Anticoagulant Activity and Mechanism of Non-Anticoagulant Sulfated Polysaccharides

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1208-1208 ◽  
Author(s):  
Michael Dockal ◽  
Susanne Till ◽  
Sabine Knappe ◽  
Hartmut J. Ehrlich ◽  
Friedrich Scheiflinger
Keyword(s):  
Structural Properties ◽  
Brown Algae ◽  
Biological Activities ◽  
Anticoagulant Activity ◽  
Procoagulant Activity ◽  
The Other ◽  
Sulfated Polysaccharides ◽  
Heparin Cofactor Ii ◽  
Thrombin Inhibition ◽  
Thrombin Generation Assay

Abstract Abstract 1208 Fucoidans are sulfated polysaccharides which are extracted from brown seaweeds and echinoderms and have a wide variety of biological activities. Described as non-anticoagulant polysaccharides (NASPs), they have been demonstrated to improve clotting in FVIII- and FIX-deficient plasma (Liu et al., 2006), making them good candidates for hemophilia treatment. However, fucoidans have also been extensively studied for their anticoagulant effects (Pereira et al., 1999), which usually occur at much higher concentrations than the procoagulant activity. This opens a large procoagulant window where procoagulant activities exceed anticoagulant effects. When analyzed by a global hemostatic thrombin generation assay in hemophilia plasma the onset of procoagulant activity is observed at concentrations as low as 0.01 μg/mL with optimal activity at about 1 μg/mL. Reversal of procoagulant activity is seen at concentrations higher than 10 μg/mL. Fucoidans and other sulfated polysaccharides activate different anticoagulant mechanisms depending on their structural properties. Branched fucoidans extracted from brown algae have been shown to directly inhibit thrombin, while linear fucoidan from echinoderms activates antithrombin III (ATIII) or heparin cofactor II (HCII)-mediated thrombin inhibition (Pereira et al., 1999). Sulfated galactans also have serpin-dependent and -independent anticoagulant activities (Glauser et al., 2009). In this study we analyzed fucoidans from several brown algae species for their anticoagulant properties and mode of action to identify the candidate with the best procoagulant and lowest anticoagulant activity. NASPs from several brown algae species including L. japonica (L.j.), F. vesiculosus (F.v.), and U. pinnatifida (U.p.) showed different anticoagulant activities in an activated partial thromboplastin time (aPTT) assay. U.p. fucoidan was about twice as anticoagulant as the other fucoidan preparations, increasing clotting time by 50% at a concentration of 4 μg/mL. In addition, NASPs were analyzed in ATIII- and HCII-thrombin model assays. L.j. fucoidan activated ATIII-mediated thrombin inhibition, whereas the other fucoidans showed no effect on ATIII. Fucoidans from L.j. and F.v. had a direct effect on thrombin, starting at about 10 μg/mL. By contrast, U.p. fucoidan did not directly affect thrombin. However, all preparations increased HCII-mediated thrombin inhibition at concentrations below 1 μg/mL. This suggests that HCII is the main target for the anticoagulant activity of fucoidans. Nevertheless, we observed substantial differences between the fucoidan candidates which will be correlated to structural properties. Our work describes the assessment of anticoagulant activities of a variety of fucoidan species to better understand their intertwined pro- and anticoagulant effects. This provides important mechanistic insights for the development of hemophilia therapies. Disclosures: Dockal: Baxter Innovations GmbH: Employment. Till:Baxter Innovations GmbH: Employment. Knappe:Baxter Innovations GmbH: Employment. Ehrlich:Baxter Innovations GmbH: Employment. Scheiflinger:Baxter BioScience: Employment.

Further Studies on the Mechanisms for the Antithrombotic Effects of Sulfated Polysaccharides in Rabbits

1988 ◽  
Vol 60 (02) ◽  
pp. 188-192 ◽  
Author(s):  
F A Ofosu ◽  
F Fernandez ◽  
N Anvari ◽  
C Caranobe ◽  
F Dol ◽  
...  
Keyword(s):  
Antithrombin Iii ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Minimum Weight ◽  
Sulfated Polysaccharides ◽  
Pentosan Polysulfate ◽  
Heparin Cofactor Ii ◽  
Thrombin Inhibition ◽  
The Relationship ◽  
Prothrombin Activation

