLE POPULATION: 6 metacarpophalangeal (fet-lock) joint specimens from 6 horses with various stages of osteoarthritis. PROCEDURE: First phalanx specimens, together with 4 gray scale reference calibration targets, were positioned in a bath with the proximal articular cartilage surface submerged in saline (0.9% NaCl) solution. Digital images were obtained from the articular surface before and after staining with Indian ink. Computer-controlled gray level analysis of the nonstained and Indian ink-stained cartilage surfaces and gray scale reference calibration targets was performed by use of the mean pixel value (based on 255-gray scale). An increase in the mean pixel value after staining was used to calculate the cartilage degeneration index (CDI). RESULTS: The CDI of the proximal articular cartilage surface of the first phalanx specimens ranged from 9.2 +/- 5.7 (early stage osteoarthritis) to 41.5 +/- 3.6% (late stage osteoarthritis). The effect of repeating the measurement 6 times in nonstained (including repositioning) and stained specimens (including repositioning and restaining) was not significant. Up to 10 measurements of nonstained specimens could be made without refreshing the bath solution. In stained specimens, mean gray level increased significantly after the sixth measurement. CONCLUSIONS AND CLINICAL RELEVANCE: The modified digital imaging technique allowed quantitative assessment of cartilage degeneration across the articular cartilage surface. The CDI is the first quantitative measure for osteoarthritis-induced cartilage degeneration over an entire joint surface in horses.     

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.     

Age-related morphometry of equine calcified cartilage

Although there are many studies in the equine literature focused on articular diseases and the aetiology of osteoarthritis, few have concentrated on normal articular structures and how they change with age. The objective of this investigation was to study the thickness and morphology of the calcified cartilage layer of the distal metacarpus over a range of ages. A parasagittal slab of bone was sectioned from the region of sesamoid contact on the medial condyle of the metacarpi from 34 horses. The slab of bone was preserved, dehydrated and embedded, undecalcified, in methylmethacrylate and then stained with toluidine blue. Six repeatable fields of interest from the distal aspect of each metacarpus were digitised and examined to determine the morphology of the calcified cartilage layer. The thickness of the calcified cartilage, range 88-426 microm, was estimated using a method of integration. The results indicate an age-related influence on the thickness of the calcified cartilage layer, generally increased in older horses. While this finding is significant, perhaps more importantly a positional relationship was also identified, indicating that pressures endured by different regions within a joint may dictate morphological development of the tissues. This study has begun to lay the groundwork to determine whether the calcified layer of the hyaline cartilage could be involved in the development of osteoarthritis.     

Use of an antineoepitope antibody for identification of type-II collagen degradation in equine articular cartilage.

OBJECTIVE: To develop an antibody that specifically recognizes collagenase-cleaved type-II collagen in equine articular cartilage. SAMPLE POPULATION: Cartilage specimens from horses euthanatized for problems unrelated to the musculoskeletal system. PROCEDURE: A peptide was synthesized representing the carboxy- (C-) terminus (neoepitope) of the equine type-II collagen fragment created by mammalian collagenases. This peptide was used to produce a polyclonal antibody, characterized by western analysis for reactivity to native and collagenase-cleaved equine collagens. The antibody was evaluated as an antineoepitope antibody by ELISA, using peptides +/- an amino acid at the C-terminus of the immunizing peptide. Collagen cleavage was assayed from equine articular cartilage cultured with interleukin-1 (IL-1), +/- a synthetic MMP inhibitor, BAY 12-9566. Cartilage specimens from osteoarthritic and nonarthritic joints were compared for antibody staining. RESULTS: An antibody, 234CEQ, recognized only collagenase-generated 3/4-length fragments of equine type-II collagen. This was a true antineoepitope antibody, as altering the C-terminus of the immunizing peptide significantly decreased competition for binding in an inhibition ELISA. The IL-1-induced release of type-II collagen fragments from articular cartilage was prevented with the MMP inhibitor. Cartilage from an osteoarthritic joint of a horse had increased staining with the 234CEQ antibody, compared with normal articular cartilage. CONCLUSIONS AND CLINICAL RELEVANCE: We generated an antineoepitope antibody recognizing collagenase-cleaved type-II collagen of horses. This antibody detects increases in type-II collagen cleavage in diseased equine articular cartilage. The 234CEQ antibody has the potential to aid in the early diagnosis of arthritis and to monitor treatment responses.     

