Update on the aetiopathogenesis of canine cranial cruciate ligament disease

Vet Comp Orthop Traumatol 2/2011

91 © Schattauer 2011Review Article

Update on the aetiopathogenesis of canine cranial cruciate ligament disease E. J. Comerford1,2; K. Smith1; K. Hayashi3 1Department of Musculoskeletal Biology, Institute of Aging and Chronic Disease, University of Liverpool, Neston, UK; 2School of Veterinary Clinical Science, University of Liverpool, Neston, UK; 3JD Wheat Veterinary Orthopedic Research Laboratory, the Department of Surgical and Radiological Sciences, School of Veterinary Medicine, University of California-Davis, Davis, CA, USA.

Keywords Canine cranial cruciate ligament disease, epidemiology, genetics, metabolism

Summary Cranial cruciate ligament disease (CCLD) is the most common cause of hindlimb lame-ness in the dog, being associated with and eventually leading to stifle osteoarthritis. Ca-nine cranial cruciate ligament disease is a gradual degeneration of the ligament extra-cellular matrix (ECM) leading to ligament rup-ture. The aetiopathogenesis of this condition is still poorly understood but several risk fac-tors have been identified such as breed, body-weight, gender and conformation. Recent de-velopments in this area include the role of genetics, stifle joint conformation, ligament ECM metabolism, and inflammation associ-ated with immune-mediated disease within the stifle joint. A genetic mode of inheritance

Correspondence to: Dr. Eithne Comerford, MVB, PhD, Diplomate ECVS, MRCVS Department of Musculoskeletal Biology Institute of Aging and Chronic Disease, and School of Veterinary Clinical Science University of Liverpool Neston, CH64 7TE United Kingdom Phone: +44 151 7956100 Fax: +44 151 7956101 E-mail: [email protected]

Vet Comp Orthop Traumatol 2011; 24: 91–98 doi:10.3415/VCOT-10-04-0055 Received: April 6, 2010 Accepted: October 27, 2010 Pre-published online: January 18, 2011

has been demonstrated in the Newfoundland which is predisposed to CCLD. Increased cellu-lar metabolism within the cranial cruciate ligament has been directly associated with in-creased craniocaudal stifle joint laxity in dog breeds at high risk of CCLD. Conformation ab-normalities, such as a narrowed distal femoral intercondylar notch, in high risk breeds have been shown to be associated with alterations in cranial cruciate ligament ultrastructure. In-creased production of inflammatory cyto-kines, such as cathepsins and interleukins, by the stifle synovial cells may occur secondary to or may be an inciting cause of ligament de-generation. Future research endeavours will focus on the association between immune-mediated response and fibrocartilaginous metaplasia and matrix degradation within the cranial cruciate ligament, and whether this can be altered in all susceptible dogs or only certain breeds.

Introduction The exact aetiopathogenesis of canine cranial cruciate ligament disease (CCLD) is undefined and controversial. Canine CCLD is recognised as degeneration of the cranial cruciate ligament (CCL) extracellu-

lar matrix (ECM), eventually leading to ligament rupture secondary to a non-con-tact injury; this is a similar situation to that found in female athletes (1). Although acute CCL rupture does occur with trauma, a number of previous studies have sug-gested that the majority of CCL ruptures

are secondary to CCLD (2, 3). In this review we will only discuss the aetiopathogenesis of degenerative non-contact CCL rupture, unless stated otherwise.

Several risk factors (breed, body weight, and neutering) have been proposed for CCLD in epidemiological studies (4, 5). Other factors, such as conformational vari-ation, inactivity, and obesity have also been associated with CCLD (6, 7). More recently the role of genetics, inflammatory factors and ECM changes within the CCL have been proposed as contributing to the aetio-pathogenesis of CCLD (8–10). In this ar-ticle we therefore aim to review recent studies on epidemiology, cruciate ligament physiology and stifle joint factors relating to canine CCLD with a view to consolidat-ing advances in this controversial area.

