Structural and Functional Properties of Tendons
Disclaimer: This dissertation has been submitted by a student. This is not an example of the work written by our professional dissertation writers. You can view samples of our professional work here.
Any opinions, findings, conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of UK Essays.
Tendons are dynamic structures; their extracellular matrices are continuously being synthesised and broken down over the course of an individual’s lifetime. The macromolecules, namely collagen, proteoglycans, hyaluronan and the non-collagenous proteins form the extracellular matrix of tendons. In normal tendon exists a fine balance between the synthesis and degradation of these macromolecules resulting in a strong healthy tendon. It is evident that damage to tendons, such as in overuse tendinopathy results in changes to the levels and types of macromolecules present in tendon with decreased levels of collagen and increased levels of proteoglycans, hyaluronan and non-collagenous proteins, causing a weakened tendon that is prone to rupture.
These degenerative features have thus far been partially characterised. By identifying the levels and various types of macromolecules present in normal tendons and tendons exhibiting overuse tendinopathy an understanding of the basis of the condition can be determined and possible ways of preventing or ameliorating tendon degeneration can be considered. The terms overuse tendinopathy and pathological tendon will be used interchangeably throughout this study.
This literature review will attempt to define and characterise the structural and functional properties of tendon and will discuss the current literature regarding the levels, types, synthesis and catabolism of macromolecules present in the extracellular matrix of tendons and also attempt to define and characterise the pathological aspects of overuse tendinopathies. Chapter Two of this thesis will dictate the materials and methodology used in these studies. Chapters Three, Four and Five will present the results of this thesis. Finally, chapter Six will include the discussion and discuss any limitations and future considerations.
1.1 Synovial Joint
Joints are articulations found between adjacent parts of bone that allow controlled frictionless movement (for review see; Mankin & Radin, 1997). In the human body there are three different types of joints and these are grouped according to the type of movement they make. They include the freely movable joints (synovial joints; i.e., most joints of the extremities such as the knee joint), slightly movable (cartilaginous joints; i.e., the vertebrae and ribs) and those that are immovable (fibrous joints; i.e., the skull). The majority of the joints found in the human body are synovial joints (for review see; Mankin & Radin, 1997).
There are six different types of synovial joints including the ball-and-socket joints, hinge joints, saddle joint, pivot joint, gliding joints and condyloid joints. A synovial joint contains a joint cavity that is enclosed by a fibrous capsule linking the adjoining bones. This joint capsule is lined by a synovial membrane that secretes a lubricating and nutritious fluid called synovial fluid that is rich in albumin and hyaluronan. The surface of each bone is typically covered with articular hyaline cartilage or in some circumstances fibrocartilage. In addition, the joint capsule is supported by accessory structures such as tendons and ligaments, which provide stability to the synovial joint (Sledge et al., 2001).
1.1.1 Articular Cartilage
Articular cartilage covers the adjoining ends of bones in joints and has a white colour (for review see; Mankin & Radin, 1997). It is a tissue that is devoid of blood and nerves and provides a wear resistant surface with low frictional properties for the joint and attains its nutrients via diffusion from the synovium into the synovial fluid (for review see; Mankin & Radin, 1997). Furthermore, articular cartilage is resilient and flexible. This allows articular cartilage to withstand large compressive and tensile forces as well as allowing it to distribute load on subchondral bone during joint loading (Kempson, 1980) even though it is only a few millimetres thick (Hardingham, 1998).
Its biomechanical properties are dependent on the structural composition of the extracellular matrix, which is comprised of water (70-80%), collagens (predominantly Type II collagen), proteoglycans (predominantly aggrecan) and non-collagenous proteins (Kuettner et al., 1991; Poole, 1997). The predominant cell type present in articular cartilage is called the chondrocyte. These cells are responsible for the maintenance, synthesis and degradation of all the extracellular matrix components (Kuettner et al., 1991; Buckwalter & Mankin, 1998).
Mature articular cartilage can be divided up into four zones including the superficial (tangential) zone, the middle (transitional) zone, the deep (radial) zone and the zone of calcified cartilage (Huber et al., 2000). The organisation and composition as well as mechanical properties of the extracellular matrix varies within these zones. The deeper zones have high proteoglycan levels and low cellularity whereas the more superficial zones contain low proteoglycan levels and increased cellularity (Aydelotte et al., 1988; Buckwalter & Mankin, 1998).
1.1.2 Joint Capsule and Ligament
The joint capsule is a fibrous connective tissue that is attached to the skeletal parts of a joint beyond their articular surfaces. The principal function of the joint capsule is to seal the joint space and to supply stability by limiting movement (for review see; Mankin & Radin, 1997). Most joint capsules are strengthened by ligaments. Ligaments act together with the joint capsule and the peri-articular muscles to provide stability to the joint preventing excessive movements. They permit free movements when lax, but can stop unwanted movements when tight by virtue of their high tensile strength.
Occasionally joint capsules are strengthened by tendons, such as the extensor tendon in the finger joint. The joint capsule and ligaments proceed to hold the bones together and to guide and limit joint movements. Ligaments attach one bone with another bone and have a limited vascular and neural supply which enable them to repair relatively well after damage (Bray et al., 1990). The knee joint is a good example of different types of ligaments. The medial collateral ligament fuses with the joint capsule, and the cruciate ligaments and the lateral collateral ligament, which are both completely independent of the joint capsule.
1.1.3 Synovial Membrane
The synovial membrane (synovium) lines the non-articular surfaces of a joint such as the joint capsule and ligaments, and is responsible for secreting and absorbing synovial fluid, which contains hyaluronan (Mason et al., 1999). Synovial fluid lubricates the joint and provides at least partly for the nutrition of articular cartilage, invertebral discs and menisci. The synovial extracellular matrix acts as a scaffolding to support synoviocytes and plays an important role in cell migration and differentiation. It is mostly composed of collagen particularly Type III collagen, with smaller amounts of proteoglycans such as decorin and biglycan (Mason et al., 1999), non-collagenous proteins such as fibronectin, elastin and lamina, hyaluronic acid as well as lipids, serum proteins and electrolytes (Hirohata & Kobayashi, 1964).
The synovial membrane has only been detected in vertebrate animals (Henderson & Edwards, 1987). Furthermore, synovial tissue is not arranged into discrete layers, but rather represents a continuum from surface to deep zones. The extracellular matrix of the synovial membrane varies in composition from its surface to its deep zones (Hirohata & Kobayashi, 1964).
Tendons are dense fibrous connective tissues found between muscles and bones (for review see; Benjamin & Ralphs, 1997). The primary role of tendon is to absorb and transmit force generated by muscle to the bone to provide movement at a joint. In addition tendons operate as a buffer by absorbing forces to limit muscle damage. Each individual muscle has two tendons, one that is proximal and the other distal. The attachment of the proximal tendon of a muscle to bone is called a muscle origin and that of the distal tendon an insertion.
A normal tendon has a bright white colour and a fibroelastic texture and enables resistance to mechanical forces. Tendons come in many shapes and this is most likely due to their function, they can be round or oval in cross section or they can come in the form of flattened sheets, fan shaped, ribbon shaped or cylindrical in shape (for review see; Benjamin & Ralphs, 1997). In a muscle like the quadriceps which creates strong forces the tendons are short and broad, while those that are involved in more delicate movements like the finger flexors, long and thin tendons are present (Kannus, 2000).
