Inhibition of thrombin generation by dermatan sulfate isolated from the skin of Oreochromis niloticus

摘要

Oreochromis niloticus has skin anticoagulant glycosaminoglycans (GAGs), but their effects on thrombin generation (TG) are unknown. This study partially characterized skin GAGs and analyzed as inhibitors of TG. Papain-extraction yield of 0.1% contained two fractions separated by DEAE-cellulose chromatography, differing on charge density and carboxylated groups by a combination of agarose/polyacrylamide electrophoresis and sequential toluidine blue/stains-all staining, presenting molecular sizes ca. 40 kDa. Depolymerization of the fractions with chondroitin ABC lyase showed dermatan sulfate (DS) as the unique GAG by agarose analysis. Both activated partial thromboplastin time (APTT) and prothrombin time tests only showed anticoagulation by fractions and mammalian DS by APTT (0.61, 0.47 and 1.72 IU, respectively) against heparin (193 IU). Fractions acted concentration- dependent on both intrinsic/extrinsic pathways in TG using 60-fold diluted human plasma, with more than 50% inactivation (41.6 and 83.3 μg), whereas DS and heparin entirely abolished at low amounts. DS from O. niloticus skin blocks in vitro TG in human plasma.