SummaryA recent study (Fernandez et al., Thromb. Haemostas. 1987; 57: 286-93) demonstrated that when rabbits were injected with the minimum weight of a variety of glycosaminoglycans required to inhibit tissue factor-induced thrombus formation by —80%, exogenous thrombin was inactivated —twice as fast in the post-treatment plasmas as the pre-treatment plasmas. In this study, we investigated the relationship between inhibition of thrombus formation and the extent of thrombin inhibition ex vivo. We also investigated the relationship between inhibition of thrombus formation and inhibition of prothrombin activation ex vivo. Four sulfated polysaccharides (SPS) which influence coagulation in a variety of ways were used in this study. Unfractionated heparin and the fraction of heparin with high affinity to antithrombin III potentiate the antiproteinase activity of antithrombin III. Pentosan polysulfate potentiates the activity of heparin cofactor II. At less than 10 pg/ml of plasma, all three SPS also inhibit intrinsic prothrombin activation. The fourth agent, dermatan sulfate, potentiates the activity of heparin cofactor II but fails to inhibit intrinsic prothrombin activation even at concentrations which exceed 60 pg/ml of plasma. Inhibition of thrombus formation by each sulfated polysaccharides was linearly related to the extent of thrombin inhibition achieved ex vivo. These observations confirm the utility of catalysis of thrombin inhibition as an index for assessing antithrombotic potential of glycosaminoglycans and other sulfated polysaccharides in rabbits. With the exception of pentosan polysulfate, there was no clear relationship between inhibition of thrombus formation and inhibition of prothrombin activation ex vivo.

scholarly journals Structural Analysis of Fucoidans

2008 ◽  
Vol 3 (10) ◽  
pp. 1934578X0800301 ◽  
Author(s):  
Maria I. Bilan ◽  
Anatolii I. Usov
Keyword(s):  
Biological Activity ◽  
Structural Analysis ◽  
Charge Density ◽  
Brown Algae ◽  
Uronic Acid ◽  
Chemical Structure ◽  
Biological Activities ◽  
Biological Properties ◽  
Sulfated Polysaccharides ◽  
New Information

Sulfated polysaccharides of brown algae (“fucoidans”) constitute a wide variety of biopolymers from simple sulfated fucans up to complex heteropolysaccharides composed of several neutral monosaccharides, uronic acid and sulfate. The increased interest in this class of polysaccharides is explained by their high and versatile biological activities, and hence, by their possible use in new drug design. Structural analysis of several fucoidans demonstrates that their biological properties are determined not only by charge density, but also by fine chemical structure, although distinct correlations between structure and biological activity cannot be formulated at present. The aim of this review is to describe the methods of structural analysis currently used in fucoidan chemistry, and to discuss some new information on the structures of fucoidans presented in recent publications.

Serpin-independent anticoagulant activity of a fucosylated chondroitin sulfate

2008 ◽  
Vol 100 (09) ◽  
pp. 420-428 ◽  
Author(s):  
Bianca F. Glauser ◽  
Mariana S. Pereira ◽  
Robson Q. Monteiro ◽  
Paulo A. S. Mourão
Keyword(s):  
Chondroitin Sulfate ◽  
Anticoagulant Activity ◽  
Factor Xa ◽  
Anticoagulant Effect ◽  
Coagulation System ◽  
Heparin Cofactor Ii ◽  
Thrombin Inhibition ◽  
Coagulation Proteins ◽  
Dependent Inhibition ◽  
Structure Versus

SummaryFucosylated chondroitin sulfate is a glycosaminoglycan from sea cucumber composed of a chondroitin sulfate-like core with branches of sulfated fucose. This glycosaminoglycan has high anticoagulant and antithrombotic activities. Its serpin-dependent anticoagulant activity is mostly due to activating thrombin inhibition by heparin cofactor II. Here, we evaluated the anticoagulant activity of fucosylated chondroitin sulfate using antithrom-bin- and heparin cofactor II-free plasmas. In contrast to mammalian heparin, the invertebrate glycosaminoglycan is still able to prolong coagulation time and delay thrombin and factor Xa generation in serpin-free plasmas. These observations suggest that fucosylated chondroitin sulfate has a serpin-independent anticoagulant effect. We further investigated this effect using purified blood coagulation proteins. Clearly, fucosylated chondroitin sulfate inhibits the intrinsic tenase and prothrombinase complexes, which are critical for thrombin generation. It is possiblethat the invertebrate chondroitin sulfate inhibits interactions between cofactor Va and factor Xa. We also employed chemically modified polysaccharides in order to trace a structure versus activity relationship. Removal of the sulfated fucose branches, but not reduction of the glucuronic acid residues to glucose, abolished its activity. In conclusion, fucosylated chondroitin sulfate has broader effects on the coagulation system than mammalian glycosaminoglycans. In addition to its serpin-dependent inhibition of coagulation protease, it also inhibits the generation of factor Xa and thrombin by the tenase and prothrombinase complexes, respectively. In plasma systems, the serpin-independent anticoagulant effect of fucosylated chondroitin sulfate predominates over its serpin-dependent action. This glycosaminoglycan opens new avenues for the development of antithrombotic agents.