Tendons are arranged in a hierarchical fashion (see Figure 1.1). A group of collagen fibres form a primary fibre bundle or subfascicle; this is the basic unit of tendon. A group of subfascicles form secondary bundles or fascicles, which form tertiary bundles constituting the tendon as a whole. The primary, secondary and tertiary bundles are encased in a thin connective tissue reticulum called the endotenon (Elliott, 1965; Kastelic et al., 1978; Rowe, 1985). The endotenon carries blood vessels, nerves and lymphatics to deeper areas of the tendon (Elliott, 1965; Hess et al., 1989). The whole tendon is surrounded by an epitenon, which is a dense fibrillar network of collagen (Jozsa et al., 1991).
The epitenon is contiguous with the endotenon and like the endotenon is rich in blood vessels, nerves and lymphatics (Hess et al., 1989). Many tendons are surrounded by a connective tissue called the paratenon. Paratenon allows free movement of the tendon against the surrounding tissues (Schatzker & Branemark, 1969; Hess et al., 1989). The myotendinous junction is the site of union with a muscle, and the osteotendinous junction is the site of union with a bone (Kannus, 2000).
In tendon, blood vessels represent between 1-2% of the entire extracellular matrix (Lang, 1960; Lang, 1963). Some blood vessels may originate from the perimysium at the musculotendinous junction and blood vessels from the osteotendinous junction (Schatzker & Branemark, 1969; Carr & Norris, 1989; Clark et al., 2000). At rest, rabbit tendons have been shown to have blood flow of around one-third that of muscle, and it is known that blood flow in tendon increases with exercise and during healing in animals (Backman et al., 1991). The oxygen consumption of tendons is 7.5 times lower than that of skeletal muscles (Vailas et al., 1978).
1.1.5 Tendon Extracellular Matrix
The major cell type present in tendon is the fibroblast (also known as tenocytes; Ross et al., 1989; Schweitzer et al., 2001; Salingcarnboriboon et al., 2003), which are embedded within an extracellular matrix (see Figure 1.2). These cells are sparsely distributed, comprising only 5% of the dry weight of adult tendon (Ross et al., 1989; Schweitzer et al., 2001; Salingcarnboriboon et al., 2003). These cells lie in longitudinal rows and have many cell extensions that extend into the extracellular matrix (McNeilly et al., 1996). Fibroblasts are responsible for the synthesis and degradation of all the macromolecular components that make up the extracellular matrix of tendon, including the most abundant macromolecule present in tendon, collagen, as well as proteoglycans, hyaluronan and non-collagenous proteins (Vogel & Heinegard, 1985; Curwin, 1997; O’Brien, 1997).
The extracellular matrix is made up of parallel bundles of collagen aligned longitudinally (60-85% of tendon dry weight) associated with elastin fibres which constitutes approximately 1-2% of the dry weight of tendon (Tipton et al., 1975; Hess et al., 1989; Jozsa et al., 1989; Curwin, 1997; Kirkendall & Garrett, 1997; O’Brien, 1997). Tendon consists of 55-70% water, most of which is associated with proteoglycans in the extracellular matrix (Elliott, 1965; Vogel, 1977; Merrilees & Flint, 1980; Riley et al., 1994b; Vogel & Meyers, 1999). The proteoglycan content of tendons is approximately 1% of dry weight of tendons (O’Brien, 1997).Water and proteoglycans have important lubricating and spacing roles in tendons that allow collagen fibres to glide over one another (Amiel et al., 1984).
The structure, composition and the organisation of the tendon matrix is crucial for the physical properties that tendons posses (Riley, 2004). The collagen component gives tendon its great tensile strength (Scott, 2003) whereas it is the proteoglycan component of the tendon matrix that enables tendons to withstand compressive load (Schonherr et al., 1995), while elastin fibres increase tendon extensibility (Scott, 2003).
1.1.6 Tendon cells
The cell population of tendon has so far been poorly characterised (for review see; Riley, 2000), the majority of tendon cells have the appearance of fibroblasts (also known as tenocytes) and constitute about 90-95% of the cells present in tendon (Ross et al., 1989; Schweitzer et al., 2001; Salingcarnboriboon et al., 2003). The remaining 5% to 10% of cells present in tendon are chondrocyte-like cells (fibrochondrocytes), which are mostly present in the fibrocartilaginous regions of tendon where tendon attaches to bone. Also present in tendon are some mast cells, capillary endothelial cells, smooth muscle cells and nerve cells (Hess et al., 1989; Jozsa & Kannus, 1997).
Fibrocartilage cells are large and have an oval shape and they are often packed with intermediate filaments (Merrilees & Flint, 1980; Ralphs et al., 1991). Tendon cells are linked to one another via gap junctions (McNeilly et al., 1996; Ralphs et al., 1998), allowing cell-to-cell interactions (McNeilly et al., 1996). Fibroblasts have a branched cytoplasm surrounding an elliptical, speckled nucleus. The rough endoplasmic reticulum and the Golgi apparatus are well developed with few mitochondria in the cytoplasm (Ippolito et al., 1980; Moore & De Beaux, 1987). Like other connective tissue cells, fibroblasts are derived from mesenchyme.
It is believed that in tendon there are a small number of mesenchymal stem cells that have the ability to differentiate into chondrogenic, osteogenic and adipogenic cells if the conditions allow (Salingcarnboriboon et al., 2003). Tendons have been shown to respond to mechanical load by modifying their extracellular matrix (Banes et al., 1988; Ehlers & Vogel, 1998; Buchanan & Marsh, 2002; Lavagnino & Arnoczky, 2005). Tendon cells receive their vascular supply from the surrounding paratenon.
Tendons were once considered almost static and unable to participate in repair. However, the activity of tendon cells has been shown to be active throughout an individual’s life as they express various matrix components (Chard et al., 1987; Ireland et al., 2001; Riley et al., 2002). Regional differences in cell morphology and activity exists in tendons, synovial-like cells that are found in the endotenon and epitenon surround the main fibre bundles (Banes et al., 1988). A greater proliferative capacity and a different matrix synthetic activity is characteristic of these synovial-like cells compared to the fibroblasts within the fibres, and are the first cells to respond following acute tendon injury (Gelberman et al., 1986; Banes et al., 1988; Garner et al., 1989; Gelberman et al., 1991; Khan et al., 1996b).
Tendon Extracellular Matrix Macromolecules
The following section will discuss the major extracellular matrix proteins and their roles in tendon. This will include the major constituent of tendon, collagen, the small and large proteoglycans and the non-collagenous proteins as well as hyaluronan. This section will also discuss the synthesis of collagens, proteoglycans and hyaluronan.
Collagen is the most copious protein present in the extracellular matrix of connective tissues and accounts for approximately 90% of the total protein of tendons, or 65% to 75% of the dry weight of tendons (von der Mark, 1981; O’Brien, 1992). There are currently 28 different collagen types (numbered I-XXVIII) present in vertebrates with at least 42 different alpha chains (Veit et al., 2006) with this number continuing to mount (Brown & Timpl, 1995; Aumailley & Gayraud, 1998). Collagen molecules can be defined as an extracellular protein that contains at least one triple helical domain (van der Rest & Bruckner, 1993). Collagen provides the tendon with its structural integrity as well as assisting in various physiological functions.
