Copyright 2007 by Saunders/Elsevier. All rights reserved.

Chapter 4:

Extracellular Matrix

Color Textbook of Histology, 3rd ed.

Gartner & Hiatt Copyright 2007 by Saunders/Elsevier. All rights reserved.

Copyright 2007 by Saunders/Elsevier. All rights reserved.

Extracellular Matrix

The extracellular matrix of connective tissue proper, the most common connective tissue of the body, is composed of a hydrated gel-like ground substance with fibers embedded in it. Ground substance resists forces of compression, and fibers withstand tensile forces. The water of hydration permits the rapid exchange of nutrients and waste products carried by the extracellular fluid as it percolates through the ground substance. Ground substance is composed of glycosaminoglycans (GAGs), proteoglycans, and adhesive glycoproteins. These three families of macromolecules form various interactions with each other, with fibers, and with the cells of connective tissue and epithelium. When sulfated GAGs form covalent bonds with a protein core, they form a family of macromolecules known as proteoglycans, many of which occupy huge domains. These large structures look like a bottle brush, with the protein core resembling the wire stem and the various sulfated GAGs projecting from its surface in three-dimensional space, as do the bristles of the brush. Proteoglycans may be of various sizes, ranging from about 50,000 daltons (decorin and betaglycan) to as many as 3 million daltons (aggrecan). The protein cores of proteoglycans are manufactured on the rough endoplasmic reticulum (RER), and the GAG groups are covalently bound to the protein in the Golgi apparatus. Sulfation, catalyzed by sulfotransferases, and epimerization (rearrangement of various groups around the carbon atoms of the sugar units) also occur in the Golgi apparatus.

For more information see the Ground substance section of Chapter 4: Gartner and Hiatt: Color Textbook of Histology, 3rd ed. Philadelphia, W.B. Saunders, 2007.

Figure 4–3  The association of aggrecan molecules with collagen fibers. Inset displays a higher magnification of the aggrecan molecule, indicating the core protein of the proteoglycan molecule to which the glycosaminoglycans are attached. The core protein is attached to the hyaluronic acid by link proteins. (Adapted from Fawcett DW: Bloom and Fawcett’s A Textbook of Histology, 11th ed. Philadelphia, WB Saunders, 1986.)

Copyright 2007 by Saunders/Elsevier. All rights reserved.

Collagen FibersCollagen fibers are formed from parallel aggregates of thinner fibrils 10 to 300 nm in diameter (Fig. 4–5). The fibrils themselves are fashioned from a highly regular assembly of even smaller subunits, tropocollagen molecules, each about 280 nm long and 1.5 nm in diameter. Individual tropocollagen molecules are composed of three polypeptide chains, called α-chains, wrapped around each other in a triple helical configuration.

Each α-chain possesses about 1000 amino acid residues. Every third amino acid is glycine, and most of the remaining amino acids are composed of proline, hydroxyproline, and hydroxylysine. It is believed that glycine, because of its small size, permits the close association of the three α-chains; the hydrogen bonds of hydroxyproline hold the three α-chains together; and hydroxylysine permits the formation of fibrils by binding the collagen molecules to each other. Although at least 15 different types of collagen are known, depending on the amino acid sequence of their α-chains, only six of them are of interest in this textbook. Each α-chain is coded by a separate messenger ribonucleic acid (mRNA). These different collagen types are located in specific regions of the body, where they serve various functions.

For more information see the Collagen Fiber section of Chapter 4: Gartner and Hiatt: Color Textbook of Histology, 3rd ed. Philadelphia, W.B. Saunders, 2007.

Figure 4–5  Components of a collagen fiber. The ordered arrangement of the tropocollagen molecules gives rise to gap and overlap regions, responsible for the 67-nm cross-banding of type I collagen. The gap region is the area between the head of one tropocollagen molecule and the tail of the next. The overlapping region is the area where the tail of one tropocollagen molecule overlaps the tail of another in the row above or below. In three dimensions, the overlap region coincides with numerous other overlap regions, and the gap regions coincide with numerous other gap regions. The heavy metals that are used in electron microscopy precipitate into the gap regions and make them visible as the 67-nm cross-banding. Type I collagen is composed of two identical a1(I) chains (blue) and one a2(I) chain (pink).

Copyright 2007 by Saunders/Elsevier. All rights reserved.

Synthesis of CollagenThe synthesis of collagen occurs on the RER as individual preprocollagen chains, which are α-chains possessing additional amino acid sequences, known as propeptides, at both the amino and carboxyl ends. As a preprocollagen molecule is being synthesized, it enters the cisterna of the RER, where it is modified Three preprocollagen molecules align with each other and assemble to form a tight helical configuration known as a procollagen molecule. It is believed that the precision of their alignment is accomplished by the propeptides. Because these propeptides do not wrap around each other, the procollagen molecule resembles a tightly wound rope with frayed ends. The propeptides apparently have the additional function of keeping the procollagen molecules soluble, thus preventing their spontaneous aggregation into collagen fibers within the cell.

The procollagen molecules leave the RER via transfer vesicles that transport them to the Golgi apparatus, where they are further modified by the addition of oligosaccharides. The modified procollagen molecules are packaged in the trans Golgi network and are immediately ferried out of the cell. As procollagen enters the extracellular environment, proteolytic enzymes, procollagen peptidases, cleave the propeptides (removing the frayed ends) from both amino and carboxyl ends. The newly formed molecule is shorter (280 nm in length) and is known as a tropocollagen (collagen) molecule. Tropocollagen molecules spontaneously self-assemble, in specific head-to-tail direction, into a regularly staggered array, fashioning fibrils that display a 67-nm banding. For more information see the Collagen synthesis section of Chapter 4: Gartner and Hiatt: Color Textbook of Histology, 3rd ed. Philadelphia, W.B. Saunders, 2007.

Figure 4–7  Sequence of events in the synthesis of type I collagen. Messenger RNA (mRNA) leaves the nucleus and attracts small and large subunits of ribosomes. As translation begins, the polysome complex translocates to the rough endoplasmic reticulum (RER), and the nascent alpha chains enter the lumen of the RER. Within the lumen, some proline and lysine residues of the a-chains are hydroxylated, and the preprocollagen molecule is glycosylated. Three a-chains form a helical configuration —the procollagen triple helix. The procollagen is transferred to the Golgi complex where further modification occurs. At the trans Golgi network the procollagen is packaged in clathrin-coated vesicles, and the procollagen is exocytosed. As the procollagen leaves the cell, a membrane-bound enzyme called procollagen peptidase cleaves the propeptides from both the carboxyl- and the amino-end of procollagen, transforming it into tropocollagen. These newly formed macromolecules self-assemble into collagen fibrils.

Copyright 2007 by Saunders/Elsevier. All rights reserved.

Elastic Fiber

Elastic fibers are manufactured by fibroblasts of connective tissue as well as by smooth muscle cells of blood vessels. They are composed of elastin, a protein that is rich in glycine, lysine, alanine, valine, and proline but that has no hydroxylysine. Elastin chains are held together in such a fashion that four lysine molecules, each belonging to a different elastin chain, form covalent bonds with each other to form desmosine cross-links. These desmosine residues are highly deformable and they impart a high degree of elasticity to elastic fibers to such an extent that these fibers may be stretched to about 150% of their resting lengths before breaking. After being stretched, elastic fibers return to their resting length.

The core of elastic fibers is composed of elastin and is surrounded by a sheath of microfibrils; each microfibril is about 10 nm in diameter and is composed of the glycoprotein fibrillin. During the formation of elastic fibers, the microfibrils are elaborated first and the elastin is deposited in the space surrounded by the microfibrils.

For more information see the Elastic fiber section of Chapter 4: Gartner and Hiatt: Color Textbook of Histology, 3rd ed. Philadelphia, W.B. Saunders, 2007.

Figure 4–11  An elastic fiber, showing microfibrils surrounding the amorphous elastin.

Copyright 2007 by Saunders/Elsevier. All rights reserved.

Basement MembraneThe interface between epithelium and connective tissue is occupied by a narrow, acellular region, the basement membrane, which is well stained by the PAS reaction and by other histological stains that detect GAGs. The basement membrane, as visible by light microscopy, is better defined by electron microscopy as having two constituents: the basal lamina, elaborated by epithelial cells, and the lamina reticularis, manufactured by cells of the connective tissue. Electron micrographs of the basal lamina display its two regions: the lamina lucida, a 50-nm-thick electron-lucent region just beneath the epithelium, and the lamina densa, a 50-nm-thick electron-dense region.

The lamina lucida consists mainly of the extracellular glycoproteins laminin and entactin as well as integrins and dystroglycans, transmembrane laminin receptors (both discussed later), that project from the epithelial cell membrane into the basal lamina.

The lamina densa comprises a meshwork of type IV collagen, which is coated on both the lamina lucida and lamina reticularis sides by the proteoglycan perlacan. The heparan sulfate side chains projecting from the protein core of perlacan form a polyanion.

The lamina reticularis aspect of the lamina densa also possesses fibronectin. The lamina reticularis, a region of varying thickness, is manufactured by fibroblasts and is composed of type I and type III collagen. It is the interface between the basal lamina and the underlying connective tissue, and its thickness varies with the amount of frictional force on the overlying epithelium. For more information see the Collagen synthesis section of Chapter 4: Gartner and Hiatt: Color Textbook of Histology, 3rd ed. Philadelphia, W.B. Saunders, 2007.

Figure 4–14  Basal lamina and lamina reticularis. (Adapted from Fawcett DW: Bloom and Fawcett’s A Textbook of Histology, 12th ed. New York, Chapman and Hall, 1994.)