Structure-Activity Relationship of Pro- and Anticoagulant Effects of Fucus Vesiculosus Fucoidan

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3353-3353
Author(s):  
Michael Dockal ◽  
Susanne Till ◽  
Zhenqing Zhang ◽  
Sabine Knappe ◽  
Sabrina Reutterer ◽  
...  
Keyword(s):  
Conflicts Of Interest ◽  
Biological Activities ◽  
Biological Effects ◽  
Monosaccharide Composition ◽  
Fucus Vesiculosus ◽  
Procoagulant Activity ◽  
Sulfated Polysaccharides ◽  
Variable Degree ◽  
Active Length ◽  
The Impact

Abstract Abstract 3353 Fucoidans are sulfated polysaccharides extracted from brown algae. Fucoidans have a wide variety of biological activities including pro- and anticoagulant activities, which occur at different concentration ranges. Therefore, fucoidans have also been described as non-anticoagulant sulfated polysaccharides (NASPs, Liu et al. Thromb Haemost 2006; 95:68). Fucoidans have complex structures due to their large molecular weight (Mw), wide Mw distribution, variable degree and pattern of sulfation, diverse monosaccharide composition, branching of the sugar chain and different monomer linkages. This structural complexity challenges identification of the components responsible for fucoidan activities. The aim of the presented study was to fractionate Fucus vesiculosus (F.v.) fucoidan by size and to de- and oversulfate it to obtain compounds of varying Mw or degree of sulfation (DS), respectively. The fucoidans were then to be analyzed for their pro-and anticoagulant activities and structure, and the effect of modified fucoidan on the target protein Tissue Factor Pathway Inhibitor (TFPI) investigated. Two approaches were applied to generate F.v. fucoidan with different structural properties to the original. Firstly, fucoidan was fractionated by size using ultrafiltration. The Mw of the resulting fractions ranged from 8–180 kD. NMR, elemental analysis and HPAEC showed that other structural features, such as sulfate content and monosaccharide composition, were similar to those of the original fucoidan. Thrombin generation (CAT) assays showed an EC50 for procoagulant activity of 0.3–3 μg/mL; aPTT increased by 50% at 4–25 μg/mL. Generally, higher Mw increased procoagulant activity. Below 15 kD, this activity was markedly reduced. The minimum active length of F.v. fucoidan was 70 carbohydrate units. De- and oversulfated F.v. fucoidans were used to investigate the impact of charge on pro- and anticoagulant activities. In the CAT assay, oversulfated fucoidans showed improved procoagulant activity with an EC50 of 0.09–0.12 μg/mL, while desulfated fucoidans demonstrated reduced procoagulant activity compared to the original. A DS of 0.5 was estimated to be the limit for procoagulant activity. Inhibition of TFPI by fucoidan was assessed with a dilute prothrombin time assay (dPT) with added full-length TFPI. TFPI-blocking activity was mainly dependent on the DS and less on the fucoidans' Mw. Interestingly, oversulfation also stimulated an undesired activation of the contact pathway. The presented study determined the minimal structural requirements for procoagulant activity of fucoidan molecules and identified features causing undesired biological effects. Disclosures: No relevant conflicts of interest to declare.

The Additive Effect of Low Molecular Weight Heparins on Thrombin Inhibition by Dermatan Sulfate

1993 ◽  
Vol 70 (03) ◽  
pp. 443-447 ◽  
Author(s):  
Benilde Cosmi ◽  
Giancarlo Agnelli ◽  
Edward Young ◽  
Jack Hirsh ◽  
Jeffrey Weitz
Keyword(s):  
Molecular Weight ◽  
Antithrombin Iii ◽  
Dermatan Sulfate ◽  
Anticoagulant Activity ◽  
Additive Effect ◽  
Low Molecular Weight ◽  
Low Molecular Weight Heparins ◽  
Heparin Cofactor Ii ◽  
Thrombin Inhibition ◽  
Sds Page

