No abstract

Proteoglycans that modulate the activities of growth factors, chemokines, and coagulation factors regulate in turn the vascular endothelium with respect to processes such as inflammation, hemostasis, and angiogenesis. Endothelial cell-specific molecule-1 is mainly expressed by endothelial cells and regulated by pro-inflammatory cytokines (Lassalle, P., Molet, S., Janin, A., Heyden, J. V., Tavernier, J., Fiers, W., Devos, R., and Tonnel, A. B. (1996) J. Biol. Chem. 271, 20458 -20464). We demonstrate that this molecule is secreted as a soluble dermatan sulfate (DS) proteoglycan. This proteoglycan represents the major form either secreted by cell lines or circulating in the human bloodstream. Because this proteoglycan is specifically secreted by endothelial cells, we propose to name it endocan. The glycosaminoglycan component of endocan consists of a single DS chain covalently attached to serine 137. Endocan dose-dependently increased the hepatocyte growth factor/scatter factor (HGF/SF)-mediated proliferation of human embryonic kidney cells, whereas the nonglycanated form of endocan did not. Moreover, DS chains purified from endocan mimicked the endocan-mediated increase of cell proliferation in the presence of HGF/SF. Overall, our results demonstrate that endocan is a novel soluble dermatan sulfate proteoglycan produced by endothelial cells. Endocan regulates HGF/SF-mediated mitogenic activity and may support the function of HGF/SF not only in embryogenesis and tissue repair after injury but also in tumor progression.In the last few years, the vascular endothelium has been shown to play a crucial role in inflammation, coagulation, angiogenesis, and tumor invasion, primarily through the fine regulation of receptor-ligand interactions and secretion of different mediators. Endothelial cells also express several proteoglycans such as decorin, biglycan, PG-100, glypican, and members of the syndecan family that regulate intercellular interactions and activation processes.Proteoglycans are complex macromolecules that consist of a polypeptide with one or more glycosaminoglycan chains covalently bound to a serine (or rarely a threonine) residue. Different families of proteoglycans have been described (1-3) as follows: heparan sulfate, chondroitin sulfate, dermatan sulfate, and keratan sulfate proteoglycans. Heparan sulfate proteoglycans have come to particular prominence recently because of their multiple regulatory interactions with growth factors, enzymes, enzyme inhibitors, and components of the extracellular matrix (4, 5). Dermatan sulfate proteoglycans (DSPGs) 1 from human fluids become of particular importance during inflammation and response to injury (6 -11), contributing, for example, to the majority of the FGF-2-dependent cell proliferation involved in the regulation of wound repair (6). DSPGs are also known to bind to many HS-binding proteins, including FGF-2, interleukin-7 (12), platelet factor 4 (13), fibronectin (14), and heparin cofactor II (15). Hepatocyte growth factor/scatter factor (HGF/SF) also binds ...

The hypothesis that neuropilin-1 (Npn-1) may interact with heparin-binding proteins other than vascular endothelial growth factor has been tested using an optical biosensor-based binding assay. The results show that fibroblast growth factor ( ). These results suggest that Npn-1 possesses a "heparin" mimetic site that is able to interact at least in part through ionic bonding with the heparin binding site on many of the proteins studied. Npn-1 was also found to potentiate the growth stimulatory activity of FGF-2 on human umbilical vein endothelial cells, indicating that Npn-1 may not just bind but also regulate the activity of heparin-binding proteins.

Glioblastomas (GBMs) are highly malignant tumors characterized by microvascular proliferation and the pseudopalisading pattern of necrosis. Investigations have, therefore, focused on vascular and endothelial cell biology in GBM. Endocan, also called endothelial cell-specific molecule-1, is a proteoglycan that is secreted by endothelial cells and upregulated by proangiogenic factors. We found that endocan is not only expressed in vitro by endothelial cells but also in the T98G and U118MG human GBM cell lines. In U118MG cells, tumor necrosis factor and fibroblast growth factor 2 upregulated endocan production, whereas exposure to hypoxia or cobalt chloride, an inducer of hypoxia inducible factor 1, increased endocan release without affecting cell viability. Endocan expression in 82 brain tumors was studied by immunohistochemistry. Endocan immunoreactivity was detected in hyperplastic endothelial cells in high-grade gliomas, mostly at the tumor margins; endothelial cells were mostly endocan negative in low-grade gliomas, and it was never detected in the cerebral cortex distant from the tumors. Tumor cells in high-grade but not low-grade gliomas also expressed endocan, and it was detected in palisading cells surrounding areas of necrosis in GBM. Endothelial cell endocan immunoreactivity also correlated with shorter survival in glioma patients. Taken together, these results suggest that endocan is associated with abnormal vasculature in high-grade gliomas.

We examine the relationship between the chain length of heparin-derived oligosaccharides, fibroblast growth factor (FGF)-2 binding kinetics and the ability of the oligosaccharides to allow FGF-2-induced proliferation of chlorate-treated rat mammary fibroblasts. First, using an optical biosensor, we show that FGF-2 did not bind disaccharides, but definitively bound to tetrasaccharides. As the chain length increased from tetrasaccharide to octasaccharide, there was a substantial increase in k(ass) (564000 M(-1) x s(-1) to 2000000 M(-1) x s(-1), respectively) and affinity (K(d) 77 nM to 11 nM, respectively) for FGF-2. From decasaccharides and longer, the k(ass) and affinity for FGF-2 was reduced substantially (tetradecasaccharide k(ass) 470000 M(-1) x s(-1), K(d) 30 nM). In chlorate-treated, and hence sulphated, glycosaminoglycan-deficient cells, FGF-2 alone or in the presence of disaccharides did not stimulate DNA synthesis and it only elicited an early transient dual phosphorylation of p42/44 mitogen-activated protein kinase (MAPK). In the same cells FGF-2 in the presence of tetrasaccharides and longer oligosaccharides was able to restore DNA synthesis and enable the sustained dual phosphorylation of p42/44(MAPK). However, the oligosaccharides from tetrasaccharides to octasaccharides were less potent in proliferation assays than deca- and longer oligosaccharides. Therefore, there was no correlation between the binding parameters and the potency of the oligosaccharides in DNA synthesis assays. These results demonstrate that tetrasaccharides are able to bind FGF-2 and enable FGF-2 to stimulate cell proliferation, which sets important boundary conditions for models of the FGF-2-heparan sulphate-FGF receptor complex.