Epidemiology

Age

Although any breed may be affected by CCLD, increased prevalence in certain breeds of dog has been well established (3, 11, 12). A study of 1.25 million dogs over 40 years showed the five most commonly af-fected breeds to be Newfoundland, Rottweiler, Labrador Retriever, Bulldog and Boxer (5). Bilateral disease is common in affected breeds with 50% of Labrador Retrievers with unilateral CCL tears rupt-uring the contralateral CCL within one year (13). Certain Terrier breeds such as the West Highland White Terrier are also over-represented (5, 14).

Recent studies have reconfirmed aetiopa-thogenic links between breeds and CCLD in breeds such as the Labrador Retriever, New-foundland and Rottweiler (3, 5).

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Vet Comp Orthop Traumatol 2/2011 © Schattauer 2011

92 E. J. Comerford et al.: Review of recent literature on canine cranial cruciate ligament disease

Genetic predisposition and risk factors

Breed-specific variation in incidence of CCLD has been considered strong evidence for a genetic basis to the condition (3, 4). A heritability of 0.27 for CCLD in New-foundlands has been proposed, suggesting a possible recessive mode of inheritance (15). However in this study CCLD had a penetrance of 51% suggesting environ-mental factors as well as genetic factors in-fluence the incidence of CCLD (15). A heri-tability estimate of 0.28 has also been ident-ified in a cohort of Boxers (16). No statisti-cal association of certain candidate genes (such as fibrillin, collagen type IX, and car-tilage oligometrix matrix protein) with CCLD and rupture in Newfoundlands has been identified to date (17). It has also been reported that collagen type IX genes are un-likely to play a role in CCLD in Boxers (18). Genotyping for 532 microsatellite markers has been recently performed in a group of Newfoundlands with and without CCL rupture. Four microsatellite markers as-sociated with CCL rupture were found in chromosomes 3, 5, 13 and 24 and there was significant trait association with CCLD in all of these microsatellite markers except 24. Further mapping of these regions may narrow the list of CCLD candidate genes (8).

A recent study examined Labrador Re-trievers and Greyhounds with OA second-ary to elbow dysplasia, hip dysplasia and CCLD for common genomic risk. Al-though 113 single nucleotide polymor-phisms in 20 candidate genes were geno-typed, no significant associations were identified (19). While this outcome would suggest that these conditions arise inde-pendently, it was proposed that the lack of accurate trait mapping significantly re-duced the power of this study.

Large breed dogs at greater risk of CCLD appear to develop it at a younger age (3). Witsberger et al calculated the relative risk for age, demonstrating that dogs older than four years were significantly more likely to develop CCL rupture (5). Another study examining a population of 328 dogs found that the mean age of CCL rupture in large breeds (body weight >15 kg) was 5.5 years, whereas the smaller breeds (body weight <

15 kg) averaged 7.4 years (20). Identifica-tion of genetic associations with CCLD in predisposed dog breeds is fundamental to the progress of discovering interventions for the problem in dogs and humans.

Gender, neuter status and hormonal influence

The prevalence of CCL rupture is higher in neutered dogs, particularly females (21–23). Neutered female dogs have the highest occurrence of CCLD according to recent studies, and they may experience in-creased weight gain and body fat composi-tion (24). In women, increased incidence of anterior cruciate ligament rupture is as-sociated with elevated oestrogen levels in the pre-ovulatory phase of the menstrual cycle (1). However, as ovariohysterectomy in dogs is associated with persistent hypo-oestrogenaemia, oestrogen may reduce the incidence of CCLD in the dog (25). There have been not been any conclusive studies determining the effect of systemic or local hormonal status on CCLD in dogs or hu-mans.

In many species, including the dog, white adipose tissue has been shown to be an important endocrine organ, elaborating an array of chemical mediators (adipo-kines) which may be pro-inflammatory (26). In humans, adipokine (adiponectin, leptin and resistin) release from intra-ar-ticular adipose and other joint tissues has been demonstrated (27). It is hypothesised that the adipose tissue of obese individuals is in a state of chronic inflammation, sug-gesting systemic or local adipokine elabor-ation, or a combination of both could be involved in the aetiopathogenesis of con-nective tissue pathology (28, 29). However, key pro-inflammatory adipokines have not been assessed and, a possible role for sys-temic adipose tissue has not been examined in CCLD and requires further investi-gation.

