Canine Knee Research Articles

Articular cartilage thickness and glycosaminoglycan distribution in the young canine knee joint after remobilization of the immobilized limb

The recovery of articular cartilage from atrophy induced by joint immobilization was investigated in immature dogs. In a previous study, we showed that 11 weeks of immobilization of the knee (stifle) joint of young dogs reduced the concentration of articular cartilage glycosaminoglycans (GAGs) by 13-47%. In the present study, right hindlimbs from six female beagles were immobilized for 11 weeks, as in the previous study, and then were remobilized for 15 weeks. Cartilage from the knee joint was compared with cartilage from nonimmobilized knees of eight age-matched control beagles. Histological samples taken from 11 different locations of the knee joint were stained with safranin O, and microspectrophotometry was used to demonstrate distribution of GAGs in the tissue. After remobilization, GAG concentration was restored in the patellofemoral region and tibial condyles. On the summits of the femoral condyles, and especially at the periphery of the femoral condyles, GAG concentration remained 8-26% less than the control values. On the summits, the thickness of the uncalcified cartilage was as much as 15% less than in the age-matched controls. Consequently, the changes induced by unloading were reversible to a great extent, but a full restoration of articular cartilage was not obtained at all sites of the knee joint within the 15 weeks of remobilization. Immobilization of the skeletally immature joint therefore may affect the development of articular cartilage in such a way that very slow recovery or permanent alterations are induced.

Structural differences between two populations of articular cartilage proteoglycan aggregates

To determine if articular cartilage contains structurally distinct populations of proteoglycan aggregates, we extracted and purified proteoglycans from canine knee cartilage under associative conditions. Equilibrium density gradient centrifugation separated three proteoglycan populations, on the basis of differences in sedimentation velocity, into groups of 21, 106, and 270 S. Electron microscopic examination showed that the 21 S samples contained free aggrecan molecules and clusters of aggrecan molecules, with a mean of five aggrecan molecules per cluster. The 106 and 270 S samples contained proteoglycan aggregates consisting of central hyaluronan filaments with multiple attached aggrecan molecules. The two populations of aggregates did not differ in mean aggrecan length or in the spacing of aggrecan molecules along the hyaluronan filaments, but the slower sedimenting aggregates (106 S) had significantly shorter hyaluronan filaments as measured by electron microscopy (mean hyaluronan length, 400 compared with 1,162 nm) and one-third as many aggrecan molecules per aggregate (mean number of aggrecan molecules per aggregate, 15 compared with 44). This study shows that articular cartilage contains aggrecan clusters and two structurally distinct populations of proteoglycan aggregates. The differences between the two types of aggregate, in particular the number of aggrecan molecules per aggregate, may reflect differences in their assembly, stability, or turnover and give them different mechanical and biological properties.

Proteoglycan and Collagen Alterations in Canine Knee Articular Cartilage Following 20 KM Daily Running Exercise for 15 Weeks

The composition of extracellular matrix was studied at 11 different sites in the knee (stifle) articular cartilage of young beagle dogs after running exercise of 15 weeks, 20 km/day. Water content was significantly elevated by 5-17% in the patellofemoral groove and in anterior and intermediate sites on the lateral condyle of femur. Collagen content was decreased by 14 to 20% in the same sites of the lateral condyle. Proteoglycan (PG) content was not significantly changed except in the posterior edge of the medial condyle of femur with a 30% decrease. The proportion of PGs capable of reaggregation with hyaluronan was increased in tibial and femoral surfaces (mean of all sites +18%). Also, the aggregating PG monomers were larger at all sites, as studied by agarose gel electrophoresis. The chondroitin-6 to 4-sulphate ratio was reduced at the summits of femoral condyles and patella by 10 to 25%, but increased in the patellar surface of femur and tibial medial condyle, fitting to a previous finding that strenuous running depleted proteoglycans at the summits of femoral condyle from the superficial zone that is rich in chondroitin-6-sulphate. The increased water content, accompanied with a decreased concentration of collagen in the lateral femoral condyle, suggests loosening of the collagenous framework, an idea compatible with an earlier notion of superficial depletion of PGs in these sites, and possibly predisposing to degeneration. The size increase of the aggregating PGs probably indicate that a larger proportion of matrix PGs were newly synthesized and hence the turnover rate of the PGs was enhanced. It is concluded that the strenuous running program induced locally restricted changes resembling early degeneration of articular cartilage, while simultaneously caused alterations that suggest a general stimulation of proteoglycan metabolism.

Primary culture of microvascular endothelial cells from canine meniscus.

