Chintalapati Madhavi, Truax Robert, Stout Rhett, Portier Ralph, Losso Jack N
Department of Food Science, Louisiana State University Agriculture Center, Louisiana State University, Baton Rouge, Louisiana 70803, USA.
J Agric Food Chem. 2009 Jun 24;57(12):5201-10. doi: 10.1021/jf803818y.
Ceramide methylaminoethylphosphonate (CMAEPn) was isolated from eastern oyster ( Crassostrea virginica ) and screened against in vitro and in vivo angiogenesis and against MCF-7 and MDA-MB-435s breast cancer cell lines. In vitro angiogenesis was evaluated by the vascular endothelial growth factor (VEGF)-induced human umbilical vein endothelial cell (HUVEC) tube formation assay. MCF-7 and MDA-MB-435s cell viability was evaluated by the CellTiter 96 AQ(ueous) One Solution Cell Proliferation assay. Apoptosis was evaluated by the caspase-9 assay, autophagy by acridine orange staining and beclin-1 level. Our study indicates that CMAEPn at 50 microM inhibited VEGF-induced tube formation by HUVEC. The viability of MCF-7 and MDA-MB-435s breast cancer cells exposed to 125 microM CMAEPn for 48 h was reduced to 76 and 85%, respectively. The viability of MCF-7 and MDA-MB-435s cells exposed to 250 microM CMAEPn for 48 h under the same conditions was reduced to 38 and 45%, respectively. CMAEPn at 125 microM inhibited VEGF-induced MDA-MB-435s cell migration and invasion. CMAEPn at 125 microM also decreased VEGF, EGF levels in the conditioned media, PI3K, IkappaB phosphorylation and degradation in the cytoplasmic extracts, and NFkappaB nuclear translocation. Both acridine orange staining and beclin-1 indicated autophagic cell death in MCF-7 and MDA-MB-435s cells, respectively. In vivo, CMAEPn at 30 mg/kg body weight inhibited bFGF-induced angiogenesis and caused a 57% reduction in hemoglobin levels in the matrigel plug assay within 7 days. This is the first report on CMAEPn-inhibited angiogenesis both in vitro and in vivo.
从美国东海岸牡蛎（Crassostrea virginica）中分离出神经酰胺甲基氨基乙基膦酸酯（CMAEPn），并对其进行体外和体内血管生成以及针对MCF - 7和MDA - MB - 435s乳腺癌细胞系的筛选。通过血管内皮生长因子（VEGF）诱导的人脐静脉内皮细胞（HUVEC）管形成试验评估体外血管生成。通过CellTiter 96 AQ（水溶液）单溶液细胞增殖试验评估MCF - 7和MDA - MB - 435s细胞活力。通过caspase - 9试验评估细胞凋亡，通过吖啶橙染色和beclin - 1水平评估自噬。我们的研究表明，50微摩尔的CMAEPn抑制VEGF诱导的HUVEC管形成。暴露于125微摩尔CMAEPn 48小时的MCF - 7和MDA - MB - 435s乳腺癌细胞活力分别降至76%和85%。在相同条件下，暴露于250微摩尔CMAEPn 48小时的MCF - 7和MDA - MB - 435s细胞活力分别降至38%和45%。125微摩尔的CMAEPn抑制VEGF诱导的MDA - MB - 435s细胞迁移和侵袭。125微摩尔的CMAEPn还降低了条件培养基中的VEGF、EGF水平，细胞质提取物中的PI3K、IkappaB磷酸化和降解以及NFkappaB核转位。吖啶橙染色和beclin - 1分别表明MCF - 7和MDA - MB - 435s细胞中存在自噬性细胞死亡。在体内，体重30毫克/千克的CMAEPn在7天内抑制碱性成纤维细胞生长因子（bFGF）诱导下的血管生成，并使基质胶栓塞试验中的血红蛋白水平降低57%。这是关于CMAEPn在体外和体内抑制血管生成的首次报道。