Exercise

An ability to sense and respond to mechan-ical force is a fundamental feature of all liv-ing tissues, including tendons and liga-

ments (30). The CCL fibroblasts appear to respond to their mechanical environment, and mechanical force may be required for cell arrangement (31, 32). Overstimulation of tendon cells through repetitive loading may initiate a degenerative cascade leading to tendinopathy (33, 34). However, under-stimulation of tendon cells can also pro-duce a pattern of catabolic gene expression that results in loss of tendon material prop-erties (35, 36). Such understimulation may arise from alterations to the cell-matrix in-teraction resulting from fibril damage at extremes of physiologic loading, and has been proposed as a mechanism for tendin-opathy (37). In the CCL, compromise of cell-matrix coupling through fibril micro-damage or cell morphology changes may lead to understimulation of fibroblasts and a degenerative cascade. There is a paucity of research examining the effect of exercise on the biochemical properties of the canine CCL. An experimental study examining the effect of regular consistent exercise on ca-nine stifle joints, its associated ligaments and cartilage did not find any injuries in these tissues or any evidence of erosions or osteophytes in the joints after lifetime exer-cise (38).

The results of recent studies suggest that inactivity may predispose tendon and liga-ments to altered and detrimental home-ostasis. However it does not appear that ex-ercise influences articular and periarticular tissue damage of canine stifle joints, and therefore it should be recommended in a regular and consistent regime.

Cruciate ligament factors

The cruciate ligaments are comprised of cells and ECM, and are mainly (60–80%) water. Of the dry weight of cruciate liga-ments, 90% is collagen (mostly type 1), with smaller amounts of elastin, proteog-lycans, glycoproteins and lipoproteins (39).

Cells and blood supply

Changes in cell phenotype and reduction in density preceding CCL rupture have been described previously (11, 40). Such changes are more common in the middle of the



© Schattauer 2011 Vet Comp Orthop Traumatol 2/2011

CCL; the common site of rupture (41). Also described is decreased cellularity in the core of ruptured CCL, with a chondroid transformation of cells and extensive dis-ruption of the ligamentous matrix (42). Loss of cells, whether through apoptosis or necrosis, may result in failure to maintain ECM integrity if they are not replaced. Using caspase-3 as a marker for apoptosis, significantly more apoptotic cells were seen in ruptured CCL compared to intact CCL (43). Another study using combined markers for caspase-3 and poly (ADP-ri-bose) polymerase did not find any differ-ence in the amount of apoptotic cells be-tween the intact areas of partially ruptured CCL, torn portion of partially ruptured CCL, and completely ruptured CCL (43). In addition, there was not a significant cor-relation between the degree of synovitis or osteophyte production and apoptotic cells (44). These studies suggest that apoptosis may be an intrinsic aetiopathogenic factor leading to CCLD rather than a con-sequence of acute rupture of the ligament. It remains unclear whether apoptosis ob-served in this study is induced extracellul-arly by extrinsic inducers such as inflam-matory mediators and nitric oxide, or in-tracellularly by intrinsic inducers such as hypoxia and nutrient deprivation. There-fore intrinsic or extrinsic inducers of cru-ciate ligament cell apoptosis may con-tribute to diminished cell communication, altered metabolic response, and ultimate ligament degeneration. Recently, however, it has been shown that apoptosis of liga-mentocytes in the canine CCL was not in-fluenced by increased nitric oxide produc-tion within the stifle joint (45).

