Evidence that platelets promote tube formation by endothelial cells on matrigel.
The involvement of platelets in neovascularization was investigated in the matrigel tube formation assay, an in vitro model of angiogenesis. Platelets promoted the formation of capillary-like structures (expressed as relative tube area) number- and time-dependently. Relative tube area increased from 0.98+/-0.02 (n = 8) in the presence of 6.25 x 10(4), to 3.21+/-0.12 (n=8) in the presence of 10(6) platelets/well compared to 0.54+/-0.04 (n=8) in their absence. This increase was unaffected by acetyl salicylic acid (ASA), apyrase, and hirudin. Photographs from representative experiments, showed that platelets adhered along the differentiating endothelium. Addition of alpha-thrombin (0.1-1 i.u. ml(-1)), the nitric oxide (NO) donor sodium nitroprusside (SNP; 1-100 microM) or the NO synthase inhibitor, L-NG-arginine-methylester (L-NAME, 30-300 microM) to the assay, had no effect on tube formation compared to that seen with platelets alone. Neuraminidase (0.01 i.u./10(7) platelets), which strips sialic acid residues from membrane glycoproteins, abolished the promoting effect of platelets on tube formation. The relative tube area in the presence of neuraminidase-treated platelets was 0.81+/-0.03 (n = 8), in the presence of untreated platelets 1.69+/-0.09, P<0.001 (n=8) and in the absence of platelets, 0.80+/-0.04 (n=8). The tetrapeptide Arg-Gly-Asp-Ser (RGDS; 20-200 microM) which inhibits von Willebrand factor, fibrinogen and fibronectin-mediated adhesion, had no effect on the promoting effect of platelets on tube formation. These results indicate that platelets promote angiogenesis in vitro. This effect is largely independent from activation by alpha-thrombin, is not modified by manipulating NO and prostaglandin metabolism and proceeds possibly through adhesion of the platelets to the differentiating endothelium.
Pipili-Synetos
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Department of Pharmacology, Medical School, University of Patras, Greece.
Papadimitriou
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Maragoudakis
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D001672:Biocompatible Materials; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D003094:Collagen; D046508:Culture Techniques; D004338:Drug Combinations; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006490:Hemostatics / Q000494:pharmacology; D006801:Humans; D007797:Laminin; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology*; D009439:Neuraminidase / Q000494:pharmacology; D009599:Nitroprusside / Q000494:pharmacology; D011509:Proteoglycans; D013917:Thrombin / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology
D001672:Biocompatible Materials; D004338:Drug Combinations; D004791:Enzyme Inhibitors; D006490:Hemostatics; D007797:Laminin; D011509:Proteoglycans; D014665:Vasodilator Agents; C059485:matrigel; D009599:Nitroprusside; D003094:Collagen; D009439:Neuraminidase; D013917:Thrombin; D019331:NG-Nitroarginine Methyl Ester