A high performance liquid chromatography (HPLC) procedure suitable for the simultaneous determination of the molecular size and concentration of macromolecular hyaluronate and proteoglycans in synovial fluid has been developed. Irrigation of the equine tarsocrural joint with 20 ml physiological saline (PSS) caused a mild inflammation with an increase of proteoglycans in the synovial fluid over the baseline arthrocentesis control sample. Proteoglycan and hyaluronate in the synovial fluid did not interact to form hyaluronate-proteoglycan aggregates, but separated as distinct chromatographic peaks. This suggests that the cartilage derived proteoglycans in synovial fluid in the inflamed joint have been proteolytically cleaved from the non-covalent aggregates containing link protein and hyaluronate. Hyaluronidase digestion completely abolished the hyaluronate peak without affecting the proteoglycans. This seems to indicate that proteoglycan in synovial fluid is unable to interact with hyaluronate in synovial fluid to form cartilage type aggregates. Proteolytic degradation and the time dependent release into the synovial fluid of such digested proteoglycan also resulted from the intra-articular injection of methylprednisolone acetate into normal tarsocrural joints and joints irrigated with PSS. These proteoglycans were insensitive to hyaluronidase but may consist of a protein moiety with attached glycosaminoglycans, as suggested by their sensitivity to proteinase and keratanase/chondroitinase digestion. These observations with cartilage treated with methylprednisolone acetate and mildly stimulated articular cartilage are inconsistent with earlier work on osteoarthritic and rheumatoid articular cartilage and have interesting implications for the pathogenesis and for the therapeutic action of intraarticular corticosteroids. A rapid HPLC procedure applicable to unprocessed small volume samples of synovial fluid gives information simultaneously on hyaluronate and proteoglycan in synovial fluid which is not attainable with immunoradiometric or isotope tracer techniques. It therefore appears to be useful for the analysis of cartilage turnover and destruction in health and disease.
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