Cells subjected to tensile load have cyto-plasmic processes which may be long and extend in all directions through the col-lagen fibres (46, 47). The detection of gap junctions in association with these pro-cesses suggests the potential for cell-to-cell communication (46). This three-dimen-sional structure has been termed the cellu-lar matrix, and has been described in many tissues including the ovine CCL (47). Re-cent unpublished observations describe cells of similar morphology in intact cru-ciate ligaments of Greyhounds and Labra-dor retrievers (K. D. Smith unpublished data, 2010). Cells frequently had cytoplas-

mic processes which varied in length and were often seen to exceed 100 μm. Processes were frequently shown to contact other cells, extending longitudinally and trans-versely through the cruciate ligaments. Cells with long processes were more com-monly noted in the cruciate ligaments of the Greyhound than the Labrador Retriev-er . These findings suggest the possibility of communication between cells (particularly in the Greyhound) which, if disrupted or absent, may result in altered metabolism of the ECM, ultimately leading to CCLD as seen in high risk dog breeds such as the La-brador Retriever. In summary, alterations in cruciate ligament cell morphology may play a role in CCLD aetiopathogenesis. Dis-ruption or lack of development of cruciate ligament cell cytoplasmic processes with rounding of cell nuclei (‘chondrocytic’ change) may alter communication between cells, possibly affecting ECM metabolism.

Although the CCL has been considered extra-articular due to the enveloping syn-ovial epi-ligament, recently it has been sug-gested that free passage of macromolecule markers from synovial fluid to the sub-stance of the CCL and blood occurs (48). As such free movement exists between the syn-ovial fluid and CCL substance, therefore, reduced intra-articular osmotic pressure, which may occur in OA, may affect blood flow to the CCL.

Extracellular matrix

The ECM changes in ruptured CCL have been well documented (49, 50). More re-cently, these changes have been further characterised biochemically, revealing that ruptured CCL have significantly higher amounts of immature collagen cross-links, total and sulphated glycosaminoglycans, water content and concentration of matrix metalloproteinase-2, compared with intact ligaments (51). These findings suggest in-creased ECM turnover in ruptured CCL. Although the ECM changes may have oc-curred before ligament rupture, it is possi-ble that these observed changes may be part of a reparative process after rupture. With regard to normal CCL, it has been shown recently that macroscopically, normal CCL from dog breeds at a high risk of CCLD

(Labrador Retrievers) compared to those from dog breeds at a low risk (Greyhounds) have altered ECM collagen turnover (in-creased pro-MMP-2) and structural prop-erties (decreased integrity of the collagen triple helices) (52). This study also sug-gested that the different metabolism of the collagenous matrix in the CCL from a high risk breed may be related to greater stifle joint laxity and sub-optimal ligament ma-terial properties (52).

An ultrastructural study of normal CCL in these two breeds of dog revealed that the collagen fibril diameters of Greyhounds were larger than those of Labrador Retriev-ers, reflecting relatively enchanced liga-ment maturity and mechanical properties (10). The same study also revealed in-creased collagen turnover in the Labrador Retriever CCL (10). There were, however, fibrocartilaginous areas discovered within the CCL ultrastructure for both breeds manifested as rounding of cell nuclei and increased staining for proteoglycans. Carti-lage-like tissue is more vulnerable to dis-ruption under normal tensile forces, there-fore any fibrocartilaginous transformation may predispose to CCLD (53). The formation of fibrocartilage does not appear to be disadvantageous to healthy racing Greyhounds as they rarely rupture their CCL, and therefore this cannot be regarded as a purely pathological degeneration in this breed. Therefore it is currently unclear whether fibrocartilaginous change is a adaptive condition in exercising and low risk breeds to CCLD, or a pathologic change (with the fibrocartilaginous change inducing an inflammatory response) in high risk breeds resulting in eventual CCLD.

Stifle joint factors

Mechanics

The material and structural characteristics of the canine CCL have been reported with a view to development of optimal method-ologies of surgical intervention in CCLD (54, 55). However until recently, few studies had compared canine stifle joint stability (craniocaudal laxity) in normal dogs at a high and low risk to CCLD. Wingfield and


others compared stifle joint laxity in Rottweilers (high risk) and Greyhounds (low risk), reporting greater craniocaudal joint laxity in the Rottweiler stifle joint compared to that of the Greyhound at stifle joint angles between 150 degrees and 110 degrees (56). In a related study, structural (load to failure) and material (ultimate stress and tangent modulus) properties were shown to be significantly higher for Greyhound stifle joints during tibial load-ing compared with those in the Rottweiler (57).

