Fibroblasts, usually the most abundant of the connective tissue cells, are responsible for producing the contents of the extracellular matrix, namely fibres and amorphous ground substance. They are found lying close to the bundles of fibres they produce and are closely related to chondroblasts and osteoblasts, the cells responsible for producing cartilage and bone matrix. The less active mature fibroblasts are known as fibrocytes.

Since fibroblasts produce the contents of the extra-cellular matrix, they play a key role in the repair process after injury. Stearns (1940b) observed that once fibre formation was initiated the fibroblast was able to produce an extensive network of fibrils in a remarkably short time.

Myofibroblasts are specialized cells which contain contractile filaments producing similar properties to smooth muscle cells. They assist wound closure after injury.

Macrophages may be resident in the connective tissues or circulating as monocytes which migrate to an area of injury and modulate into tissue macrophages (Fowler 1989). They are large cells that have two important roles. The first is phagocytosis and the second is to act as director cells, in which role they have a considerable influence on scar formation (Hardy 1989).

As a phagocyte, the macrophage acts as a housekeeper to the wound, ingesting cellular debris and subjecting it to lysosomal hydrolysis, thus debriding the wound in preparation for the fibroblasts to begin the repair process. Matter such as bacteria and cellular debris is engulfed by the phagocyte on contact.

As a director cell, the macrophage chemically activates the number of fibroblasts required for the repair process. They also play an important role in muscle regeneration leading to increased satellite cell differentiation and muscle fibre proliferation (Grefte et al 2007). A reduced number of macrophages in muscle tissue has been shown to lead to reduced muscle regeneration (Shen et al 2008). Corticosteroids can inhibit the function of the macrophage in the early inflammatory stage, resulting in a delay in fibre production (Dingman 1973, Leibovich & Ross 1974, Fowler 1989). This should be taken into consideration when exploring treatment options in the early stage.

Mast cells are large, round cells containing secretory granules that manufacture a number of active ingredients including heparin, histamine and possibly serotonin. The contents of the mast cell granules are released in response to mechanical or chemical trauma and they therefore play a role in the early stages of inflammation. Heparin temporarily prevents coagulation of the excess tissue fluid and blood components in the injured area while histamine causes a brief vasodilatation in the neighbouring non-injured area (Wilkerson 1985, Hardy 1989). Serotonins are internal nociceptive substances released during platelet aggregation in response to tissue damage. They cause contraction of blood vessels and activate pain signals (Kapit et al 1987).

Collagen is a protein in the form of fibre and is the body’s ‘glue’. It possesses two major properties, great tensile strength and relative inextensibility, and forms the major fibrous component of connective tissue structures, i.e. tendons, ligaments, fascia, sheaths, bursae, bone and cartilage.

Individual collagen fibres are normally mobile within the amorphous ground substance, producing discrete shear and gliding movement as well as dealing with compression and tension. Collagen is also the main constituent of scar tissue, in which it demonstrates its great versatility by attempting to mimic the structure it replaces.

Collagen fibres are large in diameter and appear to be white in colour. They are arranged in bundles and do not branch or anastomose. They are flexible but inelastic individually. The arrangement and weave of individual collagen fibres and collagen fibre bundles give the connective tissue structure elastic qualities; for example, an individual piece of nylon thread is inelastic, but when woven to produce tights, the weave gives the material elasticity (Peacock 1966). Collagen fibre bundles elongate under tension to their physiological length and recoil when tension is released.

The bundles of fibres are laid down parallel to the lines of the main mechanical stress, often in a wavy, sinusoidal or undulating configuration. This gives the tissue an element of crimp when not under tension. Crimp provides a buffer so that longitudinal elongation can occur without damage, as well as acting as a shock absorber along the length of the tissue, to control tension (Amiel et al 1990).

