PMID- 26969382 OWN - NLM STAT- MEDLINE DCOM- 20160921 LR - 20181202 IS - 1618-095X (Electronic) IS - 0944-7113 (Linking) VI - 23 IP - 3 DP - 2016 Mar 15 TI - Anti-angiogenic activity and mechanism of kaurane diterpenoids from Wedelia chinensis. PG - 283-92 LID - S0944-7113(16)00021-0 [pii] LID - 10.1016/j.phymed.2015.12.021 [doi] AB - BACKGROUND: Wedelia chinensis is a traditional medicinal herb used in Asia and it has been reported to possess various bioactivities including anti-inflammatory and anticancer effects. However, its anti-angiogenic activity has never been reported. PURPOSE: To determine the most potent anti-angiogenic component in W. chinensis and its molecular mechanism of action. STUDY DESIGN: Initially, the active fraction of the plant was studied. Then, we determined the active components of the fraction and explored the mechanism of the most active compound. METHODS: The ethanol extract of W. chinensis and its four fractions with different polarities were evaluated for their anti-angiogenic activity in the Zebrafish model using quantitative endogenous alkaline phosphatase (EAP) assay. The molecular mechanism of the most active compound from the active fraction was studied using the real-time polymerase chain reaction (PCR) assay on Zebrafish embryos. The inhibitory effect of the most active compound on the proliferation, invasion and tube formation steps of angiogenesis was evaluated using the vascular endothelial growth factor (VEGF)-induced human umbilical vein endothelial cells (HUVECs) model, and the influences of the active compound on tyrosine phosphorylation of VEGF receptor (VEGFR-2) and its downstream signal pathway were evaluated by western blotting assay. Moreover, its anti-angiogenic effect was further evaluated by the VEGF-induced sprouts formation on aortic ring assay and the VEGF-induced vessel formation of mice on matrigel plug assay, respectively. RESULTS: Petroleum ether (PE) fraction of the plant displayed potent anti-angiogenic activity. Twelve kaurane diterpenoids (1-12) isolated from this fraction showed quite different effects. Compounds 9-12 could dose-dependently inhibit vessel formation in the Zebrafish embryos while the others showed little inhibitory effect. Among the active diterpenoids, compound 10, 3alpha-cinnamoyloxy-9ss-hydroxy-ent-kaura-16-en-19-oic acid (CHKA), possessed the strongest effect, and it affected multiple molecular targets related to angiogenesis including VEGF and angiopoietin in Zerbrafish. Moreover, CHKA significantly inhibited a series of VEGF-induced angiogenesis processes including proliferation, invasion, and tube formation of endothelial cells. Besides, it directly inhibited VEGFR-2 tyrosine kinase activity and its downstream signaling pathways in HUVECs. CHKA also obviously inhibited sprouts formation of aortic ring, and block vessel formation in mice. CONCLUSION: Our findings demonstrate that kaurane diterpenoids is one of anti-angiogenic components in W. chinensis, and CHKA may become a promising candidate for the development of anti-angiogenic agent. CI - Copyright (c) 2016 Elsevier GmbH. All rights reserved. FAU - Huang, Weihuan AU - Huang W AD - Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, 601 Huangpu Avenue West, Guangzhou 510632, P. R. China. FAU - Liang, Yeyin AU - Liang Y AD - Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, 601 Huangpu Avenue West, Guangzhou 510632, P. R. China. FAU - Wang, Jiajian AU - Wang J AD - Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, 601 Huangpu Avenue West, Guangzhou 510632, P. R. China. FAU - Li, Guoqiang AU - Li G AD - Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, 601 Huangpu Avenue West, Guangzhou 510632, P. R. China. FAU - Wang, Guocai AU - Wang G AD - Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, 601 Huangpu Avenue West, Guangzhou 510632, P. R. China. FAU - Li, Yaolan AU - Li Y AD - Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, 601 Huangpu Avenue West, Guangzhou 510632, P. R. China. Electronic address: tliyl@jnu.edu.cn. FAU - Chung, Hau Yin AU - Chung HY AD - Food and Nutritional Sciences Programme, School of Life Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China. Electronic address: anthonychung@cuhk.edu.hk. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20160129 PL - Germany TA - Phytomedicine JT - Phytomedicine : international journal of phytotherapy and phytopharmacology JID - 9438794 RN - 0 (Angiogenesis Inhibitors) RN - 0 (Diterpenes, Kaurane) RN - 0 (Plant Extracts) RN - 0 (Vascular Endothelial Growth Factor A) RN - EC 2.7.10.1 (FLT1 protein, human) RN - EC 2.7.10.1 (Vascular Endothelial Growth Factor Receptor-1) RN - EC 2.7.10.1 (Vascular Endothelial Growth Factor Receptor-2) SB - IM MH - Angiogenesis Inhibitors/*pharmacology MH - Animals MH - Aorta/drug effects MH - Cell Movement/drug effects MH - Diterpenes, Kaurane/*pharmacology MH - Embryo, Nonmammalian/drug effects MH - Human Umbilical Vein Endothelial Cells/*drug effects MH - Humans MH - Mice MH - Mice, Inbred C57BL MH - Molecular Structure MH - Neovascularization, Pathologic/drug therapy MH - Phosphorylation MH - Plant Extracts/pharmacology MH - Rats, Sprague-Dawley MH - Signal Transduction/drug effects MH - Vascular Endothelial Growth Factor A/pharmacology MH - Vascular Endothelial Growth Factor Receptor-1/metabolism MH - Vascular Endothelial Growth Factor Receptor-2/metabolism MH - Wedelia/*chemistry MH - Zebrafish OTO - NOTNLM OT - Anti-angiogenesis OT - Kaurane diterpenoid OT - Mechanism OT - Wedelia chinensis OT - Zerbrafish EDAT- 2016/03/13 06:00 MHDA- 2016/09/23 06:00 CRDT- 2016/03/13 06:00 PHST- 2015/09/01 00:00 [received] PHST- 2015/11/10 00:00 [revised] PHST- 2015/12/12 00:00 [accepted] PHST- 2016/03/13 06:00 [entrez] PHST- 2016/03/13 06:00 [pubmed] PHST- 2016/09/23 06:00 [medline] AID - S0944-7113(16)00021-0 [pii] AID - 10.1016/j.phymed.2015.12.021 [doi] PST - ppublish SO - Phytomedicine. 2016 Mar 15;23(3):283-92. doi: 10.1016/j.phymed.2015.12.021. Epub 2016 Jan 29.