Collagen consists of three polypeptide alpha chains, which combine to form a homotrimer (three identical alpha chains) or a heterotrimer (two or three different alpha chains). Covalent bonds known as collagen cross-links develop between individual collagen molecules in a collagen fibre (Eyre et al., 1984; Bailey et al., 1998; Bailey, 2001; Brady & Robins, 2001). The collagen arrangement gives tendon its great tensile strength. Cross-links are formed from a pathway of different chemical reactions that result in divalent cross-links that join two polypeptide chains, to multivalent, i.e. tri- or even tetravalent, cross-links (Bailey & Lapiere, 1973; Eyre et al., 1984). These cross-links come about from enzymatic modification of lysine or hydroxylysine residues by the copper-dependent enzyme lysine oxidase (Robins, 1988).
Collagens are divided into two subgroups, the fibrillar and non-fibrillar collagens. Non-fibrillar collagens can be further divided into seven subfamilies including microfibril collagens, fibril-associated collagens with interrupted helices (FACIT) collagens, network collagens, MULTIPLEXIN collagens (proteins with multiple triple helix domains and interruptions), basement membrane-associated collagens, transmembrane-associated collagens and epithelium-associated collagens (von der Mark, 1999). The non-fibrillar collagens present in tendon include Types IV, VI, IX, X, XII and XIV (von der Mark, 1999).
The fibrillar collagens present in tendon include, Types I, II, III, V and XI (Kielty et al., 1993; Kadler et al., 1996; Fukuta et al., 1998; von der Mark, 1999). The fibrillar collagens contain a continuous triple helix domain, 300 nm in length, capable of undergoing the staggered, lateral associations required to form fibrils (Mayne, 1997). The resulting fibrils provide the structural support for tissues. All the fibril-forming collagens have a similar structure and size, being composed of a large, continuous central triple-helical domain (COL1) of approximately 1000 amino-acid residues
Occurs in most tissues, tendon, bone, skin etc
Main component of tendon, skin, bone, dentin, cartilage, ligament etc
Hyaline cartilage, invertebral disc
Restricted to fibrocartilage; forms less-organised meshwork
Vessels, kidney, liver, skin, tendon
Normally restricted to endotenon; forms smaller less organised fibrils
Basement membranes, tendon
Basement membrane of tendon blood vessels
Core of Type I collagen fibril - forms template for fibrillogenesis
Vessels, skin, intervertebral disc
Cell associated - found in seams between fibrils
Forms anchoring fibrils in the skin
Descements membrane in the cornea
Forms a lattice
Hyaline cartilage, vitreous humour, tendon
Cell and matrix interactions with Type II collagen fibril surface
Growth plate, tendon
Restricted to insertion fibrocartilage
Core of Type II collagen fibril - forms template for fibrillogenesis
Embryonic tendon and skin, periodontal ligament
Mediates cell/matrix interactions with Type I collagen fibril surface
Adhesion of cells to basement membranes
Foetal skin, tendon
Mediates cell/matrix interactions with Type I collagen fibril surface
Stabilizes skeletal muscle cells and microvessels
Skin, cornea, lung
Connects epithelial cells to the matrix
Endothelial cells, liver, eye
Needed for normal development of the eye
Forms radially distributed aggregates
Corneal epithelium, skin, cartilage and tendon
Binds to collagen fibrils
Matrix assembly of vascular networks in blood vessel formation
Interacts with components of microfibrils
Metastatic tumour cells, heart retina
Cell adhesion, Binds to heparin
Expressed in tissues containing Type I collagen Developing bone and cornea
Regulating Type I collagen fibrillogenesis
May play a role in adherens junctions between neurons
Testis and ovary of adult tissues
Development of the reproductive tissues
Cartilage, ear, eye and lung
Basement membranes around Schwann cells in the peripheral nervous system.
flanked by a variable amino-terminal domain of about 50-520 amino acid residues and a highly conserved non-triple-helical carboxyl-terminal domain of about 250 amino acid residues (for reviews see; Kielty et al., 1993; Fichard et al., 1995; Pihlajaniemi & Rehn, 1995; Prockop & Kivirikko, 1995; Bateman et al., 1996). The amino- and carboxyl-terminal extensions are commonly referred to as amino- and carboxyl- propeptides, respectively. The C-propeptide is called the NC1 domain, whereas the amino-propeptide is divided into sub-domains. The first is a short sequence (NC2) that links the major triple helix to the minor one (COL2) and a globular amino-terminal end (NC3) that shows structural and splicing variations.
Collagen Types II, IX, X and XI (Fukuta et al., 1998) are present at specific sites within the fibrocartilage region of tendon, found at the bone insertion and where the tendon is subjected to shear forces or compression (Fukuta et al., 1998; Waggett et al., 1998). Collagen Types II, IX, X and XI were once thought to occur only in cartilage (Visconti et al., 1996; Fukuta et al., 1998; Riley, 2000). It has now been shown that these collagens are found in the fibrocartilaginous regions of tendon, which wraps under bone. Their presumed function is to help resist compression and shear forces at these sites (Visconti et al., 1996; Fukuta et al., 1998; Waggett et al., 1998).
Collagen also plays an important role in attaching tendons to bone. Where the tendon attaches to bone, tendons commonly widen and give way to fibrocartilage, a transformation where the aligned fibres originating from the tendon are separated by other collagen fibres arranged in a three dimensional network surrounding rounded cells (Liu et al., 1995). This arrangement helps to transmit tensile forces onto a broad area and reduces the chance of failure under excessive loading. The following review will focus on the collagens that are known to exist in tendon; this includes collagen Types I-VI, IX-XII and XIV.
18.104.22.168 Type I Collagen
Type I collagen is the predominant and most studied collagen type present in the extracellular matrix of tendon, ligament and bone representing approximately 95% of the total collagen content or 60% of the tendon dry weight (Evans & Barbenel, 1975; von der Mark, 1981; Riley et al., 1994b; Rufai et al., 1995). It is synthesized by a number of cell types such as fibroblasts, osteocytes and odontoblasts. Type I collagen consists of two α1(I) chains and a shorter α2(I) chain (Kielty et al., 1993), these two chains are products of separate genes and are not a posttranslational modification of a single molecule (for review see; Kivirikko & Prockop, 1995).
The two α1(I) and one α2(I) chains of a monomer of Type I collagen are primarily comprised of approximately 338 repeating tripeptide sequences of Gly-X-Y in which X is frequently proline and Y is frequently hydroxyproline (OHPr). The ends of the α1(I) and one α2(I) chains consist of short telopeptides of between 11-26 amino acids per chain.
In longitudinal sections, the monomers are arranged in fibrils in a head-to-head-to-tail orientation. Each Type I collagen molecule consists of a long central helical region with a short non-helical domain on both the amino- and carboxyl-terminal ends. In tendon, the Type I collagen-containing fibril, organized into fibres (fibril bundles), is the major element responsible for structure stabilization and the mechanical attributes of this tissue. The fibril contains collagen molecules assembled into a quarter-staggered array, and this striated fibril has a 67 nm periodicity (for review see; Kadler et al., 1996; Orgel et al., 2006).