These findings were confirmed in an-other study examining intact CCL in two differing dog breeds at risk to CCLD (52). Craniocaudal laxity was significantly greater in the normal Labrador Retriever (high risk breed) stifle joints compared with Greyhounds (low risk breed) and this ap-peared to be related to altered CCL ECM ligament composition as indicated pre-viously. In summary, these findings suggest that altered mechanical properties of high risk CCL occur prior to ligament rupture, contributing to increased stifle joint cranio-caudal laxity and ultimate joint instability.

Proprioception

The proprioceptive functions of the nor-mal CCL are undoubtedly important for prevention of joint damage as well as for postoperative rehabilitation (58). Dener-vation impairs healing in the rabbit medial collateral ligament and alters expression of repair-associated gene mRNA (59, 60). Al-terations in neuromuscular coordination have been implicated in increased ACL rupture rates in women (61). However the contribution of these to CCLD and rupture in the dog is not understood.

Conformational variation

Conformational variation of canine hind-limbs such as a straight stifle joint angle, narrow distal femoral intercondylar notch, and steep tibial plateau slope have been as-sociated with CCLD (62–64). The presence of a straight (hyperextended) stifle joint and narrow intercondylar notch may inflict constant or intermittent impingement and

abnormal compression of the CCL against the intercondylar notch (62). A steep tibial plateau slope, or excessive internal rotation of tibia associated with medial patella lu-xation together with genu varum may also cause increased internal stress and micro-injury of the CCL, leading to CCLD (64). Altered gait as demonstrated by stifle joint kinematics may also contribute to abnor-mal loads on the CCL of dogs with a high risk of CCLD (6) and differences (lower thigh and crus mass and thigh moment of inertia in CCL- deficient limbs compared to normal limbs) in body segment parame-ters (e.g. mass, centre of mass and moment of inertia) have been noted in Labrador re-trievers with and without CCLD (65). More recently, the morphometric char-acteristics of the pelvic limbs of Labrador retrievers with and without CCL deficiency have been described, suggesting that cran-ial angulation of the proximal tibia, excess-ive tibial plateau angle (TPA), and distal fe-moral torsion appeared more likely to be associated with CCLD than femoral angu-lation, tibial torsion, and intercondylar notch stenosis (66).

There have been numerous studies evaluating the association of TPA and CCL rupture (14, 64, 67–70). Although ana-tomical differences in the shape of the proximal tibia have been documented in dogs with CCL rupture, its role in CCLD is unclear (70). The mean TPA in dogs varies between 23° and 25°, while a wide range of TPA has been reported (13° to 34°) in some studies (70). Although the correlation be-tween CCL rupture and excessive increases in TPA (TPA >55°), possibly secondary to growth plate injuries, seems established, the association between TPA in the aetio-pathogenesis of CCLD in a normal canine population remains controversial (64). Studies have shown that the TPA are not significantly different in Labrador Retriev-ers with and without CCLD, or between Greyhounds and Labrador Retrievers (69, 70). The true effect of TPA on CCL stresses in vivo is currently unknown, because mus-cular force, body size, obesity, rapid weight gain, relative inactivity, and exercise can in-fluence the amount of stress sustained by the CCL, in addition to the TPA (6).