SummaryThe aim of this study was to investigate the mechanism by which the anticoagulant activity of dermatan sulfate (DS) is increased by low molecular weight heparin (LMWH). In platelet poor plasma, LMWH enhances the effect of DS on thrombin (IIa) inhibition as determined by thrombin clotting times and with a chromogenic substrate assay. Analysis of the results of the chromogenic assays using either the algebraic fractional or the graphic isobole method suggests that LMWH has an additive effect on the anti-IIa activity of DS. This additive effect was lost when the experiments were repeated in plasma immunodepleted of antithrombin III (ATIII), indicating that the anti-IIa activity of LMWH is ATIII-dependent. To further explore the mechanism of the interaction between LMWH and DS, 125I-labeled IIa was added to plasma in the presence or absence of DS and/or LMWH and the formation of IIa-inhibitor complexes was assessed using SDS-PAGE followed by autoradiography. DS addition selectively increases the formation of heparin cofactor II (HCII)-IIa complexes, whereas LMWH enhances ATIII-IIa complex generation. Compared to plasma containing DS alone, the formation of ATIII-IIa complexes also is increased when the combination of DS and LMWH is added. These findings suggest that the additive effect of LMWH on the anti-IIa activity of DS reflects their different modes of IIa inhibition; DS potentiates IIa inhibition by HCII, while LMWH catalyses ATIII-dependent IIa inactivation. The potential clinical significance of these findings requires further investigation.

The Role of Sulfates in Fucoidan Extracted from Fucus evanescens in Proinflammatory Cytokines Production by Human Peripheral Blood Cells in vitro

2020 ◽  
Vol 65 (5-6) ◽  
pp. 3-10
Author(s):  
S. R. Khil'chenko ◽  
T. S. Zaporozhets ◽  
T. N. Zvyagintseva ◽  
N. M. Shevchenko ◽  
N. N. Besednov
Keyword(s):  
Peripheral Blood ◽  
Blood Cells ◽  
Brown Algae ◽  
Biological Activities ◽  
Molecular Structures ◽  
Sulfated Polysaccharides ◽  
Peripheral Blood Cells ◽  
Fucus Evanescens ◽  
Tnf Α

Fucoidans, sulfated polysaccharides extracted from brown algae (Phaeophyceae), have a wide spectrum of bioactivity. Studies of molecular structures of fucoidans and deciphering of molecular elements' impact on their biological activities are at their active stage. The article shows the role of sulfates and acetyl groups in fucoidan isolated from Fucus evanescens in proinflammatory cytokines production by human heparinized unfractionated peripheral blood cells. Material and Methods. The cells were incubated with native fucoidan (N) and its deacetylated (deA), partially desulfated (deS), and both deacetylated and partially desulfated (deAdeS) derivatives (100 μg/mL). Cytokine concentrations were determined in cell supernatants by ELISA in a 'sandwich' modification with commercial kits. Results. Incubation with N fucoidan led to an increase of IL-6, TNF-α, IL-8 levels in supernatants. Partial removal of sulfate groups cancelled or decreased stimulating effect for IL-6, TNF-α, cytokines, but not for IL-8. deAc fucoidan action was comparable with N polysaccharide. Native polysaccharide and its chemically modified derivatives did not change IFN-γ и IL-10 cytokine production. Conclusion. The obtained results suggest that sulfates have a significant role in cytokine-producing properties of fucoidan extracted from brown algae F.evanescens.

scholarly journals ANTITUMOR SULFATED POLYSACCHARIDES FROM BROWN ALGAE Dictyota caribaea

2020 ◽  
Author(s):  
Alexia Nathália Brígido Assef ◽  
Bianca Barros da Costa ◽  
Thamyris Almeida Moreira ◽  
Luana David do Carmo ◽  
Tamiris de Fátima Goebel de Souza ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Brown Algae ◽  
Antitumor Effect ◽  
Biological Activities ◽  
Bioactive Molecules ◽  
Sulfated Polysaccharides ◽  
Spleen Weight ◽  
Tumor Growth Inhibition ◽  
White Pulp

AbstractSulfated polysaccharides (SP) are a complex group of bioactive molecules able to inhibit tumor growth. SP increase the effectiveness of chemotherapy and reduce some side effects. Brown algae produce SP with several biological activities including antitumor. This work aimed to investigate the antitumor effect of SP from the brown algae Dictyota caribaea (Dc-SP). Dc-SP were extracted with proteolytic enzyme and supernatant was precipitated with increasing concentrations of ethanol. Antiproliferative activity of Dc-SP was tested by the MTT assay against colon cancer (HCT 116) and metastatic melanoma (B16-F10) cell lines. The antitumor effect was evaluated on Swiss mice transplanted with sarcoma 180 tumor and treated i.p. during 7 days with saline or Dc-SP (25 and 50 mg/kg/animal). Dc-SP did not exhibit cytotoxicity in vitro, however the Dc-SP-treated mice depicted up to 50% tumor growth inhibition. Dc-SP treatment induced spleen weight increasing along with intense white pulp disorganization. Furthermore Dc-SP did not depict hepatic toxicity, nephrotoxicity nor leukopenia and did induce increase of platelets count. Altogether, these results represent a promising antitumor host dependent effect induced by Dc-SP.

