Hyaluronan-binding proteins on cultured J 774 macrophages

Abstract

Cultivated macrophages of murine-line J 774 were found to bind-molecular-weight (molecular weight average approx. 5·10 6) [ 3H]hyaluronan (HA) by a saturable mechanism at 4°C. Half-maximal binding was observed at 7–8 μg/ml (1.4–1.6 nM) and the maximal binding was reached at 30–40 μg/ml. Scatchard plot analysis revealed that approx. 20 000 molecules could bind to each cell with a K d of 1.5 nM. The binding could be effectively inhibited by unlabeled HA. Also chondroitin sulphate inhibited the binding, but only to about 50%. At 37°C the J 774 cells took up and degraded the polysaccharide effectively. Affinity chromatography on HA coupled to agarose of solubilized surface-iodinated J 774 cells, revealed that a protein of approx. 60 kDa, when analyzed by sodium dodecylsulfate polyacrylamide gel electrophoresis and autoradiography, could be specifically eluted with HA-oligosaccharides. Our results suggest that J 774 macrophages can bind HA by a mechanism compatible with receptor-binding, and carry a 60 kDa HA-binding protein on their surface. This receptor-binding may mediate uptake and degradation of the polysaccharide and influence the levels and turnover of HA in interstitial fluid as well as the release of HA into the bloodstream.