Microvascular endothelial cells were isolated and cloned from canine knee menisci. Initially, the endothelial cell colonies in culture exhibited a cobblestone morphology. With passage and time, these cultures adopted a fibroblastic or fusiform morphology. The endothelial nature of the cells was demonstrated by positive immunostaining with antibodies to von Willebrand factor, laminin, and type-IV collagen and by capillary-like tube formation on Matrigel. These endothelial cells could be passaged as many as 10 times with retention of the endothelial characteristics. This population of meniscal microvascular endothelial cells will facilitate studies on the role of blood vessels in meniscal repair.

Sensory nerves only temporarily protect the unstable canine knee joint from osteoarthritis. Evidence that sensory nerves reprogram the central nervous system after cruciate ligament transection.

The slow rate at which articular cartilage degrades in dogs after transection of the anterior cruciate ligament (ACLT) has been attributed to capsular thickening and buttressing by osteophytes. We investigated the roles of the peripheral and central nervous systems in protecting knee joints with chronic ACL deficiency from breakdown. Five groups of dogs were studied; all were killed 72 weeks after left knee surgery. Group A had ACLT, group B had ACLT followed 52 weeks later by ipsilateral L4-S1 dorsal root ganglionectomy (DRG), group C had DRG followed 2 weeks later by ACLT, group D had sham DRG followed 2 weeks later by ACLT, and group E had DRG followed 2 weeks later by sham ACTL. Group E dogs did not develop knee pathology. All cruciate-deficient knees were lax at the end of the study. The osteoarthritis (OA) that developed in groups A, B, and D was comparable (P > 0.05), and was significantly greater than that in group E (P < 0.05). Group C developed much more severe OA than any of the other groups (P < 0.05). Ipsilateral sensory input is temporarily important in protecting the unstable joint from rapid breakdown. Over time, the central nervous system apparently acquires the ability to protect the unstable joint without continued ipsilateral sensory input.

The quantitation of a native chondroitin sulfate epitope in synovial fluid lavages and articular cartilage from canine experimental osteoarthritis and disuse atrophy.

Previous studies have shown the presence of a native chondroitin sulfate epitope in articular cartilage proteoglycans from canine knee joints with experimental early osteoarthritis (OA), but not in normal cartilage. The objective of this study was to quantitate the native epitope recognized by monoclonal antibody 3-B-3 in synovial fluids and articular cartilage of diseased joints. An immunoassay with monoclonal antibody 3-B-3, which recognizes a native chondroitin-6-sulfate structure, was developed and used to analyze synovial fluid lavage material and extracts of articular cartilage from canine knee joints with early experimental OA or with mild disuse atrophy, and from control animals. The concentration of epitope in the OA fluids was elevated 33-35-fold, and in the OA articular cartilage extracts it was elevated > 200-fold, compared with samples from the control group. No significant difference was detected in the levels of 3-B-3 epitope in the synovial fluid lavage material or cartilage extracts from the joints of the disuse group versus the control group. The native 3-B-3 epitope in articular cartilage and synovial fluids may be a specific marker of ongoing anabolic events in early degenerative joint disease.

In vitro load transmission in the canine knee: the effect of medial meniscectomy and varus rotation.

The purpose of this study was to determine the in vitro load-transmission characteristics of the canine knee, paying particular attention to the positioning effect of the meniscus in the coronal plane. The intact joint was first loaded and then tested under two different loading conditions after a complete medial meniscectomy. The first set of test conditions attempted to simulate those used by previous investigators, by ignoring the spacer effect of the meniscus. The second set of tests were carried out following varus rotation of the joint (to account for the loss of the meniscal spacer) to assure initial contact in both tibiofemoral compartments at the start of test cycle. It is presumed that this varus realignment occurs during weight bearing following meniscectomy in vivo. As in previous studies, the joints experienced slightly larger displacements (although not statistically significant) and had lower stiffness values following medial meniscectomy than when intact. However, following varus realignment of the joint after meniscectomy, the displacement was markedly smaller (-35% to -49%; P < 0.01) and the structural stiffness was much greater (47-123%; P < 0.05) over the range of forces analyzed, compared with the intact joint. The ratio of dissipated to input energy was 42% for the intact joint, and increased following meniscectomy to 54% (P < 0.05) with realignment and 55% (P < 0.05) without realignment. Measured contact area decreased by 17% (P < 0.05) following meniscectomy alone, and by 12% (P < 0.05) following meniscectomy with realignment.(ABSTRACT TRUNCATED AT 250 WORDS)