Relationship between synovial fluid levels of glycosaminoglycans, hydroxyproline and general MMP activity and the presence and severity of articular cartilage change on the proximal articular surface of P1

REASONS FOR PERFORMING STUDY: Osteoarthritis (OA) is one of the most prevalent and disabling chronic conditions affecting horses and leads to degeneration of articular cartilage. Diagnosis is based on clinical signs in combination with radiography, which is relatively insensitive and provides only an indication of accumulated damage. Alternative methods, such as molecular markers, are therefore needed that can quantitatively, reliably and sensitively detect osteoarthritic changes in the joints at an early stage of the disease. If such markers are to be used reliably, it is important to know the relationship between marker concentration and cartilage composition. OBJECTIVES: To study the relationship between cartilage composition, synovial fluid levels of glycosaminoglycans (GAGs), hydroxyproline (Hyp) and general matrix metalloproteinase (MMP) activity, and the presence and severity of articular cartilage damage on the articular surface of P1. METHODS: Synovial fluid (SF) was collected from the metacarpophalangeal joints of 60 mature horses, and levels of GAGs, Hyp and general MMP activity were determined. Further, GAG and denatured collagen content of the articular cartilage were determined at the dorsal articular margin of P1 (site 1) and central cavity (site 2). The presence and severity of cartilage change was quantified using the cartilage degeneration index (CDI), measured at the same 2 sites. Correlations between SF parameters, cartilage composition and degree of cartilage degeneration were sought using correlation analysis. RESULTS: There was no correlation between GAG or Hyp content of SF and the amount of GAGs or denatured collagen, respectively, in cartilage. In joints with moderate to severe cartilage damage, the GAG content of site 1 was significantly lower than in joints with no to minimal cartilage change (P = 0.005) and there was a negative correlation between the amount of denatured collagen and GAG content at site 1 in all joints (r = -039, P = 0.002). Further, in joints with moderate to severe cartilage damage, there was a significant positive correlation between MMP activity in SF and Hyp levels in SF (r = 0.72, P < 0.001) and CDI at sites 1 (r = 0.46, P = 0.03) and 2 (r = 0.43, P = 0.04). CONCLUSIONS: General MMP activity in joints with moderate to severe cartilage damage is related to the severity of those cartilage changes and to Hyp levels in SF. Glycosaminoglycan levels in SF are not directly related to MMP activity, GAG content of articular cartilage or severity of cartilage change. POTENTIAL RELEVANCE: Glycosaminoglycan levels in SF are not helpful for the early detection of cartilage lesions. In damaged joints, Hyp levels may give an indication of the severity of cartilage change as they are strongly related to MMP activity, but do not qualify as markers for the presence or absence of cartilage lesions.     

Ultrastructural immunolocalization of cartilage oligomeric matrix protein (COMP) in the articular cartilage on the equine third carpal bone in trained and untrained horses

The present study was designed to delineate the presence of COMP at the ultrastructural level comparing concentrations between two areas of articular cartilage from the equine third carpal bone, subjected to different loading, from trained and untrained horses. We also analyzed the fibril thickness of collagen type II in the same compartments and zones. Samples were collected from high load-bearing areas of the dorsal radial facet (intermittent high load) and an area of the palmar condyle (low constant load) in five non-trained and three trained young racehorses. The data show that COMP is much less abundant in the matrix in intermittent high loaded areas of articular cartilage from trained horses as compared to the untrained horses (p = 0.036). On the other hand, the untrained horses often displayed a higher immunolabeling in loaded areas compared to unloaded areas, indicating that an adequate dynamic load promotes COMP synthesis and/or retention, while an excessive load may have an opposite effect. The collagen fibril diameter showed marked variation between individuals. The present study indicates that dynamic in vivo compression at high load and frequency lowers matrix content of COMP in the articular cartilage of the third carpal bone. It also indicates that the collagen network is influenced by mechanical load following by strenuous exercise.     

