Abstract
Anthocyanins richly exist in mulberry plants and have been well characterized to have various bioactive properties. However, the antimetastasis properties of mulberry anthocyanins (MACs) remain unclear. The objectives of this study were to investigate the inhibitory effects of MACs on the metastasis of B16-F1 cells under noncytotoxic concentrations. Further investigation revealed that the antimetastatic effect of MACs was also evident in a C57BL/6 mice model. First, MACs exhibited an inhibitory effect on the migration ability by wound healing assay and Boyden chamber assay. In the cancer cell metastasis process, matrix degrading proteinases are required. B16-F1 cells treated with MACs at various concentrations showed reduced extracellular matrix (ECM) proteinases including matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) by gelatin zymography assay. The results of the Western blotting assay demonstrated that the expression levels of Ras, phosphoinositide 3-kinase (PI3K), phospho-Akt, and nuclear factor kappa B (NF-kappaB) in the MACs-treated B16-F1 cells were reduced. Therefore, it was suggested that MACs could mediate B16-F1 cell metastasis by reduction of MMP-2 and MMP-9 activities involving the suppression of the Ras/PI3K signaling pathway. Besides, B16-F1 melanoma cells were also injected into the right groin of the C57BL/6 mice, and the mice were fed with MACs at the same time. The hematoxylin-eosin stain (H&E stain) and immunohistochemistry stain showed that the MACs inhibited the mtastasis of B16-F1 cells in vivo. Taken together, the findings proved the inhibitory effect of MACs on the growth and metastasis of B16-F1 cells. These results indicated that MACs might be offered for future application as an antimetastatic agent.
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