EXTRACELLULAR MATRIX Department of Natural Sciences University of St. La Salle Bacolod City
Many animal cells are intrinsically linked to other cells and to the extracellular matrix (ECM). Cell surface molecules bind to other cells, or to other components of the ECM. They also play a role in mutual recognition of similar cell types. Bone and cartilage are mostly ECM plus a very few cells. Connective tissue that surrounds glands and blood vessels, is a gelatinous matrix containing many fibroblast cells.
The ECM contains 3 classes of molecules: structural proteins (collagens and elastins) protein-polysaccharide complexes to embed the structural proteins (proteoglycans) adhesive glycoproteins to attach cells to matrix (fibronectins and laminins).
PROTEOGLYCANS1. PROTEOGLYCANS are composed of a core protein to which glycosaminolycans (GAGs) are attached. GAGs consist of repeating disaccharide subunits. One of the two sugars in the disaccharide is often an amino sugar (N-acetyl-glucosamine or N-acetyl-galactosamine; usually with an attached sulfate group) and the other is a sugar or sugar acid (galactose or glucuronate). Chondroitin sulfate, keratan sulfate, heparan sulfate and hyaluronate are the most common GAGs.
Each of the four classes of GAGs is formed by polymerization of monomer units into repeats of a particular disaccharide and subsequent modifications, including addition of sulfate groups and inversion of the carboxyl group on carbon 5 of D-glucuronic acid to yield L-iduronic acid. Heparin is generated by hypersulfation of heparan sulfate, whereas hyaluronan is unsulfated. The squiggly lines represent covalent bonds that are oriented either above (D-glucuronic acid) or below (L-iduronic acid) the ring.
Most GAGs in the ECM are bound to proteins to form proteoglycans or mucoproteins. Numerous GAGs (1-200 per molecule, average length of 800 monosaccharide units) are attached to a core protein and different kinds of proteoglycans can be made by varying the combination of core proteins and GAGs. Proteoglycans (MW of~ 1 million) can be individual or attached to long hyaluronate molecules to form complexes (as in cartilage). They can be embedded in the plasma membrane or covalently linked to membrane phospholipids or bound to receptor proteins.
Proteoglycans and collagen may bind to receptor proteins (often integrins) which are reinforced by adhesive glycoproteins, such as fibronectins and laminins, to anchor cells to the ECM. GAGs in CT are highly sulfated which attracts water of hydration. They trap water (up to 50x their weight) to act as extracellular sponges resistant to physical forces in cartilage and joints. If fluid is injected into CT, it remains localized, walled off by a viscous ground substance. This property acts as barrier to the spread of bacteria that gains access to the tissues. Some bacteria secrete hyaluronidase (Staphylo/ Strepto/ Pneumococci), and collagenase (Clostridium perfringens) that breakdown matrix components.
EDEMA is a condition characterized by accumulation of excess tissue fluid.
Edema accompanies pathological conditions that cause: Increased hydrostatic pressure in capillaries by obstructing venous blood flow (e.g. congestive heart failure) Decreased colloid osmotic pressure in the blood caused by lack of blood proteins (e.g. starvation) Increased hydrostatic pressure in the tissue caused by blockage of lymphatic drainage by parasites or tumor cells Increased colloid osmotic pressure in the tissue caused by excessive accumulation of GAGs in the matrix. Hypothyroidism resulting from this condition is referred to as myxedema.
Principal producers of collagen fibers are fibroblasts; epithelial and smooth muscle cells also secrete their own type-IV collagen. Most numerous CT matrix, running in all directions in a wavy course; dull and opaque in appearance. Fibers bundled together branch and anastomose; individual fibers do not branch. With the EM, unit fibrils of collagen show periodic cross striations every 67 nm of their length.
(a) In tendons, type I fibrils are all oriented in the Interactions of fibrousdirection of the stress applied collagens with nonfibrous to the tendon. Proteoglycans and type VI collagen bind fibril-associated collagens. noncovalently to fibrils, coating the surface. The microfibrils of type VI collagen, which contain globular and triple-helical segments, bind to type I fibrils and link them together into thicker fibers. (b) In cartilage, type IX collagen molecules are covalently bound at regular intervals along type II fibrils. A chondroitin sulfate chain,covalently linked to the 2 type IX chains at the flexible kink, projects outward fromthe fibril, as does the globular N-terminal region.
1.INTRACELLULAR – free polysomes reading collagen mRNA attach to the rER, and protocollagen or precollagen-chains are deposited in the cisternae. Each chain has about 250 amino acids; every 3rd amino acid is glycine. The signal peptide is clipped off. Proline and lysine residues within the chains are then hydroxylated in the ER to form hydroxyproline and hydroxylysine (unusual amino acids present in large amounts in collagen). Core sugars (galactose and glucose) attach to the hydroxylysine residues in the ER. Each chain is synthesized with an extra length of peptides known as registration peptides, which ensure that the appropriate chains assemble in their correct position in the resulting triple helical molecule called procollagen. Further glycosylation may occur in the Golgi complex, where procollagen is packaged for secretion. Golgi vesicles release procollagen into the extracellular space by exocytosis.
2.EXTRACELLULAR- in the extracellular space, the enzyme procollagen peptidase cleaves the registration peptides from procollagen, converting it to tropocollagen. Catalyzed by lysyl oxidase, these become aligned in staggered fashion to form collagen fibers, possibly under the control of adjacent fiber-producing cells. The turnover of collagen is slowest in tendons, fastest in loose CT. Macrophages and neutrophils break down old collagen, and replaced by fibroblasts. As humans age, extracellular collagen becomes increasingly cross-linked, & turn-over slows down in CT.
