Fig 4. [NSIS6S]-[NSIS]₅ inhibits FGF2 secreted from cancer cells.

A to B, FGF2 concentration in HEC-1-B and FGF2-B9 cell lysates and conditioned medium was determined by ELISA. Two independent experiments were performed and the data are expressed as the mean ± SD. C, angiogenic potential of conditioned media collected from HEC-1-B and exogenous FGF2 overexpressing HEC-1-B cell line FGF2-B9 was evaluated in three dimensional fibrin gel HUVEC bead assay. Conditioned medium collected from HEC-1-B cells induced minimal endothelial tube outgrowth insensitive to blocking anti-FGF2 antibody or treatment with anti-angiogenic dodecasaccharides [NSIS]₆ and [NSIS6S]-[NSIS]₅ (upper panel). Treatment with FGF2-B9 conditioned medium resulted in increased outgrowth of endothelial tubes which was reduced by treatment with anti-FGF2 antibody, [NSIS]₆ and [NSIS6S]-[NSIS]₅. Scale bars represent 100 μm. D, quantification of a number of endothelial tubes per bead as shown in C. The number of tubes per bead induced by HEC-1-B conditioned medium is expressed as 1 (control). The effect of FGF2-B9 conditioned medium and all treatments is shown as fold change compared to the control. Two independent experiments were performed in triplicate. The data are expressed as the mean ± SD (n = 2). *, P < 0.001; ‡, P < 0.05.