Articular Cartilage Thickness and Glycosaminoglycan Distribution in the Canine Knee Joint After Strenuous Running Exercise

The influences of the strenuous running training program on the knee joint articular cartilage was studied in six female beagle dogs. At the age of 15 weeks, the dogs started running on a treadmill inclined 15 degrees uphill. Thereafter, the dogs were trained for 40 weeks, five times a week. For the final 15 weeks, the dogs ran 20 km/day. Six age-matched female beagles served as controls. The cartilage surfaces were intact after the running exercise. The training reduced the thickness of the uncalcified cartilage by 6% in the medial femoral condyle. The glycosaminoglycan concentration was reduced an average of 11% on the summits of the femoral condyles. The reduction was most pronounced (41%) in the superficial 50-micron cartilage zone. In other regions of the knee, such a decrease of glycosaminoglycans was not observed. A shift to strenuous running voided the increase in cartilage thickness and proteoglycan content previously observed after moderate running. Strenuous running induced marked depletion of proteoglycans from the superficial layer of the femoral condyles at sites subjected to highest impact loads.

Motion of the normal and anterior cruciate ligament deficient canine knee during the walk and trot

Motion of the normal and anterior cruciate ligament deficient canine knee during the walk and trot

Osteoarthritic changes in canine articular cartilage, subchondral bone, and synovium fifty‐four months after transection of the anterior cruciate ligament

Anterior cruciate ligament transection (ACLT) in the dog results in osteophyte formation and in morphologic, metabolic, biochemical, and biomechanical changes in the articular cartilage of the unstable knee that mimic those of human osteoarthritis (OA). However, in dogs studied up to 2 years after ACLT, the changes have appeared to be self-limiting, which has led to the suggestion that this is a model of cartilage damage and repair, rather than of OA. To ascertain whether changes in articular cartilage and subchondral bone of dogs subjected to ACLT lead to progressive changes of OA, we studied 3 dogs for 54 months after ACLT. Arthrotomy was performed in the dogs to visualize and then transect the anterior cruciate ligament. When the dogs were killed, full-thickness ulceration of the articular cartilage was seen on the medial femoral condyle and tibial plateau of the unstable knee, while cartilage in other regions was thicker than that of the contralateral knee, consistent with hypertrophic cartilage repair. Synovial infiltration by mononuclear cells was not more severe than that seen in dogs killed at earlier intervals after ACLT, although gross fibrotic thickening of the capsule was apparent in each dog. Histomorphometric studies revealed a marked increase in subchondral bone volume and active bone formation. These findings show that the changes that develop in the canine knee joint after ACLT are progressive and are unambiguously those of OA.

Proteoglycan alterations following immobilization and remobilization in the articular cartilage of young canine knee (stifle) joint

The distribution of proteoglycans (PGs) at 11 sites on the knee (stifle joint) cartilage of young female beagle dogs was studied following cast immobilization for 11 weeks in 90 degrees flexion and after a subsequent remobilization for 15 weeks. Immobilization induced a reduction in PG uronic acid at all sites (mean of -38%), but the greatest depletion (-64%) occurred at the anterior and posterior extremes of the femoral condyles, i.e., at locations where the immobilized cartilage lost contact to the opposing cartilage. Following remobilization, the content of uronic acid remained lower than in the age-matched controls (-18% on average), particularly at the minimum contact sites most affected by immobilization (-33%). The chondroitin-6-sulfate to chondroitin-4-sulfate ratio was reduced by immobilization in most locations (average of -14%) and returned to control values after remobilization. There was no consistent change in the percentage of aggregating PGs observed in Sephacryl S-1000 gel filtration after immobilization or remobilization. However, following remobilization, the aggregating PGs showed an enhanced proportion of the slower mobility band in agarose gel electrophoresis, indicative of a larger monomer size. In the contralateral, load-bearing knee joint, both the uronic acid content and PG monomer type distribution were identical to those observed in the experimental joint, suggesting that the state reached after the remobilization period was due to factor(s) influencing both sides. The results suggest that contact forces between articulating surfaces are required to maintain normal PG content and that the control mechanism works locally at each cartilage site. Restriction of joint mobility and loading in young animals is concluded to cause persistent changes in cartilage matrix. Furthermore, the use of the contralateral joint as the sole control in this kind of studies, although experimentally convenient, seems not to be appropriate.