Each alpha chain consists of a repeating triplet of glycine and two other amino acids marked as (Gly-X-Y)n. It is the glycine residues located in every third position that makes it possible for the three alpha chains to coil around the other. It has a molecular weight of 290 kDa. When viewing collagen fibrils under the light microscope they have a crimped appearance, during tendon loading the crimp stretches and the fibrils become aligned, and after loading the crimp will reappear, this is an important elastic component that tendon possesses (O’Brien, 1992).
The Type I collagen α chains contain approximately 290 residues of OHPr per molecule. Proline and OHPr constitute 20% to 25% of all amino acid residues of Type I collagen. The parallel arranged bundles formed by the Type I collagen fibrils gives tissues a high tensile strength with limited elasticity, and therefore is suitable for force transmission. The Type I collagen molecule has the ability to form microfibrils (filaments) as well as larger units of the fibrils or fibres (for review see; Kivirikko & Prockop, 1995). The diameter of the collagen fibril is usually between 20 nm and 150 nm but can range up to 300 nm, this depends on the stage of development (Dyer & Enna, 1976; Jozsa et al., 1984; Fleischmajer et al., 1988).
22.214.171.124 Type II Collagen
The homotrimeric Type II collagen molecule was first discovered in cartilage by Miller and Matukas in 1969 who extracted collagen from cartilage in an experiment that involved pepsin digestion. Type II collagen, although most commonly found in articular and hyaline cartilage is also expressed in tendon particularly around the fibrocartilaginous region and consists of three identical α1(II) chains (Eyre et al., 1992) which forms a meshwork structure that gives Type II collagen the ability to entrap the negatively charged proteoglycan molecules, thereby resisting the swelling pressure of proteoglycans. Each Type II collagen chain has a molecular weight of approximately 95 kDa.
The entire collagen Type II molecule is shaped like a thin rod and is 300 nm long and 1.5 nm wide and has a total combined molecular weight of 295 kDa. This molecule is essential in connective tissues that are subjected to compression such as tendon and articular cartilage. Type II collagen molecules consists of a long central helical region flanked at its amino- and carboxyl-terminus by short non-helical regions termed amino and carboxyl telopeptides (Eyre et al., 1992). As with all fibrillar collagens, Type II collagen molecules are arranged in a quarter-staggered array to form collagen fibrils. Lateral associations of these collagen fibrils forms collagen fibres (Mayne, 1997). In tendon, collagen Types IX and XI as well as the proteoglycans decorin, fibromodulin and lumican inhibit collagen Type II fibril formation reducing fibril thickness (Vogel et al., 1984; Hedbom & Heinegard, 1989; Hedbom & Heinegard, 1993).
126.96.36.199 Type III Collagen
Type III collagen is the second most abundant collagen present in tendon, representing up to 10% of the total collagen content in various tendons (Hanson & Bentley, 1983; Riley et al., 1994b). Type III collagen is a thin collagen fibre consisting of three α1(III) chains with a molecular weight of 290 kDa. In tendon most Type III collagen is found in the endotenon and epitenon (Duance et al., 1977), and is also found in between Type I collagen fibril bundles in aging tendons and at the insertion (Kumagai et al., 1994). It can also be found in skin, blood vessels, ligament and internal organs such as the gastro-intestinal tract but is not found in bone (Epstein & Munderloh, 1978; McCullagh et al., 1980; Amiel et al., 1984). It strengthens the walls of hollow structures like the intestines and uterus.
The fibrils of Type III collagen have a generally thinner diameter compared with Type I collagen fibrils (Lapiere et al., 1977; for review see; Kadler et al., 1996), however the triple helical domain is longer in length being composed of 340 amino acid repeats compared to 338 amino acid repeats in Type I collagen. In the early repair of the injured tendon, Type III collagen fibrils are quickly synthesized to restore strength and elasticity (Williams et al., 1984; Dahlgren et al., 2005). However, the fibrils do not have the same tensile strength quality as Type I collagen and so lack the functional properties needed in a tendon experiencing maximal load. The repair processes continues with Type III fibrils slowly being replaced by Type I collagen fibrils in an attempt to normalize the properties of the tendon (Duance et al., 1977; Williams et al., 1984; Dahlgren et al., 2005).
Type III collagen contains high levels of OHPr and glycine. It has been reported that these high levels of glycine may cause localised helix instability resulting in increased susceptibility to proteolytic cleavage and rapid turnover of the extracellular matrices containing this collagen (Linsenmayer, 1991). The frequency of Type III collagen is considered to be an indicator of tissue age, and is common in the early stages of healing and scar tissue formation where it provides mechanical strength to the matrix (Burgeson & Nimni, 1992).
188.8.131.52 Type IV Collagen
The non-fibrillar collagen, Type IV (Bailey et al., 1979), is a basement membrane-associated collagen (Light & Champion, 1984) composed of triple helical isoforms consisting of six genetically distinct chains [α1(IV) to α6(IV)]. Each chain is characterised by a long collagenous domain of approximately 1400 amino acid residues of Gly-X-Y repeats, that are interrupted at several sites by a short non-collagenous sequence and approximately 15 amino acid residue non-collagenous amino-terminus, and an approximately 230 amino acid residue non-collagenous domain at the carboxyl-terminus (Mayne, 1997). Type IV collagen has been reported to represent approximately 2% of the total collagen content of tendon (Ahtikoski et al., 2003). Unlike the fibrillar collagens discussed so far this collagen does not form fibrillar aggregates but are directly incorporated into the basement membrane without any prior excision of the pro-peptide extensions.
Type IV collagen is found uniquely in the basement membrane of tendon blood vessels (von der Mark, 1981) where it forms a key structural component; it is the only collagen that forms into a meshwork structure where four of these molecules are covalently joined together with their 7-S domains (Risteli et al., 1980; von der Mark, 1981). This lattice links to other basement membrane components such as laminin and entactin, which prevents the movement of cells and the migration of high molecular weight macromolecules. The carboxyl-terminus domain is not removed in post-translational processing and the fibers link head-to-head rather than in parallel. Also, Type IV collagen lacks the regular glycine in every third residue necessary for the tight collagen helix. These two features cause the collagen to form in a sheet.
184.108.40.206 Type V Collagen
Type V collagen is a member of the fibril forming subclass of collagens and has a molecular weight of 300 kDa with a triple helical domain that is 300 nm long. Type V collagen is intercalated into the core of the Type I collagen fibril, where it forms a template for fibrillogenesis and modulates fibril growth (Keene et al., 1987; Linsenmayer, 1991; Waggett et al., 1998). This function may be mediated by retention of the non-collagenous amino-terminal propeptide after Type V collagen molecules are incorporated into fibrils (Birk et al., 1990; Linsenmayer et al., 1993; Niyibizi & Eyre, 1993; Moradi-Ameli et al., 1994). The non-collagenous domain projects outward through the gap between adjacent Type I collagen molecules leaving major portions present on the fibril surface (Birk et al., 1988; Marchant et al., 1996) where they may limit lateral growth of the fibril by steric hindrance and charge interactions (Fichard et al., 1995).