Effects of polysaccharides enriched in 2,4-disulfated fucose units on coagulation, thrombosis and bleeding

2009 ◽  
Vol 102 (11) ◽  
pp. 829-836 ◽  
Author(s):  
Roberto Fonseca ◽  
Gustavo Santos ◽  
Paulo Mourão
Keyword(s):  
Chondroitin Sulfate ◽  
Marine Invertebrates ◽  
Anticoagulant Activity ◽  
Linear Chain ◽  
Sulfated Polysaccharides ◽  
Factor Xii ◽  
Heparin Cofactor Ii ◽  
Structure Activity ◽  
Sulfated Fucan ◽  
Discrete Effect

SummarySulfated polysaccharides from marine invertebrates have welldefined structures and constitute a reliable class of molecules for structure-activity relationship studies.We tested the effects of two of these polysaccharides,namely a sulfated fucan and a fucosylated chondroitin sulfate, on coagulation, thrombosis and bleeding. The compounds share similar 2,4-disulfated fucose units, which are required for high anticoagulant activity in this class of polymer.These residues occur either as branches in fucosylated chondroitin sulfate or as components of the linear chain in the sulfated fucan.These polysaccharides possess anticoagulant activity but differ significantly in their mechanisms of action.The fucosylated chondroitin sulfate inhibits thrombin by heparin cofactor II, whereas sulfated fucan inhibits thrombin by both antithrombin and heparin cofactor II. In addition, these polysaccharides also have serpin-independent anticoagulant activities. Fucosylated chondroitin sulfate, but not sulfated fucan, activates factor XII. As a result of the complex anticoagulant mechanism, the invertebrate polysaccharides differ in their effects on experimental thrombosis. For instance, the sulfated fucan inhibits venous thrombosis at lower doses than fucosylated chondroitin sulfate. In contrast, fucosylated chondroitin sulfate is significantly more potent than sulfated fucan in arterial thrombosis. Finally, fucosylated chondroitin sulfate increases bleeding, while sulfated fucan has only a discrete effect. In conclusion, the location of 2,4-disulfated fucose units in the polysaccharide chains dictates the effects on coagulation, thrombosis and bleeding.

Unbalanced Effects of Dermatan Sulfates with Different Sulfation Patterns on Coagulation, Thrombosis and Bleeding

2001 ◽  
Vol 86 (11) ◽  
pp. 1215-1220 ◽  
Author(s):  
C. P. Vicente ◽  
P. Zancan ◽  
L. L. Peixoto ◽  
R. Alves-Sá ◽  
F. S. Araújo ◽  
...  
Keyword(s):  
Dermatan Sulfate ◽  
Thrombus Formation ◽  
Anticoagulant Activity ◽  
Sulfated Polysaccharides ◽  
Heparin Cofactor Ii ◽  
Iduronic Acid ◽  
Bleeding Effect ◽  
Mammalian Counterpart

SummaryWe compared the anticoagulant, antithrombotic and bleeding effects of highly sulfated dermatan sulfates from invertebrates and their mammalian counterpart. An invertebrate dermatan sulfate containing 2-O-sulfated α-L-iduronic acid and 4-O-sulfated N-acetyl-β-D-galactosamine residues is a potent anticoagulant due to a high heparin cofactor II activity. It inhibits thrombin due to the formation of a covalent complex with heparin cofactor II, as in the case of mammalian dermatan sulfate, but the effect occurs at lower concentrations for the invertebrate polysaccharide. Surprisingly, the invertebrate dermatan sulfate has a lower potency to prevent thrombus formation on an experimental model and a lower bleeding effect in rats than the mammalian dermatan sulfate. In contrast, another invertebrate dermatan sulfate, also enriched in 2-O-sulfated α-L-iduronic acid, but in this case sulfated at O-6 position of the N-acetyl-β-D-galactosamine units, has no in vitro or in vivo anticoagulant activity, does not prevent thrombus formation but shows a bleeding effect similar to the mammalian glycosaminoglycan. Overall, these results demonstrate unbalanced effects of dermatan sulfates with different sulfation patterns on coagulation, thrombosis and bleeding, and raise interesting questions concerning the relationship among these three biological actions of sulfated polysaccharides.

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