A serpin independent inhibition of coagulation by sulfated polysaccharides from marine organisms
Glauser, B.F.1,4; Monteiro, R. Q.2; Mourão, P. A. S.1,2 and Pereira, M.S.1,3
1Laboratório de Tecido Conjuntivo, Hospital Universitário Clementino Fraga Filho; 2Instituto de Bioquímica Médica; 3Instituto de Ciências Biomédicas; 4Instituto de Química, Universidade Federal do Rio de Janeiro.
Sulfated polysaccharides from marine organisms are compounds with novel structures and a wide range of biological properties. Our laboratory has studied these compounds concerning their anticoagulant activities. Recently, we reported two sulfated polysaccharides with promising anticoagulant and antithrombotic effects. One is a sulfated galactan (SG) from the red alga Botryocladia occidentalis and the other is a fucosilated chondroitin sulfate (FucCS) from the sea cucumber Ludwigothurea grisea. The inhibitory effect of these compounds on coagulation system was also determined. The anticoagulant action of SG and FCS based on APTT coagulation assay in normal plasma is similar to unfractionated heparin (UFH) and low molecular weight heparin (LMWH), respectively. Amidolitic assays using purified coagulation proteins show that, like heparin, these polysaccharides inhibit factor Xa and/or thrombin by activation of their specific inhibitors, antithrombin (AT) and/or heparin cofactor II (HCII). In the present work we expanded the investigation about the anticoagulant mechanism of these compounds. We evaluated the ability of these polymers to prolong APTT assays using an AT and HCII depleted plasma. As expected, the activity of heparin in AT/HCII depleted plasma was totally abolished. However, polysaccharides from marine organisms retain their capacity to prolong the coagulation time even in the absence of the protease inhibitors. This observation suggests that these polysaccharides have an AT/HCII independent anticoagulant activity. In order to clarify this aspect we investigate the effect of SG and FucCS on the tenase and prothrombinase complexes, which are critical for thrombin generation and, as such, are important targets for coagulation inhibition. At concentrations up to 1ug/mL, UFH had no effect on prothrombinase function. In contrast, SG and FCS inhibited prothrombinase in a dose-dependent fashion, reaching maximal inhibition with 1 and 10ug/mL, respectively. These results suggest that SG and FCS use an alternative inhibitory activity other than the ordinary AT/HCII dependent mechanism. In the next step of our work, we will test the possible effect of marine polysaccharides on the tenase complex.
Financial Support: CNPq, FAPERJ and FUJB
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