Because collagen synthesis depends on the expression of several genes and on several post- translation events, many human diseases are associated with faulty collagen synthesis. Progressive systemic sclerosis- excessive accumulation of collagen (fibrosis) in almost all organs Keloid- local swelling caused by abnormal amounts of collagen that form in scars of skin Ehlers-Danlos type IV- aortic/ intestinal rupture due to faulty transcription of collagen type III Ehlers-Danlos type VII- increased articular motility due to decreased procollagen peptidase activity Scurvy- ulceration of gums, hemorrhages due to lack of Vit. C, a cofactor for proline hydroxylase Osteogenesis imperfecta- spontaneous fractures & cardiac insufficiency due to mutation in collagen type I
YELLOW or ELASTIC FIBERS Form gentle curves or spirals at their free ends when released from tension Do not form bundles; individual fibers branch and anastomose to form networks They can be stretched to 150% of their length without breaking, but lose their resiliency with advancing age. Appears yellowish, highly refractile, homogenous and are not made up of fibrillar subunits that are visible with the light microscope. Each fibril is made up of still smaller fibrils united by a small amount of ground substance. These smaller “microfibrils” have periodic cross bandings.
Synthesis and Assembly of Elastin:1.Intracellular- microfibrillar proteins containing mostly hydrophilic amino acids, and proelastin (contains large amounts of the hydrophobic amino acids glycine, proline and valine, thus accounting for elastin’s insolubility) are synthesized on rER and secreted separately.2.Extracellular- proelastin molecules polymerize extracellularly to form elastin chains. Lysyl oxidases then catalyze the conversion of certain lysine residues of elastin to aldehydes, 3 of which condense with a 4th unaltered lysine residue to form desmosine and isodesmosine. These very rare amino acids found in elastin cross- link individual chains, which then associate with numerous microfibrils to form a branching and anastomosing network of elastic fibers.
ARGYROPHYL or RETICULAR FIBERS Fibers are not branched, and are not so wavy as the collagenous fibers when released from tension. They are chemically identical to collagen, hence these fibers are considered as precursors of type I and III collagen; however, they are thinner and form delicate networks instead of thick bundles Chemical characteristics- show affinity to silver (black) stains, hence argyrophyl; do not yield gelatin on boiling; not easily dissolved by dilute acids and alkali; not so easily digested by gastric juice; not so resistant to solutions of alkaline pancreatic juice. Distribution- abundant in regions around blood vessels, muscle fibers, fat cells, basement membrane of epithelia, endoneurium, lymphoid organs and red bone marrow.
Glycoproteins are globular proteins to which shorter, branched oligosaccharide chains are covalently bound. These so-called adhesion glycoproteins mediate attachment of cells to their matrix, influence the state of differentiation of cells, and organization of their cytoskeleton. Examples are fibronectin, laminin, thrombospondin, chondronectin and fibrillin.
FIBRONECTINS, a family of closely related glycoproteins, are soluble in body fluids (blood), insoluble in the ECM and partially soluble at the cell surface. The fibronectins bind cells to the matrix and guide cellular movement. The RGD (arginine-glycine-aspartate) sequence binds to the integrin fibronectin receptor. The fibronectins bind cells to the ECM by bridging cell-surface receptors to the ECM. The intracellular cytoskeleton will align with the extracellular fibronectin to detemine cell shape. In many kinds of cancer, cells unable to make fibronectins loose shape and detach from the ECM to become malignant.
During cell movement (as during embryogenesis), pathway s of fibronectins guide cells to their destinations. Soluble plasma fibronectin promotes blood clotting by direct binding of fibrin. Fibronectins guide immune cells to wounded areas and thus promote wound healing.
LAMININS bind cells to the basal lamina of epithelial and connective tissues, and to their surrounding muscle cells, fat cells, and Schwann cells. The basal lamina serves as a structural support for tissues and as a permeability barrier to regulate movement of both cell and molecules.
Laminin is a very large protein comprised of three proteins that form a cross. The domains of laminin bind type IVcollagen, heparin, hep arin sulfate, entactin and laminin receptor proteins in overlyingcells to allow bridgingbetween the cells and the ECM. Progeria (early onset ofaging), is possibly dueto a defective laminin.
THROMBOSPONDIN - activated platelet-product. It binds to fibrinogen, plasmalogen and its activator; a participant in blood clotting. Its function is poorly understood. CHONDRONECTIN- a component of cartilage matrix that mediates attachment of chondrocytes to their matrix FIBRILLIN- a nonsulfated glycoprotein speculated to be essential for normal development. It is often associated with elastic fibers or with epithelial basal laminae. Marfan syndrome is due to a defective fibrillin gene on chromosome 15, characterized by excessively long arms and legs and progressive dilatation and fatal rupture of the ascending aorta (Pres. Abraham Lincoln).
The DGC comprises 3 subcomplexes: Molecular Connections Between ECM dystroglycan, sarcoglycan/ sarcospan of integral membrane and Disease: Muscular Dystrophy proteins; and the cytosolic adaptercomprising dystrophin, other adapter proteins, and signaling molecules. Through its O-linked sugars, dystroglycan binds to components of the basal lamina, such as laminin. Dystrophin- the protein defective in Duchenne muscular dystrophy, links dystroglycan to the actin cytoskeleton, and dystrobrevin links dystrophin to the sarcoglycan/ sarcospan subcomplex. Nitric oxide synthase (NOS) produces nitric oxide, a gaseous signaling molecule, and GRB2 is a component of signaling pathways activated by certain cell-surface receptors. Mutations in dystrophin, other DGC components, laminin, or enzymes that add the O-linked sugars to Schematic model of the dystrophin dystroglycan disrupt the DGC- glycoprotein complex (DGC) in skeletal mediated link between the exterior and the interior of muscle cells and muscle cells. cause muscular dystrophies.