Low molecular weight fucoidan promotes FGF-2-induced vascular tube formation by human endothelial cells, with decreased PAI-1 release and ICAM-1 downregulation

Abstract

Basic fibroblast growth factor (FGF-2) is a member of the heparin-binding growth factor family involved in neovascularization [ [1] Ribatti D. Urbinati C. Nico B. Rusnati M. Roncali L. Presta M. Endogenous basic fibroblast growth factor is implicated in the vascularization of the chick embryo chorioallantoic membrane. Dev. Biol. 1995; 170: 39-49 Crossref PubMed Scopus (148) Google Scholar ]. Compounds compatible with therapeutic use and able to modulate FGF-2 activity could be of interest for revascularization of ischemic areas. During neovascularization, endothelial cells first acquire invasive properties by secreting proteases that degrade the basal membrane [ [2] Schnaper H.W. Barnathan E.S. Mazar A. Maheshwari S. Ellis S. Cortez S.L. et al. Plasminogen activators augment endothelial cell organization in vitro by two distinct pathways. J. Cell. Physiol. 1995; 165: 107-118 Crossref PubMed Scopus (80) Google Scholar ]. Endothelial cells synthesize and release fibrinolytic system proteases, such as tissue-type and urinary plasminogen activators (t-PA and u-PA, respectively), as well as their physiological inhibitor, plasminogen activator inhibitor-1 (PAI-1). They also express urokinase receptor (uPAR) on their surface. Endothelial cells then migrate and differentiate, leading to the formation of new vascular tubes. These steps in angiogenesis alter the expression and/or activity of integrins and cellular adhesion molecules such as PECAM-1 and ICAM-1, allowing new cell–cell interactions and binding to ligands of the extracellular matrix [ 3 DeLisser H.M. Christofidou-Solomidou M. Strieter R.M. Burdick M.D. Robinson C.S. Wexler R.S. et al. Involvement of endothelial PECAM-1/CD31 in angiogenesis. Am. J. Pathol. 1997; 151: 671-677 PubMed Google Scholar , 4 Griffioen A.W. Damen C.A. Martinotti S. Blijham G.H. Groenewegen G. Endothelial intercellular adhesion molecule-1 expression is suppressed in human malignancies: the role of angiogenic factors. Cancer Res. 1996; 56: 1111-1117 PubMed Google Scholar ]. In a previous study, we reported that a low molecular weight fucoidan (LMWF) of marine plant origin (brown algae Ascophyllum nodosum) potentiated FGF-2-induced tube formation by human endothelial cells from umbilical vein (HUVEC) involving the overexpression of the α6 integrin subunit [ [5] Chabut D. Fischer A.M. Colliec-Jouault S. Laurendeau I. Matou S. LeBonniec B. et al. Low molecular weight fucoidan and heparin enhance the FGF-2-induced tube formation of endothelial cells through heparan sulfate-dependent alpha 6 over-expression. Mol. Pharmacol. 2003; 64: 696-702 Crossref PubMed Scopus (75) Google Scholar ]. HUVEC, when stimulated with FGF-2 and a low molecular weight heparin (dalteparin), also formed tubes, although to a lesser extent [ [5] Chabut D. Fischer A.M. Colliec-Jouault S. Laurendeau I. Matou S. LeBonniec B. et al. Low molecular weight fucoidan and heparin enhance the FGF-2-induced tube formation of endothelial cells through heparan sulfate-dependent alpha 6 over-expression. Mol. Pharmacol. 2003; 64: 696-702 Crossref PubMed Scopus (75) Google Scholar ]. The purpose of the present study was to determine whether LMWF, as compared to dalteparin (concomitant with α6 overexpression) modulates the expression of cellular adhesion molecules and the release of proteins involved in pericellular lysis. Compared to untreated cells, HUVEC pretreated for 72 h with FGF-2 (5 ng/ml) differentiated in a partially organized capillary network (Fig. 1) on a basal membrane substitute (Matrigel). Flow cytometry showed that ICAM-1, although expressed on untreated cells, was no longer detectable on HUVEC treated with FGF-2 (Fig. 1). No change was apparent with respect to PECAM-1 labeling. As shown in Fig. 1, HUVEC, when incubated simultaneously with FGF-2 and 10 μg/ml of LMWF or dalteparin, branched and formed tubes with closed areas, denoting intense proangiogenic activity that was more pronounced with LMWF. No further effect on ICAM-1 or PECAM-1 expression was detected after addition of polysaccharides to FGF-2. However, a direct relationship between FGF-2-induced angiogenesis and downregulation of ICAM-1 on tumor-infiltrating endothelial cells has been reported [ [4] Griffioen A.W. Damen C.A. Martinotti S. Blijham G.H. Groenewegen G. Endothelial intercellular adhesion molecule-1 expression is suppressed in human malignancies: the role of angiogenic factors. Cancer Res. 1996; 56: 1111-1117 PubMed Google Scholar ]. The absence of detectable ICAM-1 expression on HUVEC surface should potentiate cell migration and vascular tube formation on Matrigel.