In_uence of Sulfation on Platelet Aggregation and Activation with Differentially Sulfated Hyaluronic Acids

A number of sulfated hyaluronic acid derivatives (HyalS2.5 , HyalS3 , and HyalS4 ) were prepared by sulfation of the -OH groups present on hyaluronic acid and were generi- cally termed HyalSx. The anticoagulant properties of this series of compounds has previously been shown to be good in terms of their whole blood clotting inhibition and factor Xa and thrombin inactivation. The purpose of the present study was to investigate whether the use of these com- pounds would be bene~cial to patients who would normally be given heparin, and to perform some preliminary investi- gations into their effects on platelets. The three compounds were thus studied by investigating their ability to inhibit von Willebrand factor–dependent platelet agglutination in comparison with unfractionated heparin. Agglutination was determined turbidometrically after the addition of risto- cetin to stirred formaldehyde-~xed platelets and was dem- onstrated to be dependent on the presence of sulfate groups on the polysaccharide chain and correlated with the degree of HyalSx sulfation. Interactions possibly important in low shear environments were investigated by measuring the pharmacological action of the HyalSx on spontaneous platelet activation and aggregate formation by _ow cy- tometry. The data indicate that platelet activation is not correlated with the number of sulfate or hydroxyl groups on HyalSx, suggesting that activation occurs not via electro- static interactions or H bonding, but via some other mecha- nism. A differentiation between low and high glycosamino- glycan sulfation densities is observed with respect to platelet aggregation, which is correlated with the number of sulfated groups per disaccharide unit. The ability of HyalSx to inhibit platelet aggregation induced by ADP and throm- bin was measured by aggregometry. HyalS4 resisted throm- bin stimulation to a similar extent as heparin. All Hyal derivatives, however, were better at inhibiting ADP-induced aggregation than was heparin. We conclude, therefore, that clinical use of HyalSx in place of heparin may be bene~cial because ristocetin-dependent agglutination, and therefore resistance to platelet aggregation in high shear environ- ments, in addition to resistance to stimulation by ADP, has been shown to be superior to heparin. Spontaneous platelet activation and aggregation are induced at an overall low level, even at high HyalSx concentrations, and are compara- ble with that of heparin