Abstract: Background and purpose: Dihydromyricetin (DHM) exerts anti-cancer effects by inhibiting cell cycle, promoting apoptosis and decreasing angiogenesis. The aim of this study was to investigate the anti-angiogenic effect of DHM on the gastric cancer in vitro and in vivo and its mechanisms. Methods: Human umbilical vein endothelial cells (HUVECs) and human gastric cancer MKN28 cells were exposed to increasing doses of DHM. Nontoxic doses of DHM were screened by cell counting kit-8 (CCK-8) assay. The migratory capacity of HUVECs was analyzed using transwell migration chambers. The capillary tube formation assay was used to evaluate the angiogenic activity of HUVEC. Western blot was employed to analyze the levels of vascular endothelial growth factor A (VEGFA), phospho-vascular endothelial growth factor receptor (VEGFR) 2, phospho-extracellular signal-regulated kinase (ERK), phospho-c-Jun NH2-terminal kinase (JNK) and phospho-p38 in the HUVEC and MKN28 cells. Matrigel plug assay was employed to analyze the anti-angiogenic potential of DHM in vivo. MKN28 cells were implanted subcutaneously into right flank of nude mice to observe the anti-cancer effect of DHM in vivo. Immunohistochemistry was employed to analyze the levels of Ki-67, CD34 and VEGFA in the xenografts after DHM treatment. Results: DHM exerted no cytotoxic effects on the growth of both HUVECs and MKN28 cells from the concentration of 0.5 μmol/L to 2.5 μmol/L. However, nontoxic doses of DHM treatment significantly inhibited HUVEC migration and tube formation, and the suppressive effects of DHM on HUVEC migration and tube formation were partly reversed by the addition of VEGFA. In MKN28 cells, the levels of VEGFA and phospho-ERK declined in a dosage-dependent manner after DHM treatment, whereas the addition of DHM did not change the expression levels of phospho-JNK and phospho-p38. DHM treatment also triggered a dosage-dependent phospho-VEGFR2 down-regulation in HUVEC. In vivo experiment showed the angiogenesis and the growth of subcutaneously implanted tumors were notably inhibited by DHM at low dosage with almost no chemotoxic side effects. The protein level of CD34 and VEGFA in tumor tissues were obviously decreased by the treatment of DHM. Conclusion: A low dosage of DHM inhibits the angiogenesis of gastric cancer both in vitro and in vivo. And the anti-angiogenic function of DHM, at least partly, is realized by inactivating ERK/VEGFA/VEGFR2 signaling pathway.

Key words: Dihydromyricetin, Gastric cancer, Extracellular signal-regulated kinase, Angiogenesis