Open Meniscus Repair: Indications, Technique, and Results

As noted through the work of Fairbank, total meniscectomy is not a benign operation. Cox et al found that meniscectomies in canine knees lead to gross and microscopic degenerative changes. They also noted that partial meniscectomies lead to less severe degenerative changes. They believed that there was a direct relationship between the degree of degenerative change and the amount of meniscus removed. McGinty et al reported early return to function with decreased morbidity and decreased complication rates after partial meniscectomy; this was compared to both open and arthroscopic techniques for total meniscectomy. Jackson and Dandy have documented improved results of partial meniscectomy when compared with total meniscectomy in an intermediate range (average 5 to 10 years) follow-up study. Earlier studies by the senior author, Hamberg et al, and Cassidy and Shaffer have documented that meniscal repairs can heal. The more recent follow-up study of DeHaven et al has documented that repaired menisci not only heal satisfactorily but that durability and biomechanical function can also be maintained. Of course, long-term review of these same patients will be necessary to prove that successful repair will prevent the degenerative changes noted in knees following meniscectomy.

Iodide and albumin kinetics in normal canine wrists and knees

The clearance rates of free iodide and of radioiodinated serum albumin were measured in the knee and wrist joints of 9 normal adult dogs. Iodide clearance from the knee was 3 times greater than that from the wrist. In contrast, radioiodinated serum albumin clearance from the knee was only slightly greater than that from the wrist. Interpreted as respective indices of effective synovial plasma flow and lymphatic drainage, these values indicate that the filtration fraction is normally greater in microvessels of the wrist than in those of the knee. These findings complement the results of companion studies of Starling forces that indicate a higher pressure microvascular bed in the wrist than in the knee.

Indentation stiffness of young canine knee articular cartilage—Influence of strenuous joint loading

The indentation stiffness of knee articular cartilage subjected to strenuous physical training (SPT: treadmill running 20 km day −1 for 15 weeks, n = 6) of young Beagles was tested and compared to that obtained from age-matched (55 weeks, n = 9) controls. The mathematical solution for the shear modulus, as determined from indentation of an elastic layer bonded to a rigid half space, was extended to small Poisson's ratios and applied to the analysis of cartilage response after a step stress (0.39 MPa) application. In these measurements with an impervious, plane-ended indenter, the equilibrium deformation was systematically greater than values predicted from the instant response by the linear biphasic theory. Therefore, the accurate determination of Poisson's ratio from the creep curves was not possible. The mean shear modulus (calculated by using the deformation at 900 s after load application and assuming a constant Poisson's ratio of 0.40 for the matrix) of canine knee articular cartilage was 0.37 MPa. While the cartilage thickness was not affected by SPT, the cartilage of the lateral tibial plateau was stiffer (13.3%, p<0.05) than that in controls. However, in the femoral condyles, the stiffness was at the control level or even below. Our results on cartilage structure and properties suggest that SPT, in contrast to our previous findings with moderate training, does not necessarily improve the biological properties of articular cartilage in young animals.

Bone ingrowth into the tibial component of a canine total condylar knee replacement prosthesis.

The purpose of this study was to evaluate the occurrence of bone ingrowth in a porous coated tibial component of a canine total condylar design knee replacement model. The entire undersurface of the tibial prosthesis was covered by a titanium fiber metal porous composite pad. Projecting from this surface were three short, cylindrical, fiber metal coated pegs that along with a posterior screw provided initial stabilization of the device. Left total knee replacements were performed on six dogs using alignment and cutting jigs to prepare the bony surfaces. The dogs were killed 6 months following surgery. Extensive bone ingrowth was present in all pegs of every tibial component. Bone ingrowth was present in 12% of the pad area of one component, 22.7 to 41.6% in four components, and 81.1% in another. The void spaces in other areas were filled with fibrous tissue or in the periphery of the device with fibrocartilage. Bone ingrowth in the pad was consistent in the vicinity of the pegs and was variable in regions not adjacent to the pegs, suggesting that the pegs exerted a strong influence on the pattern of bone ingrowth into the pad.

Experimental models to promote healing of tears in the avascular segment of canine knee menisci.

Longitudinal tears were created in canine lateral menisci and techniques were applied to induce healing by removal of a core of tissue from the periphery of the meniscus to the tear or by implantation of a vascularized synovial flap into the tear. The meniscal tears did not heal in knees that were not immobilized, and they healed poorly and sporadically in knees that were immobilized in a cast but bore some weight. However, a higher percentage of tears that were treated by the core-removal or synovial-flap technique healed when the knee was firmly immobilized and weight-bearing was prevented by the use of an external skeletal fixator across the joint for eight to twelve weeks. Neovascularization and access to a source of reparative cells appear to be important in the healing process.