Collagen fibres are strong under tensile loading but weak under compressive forces, when they have a tendency to buckle. The orientation of the collagen fibres determines the properties that a structure will have. Crimp patterns are dependent upon function and therefore differ in different tissues, i.e. the arrangement of collagen fibres perpendicular to the surface in articular cartilage provides a cushioning force for weight-bearing, while the parallel arrangement in tendons provides great tensile strength for transmitting loads and resisting pull. Crimp patterns may also vary between different ligaments and different tendons.

Procollagen, the first step in collagen fibre formation, is produced intracellularly by the fibroblast. Amino acids are assembled to form polypeptide chains, which are attracted and held together by weak intramolecular hydrogen bonds known as cross-links (Fig. 2.2). Three polypeptide chains bond to form a procollagen molecule in the form of a triple helix which is exocytosed by the fibroblast into the extracellular space.

Once outside the cell the chains are known as tropo-collagen molecules and several tropocollagen molecules become bonded by intermolecular cross-links to form a filament or microfibril (Fig. 2.3). With the maturation into tropocollagen, the cross-links are stronger covalent bonds and occur at specific nodal intercept points, making the structure more stable (Nimni 1980, Donatelli & Owens-Burkhart 1981, Hardy 1989). Cross-links exist at every level of organization of collagen acting to weld the units together into a rope-like structure (Fig. 2.4). Intermolecular cross-linking in particular gives collagen its great tensile strength as it matures. The greater the intermolecular cross-linking, the stronger the collagen structure, with bone being considered to be the most highly cross-linked tissue (Hardy 1989).

Collagen turnover, the dynamic state of the tissue, may be related to the number of cross-links with fibres being continuously and simultaneously produced and broken down. When collagen production exceeds breakdown, more cross-links develop and the structure resists stretching. If collagen breakdown exceeds production there is a reduction in the number of cross-links and the structure stretches more easily (Alter 2000). Immature collagen tissue possesses reducible cross-links and as collagen matures these reducible links stabilize to form stronger non-reducible cross-links. Excessive cross-link formation can be prevented in immature scar tissue by the application of transverse frictions and graded mobilization techniques. Established cross-links in adhesive scar tissue are mobilized by transverse frictions before the longitudinal orientation of the fibres can be encouraged through the application of graded stress. In some instances manipulative rupture of adhesive scar tissue is indicated (see Ch. 3).

Many microfibrils make up a collagen fibril and many collagen fibrils make up a collagen fibre. Collagen fibres continue to aggregate together into larger and larger bundles and the production, aggregation and orientation of collagen are strongly influenced by mechanical tension and stress. Bundles of collagen are arranged in a specific pattern to accommodate to the function of each individual connective tissue structure (Chamberlain 1982).

It has been shown that when fibroblasts grown in tissue culture are subjected to regional tension, the cells exposed to the tensile forces multiply more rapidly and orient themselves in parallel lines in the direction of the tension (Le Gros Clark 1965).

Stearns (1940a, 1940b) identified that internal and external mechanical factors influence fibre orientation. Cell movement and occasional cytoplasmic retraction produced early local orientation of fibres, while a period of secondary orientation of fibres into heavy parallel layers was probably the result of external mechanical factors. This secondary orientation of fibres appeared to take place during the remodelling phase of wound healing and is the way in which soft tissue structures develop in response to intermittent stress and mechanical tension.

Immobilization produces rapid changes of collagen tissue as it adapts to its new resting length. Collagen which develops in the absence of mechanical stress (i.e. in the absence of movement) has a random orientation, a change in the numbers and thickness of the fibres and loss of ground substance. This reduction in the lubricating interfibular gel allows greater adherence at the fibre–fibre interface (Hardy & Woodall 1998).

Collagen takes on many forms and functions. In tendons it is tough and inelastic, in cartilage it is resilient, while in bone it is hard. This difference in structure is related to the diameter, orientation and concentration of the fibres. Collagen fibres have been classified into groups (Nimni 1980). The most common form is type I collagen consisting of large-diameter fibres, found abundantly in structures subjected to tensile forces. Type II collagen consists of a mixture of large- and narrow-diameter fibres and is abundant in structures subjected to pressure or compressive forces. Reticulin is considered to be a delicate supporting network of fragile type III collagen fibres; it may be present in the earliest stages of soft tissue repair.