Differences in the topographical distribution of articular cartilage degeneration between equine metacarpo- and metatarsophalangeal joints

REASONS FOR PERFORMING STUDY: The equine metacarpophalangeal (MCP) and metatarsophalangeal (MTP) joints, although having virtually the same geometrical appearance, differ in the prevalence of joint pathologies, such as osteochondral fragmentation, and in biomechanical behaviour. The recently developed cartilage degeneration index (CDI) technique offers a possibility to assess quantitatively differences in cartilage degeneration between these joints and to compare these with known differences in biomechanics and clinical observations. OBJECTIVES: To compare the topographical distribution of articular cartilage degeneration across the proximal articular surface of the proximal phalanx (P1) in the equine fore- and hindlimb. METHODS: In 24 distal hindlimbs from 24 horses, articular cartilage degeneration of the proximal articular surface of P1 was quantified using the CDI. Overall CDI value (CDI(P1)) and CDI values of 6 areas of interest were determined: the medial dorsal surface (mds), lateral dorsal surface (lds), medial central fovea (mcf), lateral central fovea (lcf), medial plantar surface (mps) and lateral plantar surface (lps). The joints were divided into 4 equally sized groups of increasing CDI(P1) values. From an existing CDI database of MCP joints, 24 joints were selected with matching CDI(P1) values to the MTP joints and CDI values for the same areas of interest were determined. RESULTS: In both the MCP and MTP joints, highest CDI values were determined at the dorsal articular surfaces. Values were not significantly different between fore- and hindlimbs. In contrast to the MCP joint, CDI values at the plantar joint margin were significantly higher compared to CDI values in the central sites in the MTP joint. CDI values for the plantar surfaces of P1 were significantly higher than those for the palmar surfaces in the forelimb in joints with advanced stages of OA; and values for the central regions of P1 were significantly lower in the hindlimb compared with the forelimb in joints with severe OA. CONCLUSIONS: In both fore- and hindlimbs, initial cartilage degeneration started at the dorsal articular margin of P1. There was a major difference in the spread of cartilage degeneration; in the forelimb both the central and palmar parts are about equally involved, whereas in the hindlimb the plantar parts were significantly more and the central parts significantly less involved. These differences can be linked to differences in biomechanical loading reported elsewhere. POTENTIAL RELEVANCE: This study supports the hypothesis that differences in biokinematics between fore- and hindlimbs are associated with differences in the development of cartilage degeneration and other joint pathologies such as osteochondral fragmentation in the MCP and MTP joints. This information is indispensable for a better understanding of the dynamic nature and progression of these joint disorders and may be of help when monitoring the effects of therapeutic interventions and preventative measures.     

Cylindrical press-fit osteochondral allografts for resurfacing the equine metatarsophalangeal joint

OBJECTIVE: To investigate the feasibility of resurfacing the equine fetlock joint using cylindrical, orthotopic, press-fit, osteochondral allografts. STUDY DESIGN: Experimental study. ANIMALS: Ten mature, mixed-breed horses. METHODS: Cylindrical, osteochondral grafts (6.5-mm diameter) were harvested aseptically from cadaveric equine metatarsophalangeal joints. Allografts were transplanted into 6 horses; 4 horses were sham operated. The surgical approach involved creation of a bone block at the origin of the medial collateral ligament and luxation of the metatarsophalangeal joint. Grafts were placed into the medial and lateral metatarsal condyles. Radiographs were taken at 8 and 25 weeks, and lameness was evaluated at 25 weeks. Horses were killed at 25 weeks. Analyses included gross evaluation, microradiography, paravital staining, light microscopy, and cartilage biochemistry. RESULTS: No complications occurred that could be attributed to the surgical procedure. Graft congruency with the surrounding articular cartilage was fair to excellent. Two horses were sound at 25 weeks. Most grafts had more than 90% articular cartilage coverage, and histologic and microradiographic analysis revealed good graft incorporation and articular cartilage survival. Sulphated glycosaminoglycan concentration was decreased in grafted tissue. CONCLUSIONS: We attribute the viability of osteochondral allografts in the equine fetlock to adequate congruency, stable graft fixation, and the use of orthotopic tissue. Host response to the allograft bone tissue did not affect cartilage viability. CLINICAL RELEVANCE: Before clinical use, improvements to instrumentation are required that would decrease damage to grafts and minimize technique-associated incongruencies of the articular surface at the time of grafting. Larger grafts would also likely be required to resurface a greater surface area.     