Histological and biomechanical assessment of articular cartilage from stored osteochondral shell allografts

Normal and stored articular cartilage from the medial tibial plateaus of mature canine knee joints were evaluated histologically and biomechanically. The medial plateaus from the right knee (control) were assessed fresh, while the left (stored) were preserved in culture media at 4 degrees C for 3, 7, 14, or 28 days and then evaluated. Biomechanically, confined compression tests were performed on all specimens to determine the aggregate modulus and apparent permeability of the articular cartilage. Histologically, Safranin O- and hematoxylin and eosin (H&E)-stained sections were evaluated. All stored cartilage specimens had an aggregate modulus on average lower than normal, but the differences were not significant (p greater than 0.10). The apparent permeability was on average higher than but also not significantly different from normal (p greater than 0.10). Time in storage (up to 28 days) did not have a significant effect on the biomechanical properties of stored cartilage normalized by control values (p greater than 0.50). Safranin O and H&E histological evaluation also showed no overall changes in cell appearance or staining of the stored cartilage when compared with control for the time periods studied.

Suppression of glycosaminoglycan synthesis by articular cartilage, but not of hyaluronic acid synthesis by synovium, after exposure to radiation.

We recently found that injection of 2 mCi of yttrium 90 (90Y; approximately 23,000 rads) into normal canine knees stimulated glycosaminoglycan (GAG) synthesis by femoral condylar cartilage. The present investigation was conducted to determine whether radiation affects cartilage metabolism directly. Rates of GAG synthesis and degradation in normal canine articular cartilage were studied following irradiation. Cultured synovium from the same knees was treated similarly, to determine the effects of irradiation on hyaluronic acid synthesis. Twenty-four hours after exposure to 1,000 rads, 10,000 rads, or 50,000 rads, 35S-GAG synthesis by the cartilage was 93%, 69%, and 37%, respectively, of that in control, nonirradiated cartilage. The effect was not rapidly reversible: 120 hours after exposure to 50,000 rads, GAG synthesis remained at only 28% of the control level. Autoradiography showed marked suppression of 35S uptake by chondrocytes after irradiation. Cartilage GAG degradation was also increased following irradiation: 4 hours and 8 hours after exposure to 50,000 rads, the cartilage GAG concentration was only 66% and 54%, respectively, of that at time 0, while corresponding values for control, nonirradiated cartilage were 90% and 87%. In contrast to its effects on cartilage GAG metabolism, radiation at these levels had no effect on synovial hyaluronic acid synthesis.

Evaluation of Dacron-covered and Plain Bovine Xenografts as Replacements for the Anterior Cruciate Ligament

Surgical repair of the anterior cruciate ligament often involves the use of a suitable autograft. As alternatives to sacrificing these normal structures, various allografts, xenografts, and synthetic materials have been investigated as ligament replacement materials. This study investigates Dacron fabric-covered and plain bovine xenograft tendon as such materials in the canine knee. The implants were tested to failure in an MTS machine following 13 weeks of implantation in a canine knee. Dacron woven fabric-covered implants became more firmly attached than those covered by Dacron mesh fabric or plain xenografts. The implants were also analyzed according to their method of attachment (fixation staples or sutures). Overall, the sutured implants failed at slightly higher forces than did the stapled ones. Histologically, limited vascular invasion of the xenograft was observed. No host fibrous or osseous tissue could be identified within the graft. Fibrous tissues did form between the bone and xenograft. The implants exhibited extreme intraarticular wear, which suggests a low potential for intraarticular ligament replacement.

Biomechanical properties of the canine knee articular cartilage as related to matrix proteoglycans and collagen.

The instant, creep and equilibrium responses of canine knee articular cartilages were determined after a constant load application with an in situ indentation creep test and related to the chemical composition of the tissue. Instantly, the cartilage stiffness correlated inversely with the proportion of proteoglycans (PGs) extractable with guanidium chloride. The tibial cartilage, rich in PGs but relatively poor in collagen, showed a low resistance to instant rearrangement of the solid matrix after load application. However, the resistance of the tibial cartilage to water flow during creep deformation was similar or even higher than in the femur. The rate of creep correlated inversely with the PG content. The equilibrium modulus of the femoral cartilage (0.40 MPa), 29 per cent higher than in the tibia (0.31 MPa), was related to the content of PGs, while in the tibia the direct correlation between PGs and modulus was not observed. Our results suggest that while PGs control the fluid flow in articular cartilage, a high PG content alone does not guarantee high stiffness of the cartilage. Instead, the properties of the collagen network are suggested to control particularly the instant shape alterations of the articular cartilage under compression.