This collagen contains three distinct alpha chains [α1(V), α2(V) and α3(V)] and is a quantitatively minor fibrillar collagen present in tissues where Type I collagen is expressed such as tendon, bone, placenta and skin. This collagen type can form very small diameter fibrils, where Type V collagen triple helical epitopes are exposed, adjacent to cells or basement membranes (Modesti et al., 1984; Gordon et al., 1994). There are several Type V collagen isoforms that differ in chain composition (Fichard et al., 1995). Type V collagen alpha chains also form heterotypic molecules with Type XI collagen alpha chains (Niyibizi & Eyre, 1989; Kleman et al., 1992; Mayne et al., 1993; Mayne et al., 1996) and is found in small amounts inside the fascicles and in the endotenon.
220.127.116.11 Type VI Collagen
Type VI collagen (Furthmayr et al., 1983) is a microfibril collagen consisting of α1(VI), α2(VI) and α3(VI) chains and has a molecular weight of greater than 420 kDa (Jander et al., 1983; Chu et al., 1987; Mayne & Burgeson, 1987) and forms multi molecular filamentous beaded structures after secretion from the cell (Timpl & Chu, 1994). It contains a short triple helical domain of 335–336 amino acids with repeating Gly-Xaa-Yaa sequences flanked by two large globular domains located at the carboxyl- and amino-termini (Chu et al., 1987). These are composed primarily of approximately 200 amino acid subdomains with homology to von Willebrand factor type A domains (Chu et al., 1990).
The amino-terminal globular domain is larger than the carboxyl-terminal domain and consists almost exclusively of the α3(VI) chain, which has nearly twice the mass of the α1(VI) and α2(VI) chains (Kielty et al., 1993; Timpl & Chu, 1994). This collagen forms a sheet-like structure and is usually found co-distributed with Type I collagen fibres in normal tendons (von der Mark, 1981; Waggett et al., 1998). This collagen is present in small amounts in tendon and is known to play a role in binding cells to matrix molecules, including fibrillar collagens, hyaluronan and decorin (Pfaff et al., 1993) as its chains appear to be recognised by cellular receptors (Bonaldo et al., 1990).
Collagen Type VI shows a high affinity, specific interaction with biglycan and decorin by binding to its core protein (Wiberg et al., 2001). The interaction is with the amino-terminal globular domain of the collagen Type VI (Specks et al., 1992; Burg et al., 1996). It may have a role in the development of the matrix supramolecular structure as well as in tissue homeostasis by mediating interactions of cells with the extracellular matrix. More specifically, interactions of collagen Type IV with collagen Type XIV, collagen Type IV and the fibrillar collagens Type I and II, decorin, microfibril-associated glycoprotein-1 and hyaluronan as well as the α1β1 and α2β1 integrins and the cell surface proteoglycan NG2 have been demonstrated (Bonaldo et al., 1990; McDevitt et al., 1991; Bidanset et al., 1992; Brown et al., 1993; Kuo et al., 1997; Pfaff et al., 1993; Burg et al., 1996; Finnis & Gibson, 1997) .
Collagen Type VI also interacts via its triple helical domain with perlecan and fibronectin (Tillet et al., 1994). It is also involved in cell migration and differentiation, and may play a role in bridging cells with the extracellular matrix. Collagen VI assembles intracellularly into antiparallel, overlapping dimers that then align and form tetramers (Engvall et al., 1986).
18.104.22.168 Type IX Collagen
Type IX collagen forms thin-beaded filaments that may interact with fibrils and cells and has a molecular weight of 250 kDa. Type IX collagen is a heterotrimer comprised of three chains [α1(IX), α2(IX) and α3(IX)] and contains three collagenous domains (COL-1, -2 and -3) separated by four non-collagenous domains (NC-1, -2, -3 and -4; Shimokomaki et al., 1980; Ninomiya & Olsen, 1984; Ninomiya et al., 1985; van der Rest et al., 1985; Har-El et al., 1992), these are numbered from the carboxyl-terminus. Present on the α2(IX) chain (NC-3 domain) is a single chondroitin sulphate chain, therefore this collagen can be considered a proteoglycan (Bruckner et al., 1985; Huber et al., 1986; Konomi et al., 1986; McCormick et al., 1987; van der Rest & Mayne, 1987; Olsen, 1989; Ayad et al., 1991; Yada et al., 1992).
The amino-terminal NC-4 domain of the α1(IX) chain is a large globular domain of 266 amino acid residues (Vasios et al., 1988). Type IX collagen is a minor constituent of articular cartilage and tendon (Fukuta et al., 1998). This collagen is a FACIT collagen as its resides on the exterior surface of collagen fibrils and cannot self-associate (Vaughan et al., 1988) and is found covalently cross-linked to collagen Type II (Wu et al., 1992). FACIT's are relatively short collagens, have interruptions in the triple helical domain and can be found at the surfaces of collagen fibrils. COL-1 and COL-2 appear to be involved in interactions with fibrils and COL-3 serves as the arm sticking out of the fibril. Two forms of Type IX collagen exist, with major differences in the NC4 domain (Svoboda et al., 1988).
22.214.171.124 Type X Collagen
Type X collagen was first described in 1982 as a collagenous molecule of 59 kDa per chain in cultures of chondrocytes from developing bone (Schmid & Conrad, 1982). This molecule is a homotrimer [α(X)]3. It forms a meshwork structure and is found in the fibrocartilaginous region of tendon. The amino acid sequence, gene structure and molecular organization of Type X collagen is extremely similar to that of Type VIII collagen.
The triple helical domain is 460 residues long and is also marked by the presence of eight imperfections at locations similar to that of Type VIII collagen. The carboxyl-terminal domain is 162 residues long and the amino-terminal domain is only 52 residues long. The Type X collagen molecule consists of a putative signal-peptide sequence (18 amino acids), an amino-terminal non-collagenous domain (38 amino acids), a triple helix (463 amino acids) and a carboxyl-terminal non-collagenous domain (161 amino acids; Thomas et al., 1991).
126.96.36.199 Type XI Collagen
Type XI collagen is a heterotrimer (Morris & Bachinger, 1987) composed of (α)1, (α)2 and (α)3(XI) chains and is 300 kDa in size with a helical domain length of 320 nm. Type XI collagen is found in small amounts in tendon and is associated with the more abundant Type II collagen fibrils (Burgeson et al., 1982; Mendler et al., 1989; Fichard et al., 1995). Type XI collagen is found in cartilage and vitreous humor of the eye. The cDNA and gene sequences have been identified for all the chains, as well as the chromosomal localizations of the genes: the gene for the α1(XI) chain (COL11A1) is located on chromosome 1p21, the gene for the a2(XI) chain (COL11A2) in chromosome 6p21.2, and the α3(XI) chain on chromosome 12q13-q14 (Prockop & Kivirikko, 1995).
188.8.131.52 Type XII Collagen
Type XII collagen forms a large molecule of 660 kDa. Type XII collagen is a FACIT collagen that is associated with the surface of Type I collagen fibrils (Keene et al., 1991), in particular at the insertion and it also interacts with decorin and fibromodulin (Font et al., 1996). It is highly expressed in fibrous connective tissues such as ligament and tendon and is also found in the dermis, cornea, blood vessel walls, skin, meninges and developing membranous bones containing Type I collagen (Sugrue et al., 1989; Oh et al., 1993), but also in connective tissue of cartilage containing Type II collagen (Lunstrum et al., 1991; Watt et al., 1992).
