In vitro and in vivo anti-angiogenic activities of milk sphingolipids

Anti-angiogenic therapies aimed at halting new blood vessel formation are now being extensively studied as inhibitors of excessive angiogenesis. Conversely, compounds with ability to stimulate angiogenesis are being considered as a therapeutic approach for insufficient angiogenesis. Food-borne bioactive compounds such as genistein, resveratrol, curcumin, the Bowman-Birk inhibitor, and catechins are being potentially established as good candidates for angioprevention.; The aim of our study was to determine the anti- or pro-angiogenic activity of milk-based glycosphingolipids such as C6-ceramide (Cer), Sphingomyelin (SPM) and Glucosylceramide (GluCer), in vitro, using breast cancer (MCF-7), colon cancer (Caco-2) and prostate cancer (DU-145) cell-lines, on angiogenic factors such as vascular endothelial growth factor (VEGF), cathepsin-D and hypoxia inducing factor-1alpha (HIF-1alpha) expression and cell migration under normoxia and hypoxia. Another aim was to conduct an in vivo study using chorioallantoic membrane (CAM) and zebrafish model system to substantiate the in vitro results.; Breast cancer cells (MCF-7) treated with SPM had reduced cell migration under hypoxic conditions. Cathepsin-D expression under SPM treated MCF-7 cells was significantly lower under both conditions. GlcCer had significant apoptotic activity under hypoxic MCF-7 cells. Colon cancer cells (Caco-2) treated with Cer had reduced cell growth at >50 muM under normoxic as well as hypoxic conditions. Cathepsin-D, cell migration and HIF-1alpha expression were significantly reduced under hypoxic condition. SPM had low cathepsin-D levels and cell migrations in normoxic and hypoxic conditions as well as low HIF-1alpha at hypoxic condition. In GlcCer treated cells, the levels of cathepsin-D and cell migration were reduced under normoxic and hypoxic conditions.; Prostate cancer cells (DU-145) exposed to SPM had reduced cell viability. All the compounds had lower levels of VEGF expression at normoxic conditions at 50 muM exposure; only GlcCer had lower VEGF expression under hypoxic condition. The cell migration was reduced under normoxic condition and also for cells exposed to Cer under hypoxic condition. In vivo results showed ceramide was anti-angiogenic as confirmed by both CAM assay as well as zebrafish model. SPM proved to facilitate sprouting, however, the blood vessels looked dilated. GlcCer disrupted the neovascularization in CAM model and restricted the ISV formation in zebrafish.