Intercondylar notch stenosis and its as-sociation with CCL and ACL rupture in dogs

and humans respectively has been well de-scribed (71–73). The intercondylar notch width indices were found to be greater in the stifle joints of low risk dogs (Greyhounds) compared to those of high risk (Labrador and Golden Retrievers) (62). The authors concluded that impingement by the inter-condylar notch on the CCL of high risk breeds may result in altered biochemical composition and reduced structural integ-rity of the ligament, predisposing the liga-ment to increased laxity and degeneration. Guerrero et al. evaluated the effect of con-formation of the distal portion of the femur and proximal portion of the tibia on CCL rupture, and reported that no anatomical differences were detected in the distal por-tion of the femur between dogs with and without CCL rupture (74). The authors pro-posed that development of the tibial tube-rosity and shape (convexity) of tibial con-dyles may be relevant in the pathogenesis of CCL rupture. Tibial tuberosity con-formation may also be a risk factor for CCL rupture, with a recent study reporting that smaller tibial tuberosity widths were associ-ated with increased cranial tibial thrust, which yielded propensity for more rapid de-velopment of CCLD, thus leading to rupture in a younger population of dogs (75).

Subchondral bone pathology

Increased femoral and tibial subchondral bone stiffness may be involved in the pa-thogenesis of OA (76). However, published and preliminary data suggests that sub-chondral bone stiffness preceding knee joint laxity and OA does not occur in guinea pigs (77, 78). Altered joint balance, as reflected in increased tibial subchondral bone mineral density, has been demon-strated in breeds deemed at high risk of CCLD compared to dogs at low risk to CCLD (E. J. Comerford 2010, unpublished data). Gait asymmetry and “handedness” in the dog may lead to this imbalance in high risk dogs contributing to increased sub-chondral bone mineral density and joint degeneration (E. J. Comerford 2010, un-published data). Recently, magnetic reson-ance imaging studies suggested the signifi-cant role of subchondral bone lesions in CCLD mechanisms (79, 80). It is therefore




currently unclear whether altered sub-chondral bone mineral density plays an ae-tiopathogenic role in CCLD or may be in-trinsic to evolution of the disease process in dogs or in humans

Inflammation and immunity

The hostile intra-articular environment created by arthropathies such as immune-mediated arthritis, immune synovitis and joint sepsis may result in CCLD (81–84). Some authors have suggested that there is an immunologic component to rupture of the CCL, because of the demonstration of immune-complexes in synovial fluid and sera and immunoglobulin (mainly IgM) in the synovial membrane (83–85). A recent study evaluated anticollagen type I anti-bodies in synovial fluid of the affected stifle joint and the contralateral stifle joint of dogs with unilateral CCL rupture, and con-cluded that synovial fluid antibodies against collagen type I alone do not initiate CCL rupture (86).

There is also considerable interest in the pro-inflammatory cytokines and their role in the catabolic processes occurring in pathological connective tissues, but their role in the pathogenesis of CCLD is still unclear (85–87). The expression of mRNA of cytokines in synovial fluid cells from multiple joints in dogs with unilateral CCL rupture was measured and this revealed that that IL-8 expression tended to be higher in stifle joints that will sustain rup-ture of the CCL during the subsequent six months by comparison with those joints where rupture will not occur (87). Collage-nolytic enzyme expression has been found in the ruptured CCL and synovial fluid, and synovial macrophage-like cells that produce matrix-degrading enzymes have been identified (50, 88, 89). These findings suggest that inflammatory process may predispose to CCLD, by release of inflam-matory mediators and proteolytic enzymes during inflammatory process. Release of collagenolytic proteases from the synovium into the stifle synovial fluid can significantly degrade the structural properties of the CCL and increase the likelihood of a pathological rupture (50, 88, 89).

In a recent study, a rat model of ACL in-jury demonstrated increased collagenolytic enzymes in synovial fluid (90). Of all tis-sues within the joint, the synovium con-tributed most to this increase. Thus inflam-mation of the epiligament may have an in-fluence on the physiology of the cruciate ligaments through release and mediation of growth factors and cytokines and release of collagenolytic enzymes. Furthermore, as the epiligament does not appear to form a barrier to macromolecule passage between synovium and CCL, these molecules may act directly on the ligament (48).

