Human ryudocan from endothelium-like cells binds basic fibroblast growth factor, midkine, and tissue factor pathway inhibitor.
Ryudocan, a heparan sulfate proteoglycan, was isolated from human endothelium-like EAhy926 cells by a combination of ion-exchange and immunoaffinity chromatography. Purified human ryudocan has biochemical properties similar to those of rat ryudocan isolated from microvascular endothelial cells. Human ryudocan contains only heparan sulfate (HS) glycosaminoglycan chains along with a core protein with an apparent molecular mass of 30 kDa. We evaluated the interactions between purified human ryudocan and several extracellular ligands by using a solid-phase binding assay. We found that basic fibroblast growth factor (bFGF), midkine (MK), and tissue factor pathway inhibitor (TFPI) exhibit significant ryudocan binding. Heparitinase (but not chondroitin ABC lyase) treatment destroyed the ability of ryudocan binding to bFGF, MK, and TFPI. Heparin and HS, but not chondroitin sulfate, inhibited such ryudocan binding. Thus, the HS chains of ryudocan appear to be responsible for its binding to bFGF, MK, and TFPI. The apparent dissociation constants for purified ryudocan were as follows: bFGF, 0.50 nM; MK, 0.30 nM; and TFPI, 0.74 nM. Immunohistochemical analysis revealed that ryudocan was expressed in fibrous connective tissues, peripheral nerve tissues, and placental trophoblasts. These findings suggest that ryudocan may possess multiple biological functions, such as bFGF modulation, neurite growth promotion, and anticoagulation, via HS-binding effectors in the cellular microenvironment.
Kojima
T
T
First Department of Internal Medicine, Nagoya University School of Medicine, Nagoya 466, Japan.
Katsumi
A
A
Yamazaki
T
T
Muramatsu
T
T
Nagasaka
T
T
Ohsumi
K
K
Saito
H
H
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001345:Autoradiography; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D002852:Chromatography, Ion Exchange; D017077:Culture Media, Conditioned; D016207:Cytokines / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000494:pharmacology*; D006801:Humans; D006822:Hybrid Cells; D007150:Immunohistochemistry; D007457:Iodine Radioisotopes; D007700:Kinetics; D008024:Ligands; D008074:Lipoproteins / Q000378:metabolism*; D008175:Lung Neoplasms; D008562:Membrane Glycoproteins; D000078792:Midkine; D008969:Molecular Sequence Data; D011485:Protein Binding; D011509:Proteoglycans / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D011817:Rabbits / Q000276:immunology; D051381:Rats; D015842:Serine Proteinase Inhibitors / Q000378:metabolism; D053671:Syndecan-4; D014407:Tumor Cells, Cultured
D000906:Antibodies; D002352:Carrier Proteins; D017077:Culture Media, Conditioned; D016207:Cytokines; D006025:Glycosaminoglycans; D007457:Iodine Radioisotopes; D008024:Ligands; D008074:Lipoproteins; D008562:Membrane Glycoproteins; D011509:Proteoglycans; C508304:SDC4 protein, human; C508307:Sdc4 protein, rat; D015842:Serine Proteinase Inhibitors; D053671:Syndecan-4; C051928:lipoprotein-associated coagulation inhibitor; D016222:Fibroblast Growth Factor 2; D000078792:Midkine