Sulfated polysaccharides are required for collagen-induced vascular tube formation.
We have previously shown that soluble type I collagen can induce vascular tube formation when it contacts the apical side of a confluent endothelial monolayer. In this study we have examined which soluble agent(s) are required for collagen-induced tube formation. Human neonatal foreskin microvascular endothelial cells, maintained in basal medium, were preincubated with each test agent for 2 h prior to the addition of solubilised type I collagen (100 micrograms/ml). After 6 h, tube formation was quantitated using image analysis and expressed as the mean area of tube formation (mm2) per microscopic field of view. Collagen-induced tube formation did not occur in the presence of endothelial cells growth supplement, basic fibroblast growth factor, or normal pooled human serum. In contrast, the addition of heparin at 5 or 50 micrograms/ml caused extensive tube formation (0.22 +/- 0.07 and 0.30 +/- 0.12 mm2, respectively) whereas at 500 micrograms/ml little tube formation occurred (0.03 +/- 0.02 mm2). Protamine sulfate, an antagonist of heparin, inhibited collagen-induced tube formation in a dose-dependent manner. Pentosan polysulfate, dextran sulfate, heparan sulfate, and chondroitin sulfate mimicked the action of heparin. Partially sulfated heparin (de-N-sulfated heparin) stimulated less tube formation compared to heparin (0.15 +/- 0.06 mm2 at 50 micrograms/ml). The nonsulfated polysaccharides, xylan and dextran, had no effect on tube formation. In summary, sulfated polysaccharides are required for collagen-induced vascular tube formation in vitro. The sulfation of these molecules appears to be vital for collagen-induced tube formation.
Jackson
C J
CJ
Sutton Rheumatism Research Laboratory, Department of Rheumatology, Royal North Shore Hospital, St. Leonards, New South Wales, Australia.
Giles
I
I
Knop
A
A
Nethery
A
A
Schrieber
L
L
D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology*; D016264:Dextran Sulfate / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D008297:Male; D009389:Neovascularization, Pathologic / Q000378:metabolism / Q000473:pathology; D010426:Pentosan Sulfuric Polyester / Q000494:pharmacology; D011134:Polysaccharides / Q000378:metabolism / Q000494:pharmacology*; D011479:Protamines / Q000494:pharmacology; D012995:Solubility; D013431:Sulfates / Q000378:metabolism / Q000494:pharmacology
D011134:Polysaccharides; D011479:Protamines; D013431:Sulfates; D010426:Pentosan Sulfuric Polyester; D006493:Heparin; D003094:Collagen; D016264:Dextran Sulfate