The role and function of the various connective tissues is reflected in the kinds of cells present, the types of fibres and the characteristics of the ground substance. For example, in what is described as loose connective tissue or areolar tissue, a relatively large number of different cell types is present. One type, the fibroblast, is responsible for producing the extracellular fibres that serve a structural role in the tissue and for producing and maintaining the ground substance. Other cell types, such as plasma cells, macrophages and eosinophils function within the ground substance of the tissue. In contrast, bone, another variety of connective tissue characterised by a mineralised extracellular matrix, contains less different cell types all associated with the production or resorption of fibres organised in a very specific pattern and calcified to create the necessary hardness. Similarly, in tendons and ligaments, the fibres are the prominent feature of the tissue. They are arranged in parallel array and are densely packed to provide maximum strength.
The primary location of loose connective tissue is beneath those epithelia that cover the body surfaces and line the internal surfaces of the body. It is also present in association with the epithelium of glands and surrounds the smallest vessels. It thus represents the initial site in which antigens and other foreign substances or organisms having breached an epithelial surface can be challenged and destroyed. It is thus the site of inflammatory and allergic of immune reactions.
The main constituent of connective tissues on a dry weight basis is collagen, 16 different types of which are known at present (Ross et al., 1995) that are classified by Roman numerals on the basis of chronology of discovery.
The collagen fibres are flexible and have a remarkably high tensile strength.
With the light microscope, collagen fibres appear as wavy structures of variable width and indeterminate length.
When examined with the electron microscope, the collagen fibre appears as a bundle of fine, thread-like subunits, the collagen fibrils. Within a fibre, these fibrils are relatively uniform in diameter. In different locations and in different stages of development, however, the fibrils differ in size. In developing or immature tissues, the fibrils may be as small as 20nm in diameter, whereas those in dense regular connective tissue of tendons or in other tissues that are subject to considerable stress may be up to ten times thicker. When stained with osmium they exhibit a sequence of closely spaced transverse bands that repeat every 68 nm. This banding is a reflection of the fibril's subunit structure (fig. 1), specifically, the size and shape of the collagen molecule and the arrangement of the molecules in forming the fibrils (Bailey, 1990; Ross et al., 1995).
The alpha chains are not all alike. These small biochemical differences result in dramatic differences in the self-assembly of the molecules thus providing a wide range of supramolecular structures (Bailey 1990). Each type of collagen plays a specific biological role.
Type I collagen is the most abundant structural component of skin, tendons and bones. It represents 90 % of the total collagen content. Two of its alpha chains (alpha 1) are identical, and one is different (alpha 2). Thus in collagen nomenclature it is designed [alpha1(I)]2alpha2(I).
Type II makes the structural framework of cartilage and intervertebral disks. It occurs as very fine fibrils. It is designated as [alpha1(II)]3.
Type III is present in many tissues: 1 to 2 % in tendons, 10 % in the skin and even 50 % in the vascular system (Ross et al., 1995). It is designated as [alpha1(III)]3. It represents less than 5 % of the collagen content of the normal palmar aponeurosis (Menzel et al 1979). It is more abundant in foetal or remodelling or distensible tissues (Bailey, 1990). Its increased concentration indicates tissue immaturity of persistent connective tissue remodelling. It is found in increased amounts in the early stages of wound healing and in hypertrophic scars. Its persistence at supranormal levels in Dupuytren's nodules is indicative of the immaturity of this tissue and it is not an indicator of Dupuytren's disease nor an indicator of some genetic malfunction.
Type V is is distributed uniformly throughout connective tissue stroma of which it is a minor component. It forms fine fibres but the typical striations are not observed. Its precise role is not yet determined.
Type IX appears to be associated with type II collagen of cartilage. It seems to contribute to the stabilisation of the network of cartilage collagen fibres by interaction at their intersections.
Type VI seems to have a wide distribution (Hessle et al., 1984). It has been identified in cornea, skin and tendon but its functional role has not been established.
One must note that vitamin C is required for the function of prolylhydroxylase and lysylhydroxylase. Without the posttranslational hydroxylation of proline and lysine, the hydrogen bonds essential to the final structure of the collagen molecule cannot form. This explain why wounds fail to heal and bone formation is impaired in scurvy.
They are closely related to collagenous fibres in that they both consist of collagen fibrils. The reticular fibre is composed of type III collagen (type IV may also be associated with reticular fibres). The individual fibrils are always of narrow diameter (20 nm) and typically they do not bundle to form thick fibres.
They contain a greater relative content of sugar groups than collagen fibres.
In loose connective tissue, networks of reticular fibres are found at the boundary of connective tissue with epithelium and around adipocytes, small blood vessels, nerve and muscles cells.
In most locations, the reticular fibre is produced by fibroblasts. Important exceptions to this general rule include endoneurium of peripheral nerves, where Schwann cells secrete reticular fibres, and the reticular and other collagenous fibres secreted by smooth muscle cells of the tunica media of blood vessels and the muscularis of the alimentary canal.
Elastic fibres are produced by fibroblasts and smooth muscle cells. They are composed of two structural components, elastin and microfibrils.
Elastin is a protein that, like collagen, is rich in proline and glycine but, unlike collagen, is poor in hydroxyproline and completely lacks hydroxylysine. Microfifrils are a fibrillar glycoprotein that is relatively straight and thin, measuring 12 nm in diameter.
The random coiling of the elastin molecule gives the elastic fibre its ability to be stretched and then recoil back to its original state. The configuration of the individual molecules continuously changes, oscillating from one form to another. Elastin also contains desmosine and isodesmosine. These large amino acids are responsible for the covalent bonding of the elastin molecules to one another to form a cross-linked network.