Type XII collagen is a homotrimer (Dublet et al., 1989) with two triple-helical domains (COL1-2) and three non-triple-helical domains (NC1-3; Gordon et al., 1989; Yamagata et al., 1991). The globular amino-terminal domain (NC3) contains several distinct subdomains homologous to domains found in other molecules, i.e. fibronectin type III repeats, von Willebrand factor A domains and the amino-terminal globular domain found in α1(IX) collagen (the NC4 domain of the long form of the α1(IX) chain). These non-collagenous subdomains make up most of the total length of the Type XII collagen molecule, while the triple-helical domains contribute only a small amount of the entire molecule.
184.108.40.206Type XIV Collagen
Type XIV collagen is a homotrimeric molecule composed of two collagenous domains (COL1-COL2) and three non-collagenous domains (NC1-NC3) and is closely related to Type XII collagen (Dublet & van der Rest, 1991; Gordon et al., 1991; Gerecke et al., 1993; Walchli et al., 1993). This multi-domain collagen can interact with more than one extracellular component simultaneously allowing integration of the developing matrices. Type XIV collagen has a developmental expression pattern in tendon consistent with a role in linear fibril growth.
Type XIV collagen can interact with GAG chains of other proteoglycans, namely with dermatan sulphate of the small proteoglycan decorin (Font et al., 1993), with the heparin sulphate chains of the basement membrane proteoglycan perlecan and with heparin (Brown et al., 1993). This collagen forms a large molecule of 660 kDa.
Type XIV collagen is also able to bind to the small proteoglycan fibromodulin (Font et al., 1996) and to the triple-helical domain of Type VI collagen (Brown et al., 1993). Type XIV has been shown to be commonly expressed in the dermis, tendon, perichondrium, perimysium, the stroma of the lungs and liver and in blood vessels, and also in virtually every tissue containing collagen Type I (Castagnola et al., 1992; Walchli et al., 1994), but also, as with Type XII collagen, in cartilage tissue containing Type II collagen (Lunstrum et al., 1991; Watt et al., 1992). The GAG chain of decorin has been shown to be crucial for binding to this collagen (Ehnis et al., 1996).
Shatton and Schubert first discovered the proteoglycans in 1954. Proteoglycans are defined as containing a protein core with one or more glycosaminoglycan/s (GAG) covalently attached (for review see; Iozzo, 1998; Watanabe et al., 1998; Culav et al., 1999). The function of the different proteoglycans is dictated by the structure of their protein core and GAG side chains. The GAG side chains of the proteoglycans are linear polysaccharides that can attract water and thus contribute to tissue hydration (Iozzo, 1998). They play an essential role in the biochemical, biomechanical and structural properties of the tendon matrix (Roughley & Lee, 1994; Iozzo, 1998; Watanabe et al., 1998; Wight & Merrilees, 2004). They have important roles as cell surface receptors and coreceptors, mediating cell-cell signalling, recognition and binding (Rauch et al., 2001; Selva & Perrimon, 2001; Nakato & Kimata, 2002; Stallcup, 2002; Couchman, 2003; Yoneda & Couchman, 2003).
Proteoglycans make up less than 1% of the dry weight of most tensile regions of tendons. In tendons, they are most commonly found in the extracellular matrix where they are associated with collagen fibrils and matrix proteins, stabilising the extracellular matrix of the tendon (Iozzo & Murdoch, 1996). The proteoglycans found in tendon include decorin, biglycan, fibromodulin, lumican, aggrecan and versican.
Proteoglycans have been classified into two subfamilies according to their molecular weight: 1) the small leucine-rich repeat proteoglycans (SLRPs) and 2) the large aggregating proteoglycans (for review see; Iozzo & Murdoch, 1996).
220.127.116.11 Small Leucine-Rich Repeat Proteoglycans (SLRPs)
All SLRPs contain a small core protein (36-42 kDa) with an amino-terminal domain where GAGs attach, a carboxyl-terminal domain and a central region which contains leucine-rich repeats (LRRs; Iozzo & Murdoch, 1996; Iozzo, 1997; Iozzo, 1999). These small proteoglycans form a horseshoe shaped core protein that is thought to be involved in protein-protein interactions (Scott, 1996). Indeed, these proteoglycans have a role in the mechanical behaviour of tendon through their binding with macromolecules of the tendon matrix. They have been shown to bind to collagen fibrils, growth factors and inhibit collagen fibrillogenesis. The majority of the SLRPs that are present in tendon are substituted with between 1 and 4 chains of GAG which may be chondroitin sulphate, keratan sulphate or dermatan sulphate (for review see; Yoon & Halper, 2005). Occasionally the SLRPs exist in non-glycosylated forms. All connective tissues contain at least one member of the SLRPs (Iozzo, 1997; Iozzo, 1999).
The SLRP gene family contains at least 13 members with more being discovered as time passes (Ameye & Young, 2002). SLRPs interact with various parts of the extracellular matrix having a role in the modulation of matrix formation and integrity, regulation of cell growth, migration and adhesion (Iozzo & Murdoch, 1996; Iozzo, 1998). The core proteins of the SLRPs are characterised by leucine-rich repeats (LRRs; Henry et al., 2001), these structures are involved in the binding of the SLRPs to collagen (Vogel et al., 1984; Hedbom & Heinegard, 1989; Rada et al., 1993; Henry et al., 2001). The central domain of the small proteoglycans can represent up to approximately 80% of the protein moiety and contains approximately 10 fold repeats (with the exception of class III SLRPs) of between 20 to 29 amino acid residue LRR with asparagine and leucine residues (Iozzo, 1999).
Proteoglycans such as fibromodulin also contain 10 LRRs and in addition to this, with the exception of proline arginine-rich end leucine-rich repeat protein (PRELP) have keratan sulphate chains attached to the LRRs (Iozzo, 1999). Proteoglycans in class III such as epiphycan are much smaller and have just six LLRs and contain sulphated tyrosine residues in the amino-terminal extension. The remaining two proteoglycans do not belong to any of the classes mentioned, they have 11 LLRs and based on their amino acid sequence they are more related to one another than any other SLRPs (Iozzo, 1999).
The most abundant proteoglycan present in tendon is decorin (see Figure 1.3; Vogel & Heinegard, 1985; Krusius & Ruoslahti, 1986; Samiric et al., 2004b) representing approximately 80% of the total proteoglycan content (Samiric et al., 2004b). Other members of the SLRPs present in tendon include biglycan (see Figure 1.3; Fisher et al., 1989; Samiric, 2003), lumican (Blochberger et al., 1992; Funderburgh et al., 1993) and fibromodulin (Oldberg et al., 1989). This review will focus on the small proteoglycans that are known to be present in tendon including decorin, biglycan, fibromodulin and lumican.
