Master of Science (MS)
School of Nutrition and Food Sciences
The purpose of this research was to investigate the in vitro and in vivo anti-angiogenic properties of ceramide isolated from oyster (Crassostrea virginica). Ceramide was isolated from oyster by silicic acid chromatography, purified by alkaline hydrolysis, and analyzed by electrospray ionization mass spectrometry (ESI-MS). In vitro, the activity of ceramide on human hormone-dependent (MCF-7) and hormone-independent (MDA-MB-435s) breast cancer cells was evaluated for cell viability and proliferation, VEGF-induced cell migration and invasion, apoptosis, and autophagy. The activity of ceramide was also evaluated for VEGF-induced human umbilical vein endothelial cells (HUVEC) tube formation. The protein levels of vascular endothelial growth factor (VEGF), epidermal growth factor (EGF) in MCF-7 and MDA-MB435s breast cancer cells treated with ceramide were examined by ELISA. Autophagy was observed by the presence of autophagic vacuoles, and monodansylcadaverine was used as a marker for the detection of autolysosomes. The levels of the transcription factor NF-kB and the activity of phosphatidylinositol kinase were determined by ELISA and Western blot respectively from the cellular extracts of control and ceramide treated breast cancer cells. The activity of ceramide, in vivo, was evaluated for inhibition of bFGF-induced angiogenesis with a matrigel plug assay. The yield of ceramide methyl aminoethylphosphonate from deshelled oysters was 0.16% on a dry weight basis. The molecular size of isolated ceramide was 413 daltons. The viability of MCF-7 and MDA-MB-435s cells exposed to 125 μM of ceramide for 48 h was reduced to 76% and 85%, respectively. The viability of MCF-7 and MDA-MB-435s cells exposed to 250 μM of ceramide for 48 h was reduced to and 38% and 45%, respectively. Ceramide at 50 μM inhibited tube formation by HUVEC in the presence of VEGF at 10ng/ml. Ceramide at 125 μM inhibited VEGF-induced MDA-MB-435s cell migration and invasion and also decreased VEGF, EGF levels, and NF-kB activity in the conditioned media. Ceramide from oyster mediated MCF-7 and MDA-MB-435s breast cancer cell death by autophagy, respectively. In vivo, ceramide at 30mg/kg body weight caused a 57% reduction in hemoglobin levels in the matrigel plug assay within seven days. Ceramide from oyster inhibited angiogenesis in vitro and in vivo.
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Chintalapati, Madhavi, "Inhibition of Hormone Dependent and Independent Breast Cancer Cells by Oyster(Crassostrea virginica) Ceramide" (2006). LSU Master's Theses. 2153.
Jack N Losso