**2.1 Histology of cartilage**

130 Modern Arthroscopy

ECM are water, collagen, and proteoglycans. These combine with the chondrocytes to form the complex structure of articular cartilage which varies throughout its depth. The structure of articular cartilage is typically divided into 4 zones (superficial, middle, deep, and zone of calcified cartilage). The superficial zone or tangential zone is adjacent to the joint cavity and forms a gliding surface. The superficial zone is characterized by thin collagen fibrils that are aligned parallel with the articular surface. This zone also has disk shaped chondrocytes, low proteoglycan content, with high collagen and water contents. The middle (transitional) zone is characterized by large diameter collagen fibers which are oriented obliquely, round chondrocytes, and an increased proteoglycan content. The deep (radial) zone has the highest proteoglycan content with collagen fibers oriented perpendicular to the joint surface. The chondrocytes in the deep zone are round and organized into columns. The deepest layer is the zone of calcified cartilage and separates the hyaline cartilage from the subchondral bone. This zone is characterized by collagen fibrils that are radially aligned with round chondrocytes that are buried in calcified matrix. This zone has a low concentration of

Fig. 1. This figure illustrates the zones of hyaline cartilage: superficial, middle, deep, and zone of calcified cartilage. Lacunae are present in hyaline cartilage and typically contains 1-2

Articular cartilage is avascular and obtains its nutrition from the diffusion of synovial fluid through the ECM and from underlying bone. Chondrocytes produce ECM components in response to chemical (growth factor and cytokines) and physical (mechanical load, hydrostatic pressure) stimuli. The ECM is composed primarily of water 65-80%, collagen (type II) 10-20%, and aggrecan 4-7%. Other components of the ECM make up less than 5% of articular cartilage. These include proteoglycans, biglycan, decorin, fibromodulin, various collagen types (V, VI, IX, X, XI), link protein, hyaluronate, fibronectin, and lipids. The role of each of these molecules is not fully understood. Collagen functions to provide shear and tensile strength to the cartilage. Proteoglycans are produced and secreted by the chondrocytes. Proteoglycans are tangled in between collagen fibers creating an ECM that inhibits the movement of water and provides compression strength of cartilage. Aggrecan

chondrocytes. There is a periphery of increased proteoglycan content around each

chondrocyte and lacuna.

proteoglycans and a high concentration of calcium salts (Fig 1).

To image articular cartilage, standard hematoxylin and eosin (H&E) is sufficient to visualize cartilage damage and clinical use (Fig 2). However, additional stains can provide more specific information about the ECM, proteoglycans, and chondrocytes. For proteoglycans cationic dyes such as Safranin-O and Alcian blue are typically used. Safranin-O stains polysaccharides (both carboxylated and sulfated) orange. Alcian blue can stain for both types of polysaccharides (pH 2.5) or be more specific for sulfated polysaccharides (pH 1.0) such as chondroitin sulfate, turning them turquoise (Horvai 2011). A Trichrome (Gomori or Masson's) stain highlights the orientation of collagen fibrils with a bright blue stain, while staining cytoplasm and other proteins red. Additionally, elastin fibers which are abundant in elastic cartilage are typically visualized with a silver stain such as a Verhoff stain where the fibers stain black. Cartilage staining can provide clear visualization of the cartilage profile in a specific location, however it is more qualitative than quantitative. To obtain more quantitative measurements of protein content Polymerase Chain Reaction (PCR) and other molecular techniques are needed.
