Localization of proteoglycan monomer and link protein in the matrix of bovine articular cartilage: An immunohistochemical study.

Abstract

Using monospecific antisera and immunofluorescence microscopy, proteoglycan monomer (PG), and link proteins were demonstrated throughout the extracellular matrix of bovine articular cartilage. A narrow band of strong pericellular staining was usually observed for both molecules, indicating a pericellular concentration of proteoglycan monomer: this conclusion was supported by dye-binding studies. Whereas PG was evenly distributed throughout the remaining matrix, more link protein was detectable in interterritorial sites in middle and deep zones. Well-defined zones of weaker territorial staining for link protein stained strongest for chondroitin sulfate. Trypsin treatment of cartilage resulted in a loss of most of the PG staining, but some selective retention of link protein, particularly around chondrocytes in the superficial zone at and near the articular surface. This residual staining was largely removed if sections were fixed after chondroitinase treatment. After extraction of cartilage with 4M guanidine hydrochloride, only PG remained and this was concentrated in the superficial zone. These observations are shown to support the concept of aggregation of PG and link protein with hyaluronic acid (HA) in cartilage matrix, and the binding of PG and link protein to HA, which is attached to the chondrocyte surface. Culture of cartilage depleted of PG and link protein by trypsin demonstrated that individual chondrocytes can secrete both PG and link proteins and that the organization of cartilage matrix can be regenerated in part over a period of 4 days.

Mechanisms of cartilage growth: Modulation of balance between proteoglycan and collagen in vitro using chondroitinase ABC

“…Remarkably, chondroitinase ABC-treated and subsequently cultured explants were able to restore GAG content to native levels ( Figures 2B and 3), a finding similar to that in previous studies that demonstrated the ability of cartilage to restore proteoglycan content and organization within 4-10 days after enzymatic digestion by increasing the rate of proteoglycan synthesis (45)(46)(47)(48). Although chondroitinase ABC treatment stimulated sulfate incorporation in the superficial layer, the level of sulfate incorporation in the middle layer was similar to that of the untreated explants ( Figure 4A), despite marked GAG deposition ( Figure 2B).…”

Enhanced cleavage of type II collagen by collagenases in osteoarthritic articular cartilage.

“…Animals were exsanguinated by cardiac puncture and ‫ف‬ 50 ml of serum was obtained. For immunoassays, F(ab Ј ) 2 preparations were obtained by pepsin digestion of each antiserum, as previously described (51). The Fc portion and undigested IgG were removed by AH-Sepharose-Protein A chromatography (Pharmacia Fine Chemicals, Uppsala, Sweden).…”

Extracellular presence of the lysosomal proteinase cathepsin B in rheumatoid synovium and its activity at neutral pH

“…The sections were stained with fluoresein-labeled rabbit anti-sheep IgG, diluted at 1/10 with PBS, for 30 minutes, and then washed in PBS as above. Cell cytoplasms were counterstained with eriochrome black (1150 dilution in PBS) and viewed for green and red fluorescence with a Zeiss Photomicroscope 111 (13). Sections from tissue cultured with or without '251-labeled cathepsin B were placed onto gelatincoated glass slides and then dipped in Ilford Nuclear Research Emulsion gel and stored frozen at -20°C.…”