Decorin is composed of a 36-40 kDa core protein (Lorenzo et al., 2001) that was deduced by cDNA analysis from human fibroblasts (Krusius & Ruoslahti, 1986). This small proteoglycan regulates numerous functions in the extracellular matrix including collagen fibrillogenesis (Reed & Iozzo, 2002), collagen degradation (Bhide et al., 2005), cell growth (Kinsella et al., 2004) and extracellular matrix signalling (Seidler et al., 2006). It is believed that decorin allows collagen fibrils to increase in diameter up to a certain point and then prevents further enlargement (Canty & Kadler, 2002). The core protein contains a 30 amino acid residue propeptide of which the first 16 residues are likely to represent a signal peptide, a central domain containing ten LRRs flanked by disulfide-bonded terminal sequences (Day et al., 1986; Krusius & Ruoslahti, 1986).
A feature of decorin is that it contains one GAG chain of either chondroitin or dermatan sulphate (depending on the tissue) attached to the serine residue at position 4 of the amino-terminal amino acid sequence (Chopra et al., 1985) and the central domain has three attachment sites for amino-linked oligosaccharides. Removal of the GAG chain or the amino-terminal 17 amino acid residues of the decorin protein does not affect the ability of decorin to inhibit fibrillogenesis (Vogel et al., 1987) suggesting this interaction is core protein orientated (Vogel et al., 1987).
Decorin was named due to its electron microscopic appearance on the collagen network, decorating the Type I collagen fibres at the ‘d’ and ‘e’ bands (Scott, 1980; Scott et al., 1981; Scott & Orford, 1981). Decorin can be found in many connective tissues where it is found in great abundance such as tendon, bone, sclera, skin, aorta and cornea, although originally it was isolated from the cartilage and bone. It has been shown that decorin represents up to 1% of tendon dry weight (Vogel & Meyers, 1999; Derwin et al., 2001).
In tendon, decorin constitutes approximately 80% of the total proteoglycan content in the proximal region or the tensile region where tendon attaches to muscle (Samiric, 2003). As a consequence of the binding of decorin to the surface of collagen fibrils, the lateral assembly of triple helical collagen molecules is delayed (Vogel et al., 1984) and the final diameter of the collagen fibrils becomes thinner (Vogel & Trotter, 1987). However, it may be important in the organogenesis and regulation of cell division and differentiation, since its protein core has been shown to bind transforming growth factor-β (TGF-β; Iozzo, 1997). The core protein contains 10 LRRs flanked by disulphide bond stabilised loops on both sides. The core protein contains additional sites for glycosylation (amino-linked glycosylation sites) within the LRRs (Krusius & Ruoslahti, 1986).
Decorin was shown to interact with collagen via its core protein and influence collagen fibrillogenesis (Vogel et al., 1984). Decorin interacts with collagen Types I, II, III, V, VI, XII and XIV to modulate collagen fibrillogenesis (Vogel et al., 1984; Bidanset et al., 1992; Font et al., 1993; Hedbom & Heingard, 1993; Whinna et al., 1993; Thieszen & Rosenquist, 1994; Vogel et al., 1994; Font et al., 1996) as well as fibronectin (Schmidt et al., 1987), thrombospondin (Winnemoller et al., 1992), the complement component C1q (Krumdieck et al., 1992), low-density lipoprotein, the receptor required for its endocytosis (Hausser et al., 1989), epidermal growth factor receptor (EGFR) and TGF-β (Yamaguchi et al., 1990; Hilderbrand et al., 1994; Kovanen & Pentikainen, 1999). Decorin neutralises the growth stimulating effect of TGF-β, thereby regulating various cell processes including cell proliferation, differentiation, adhesion and deposition of the extracellular matrix and has been shown to suppress tumour cell growth by binding to an EGFR (Moscatello et al., 1998). The LRR motif of decorin enables the proteoglycan to take the shape of a horseshoe has been shown to bind to a single triple helix of collagen (Weber et al., 1996).
Decorin binds to collagen primarily via LRRs 4-5, which is composed of 40 amino acid residues (Svensson et al., 1995). The protein core of decorin is compact and horseshoe-shaped, a shape suitable for protein-protein interactions (Scott, 1996). The collagen-binding site is located in the cysteine-free central domain of the decorin core protein (Svensson et al., 1995). Decorin is found primarily at a distance from cells (Bianco et al., 1990). Recent studies have shown that decorin may be capable of forming a dimeric structure (Scott et al., 2003). At specific sites every 64-68 nm, decorin is found connected to collagen fibres where it modulates collagen fibril formation (Scott et al., 1981; Hedbom & Heinegard, 1993). A recent study has identified the sixth leucine motif of decorin as the major collagen I-binding site in vitro (Kresse et al., 1997).
Biglycan is a small proteoglycan and its primary structure has been deduced by cDNA analysis in humans (Fisher et al., 1989). The biglycan core protein is substituted with two GAG chains consisting of either chondroitin sulphate or dermatan sulphate. These GAGs are attached to the amino-terminus of biglycan. Dermatan sulphate is more abundant in most tissues and is attached to serine residues at position 5 and 10 of the amino-terminal amino acid sequence on its 38-42 kDa core protein (Kresse et al., 2001).
Biglycan was originally found like decorin, in cartilage and bone and their amino acid sequences have a high homology (Fisher et al., 1983 Rosenberg et al., 1985), it is also found in tendon, capillary endothelium, skeletal muscle, dermis and skin (Fisher et al., 1989; Bianco et al., 1990; Schonherr et al., 1993; Ameye et al., 2002). In humans the core protein consists of 331 amino acids and is encoded by an exon gene on chromosome X (Danielson et al., 1993).
Interactions of biglycan with collagen Types I, V, VI and XIV have been shown both via its core protein or its GAG chains, however with lower affinity than decorin (Schonherr et al., 1995). The binding affinity for Type I collagen is low, but much higher with Type VI collagen, with which it forms hexagonal networks (Wiberg et al., 2002). Like decorin the core protein contains 10 LRRs flanked by disulphide bond stabilised loops on both sides. Biglycan interacts with collagen Type VI and the complement component C1q (Krumdieck et al., 1992). Biglycan is a Zn2+-binding protein (Yang et al., 1999).
Like decorin, biglycan binds to TGF-β, and therefore has the ability to participate in modulation of cell proliferation (Ruoslahti & Yamaguchi, 1991; Border et al., 1992). Biglycan has the ability to bind to Type I collagen, this interaction is dependent on the presence of amino-linked oligosaccharides of the biglycan core protein. Biglycan has been shown to most commonly occur on the cell surface or in the pericellular space of connective tissues (Bianco et al., 1990; Fedarko et al., 1992).
Another member of the small proteoglycan family is fibromodulin. Fibromodulin was originally identified in cartilage as a 59 kDa protein and contains 10 internal repeats of 24 amino acid residues rich in leucine and is a member of class II proteoglycans (Lorenzo et al., 2001). These repeats are located on the central part of the core protein and make up approximately 80% of all amino acid residues (Ruoslahti & Yamaguchi, 1991; Iozzo & Murdoch, 1996; Lorenzo et al., 2001). It is also found in other connective tissues including tendon and ligament. It contains five asparagine residues in the central region of the LRRs (Asn-X-Thr/Ser; Oldberg et al., 1989) four of which may simultaneously have keratan sulphate (type I keratan sulphate) or amino-linked oligosaccharide bound covalently (Plaas et al., 1990). At the amino-terminal end of the fibromodulin core protein is a tyrosine rich region that is not present in decorin or biglycan (Antonsson et al., 1991).
