Ground substance is a viscous, clear substance with a slippery feel and high water content. In the light microscope, ground substance appears amorphous in sections of tissue preserved by freeze-drying or in frozen sections stained with basic dyes or by the PAS method. In routine H&E preparations, ground substance is always lost because of its extraction during fixation and dehydration of the tissue. The result is an empty background; only cells and fibers are evident. Thus, in most histologic preparations, the appearance of ground substance—or its lack of appearance—belies its functional importance. Ground substance consists predominately of three groups of molecules: Proteoglycans, very large macromolecules composed of a core protein; glycosaminoglycan molecules (GAGs), which are covalently bound to the proteoglycans; and multiadhesive glycoproteins. The size and structure of the three groups of molecules vary enormously.
GAGs are responsible for the physical properties of ground substance.
The GAGs are the most abundant heteropolysaccharide components of ground substance. These molecules represent long-chain unbranched polysaccharides composed of repeating disaccharide units. The disaccharide units contain either of two modified sugars—N-acetylgalactosamine (GalNAc) or N-acetylglucosamine (GlcNAc)—and a uronic acid such as glucuronate or iduronate. GAGs (except hyaluronan) are synthesized by connective tissue cells as a covalent, posttranslational modification of proteins called proteoglycans. For example, heparin is formed by enzymatic cleavage of heparan sulfate; dermatan sulfate is similarly modified from chondroitin sulfate.
GAGs are highly negatively charged because of the sulfate and carboxyl groups located on many of the sugars, hence their propensity for staining with basic dyes. The high density of the negative charge (polyanions) also attracts water, forming a hydrated gel. The gel-like composition of ground substance permits rapid diffusion of water-soluble molecules. At the same time, the rigidity of the GAGs provides a structural framework for the cells. GAGs are located primarily within the ground substance as well as on the surface of cells within the ECM. On the basis of differences in specific sugar residues, the nature of their linkages, and the degree of their sulfation, a family of seven distinct GAGs is recognized. They are listed and partially characterized in Table 6.3.
Hyaluronan is always present in the extracellular matrix as a free carbohydrate chain.
The GAG hyaluronan (hyaluronic acid) deserves special note because it differs from the other GAGs in several respects. It is an exceedingly long, rigid molecule composed of a carbohydrate chain of thousands of sugars rather than the several hundred or fewer sugars found in other GAGs. Hyaluronan polymers are very large (100 to 10,000 kilodaltons) and can displace a large volume of water. They are synthesized by enzymes on the cell surface; therefore, they are not posttranslationally modified like all other GAGs. Hyaluronan is also unique among the GAGs in that it does not contain any sulfate.
Each hyaluronan molecule is always present in the form of a free carbohydrate chain; in other words, it is not covalently bound to protein, so it does not form proteoglycans. By means of special link proteins, however, proteoglycans indirectly bind to hyaluronan, forming giant macromolecules called proteoglycan aggregates (Fig. 6.16). These molecules are abundant in the ground substance of cartilage. The pressure, or turgor, that occurs in these giant hydrophilic proteoglycan aggregates accounts for the ability of cartilage to resist compression without inhibiting ﬂexibility, making them excellent shock absorbers.