Inhibition of the association of proteoglycans with cultured vascular endothelial cell layers by calcium and sodium spirulan

Vascular endothelial proteoglycans exhibit an antithrombogenic activity by activating antithrombin III and heparin cofactor II on the luminal surface of the vascular wall. Calcium spirulan (Ca-SP) is a novel sulfated polysaccharide isolated from the blue-green alga Spirulina platensis. Since Ca-SP exhibits anti-thrombin activity by activation of heparin cofactor II, we hypothesized that the polysaccharide may influence the metabolism of anticoagulant proteoglycans synthesized by endothelial cells. When cultured bovine aortic endothelial cells were treated with Ca-SP (50 μg/ml or less) in the presence of [³⁵S]sulfate for 24 hr, the accumulation of labeled proteoglycans in the cell layer was decreased but that in the conditioned medium was significantly increased, indicating that Ca-SP inhibits the association of proteoglycans with vascular endothelial cell layers. Na-SP, which was prepared by replacement of calcium ion with sodium ion, showed an equal effect. When the endothelial cells were labeled with [₃₅S]sulfate and then treated with Ca-SP (5 μg/ml or more) for 1 hr in the absence of [₃₅S] sulfate, the percentage of [³⁵S]sulfate-labeled proteoglycans released into the medium was markedly increased by Ca-SP. DEAE-Sephacel ion exchange chromatography of [³⁵S]sulfate-labeled proteoglycans released into the medium from Na-SP-treated cells indicated that Na-SP stimulates the release of both heparan and chondroitin/dermatan sulfate proteoglycans from the cell layer. Taking these results together it is suggested that Ca-SP and Na-SP promote the release of proteoglycans from vascular endothelial cells by inhibiting the association of the macromolecules with the cell layer.