COMPONENTE INSTITUCIONAL

Although the OA disease process is distinctive from normal ageing, the relationship between age and OA is important. Not only is age a major risk factor for OA but treatments aimed at delaying cartilage ageing provide potential therapeutics. The prevalence of the OA increases with age with between 30 and 50% of adults over 65 years experiencing the condition (Felson, 2004). However, OA is not an inevitable consequence of ageing (Loeser, 2010). Consistent with the heterogenous nature of OA, ageing is just one (though the greatest) of the many risk factors involved (Suri et al., 2012). The interactions between other OA risk factors and age in ascertaining the sites and severity of OA are illustrated in Figure 1.8.

There have been a number of theories as to why ageing and in particular cartilage ageing plays such a major role in OA pathogenesis. Joint health is dependent on the normal structure and function of all the constituent tissues and OA is a disorder of the entire joint. Cartilage is the most susceptible tissue within the joint to damage and demonstrates the most intense age related changes.

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Figure 1.7. Schematic diagram depicting major human aggrecan cleavage sites. G1, G2, G3 represents the respective globular domains, IGD; interglobular domain, KS; keratan sulphate-rich region, CS-1, -2; chondroitin sulphate-rich regions. Aggrecanase (AGG) and MMP cleavage site sequences in the IGD and CS-2 domains are shown. Numbering of amino acids corresponds with the human sequence.

With advancing age changes occur in both in the ECM and chondrocytes. Mitotic activity and synthetic activity of chondrocytes alter with age whereas in OA chondrocytes are characterised by increased cell activation, proliferation and gene expression (Aigner et al., 2004b). A number of theories postulate as to why age contributes to cartilage degeneration. One established theory is that OA develops due to continuous accruing of repetitive load cycles which instigates constant microtrauma due to physiological loading, resulting in a loss of structure and function (Loeser, 2009). Another theory relates to ECM modifications including collagen and aggrecan with age. Increased cross-linking of collagen over time causes cartilage to stiffen, thus reducing flexibility during physiological deformation. Aggrecan structure also changes with age due to degradation and

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impaired synthesis. The molecule becomes smaller and there is a reduction in sugar side chains (Glant et al., 1998). This affects the ability of aggrecan to bind water and it is this feature that gives aggrecan and therefore cartilage the compressive stiffness required for function. Finally glycation end products increase with age (known as advanced glycation end products (AGEs)) (Verzijl et al., 2002). These are non-enzymatic protein modifications that affect both matrix integrity and chondrocyte biology and therefore affect the mechanical properties of the cartilage.

1.5.1. Mechanisms of ageing in cells and tissues in the development of OA

Three main areas have been demonstrated as contributing to ageing in cartilage in relation to the development of OA. These are cell senescence, ECM ageing and age related oxidative stress.

a. Chondrocyte senescence

The senescence model for ageing theorises that chondrocytes become senescent due to proliferation and/or oxidative cell stress resulting in the inability of chondrocytes to maintain matrix turnover (Aigner et al., 2004b).

There is little evidence of chondrocyte turnover in adult articular cartilage (Martin and Buckwalter, 2001). Although they can divide occasionally, adult articular cartilage is classified as post-mitotic with trivial cell turnover. Consequently they are long lived and disposed to the accumulation of age-related changes over time. There is limited evidence for the existence of progenitor cells in cartilage which would allow senescent cells to be replaced. One study identified mesenchymal progenitor cells in human normal and OA cartilage (Alsalameh et al., 2004) whilst a study in young bovine tissue identified a progenitor cell population on the articular surface (Williams et al., 2010). Additionally equine derived cartilage progenitor cells are capable of functional cartilage repair (McCarthy et al., 2012). Should a local pool of progenitor cells exist they seem unable to replaced senescent chondrocytes.

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Studies have demonstrated cellular degenerations including DNA damage in OA chondrocytes (Helmick et al., 2008) which normally results in apoptosis. The extent of apoptosis is debated. Although some believe it is rare in OA cartilage (Horton et al., 1998) there is evidence for both an age-related (Adams and Horton, 1998) and OA (Morgenroth et al., 2012) induced loss of chondrocytes from apoptosis. There does, in humans at least, appear to be a reduction in chondrocyte number with age progression (Dieppe, 1995).

A reduction in the chromatin protein high mobility box group B2 (HMGB2) in the cartilage superficial zone chondrocytes of animals and man with age occurs which may contribute to chondrocytes death (Taniguchi et al., 2009). The protein is a transcriptional regulator which maintains superficial zone chondrocytes survival through β-catenin signalling and in addition controls gene expression profile of the superficial zone cells. HMGB2 null mice display early-onset OA-like alterations associated with increased susceptibility to cell death (Taniguchi et al., 2009).

Telomere shortening, unrelated to replicative senescence, has been identified in chondrocytes with advancing age (Martin et al., 1997). This is possibly due to oxidative damage or inflammation (Grahame and Schlesinger, 2012). These finding may contribute to chondrocyte senescence in-vivo (Aigner et al., 2004b). There are a number of concepts evident in cell senescence. In classic replicative senescence there is an inability of the cells to experience further cell division. However there is also evidence for phenotypic alterations; the ‘senescent secretory phenotype’ (Campisi, 2005). Accumulation of these cells, which secrete increased amounts of MMPs and cytokines, contributes to cell ageing. Given the enhanced production of cytokines and MMPs in OA this provides a direct link between ageing and OA (Loeser, 2010).

Finally cell senescence has been linked to a reduction in the capacity of chondrocytes to respond to growth factors with age and OA (Campisi, 2005) including insulin growth factor-1 (IGF-I) and TGF-β, which may contribute to the imbalance between anabolic and catabolic activity in OA.

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Since IGF-I has an important autocrine survival role in cartilage this age related reduction may be pivitol in age-related cell death (Loeser and Shanker, 2000).

b. Cartilage matrix ageing

There is evidence of a gradual loss of cartilage matrix with ageing as demonstrated in knee magnetic resonanace imaging (MRI) studies (Connie et al., 2011). This is due to a loss of chondrocytes, reduced growth factor activity, and cartilage water loss. The water content of cartilage is largely controlled by the presence of aggrecan. This changes in its structure, glycosylation extent and size with age (Buckwalter et al., 1994). The age-related matrix protein modifications AGEs have a role in OA development (Loeser, 2009). As increased collagen cross-links are evident as a result of AGEs, cartilage biomechanical properties are effected resulting in susceptibilty to failure (Verzijl et al., 2000b). Thus overall these changes result in collagen that is less flexible, aggrecan that is smaller and less able to ‘hold’ water and altered phenotype leading to alterations in the anabolic/catabolic balance.

c. Age-related oxidative stress

Oxidative damage from chronic formation of reactive oxygen species (ROS) results in age related tissue changes (Carlo and Loeser, 2003). Both ROS and reactive nitrogen species such as nitric oxide are produced by HAC. Furthermore, rat studies identified increased ROS with age (Jallali et al., 2005). Studies have also found increased oxidised glutathione (an intracellular anti-oxidant) with age in human chondrocytes (Jallali et al., 2005). Other anti-oxidants are detected at reduced levels with age (Jallali et al., 2005) and in OA. Whilst increased ROS results in deoxyribonucleic acid (DNA) damage leading to reduced chondrocyte viability and matrix production. Interestingly ROS interferes with IGF-I signalling leading to reduced matrix production (Yin et al., 2009). Further ROS may be stimulated by cytokines such as IL-1 and TNF-α which are elevated in OA. This increase in ROS can then increase MMP production (Forsyth et al., 2005).