A further unique chondroitin sulfate from the shrimp Litopenaeus vannamei with antithrombin activity that modulates acute inflammation.

The detailed structure of a further Chondroitin Sulfate from Litopenaeus vannamei shrimp (sCS) is described. The backbone structure was established by 1H/13C NMR, which identified 3-O-sulfated GlcA, 4-O-sulfated GalNAc, 6-O-sulfated GalNAc, and 4,6-di-O-sulfated GalNAc residues. GlcA is linked to GalNAc 4,6 di S and GlcA 3S is linked to GalNAc 4S, GalNAc 4,6 di-S and GalNAc6S residues. The anticoagulant properties of this sCS were evaluated by activated partial thromboplastin time, anti-IIa, anti-Xa and anti-heparin cofactor II-mediated activities, and sCS failed to stabilise antithrombin in a fluoresence shift assay. The anti-inflammatory effect of sCS was explored using a model of acute peritonitis, followed by leukocyte count and measurement of the cytokines, IL-1ß, IL-6 and TNF-α. The compound showed low clotting effects, but high anti-IIa activity and HCII-mediated thrombin inhibition. Its anti-inflammatory effect was shown by leukocyte recruitment inhibition and a decrease in pro-inflammatory cytokine levels. Although the biological role of sCS remains unknown, its properties indicate that it is suitable for studies of multi-potent molecules obtained from natural sources.

Anti-IIa activity and antitumor properties of a hybrid heparin/heparan sulfate-like compound from Litopenaeus vannamei shrimp.

In this present study, the anti-IIa activity and the antitumor properties of a hybrid heparin/heparan sulfate-like compound (sH/HS) from Litopenaeus vannamei shrimp heads are related. In addition to inhibiting 90.7% of thrombin activity at the lowest tested concentration (0.5 µg/mL), sH/HS compound stimulated the synthesis of antithrombotic heparan sulfate by endothelial cells in a dose-dependent manner. In vitro experiments demonstrated that the molecule from shrimp displayed a potent anti-angiogenic effect, reducing over 80% of the tubular structures formation at 50 and 100 µg/mL. In addition, sH/HS compound was able to inhibit the migration of B16F10 cells at all tested concentrations without affecting the cell viability. Although the studied compound had no effect on the proliferation of such cells during a period of 24 h, it had a significant long-term anti-proliferative effect, reducing about 80% of colony formation and anchorage-independent growth at 50 and 100 µg/mL concentrations. When its effectiveness was tested in vivo, it was demonstrated that sH/HS promoted a reduction of more than 90% of tumor growth. In the context of thromboembolic disorders associated with cancer, such findings make the sH/HS compound an excellent target for studies on inhibiting of development and tumor progression, and the prevention of coagulopathies.

A non-hemorrhagic hybrid heparin/heparan sulfate with anticoagulant potential.

The structural characterization and the anticoagulant potential of a novel heparin/heparan sulfate-like compound from the heads of Litopenaeus vannamei shrimp are described. While it is distinct from either heparin or heparan sulfate, enzymatic depolymerization and nuclear magnetic resonance spectroscopy analyses revealed that this molecule does share some structural features with heparin, such as the high degree of N- and 6-O-sulfation and minor N-acetylation, and with heparan sulfate, in the glucuronic acid content. Its ability to stabilize human antithrombin explains its significant anticoagulant activity in aPTT and Factor-Xa inhibition assays. Interestingly, in contrast to mammalian heparin, the shrimp compound displayed negligible hemorrhagic effect. Together, these findings have particular interest since they reveal a novel molecule with significant anti-Xa activity coupled with low bleeding effects which make the shrimp heparin/HS-like compound a potential alternative for mammalian heparin.

A heparin-like compound isolated from a marine crab rich in glucuronic acid 2-O-sulfate presents low anticoagulant activity.

A natural heparin-like compound isolated from the crab Goniopsis cruentata was structurally characterized and its anticoagulant and hemorrhagic activities were determined. Enzymatic and nuclear magnetic resonance analysis revealed that its structure is rich in disulfated disaccharides, possessing significant amounts of 2-O-sulfated-ß-D-glucuronic acid units. Furthermore, low amounts of trisulfated disaccharide units containing 2-O-sulfated-α-L-iduronic acid were detected, when compared to mammalian heparin. In addition, this heparin-like structure showed negligible in vitro anticoagulant activity and low bleeding potency, facts that make it a suitable candidate for the development of structure-driven, heparin based therapeutic agents with fewer undesirable effects.

Anti-inflammatory properties of a heparin-like glycosaminoglycan with reduced anti-coagulant activity isolated from a marine shrimp.

The anti-inflammatory properties of a heparin-like compound from the shrimp Litopenaeus vannamei are related. Besides reducing significantly (p<0.001) the influx of inflammatory cells to injury site in a model of acute inflammation, shrimp heparin-like compound was able to reduce the matrix metalloproteinase (MMPs) activity in the peritoneal lavage of inflamed animals. Moreover, this compound also reduced almost 90% the activity of MMP-9 secreted by human activated leukocytes. Negligible anti-coagulant activities in aPPT assay and a poor bleeding potential make this compound a better alternative than mammalian heparin as a possible anti-inflammatory drug.

Mast cells are present in epithelial layers of different tissues of the mollusc Anomalocardia brasiliana. In situ characterization of heparin and a correlation of heparin and histamine concentration.

Heparin, other sulphated glycosaminoglycans and histamine were extracted from various dissected organs of Anomalocardia brasiliana, a mollusc from the South Atlantic, and quantified. A good correlation between heparin and histamine content was found in the labial palp, intestine, ctenidium, mantle and foot tissues. The tissue location of metachromatic cells, putatively containing heparin, was identified histologically with Alcian Blue, Toluidine Blue, Masson trichrome, Haematoxylin-Eosin and PAS. Except for the foot, cells containing metachromatic granules were found in the epithelium surfaces of all the organs analysed. An in situ identification of heparin using nitrous acid and heparinase degradation has established unequivocally the presence of this compound in the metachromatic cells. The location of 'mast-like' cells at the epithelium surface of mollusc tissues exposed to the environment are very similar to the distribution of mammalian and other vertebrate mast cells and gives support to the suggestion for a role of mast cells in defense mechanisms.