Fig. 5: SSO treatment diminishes tube formation and angiogenesis in RMS cells.

A A protein profiler array was used to assess the expression of angiogenic factors, and the expression was quantified. B Quantification for n = 2, statistics were done using 2-way ANOVA ****p < 0.0001. C Rh30 cells were treated with NS and SSO55 compounds and lysed for Western blot analysis. MMP-9 and Actin are shown. Quantification of MMP-9 expression, n = 3, Mann–Whitney test. D Human umbilical vein endothelial cells (HUVEC) were incubated with either PBS or media from Rh30 cells treated with either NS or SSO55 alone or in combination with IGF-1R antibody. E The vessels/branch formation was measured and quantified using CD31 antibody staining. F Rh30 cells were seeded in a six-well plate, transfected with NS or SSO55. After 24 h, all wells but the controls were treated with IGF-1R antibody (depicted in red). After 30 min, one NS and SSO55 treated well was harvested and IGF-2 was added to the remaining wells. The wells were harvested with RIPA buffer supplemented with protease and phosphatase inhibitors after 2, 5, and 10 min. The blots for p-AKT (MW ~60 kDa), total AKT (MW ~60 kDa), and GAPDH (MW ~37 kDa) are depicted. The relative quantification (pAKT/ loading control) was done using ImageJ and is shown in the graph (G). Statistics for n = 3, paired t-test, p-value = 0.357 and 0.417, respectively. Results are shown as the standard error of the mean (±SEM).