Differential distribution of cathepsins B and L in articular cartilage during skeletal development in the horse

REASONS FOR PERFORMING STUDY: This study was designed to examine a new role for cysteine proteinases in the process of endochondral ossification. OBJECTIVES: The aim of the present study was to investigate the presence and distribution of cathepsin B and cathepsin L in equine articular cartilage during development. METHODS: Full-depth cartilage samples from a total of 40 horses (age range: 4 month fetuses to 2 years) were examined and enzymes detected by immunocytochemical localisation. RESULTS: Observations on the presence of cathepsins B and L revealed significant age-related differences, resulting in clear division of the animals into 2 age groups: i) fetuses and neonates; ii) young growing horses (age 4 weeks to 2 years). Cathepsin B was not detected in cartilage from the majority of fetuses and neonates but was located characteristically in chondrocytes at the articular surface and hypertrophic zone in all growing horses. In contrast, cathepsin L was predominantly present in fetal and neonatal cartilage, located primarily in proliferating chondrocytes. CONCLUSIONS: This study is the first to demonstrate differential and site-specific roles for cathepsin B and cathepsin L in skeletal development in the horse. Potential relevance: The demonstrated involvement of cathepsins B and L in endochondral ossification is of relevance to developmental orthopaedic diseases such as osteochondrosis in which there is a focal failure of bone formation.     

Spontaneous production of nitric oxide (NO), prostaglandin (PGE2) and neutral metalloproteinases (NMPs) in media of explant cultures of equine synovial membrane and articular cartilage from normal and osteoarthritic joints

Nitric oxide (NO), prostaglandin E2 (PGE2), and the activity of neutral metalloproteinases (NMPs) were measured in conditioned media of equine synovial membrane and articular cartilage explant cultures from horses with normal joints (n = 7) and from horses affected with moderate (n = 7) or severe osteoarthritis (n = 14) as judged by macroscopic appearance. Normal articular cartilage appeared glossy and bluish-white, was of normal thickness and showed no evidence of discolouration, fibrillation or other cartilage discontinuity. Slight discolouration and fibrillation or minor clefts of the cartilage were considered as moderate OA, whereas erosions of articular cartilage down to the subchondral bone were considered as cases of severe OA. Explant cultures of equine synovial membrane and articular cartilage released the local mediators, NO and PGE2, as well as detectable levels of NMP activity into culture media. Concentrations of NO were higher in articular cartilage explants compared to synovial membrane explants, whereas concentrations of PGE2 were higher in synovial membrane explants. The NMPs with collagenolytic activities were similar in both explant cultures, whereas gelatinolytic activities were higher in synovial membrane explant cultures and caseinolytic activities were generally higher in articular cartilage explant cultures. Furthermore it was shown that concentrations or enzyme activities increased according to the severity of disease of the joints. Concentrations for NO, collagenolytic and gelatinolytic NMPs were relatively stable, whereas PGE2 and caseinolytic NMP concentrations increased over time in culture.     

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.     

Validation of magnetic resonance imaging for measurement of equine articular cartilage and subchondral bone thickness

OBJECTIVE: To validate use of magnetic resonance images (MRIs) for measurement of equine articular cartilage and subchondral bone thickness by comparison with measurements in histologic specimens. SAMPLE POPULATION: 32 cadaveric carpal joints from 16 horses. PROCEDURE: Magnetic resonance imaging was performed by use of 3-dimensional fast spoiled gradient echo (SPGR) and T2* 3-dimensional fast gradient echo (GRE) pulse sequences with and without fat saturation. Standard sites on the medial and lateral facets of the intermediate, radial, and third carpal bones were used for subchondral bone and articular cartilage thickness measurements. Digital image analysis software was used for MRI measurements 10 mm from the dorsal extent and perpendicular to the articular surface. Histomorphometric measurements of hyaline, calcified cartilage, and subchondral bone thickness were obtained at selected sites. Comparisons between histomorphometric and MRI measurements and between magnetic resonance pulse sequences were evaluated. RESULTS: There were significant correlations between GRE and SPGR and SPGR and histologic measurements of articular cartilage, with no significant difference between measurements and good agreement. When calcified cartilage was excluded from the histologic measurement, MRI measurements were significantly greater than histologic measurements. For subchondral bone thickness, there was significant correlation between GRE and SPGR but GRE was significantly greater than SPGR measurements. Histomorphometric and MRI measurements were strongly correlated and not significantly different. CONCLUSIONS AND CLINICAL RELEVANCE: Magnetic resonance imaging provides a good representation of cartilage and subchondral bone thickness, supporting its use in the study and clinical diagnosis of osteochondral structure and alteration.