Characterization of cathepsins in cartilage

Abstract

The presence of a cathepsin B-like enzyme in rabbit ear cartilage was established by the use of the synthetic substrates benzoyl-l-arginine amide and benzoyl-dl-arginine 2-naphthylamide. This was facilitated by using a technique that permits the incubation of a fixed weight of thin (18mu) cartilage sections with an appropriate exogenous substrate. The enzymic properties of cathepsin B in cartilage have been compared with an endogenous enzyme that liberates chondromucopeptide by degrading the cartilage matrix autocatalytically at pH5. Besides being maximally active at pH4.7, these cartilage enzymes are enhanced in activity by cysteine and inhibited by arginine analogues, iodoacetamide, chloroquine and mercuric chloride. They are not inhibited by EDTA, di-isopropyl phosphorofluoridate and diethyl p-nitrophenyl phosphate. When inhibiting the release of chondromucopeptide from cartilage at pH5, the arginine-containing synthetic substrates are hydrolysed simultaneously. These enzymes also share the same heat-inactivation characteristics at various pH values, being stable at acid pH and unstable at neutral and alkaline pH. The experimental evidence indicates that a cathepsin B-like enzyme may be partly responsible for the autolytic degradation of cartilage matrix at pH5.

Collagen synthesis in normal and osteoarthritic human cartilage.

“…This suggests that the synthetic activity of collagen in osteoarthritis is markedly increased and further, that this increase varies directly with the severity of the disease process through grade [6][7][8] (14)(15)(16) and that the increment varies directly with the severity of the process as determined by the histological-histochemical grade (16). The shape of the curve reported for 35SO4 incorporation into osteoarthritic cartilages of different grades of severity is quite similar to that seen in Fig.…”

“…These matrix components are synthesized locally by the chondrocytes and despite the "inert" appearance of the tissue, there is now ample evidence to suggest that at least a small portion of the proteoglycan has a rapid turnover (5,6). This view is further confirmed by recent-reports that several enzyme systems are present within the tissue, which have as their substrate the proteoglycan and its component biochemical moieties, and are presumed responsible for the catabolic phase of the metabolic cycle (7)(8)(9)(10)(11).…”

Cathepsin D. Characteristics of immunoinhibition and the confirmation of a role in cartilage breakdown

“…Recent work has suggested that the lysosomal system plays a key role in the catabolism of the proteoglycans of cartilage (for reviews see Barrett, 1968;Dingle, 1969), and there is strong, though circumstantial, evidence that this process is mediated primarily by lysosomal cathepsin D. Other workers have proposed the involvement of hyaluronidase (Bollet, Handy & Sturgill, 1963;Weissman & Spilberg, 1968), cathepsinB (Ali, 1964;Ali, Evans, Stainthorpe & Lack, 1967) and fixylosidase (Fisher, Higham, Kent & Pritchard, 1966;Fisher, Whitehouse & Kent, 1967), whereas Woessner (1967) has added further weight to the indirect evidence in favour of the importance of cathepsin D. If the hypothesis that cathepsin D is the enzyme primarily responsible for the breakdown of cartilage matrix proves to be correct, it has important implications in the pathology of connective-tissue disease, particularly arthritis (Dingle, 1962(Dingle, , 1968Hamerman & Barland, 1966;PageThomas, 1969;Weissman & Spilberg, 1968). One way in which this hypothesis could be tested would be to inhibit the enzyme in a biological test system.…”

Effect of Increased Weight Stress on Joint Integrity, Synovial Fluid Composition and Cathepsin B-Like Activity of Articular Cartilage in Growing Swine

“…The temperature of the barn was maintained at 22'C. Mechanical stress was achieved by placing an initial load of l57o of body weight in shoulder bags on the animals (Walker et al 1966 (Ali et al 1967). These were mounted and sectioned at l8 prm on a cryostat at -20"C cabinet temperature.…”