Fibromodulin is involved in regulating the orientation of collagen fibres and is known to bind to collagen Types I, II, VI, XI and XII as well as TGF-β and inhibits fibrillogenesis of both Type I and Type II collagens. There is some evidence to suggest that fibromodulin binds to these collagens in a different position to decorin (Hedbom & Heinegard, 1993; Hildebrand et al., 1994; Font et al., 1996). This suggests that fibromodulin may have a similar role to decorin within the tendon extracellular matrix. This proteoglycan contains four keratan sulphate chains attached to the amino-terminus end attached via an N-glycosidic linkage to asparagines.
Its molecular weight varies with age, tissue and animal (Ezura et al., 2000). It plays a key role in the formation of mature large collagen fibrils (Jepson et al., 2002). Fibromodulin is expressed at high levels in tendon (Oldberg et al., 1989) and binds to the exact same region on Type I collagen as lumican does (Svensson et al., 1999). This site is however, separate from the decorin-binding site (Hedbom & Heinegard, 1993). Like decorin the protein core of fibromodulin is compact and horseshoe-shaped, a conformation suitable for protein-protein interactions (Scott, 1996). As with decorin, fibromodulin has the ability to decorate the surface of collagen fibres and therefore may have a role in regulating collagen fibril diameter.
Lumican is a highly sulphated keratan sulphate containing proteoglycan that was initially found in the cornea containing two to three keratan sulphate chains (Chakravarti et al., 1995). It is also present in tendon (Funderburgh et al., 1987), articular cartilage (Knudson & Knudson, 2001), intestine (Blochberger et al., 1992), skin, lung (Dolhnikoff et al., 1998) as well as the kidney (Schaefer et al., 2000). There is a lot of information on corneal lumican, much less is known about this molecule in other connective tissues including tendon. Lumican most closely resembles fibromodulin (Blochberger et al., 1992; Grover et al., 2000) and like fibromodulin belongs to the class II proteoglycans (Iozzo, 1997; Iozzo, 1999; Jepsen et al., 2002).
Like fibromodulin it also contains 10 LRRs (Iozzo & Murdoch, 1996; Henry et al., 2001; Lorenzo et al., 2001) with which it shares the same collagen binding site (Jepson et al., 2002), but is slightly smaller with a molecular weight of 38 kDa and contains fewer keratan sulphate chains (2-3; Iozzo & Murdoch, 1996).
Lumican binds to the fibrillar collagens in connective tissues (Blochberger et al., 1992; Chakravarti et al., 1995). It binds to Type I collagen, inhibits the size of collagen fibrils and is involved in the modulation of tendon strength (Rada et al., 1993). The lumican core protein, without the keratan sulphate side chains, is equally efficient in inhibiting collagen fibrillogenesis in vitro, suggesting this function to be entirely core protein mediated (Rada et al., 1993). The protein core of lumican is compact and horseshoe-shaped and is involved in protein-protein interactions (Scott, 1996). Lumican contains additional clusters of tyrosine-sulphate residues in its core protein (Sandy et al., 1997).
1.2.3 Large Aggregating Proteoglycans
A characteristic feature of the large aggregating proteoglycans is the presence of globular domains separated by a GAG attachment region (Abbaszade et al., 1999). The large proteoglycans can be divided into two sub families, the first is the hyalectins which have the ability to bind to hyaluronan (Iozzo & Murdoch, 1996; Aumailley & Gayraud, 1998; Iozzo, 1999). Members of this group of proteoglycans include aggrecan, versican, neurocan and brevican (Iozzo & Murdoch, 1996; Iozzo, 1998). All of the hyalectins have a three-domain structure which includes an amino- terminal domain for hyaluronan binding, a central domain that contains the GAG chains and a carboxyl-terminal lectin-like domain (Iozzo & Murdoch, 1996; Iozzo, 1998).
The second family of large proteoglycans include the non-hyaluronan binding proteoglycans, which include perlecan, agrin and testican which have been less studied than the hyalectins. The large proteoglycans are sometimes called the modular proteoglycans or leticans and are rich in chondroitin and keratan sulphate chains (Iozzo & Murdoch, 1996). They have a large core protein (100-370 kDa) and are mostly entrapped within and between collagen fibrils and fibres (Iozzo, 1998). Due to the large proteoglycans high fixed charge density they are stiffly expanded to provide the collagen fibres with a high capacity to resist high compressive and tensile forces
(Iozzo, 1998). During domain stress these molecules are compressed by approximately 20% (Iozzo, 1998).
The most studied large proteoglycans are aggrecan (see Figure 1.4; Hardingham & Fosang, 1995) and versican (see Figure 1.4; Margolis & Margolis, 1994). Although aggrecan and versican have primarily been studied in articular cartilage and blood vessels respectively, they have also been identified in tendon (Vogel & Heinegard, 1985; Vogel & Thonar, 1988; Vogel et al., 1994; Rees et al., 2000; Samiric, 2003). This review will focus on the large proteoglycans that are present in tendon.
Aggrecan was first described by Shatton and Schubert in 1954 as a mucoprotein. As many as one hundred individual aggrecan monomers interact with hyaluronan to form an aggregate of up to several hundred million in molecular weight (Heinegard & Hascall, 1974; Buckwalter & Rosenberg, 1982). The aggrecan monomer consists of a large core protein (MW 220-250 kDa) containing three globular domains (G1, G2 and G3; Iozzo & Murdoch, 1996). At the amino-terminus end of the core protein is the G1 domain (Heinegard & Hascall, 1974), which non-covalently and specifically binds to hyaluronan (Fosang & Hardingham, 1989). The G1 domain of aggrecan is homologous to link protein (Hering et al., 1997) and is separated from the second homologous domain (G2) by the linear interglobular domain (IGD; Fosang & Hardingham, 1989).
The IGD has been shown to be susceptible to proteolytic degradation by many different classes of proteinases (Hardingham & Fosang, 1995). The G2 domain of aggrecan has no known function. This domain is unique to the aggrecan protein and consists of tandem repeats just like the G1 domain but does not interact with hyaluronan or link protein (Fosang & Hardingham, 1989). At the G3 carboxyl-terminus end are two alternatively spliced epidermal growth factor (EGF-1) like regions (Baldwin et al., 1989; Fulop et al., 1993) a lectin like G3 domain (Doege et al., 1986) and an alternatively spliced domain which has sequence similarity to complement-regulatory proteins.
These motifs suggest that the G3 globular domain has a role in cell adhesion (Siegelman et al., 1990) and is involved in GAG synthesis and secretion of the proteoglycan (Kiani et al., 2002). The C-type lectin motif has been shown to bind to fructose and galactose (Halberg et al., 1988; Saleque et al., 1993; Hardingham et al., 1994) and various proteins (Watanabe et al., 1998), indicating binding to cellular or matrix ligands. Some aggrecan monomers lack the G3 domain possibly due to proteolytic cleavage (Weidemann et al., 1984; Dennis et al., 1990).
Attached to the core protein are many GAG chains. Aggrecan is a highly glycosylated proteoglycan with numerous chondroitin and keratan sulphate chains attached (Iozzo & Murdoch, 1996). A typical aggrecan molecule may contain up to 100 chondroitin sulphate cha
Cite This Dissertation
To export a reference to this article please select a referencing stye below: