The distribution and function of elastin and elastic fibres in the canine cruciate ligament complex

Anterior cruciate ligament rupture (ACL) is a major source of morbidity in the dog, leading to severe osteoarthritis of the knee joint and marked lameness. Following rupture, the ACL will not heal and in the dog, ACL rupture is thought to be the end stage of degenerative ligament disease (non-contact ACL injury). The extracellular matrix (ECM) of CLs has been extensively studied but little is known of the role of elastic fibres in the physiology of the ECM, the mechanics of CL function and in CL degeneration. Elastic fibres include polymers of fibrillins (microfibrils), bundles of microfibrils (oxytalan fibres) and elastin fibres (bundles of microfibrils with an elastin core). The hypothesis of this thesis is that elastin has a mechanical and a biological role in the canine cruciate ligament complex. It is further hypothesised that the distribution and function of elastic fibres will vary between three breeds of dog with differing risk of ACL rupture are: the greyhound with a low risk, the beagle with a low-to-moderate risk and the Labrador retriever with a high risk. The distribution of elastic fibres, fibrillins and cells was investigated throughout the CL complex using a combination of histochemical staining and immunofluorescence. CL microanatomy was studied using Nomarski differential interference microscopy. Elastin was measured biochemically and compared to histologic assessment of tissue architecture, elastic fibre staining and other biochemical parameters. The biological effect of elastin degradation products (EDPs) was assessed in an in vitro ACL cell culture model. A low risk dog breed to ACL rupture (greyhound) was used in all investigations and comparisons were made with other breeds with regard to cellular and elastic fibre anatomy. Differences in cell morphology between breeds with differing risk of ACL rupture may reflect fundamental differences in CL physiology possibly through altered cell-to-cell communication. Cellular and matrix changes, considered degenerative, were seen throughout the CL complex and may reflect adaptation rather than degeneration in certain dog breeds such as the greyhound. Elastin content ranged from 5.9 to 19.4% of ligament dry weight. This was a far greater proportion of canine CLs than previously. Elastin fibres may have a mechanical role in bundle reorganization following ligament deformation. The distribution of fibrillins 1 and 2 was different from the pattern previously reported in tendon and may represent a fundamental difference between ligament and tendon. In the greyhound CL there was a significant proportional increase in oxytalan fibre staining with advancing CL degeneration. This response was seen also in the Labrador retriever and the beagle but the increase in oxytalan fibre staining was less marked with advancing degeneration. Therefore production of oxytalan fibres may reflect a healing response in damaged CL tissue in breeds at a low risk of ligament rupture. Fragments of elastin containing the VGVAPG motif affect canine ACL cells in vitro resulting in increased transcription of fibrillin 2 mRNA. Additionally, there was synergism with TGF-β1 resulting in upregulation of the elastin laminin receptor 1, through which EDPs are transduced. EDPs may thus have a role in response to injury in the CL.