Osteoarthritis is a pathological condition characterized, in part, by an excessive breakdown of the articular cartilage. A decreased content and reduced state of aggregation of proteoglycans, major components of this tissue, are found early on in the disease process. The purpose of the described three related lines of investigation is directed toward a better understanding of the regulation of proteoglycan metabolism. The structure, biochemical properties, mechanisms of action, and the systems of regulation of human cartilage arylsulfatases A and B will be studied with specific reference to their role in proteoglycan catabolism. Production of monospecific antibodies will allow development of an immunoassay to quantitate and localize enzyme production and to determine whether increased activities result from increased synthesis or activation. Conditions will be sought to increase enzyme production by chondrocytes and thus permit the elucidation of their primary sequence with the use of recombinant DNA techniques. In this way, it will be possible to compare their enzymology in normal and osteoarthritic cartilage. We have shown that proteoglycans from human articular cartilage are phosphorylated on serine residues of the core protein. The location of phosphoserine sites along this protein core, the """"""""in vitro"""""""" rates of phosphorylation, and the number of phosphoserine residues per subunit will be determined and compared in normal and osteoarthritic cartilage. The phosphorylating and dephosphorylating enzymes present in cartilage will be isolated and characterized. The role of phosphorylation on aggregate formation, chain termination and catabolism via endocytosis will be tested with a view to defining how phosphorylation modulates proteoglycan metabolism. The development of the application of HPLC to the study of proteoglycans will be continued. Various combinations of stationary and mobile phases will be tested to achieve efficient separation of proteoglycan monomers and aggregates as well as other cartilage matrix components from each other. Findings from these studies should increase our knowledge of cartilage metabolic regulation and thus facilitate early diagnosis and provide a rationale for non-surgical therapy in osteoarthritis.
