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S U M M A R YCartilage is categorized into three general subgroups, hyaline, elastic, and fibrocartilage, based primarily on morphologic criteria and secondarily on collagen (Types I and II) and elastin content. To more precisely define the different cartilage subtypes, rabbit cartilage isolated from joint, nose, auricle, epiglottis, and meniscus was characterized by immunohistochemical (IHC) localization of elastin and of collagen Types I, II, V, VI, and X, by biochemical analysis of total glycosaminoglycan (GAG) content, and by biomechanical indentation assay. Toluidine blue staining and safranin-O staining were used for morphological assessment of the cartilage subtypes. IHC staining of the cartilage samples showed a characteristic pattern of staining for the collagen antibodies that varied in both location and intensity. Auricular cartilage is discriminated from other subtypes by interterritorial elastin staining and no staining for Type VI collagen. Epiglottal cartilage is characterized by positive elastin staining and intense staining for Type VI collagen. The unique pattern for nasal cartilage is intense staining for Type V collagen and collagen X, whereas articular cartilage is negative for elastin (interterritorially) and only weakly positive for collagen Types V and VI. Meniscal cartilage shows the greatest intensity of staining for Type I collagen, weak staining for collagens V and VI, and no staining with antibody to collagen Type X. Matching cartilage samples were categorized by total GAG content, which showed increasing total GAG content from elastic cartilage (auricle, epiglottis) to fibrocartilage (meniscus) to hyaline cartilage (nose, knee joint). Analysis of aggregate modulus showed nasal and auricular cartilage to have the greatest stiffness, epiglottal and meniscal tissue the lowest, and articular cartilage intermediate. This study illustrates the differences and identifies unique characteristics of the different cartilage subtypes in rabbits. The results provide a baseline of data for generating and evaluating engineered repair cartilage tissue synthesized in vitro or for post-implantation analysis.

Dramatic changes occur in skin as a function of age, including changes in morphology, physiology, and mechanical properties. Changes in extracellular matrix molecules also occur, and these changes likely contribute to the overall age-related changes in the physical properties of skin. The major proteoglycans detected in extracts of human skin are decorin and versican. In addition, adult human skin contains a truncated form of decorin, whereas fetal skin contains virtually undetectable levels of this truncated decorin. Analysis of this molecule, herein referred to as decorunt, indicates that it is a catabolic fragment of decorin rather than a splice variant. With antibody probes to the core protein, decorunt is found to lack the carboxyl-terminal portion of decorin. Further analysis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry shows that the carboxyl terminus of decorunt is at Phe 170 of decorin. This result indicates that decorunt represents the amino-terminal 43% of the mature decorin molecule. Such a structure is inconsistent with alternative splicing of decorin and suggests that decorunt is a catabolic fragment of decorin. A neoepitope antiserum, anti-VRKVTF, was generated against the carboxyl terminus of decorunt. This antiserum does not recognize intact decorin in any skin proteoglycan sample tested on immunoblots but recognizes every sample of decorunt tested. The results with anti-VRKVTF confirm the identification of the carboxyl terminus of decorunt. Analysis of collagen binding by surface plasmon resonance indicates that the affinity of decorunt for type I collagen is 100-fold less than that of decorin. This observation correlates with the structural analysis of decorunt, in that it lacks regions of decorin previously shown to be important for interaction with type I collagen. The detection of a catabolic fragment of decorin suggests the existence of a specific catabolic pathway for this proteoglycan. Because of the capacity of decorin to influence collagen fibrillogenesis, catabolism of decorin may have important functional implications with respect to the dermal collagen network.The mechanical properties of the dermis are determined primarily by the extracellular matrix. These mechanical properties change dramatically as a function of age (1, 2), perhaps as a direct result of the known age-related changes in the molecules of the dermal extracellular matrix. Age-related differences have been shown for fibrillar collagens (3-7), which are the major extracellular matrix components of the dermis (8). In addition to collagen, the dermal extracellular matrix also contains proteoglycans, which show age-related differences (9 -16). Perhaps related to these changes in proteoglycans are age-related increases in the water content of the dermis (11) and in the content of mobile water (17).Although dermal proteoglycans are present in much lower abundance than collagen, evidence indicates that these molecules are important in the physiology of skin. For example, the small proteogl...

The cell and its glycosaminoglycan-rich pericellular matrix (PCM) comprise a functional unit. Because modification of PCM influences cell behavior, we investigated molecular mechanisms that regulate PCM volume and composition. In fibroblasts and other cells, aggregates of hyaluronan and versican are found in the PCM. Dermal fibroblasts from Adamts5 ؊/؊ mice, which lack a versican-degrading protease, ADAMTS5, had reduced versican proteolysis, increased PCM, altered cell shape, enhanced ␣-smooth muscle actin (SMA) expression and increased contractility within three-dimensional collagen gels. The myofibroblast-like phenotype was associated with activation of TGF␤ signaling. We tested the hypothesis that fibroblast-myofibroblast transition in Adamts5 ؊/؊ cells resulted from versican accumulation in PCM. First, we noted that versican overexpression in human dermal fibroblasts led to increased SMA expression, enhanced contractility, and increased Smad2 phosphorylation. In contrast, dermal fibroblasts from Vcan haploinsufficient (Vcan hdf/؉ ) mice had reduced contractility relative to wild type fibroblasts. Using a genetic approach to directly test if myofibroblast transition in Adamts5 ؊/؊ cells resulted from increased PCM versican content, we generated Adamts5 ؊/؊ ;Vcan hdf/؉ mice and isolated their dermal fibroblasts for comparison with dermal fibroblasts from Adamts5 ؊/؊ mice. In Adamts5 ؊/؊ fibroblasts, Vcan haploinsufficiency or exogenous ADAMTS5 restored normal fibroblast contractility. These findings demonstrate that altering PCM versican content through proteolytic activity of ADAMTS5 profoundly influenced the dermal fibroblast phenotype and may regulate a phenotypic continuum between the fibroblast and its alter ego, the myofibroblast. We propose that a physiological function of ADAMTS5 in dermal fibroblasts is to maintain optimal versican content and PCM volume by continually trimming versican in hyaluronan-versican aggregates.In addition to the role of cells in building specialized structural extracellular matrix, they maintain a pericellular matrix (PCM) 2 that is dynamic, provisional, and intimately associated with the cell surface (1, 2). Thus, it is appropriate to view a cell and its PCM as a functional unit. Processes that could potentially be regulated by PCM include macromolecular assembly (e.g. assembly of collagen, fibronectin fibrils, or fibrillin microfibrils), cell surface receptor-ligand interactions, binding of pathogens, and cell-matrix and cell-cell adhesion. PCM is sometimes referred to as the glycocalyx because of its abundant carbohydrate content (3). The PCM in many cell types such as chondrocytes, ova, neurons (in which PCM is also termed the perineuronal net), vascular smooth muscle cells, and fibroblasts contains aggregates of hyaluronan (HA) with chondroitin sulfate proteoglycans (CSPG) (4 -7). These aggregates are formed from non-covalent association of HA with large CSPGs such as aggrecan, versican, brevican, and neurocan. In contrast to the aforementioned cell types, heparan sulfa...

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