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Fig. 5
Simultaneous application of Tsg and BMPER in functional in vitro and in vivo angiogenesis assays abolishes induction of proangiogenic effects. (A–C) Tsg-stimulated HUVEC sprouting was inhibited when recombinant BMPER protein was added in equal amounts. (A) HUVECs were incubated with the indicated proteins and subjected to a tube formation assay. Representative micrographs of control, Tsg, BMPER and Tsg + BMPER simultaneous application are shown. Scale bars: 200µm. (B) Quantification of cumulative sprout length of capillary-like structures. (C) Number of branch points. (D,E) BMPER-stimulated HUVEC sprouting was inhibited when recombinant Tsg protein was added in equal amounts. (D) Quantification of cumulative sprout length of capillary-like structures. (E) Number of branch points. Values are means ± s.e.m.; n = 3; *P<0.001 versus control. (F) Quantification of mouse Matrigel plug assay with the indicated concentrations of recombinant Tsg and BMPER protein. Values are means ± s.e.m.; n = 3. (G) Western blot analysis of HUVECs stimulated with BMPER, Tsg and the combination of BMPER + Tsg (concentration = 0.9nM) performed with the indicated antibodies. Representative western blots of one of three independent experiments are shown.