Elastic fibres, consisting of the protein elastin, are yellow in colour and much thinner and less wavy than collagen fibres. Elastic fibres run singly, never in bundles, and freely branch and anastomose.

Elastic fibres provide the tissue with extensibility so that it can be extended in all directions but if tension is constantly exerted in one direction the elastic fibres may be laid down in sheets known as lamellae, e.g. ligamentum flavum (the ‘yellow ligament’). Elastic fibres make up some of the connective tissue fibres of ligaments, joint capsules, fascia and connective tissue sheaths.

The connective tissue extracellular matrix comprises the interfibrillar amorphous ground substance, with its fibrous content. As well as maintaining the mobility and integrity of the tissue structure at a macrostructural level, the amorphous ground substance is responsible for nourishing the living cells by facilitating the diffusion of gases, nutrients and waste products between the cells and capillaries.

It contains carbohydrate bound to protein (Standring 2009). The carbohydrate is in the form of polysaccharides, hexuronic acid and amino sugars, alternately linked to form long-chain molecules called glycosaminoglycans (GAGs). The main GAGs in connective tissue matrix are hyaluronic acid, chondroitin-4-sulphate, chondroitin-6-sulphate and dermatan sulphate (Donatelli & Owens-Burkhart 1981).

When GAGs are covalently bonded to proteins, the molecules are called proteoglycans (Cormack 1987). These proteoglycan molecules have the property of attracting and retaining water (Bogduk 2005). This hydration of structures depends on the proportion of proteoglycans and the flow of water into the extracellular matrix. Increased hydration creates rigidity in the extracellular matrix, allowing it to exist as a semisolid substance or gel, which improves the tissue’s ability to resist compressive forces. Therefore tissues which are subjected to high compressive forces, such as bone and articular cartilage, have a high proteoglycan content. The proteoglycans also form a supporting substance for the fibre and cellular components. Decreased hydration allows it to exist as a viscous semisolution or sol, which improves the tissue’s ability to resist tensile forces. Therefore tissues which are subjected to high tensile forces, such as tendons and ligaments, have a low proteoglycan concentration (Levangie & Norkin 2001).

The concentration of GAGs present in tissues is related to their function and gives connective tissue structures viscous properties. More water is associated with a higher GAG concentration and rabbit ligamentous tissue has a significantly greater water content than that in rabbit tendinous tissue (Amiel et al 1982). This increase in GAG and water content alters the viscoelastic properties and may provide the ligament with an additional shock-absorbing feature that is unnecessary in most tendons.

The amorphous ground substance forms a lubricant, filler and spacing buffer system between collagen fibres, fibrils, microfibrils and the intercellular spaces (Akeson et al 1980). It reduces friction and maintains distance between fibres as well as facilitating the discrete shear and gliding movement of individual collagen fibres and fibrils. It is the lubrication and spacing at the fibre–fibre interface that are crucial to the gliding function at nodal intercept points where the fibres cross in the tissue matrices (Amiel et al 1982). If the tissues are allowed to adopt a stationary attitude (i.e. becoming immobile), anomalous cross-links form at the nodal intercept points.

A balance between the cross-link formation relative to the tissue’s tensile strength and mobility is important to normal connective tissue function. Excessive cross-linking and loss of GAGs and water volume result in loss of the critical distance between the fibres. The fibres come into contact with each other and stick together leading to altered tissue function and pain resulting from loss of extensibility and increased stiffness.

The elasticity of connective tissue fibres together with the viscosity of the amorphous ground substance gives connective tissue structures viscoelastic properties which ensure that normal connective tissues are mobile.

The biomechanical properties of connective tissue depend on the number and orientation of collagen fibres and the proportion of amorphous ground substance present. Each connective tissue structure is specifically designed for function but the tissues can be grouped simply into irregular and regular connective tissue.