In developing tissues where elastic fibres are being formed, the microfibrils appear first. They are believed to serve as an organising structure for the growing elastic fibre. In mature fibres, the microfibrils are located within the elastic fibre (they are entrapped within the newly deposited elastin) and at its periphery.
A known component of the microfibril is the protein fibrillin. In Marfan's syndrome, there is a defect in the fibrillin gene. One of the consequences of the disease is abnormal elastic tissue (Ross et al., 1995).
In the same way, the development and integration of the collagen fibres and thereby their biophysical properties are dependant upon the presence of proteoglycans.
Proteoglycans are very large macromolecules comprised of a core protein to which at least one glycosaminoglycan side chain is covalently bound (Gurr et al., 1993). Glycosaminoglycans (GAGs), formerly referred to as mucopolysaccharides, are long-chained polysaccharides made up of repeating disaccharides units. One of the sugars in each disaccharides is a hexosamine (glycosamine), hence the name. GAGs are highly negatively charged due to sulfate and carboxyl groups located on many of the sugars. The negative charges attract water, forming a hydrated gel.
To a large extent the structure of the glycosaminoglycans was established by the work of Meyer (1970). Glycosaminoglycans are linear polymers of repeated disaccharides. The number of repeated disaccharides varies, but typical values are in the order of 50. The constituent monosaccharide residues usually show the chair C-1 conformation (Gässler, 1993). The reason this conformation is favored by the D-monosaccharides lies within the position of the substituents. In the C-1 conformation, most will occupy equatorial positions and maintain the largest possible distance from one another.
Due to differences in specific sugar content, the nature of their linkages and the degree of sulfatation, several different GAGs can be recognized: hyaluronic acid, chondroitin 4- and 6- sulfate, dermatan sulfate, heparan sulfate, heparin, keratan sulfate (table 1).
|Name||Approximate molecular weight ( in Daltons)||Disaccharide composition|
|Hyaluronic acid||1,000,000||D-Glucuronic acid + N-acetylglucosamine|
|Chondroitin 4-sulfate||25,000||D-Glucuronic acid + N-acetylgalactosamine 4-sulfate|
|Chondroitin 6-sulfate||25,000||D-Glucuronic acid + N-acetylgalactosamine 6-sulfate|
|Dermatan sulfate||35,000||L-Iduronic (or D-glucuronic) acid + N-acetylgalactosamine 4-sulfate|
|Keratan sulfate||10,000||Galactose or galactose 6-sulfate + N-acetylglucosamine 6-sulfate|
|Heparan sulfate||15,000||D-Glucuronic (or L-iduronic) acid 2-sulfate + N-acetylglucosamine or N-acetylgalactosamine|
|Figure 5-3: Structure of hyaluronic acid|
|Figure 5-4: Structure of large , hyaluronan aggregating proteoglycans (PG-LA)|
Tissues submitted to compressive loading or to multidirectional tensional forces have a higher content of chondroitin sulfate. Developing embryonic tissues or wound repair tissues also contain larger proportions of chondroitin sulfate than do their more mature counterparts. Collagen fibres which are normally associated with higher levels of chondroitin sulfate are generally finer than those associated with dermatan sulfate but thicker than those associated with hyaluronic acid (Flint, 1990).
At the beginning of the healing process, the concentration of hyaluronic acid and chondroitin sulfate is relatively high. Later, the chondroitin sulfate gives progressively way to a higher concentration of dermatan sulfate as the wound matures. It is noteworthy that in hypertrophic scarring and keloids the total concentration of glycosaminoglycans and the concentration of chondroitin sulfate are much higher than normal (Honda 1986).
Collagen fibrils associated with dermatan sulfate tend to be thicker and have a higher tensile strength. Dermatan sulfate is thus found as the dominant polymer in tension-transmitting structures such as tendon (Flint, 1990).
Small proteoglycans do not aggregate with hyaluronan and their structure differ distinctly from the large ones. Their function is currently unknown (Gurr et al., 1993).
Although it has been known since a long time that bones are continuously remodelled during growth or after injury, it is only recently that one has realised that most if not all connective tissues, including tendons and ligaments, are continuously remodelled in response to the needs of their environment (Flint 1980, 1990).
Numerous publications summarised by Flint (1990) have shown that the various forms of connective tissues are remarkably constant in their response to physical changes in their environment. Any force applied, say tension or pressure, induces a homeostatic response in the cells leading to a metabolic shift and a consequent reorganisation of the cell's extracellular environment.
For example, in functionally active tendons, the total GAG content is very low, usually of the order of 0.2 % of the dry weight (Gillard et al 1977). However, when the tendons are relieved of their tensional load, or if they are subjected to additional compressional loads, the amount of GAG may be markedly increased. In some specialised instances small intratendinous cartilagineous sesamoids may develop where the tendon is subjected to additional focal compressive loading. In these pressure-bearing regions, the total GAG content may be 15 - 20 times as much as in the tension-transmitting parts of the tendon (Gillard et al 1979). The metabolic changes can already be quite considerable after 24 hours and the architecture of the extracellular matrix can be profoundly reorganised after 10 days. The nature of the adaptive response is basically the same for all connective tissues but the specific changes are related to their biochemical composition.
Connective tissues react in predictable ways to the constraints of their environment. The study of the total quantity and the proportions of the various types of collagen, the quantity and the proportions of the various proteoglycans and the tridimensional relationships of these elements in Dupuytren's disease could thus give some insight into the mechanisms involved in the development of the contracture.