The role of bacteria and associated syn-ovial inflammation in the aetiopathogen-esis of CCLD has also been speculated (82). Muir and others investigated the role of mixtures of bacterial nucleic acids in the pa-thogenesis of arthritis in naturally occur-ring CCLD using PCR, compared with that of normal stifles and stifles with experi-mentally induced CCL rupture (91). The presence of bacterial DNA within the syn-ovium was significantly associated with naturally occurring CCL rupture and DNA from environmental bacteria was only found in dogs with the naturally occurring CCL rupture. The authors hypothesise that mixtures of bacterial DNA are an important causative factor in the pathogenesis CCL rupture. In addition, Muir et al. determined expression of a panel of immune response genes and matrix turnover genes in synovial fluid collected from dogs with CCL rupture, and suggest that antigen-specific immune responses within the stifle joint may be in-volved in the pathogenesis of persistent syn-ovitis and associated joint degradation in dogs with degenerative CCL rupture (82).

Despite these recent findings, it still re-mains greatly controversial whether in-flammatory changes are primary contribu-tors or are secondary phenomena in re-sponse to the tissue damage during CCL rupture, stifle instability and OA. To date little work has investigated for evidence of these changes in normal CCL from dogs at a high risk to CCLD.

Diminished healing capacity

Poor healing in the CCL has been well documented (39, 92). The canine CCL, in

comparison with the medial collateral liga-ment, demonstrated the lack of provisional scaffold (fibrin-platelet plug) formation as well as reduced levels of key ECM proteins and cytokines within the wound (93, 94). The failure of the provisional scaffold to form in the CCL may result from increased urokinase plasminogen activator within the joint following trauma, a response thought to prevent arthrofibrosis in hu-mans (95). This premature loss of the pro-visional scaffold has been proposed as a mechanism for failure of the CCL to heal (96).

The inorganic free radical nitric oxide has also been a topic of recent research. Tis-sue concentrations have been shown to in-crease in CCL compared to other articular stifle ligaments and nitric oxide has also been demonstrated in ruptured CCL (97, 98). Nitric oxide may inhibit collagen and PG production, and thus contribute to poor healing in humans (99). The canine CCL has been shown to produce more ni-tric oxide in vitro when exposed to an indu-cible nitric oxide synthase cocktail than ex-plants of the canine medial collateral liga-ment or ligament of femoral head (100). However, in the same study a correlation between matrix metalloproteinase produc-tion and nitric oxide was not demon-strated, and its role in CCL physiology is largely unknown.

Other mechanisms proposed for poor CCL healing include deficiencies in stimu-lation or intrinsic deficiencies of cell mi-gration and proliferation (101–104). Inad-equate blood supply may also contribute to the development of CCL rupture (11, 41, 49, 92, 105, 106). An in vitro model of CCL injury showed increased matrix metallo-proteinase expression, and a 6.3 fold in-crease in matrix metalloproteinase-2 activ-ity in the CCL compared to the medial col-lateral ligament. This may contribute to poor healing of the CCL by comparison with the medial collateral ligament which heals readily (90).

Conclusion

In summary, the aetiopathogenesis of ca-nine CCLD is unknown, but it certainly ap-pears to be a multifactorial condition re-




sulting in overall stifle joint ‘organ’ failure (107). One working hypothesis is that gen-etic and breed factors may contribute to ab-normal stifle conformation, kinematics and gait, which then in turn may lead to cy-tokine and protease release from stifle joint synovial fluid and membrane, and possibly the CCL itself. However abnormal mech-anics such as altered periarticular stifle joint conformation may also result in CCL compression at extremes of joint motion leading to structural changes within the ligament ECM (abnormal biology) which in turn can contribute to ligament laxity and joint instability. The ECM composi-tional changes (such as increased fibrocartilage) may in turn induce an inflamma-tory reaction, switching on an inflamma-tory cascade within the stifle joint con-tributing to CCLD and ligament rupture. Whether abnormal biology or mechanics initiate CCLD as has been discussed by Cook, the authors of this review agree that both factors play a significant overlapping role in the progression of stifle joint OA and ultimate joint failure (107).



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Documents

Update on the aetiopathogenesis of canine cranial cruciate ligament disease