Ethanol extract of Gleditsia sinensis thorn suppresses angiogenesis in vitro and in vivo

Abstract

BackgroundGleditsia sinensis thorns have been widely used in traditional Korean medicine for the treatment of several diseases, including obesity, thrombosis, and tumor-related diseases. The aim of the study is to determine the antiangiogenic effect of Gleditsia sinensis thorns in vitro and in vivo in a bid to evaluate its potential as an anticancer drug.MethodsEthanol extract of Gleditsia sinensis thorns (EEGS) were prepared and used for in vitro and in vivo assays. In vitro antiangiogenic effect of EEGS was determined in HUVEC primary cells by cell migration and tube formation assays. In vivo antiangiogenic effect of EEGS was determined by measuring vessel formation and vascular endothelial cells migrating into the implanted matrigels in nude mice. The angiogenesis-related proteins of which expression levels were altered by EEGS were identified by proteomic analysis.ResultsEEGS exerted a dose-dependent antiproliferative effect on HUVEC cells without significant cytotoxicity. Angiogenic properties, such as cell migration and tube formation, were significantly inhibited by EEGS in a dose-dependent manner. New vessel formation was also suppressed by EEGS, as determined by the directed in vivo angiogenesis assays in nude mice. EEGS reduced the expression of proangiogenic proteins, endothelin 1 and matrix metallopeptidase 2, in HUVEC cells.ConclusionsOur findings suggest that EEGS can inhibit angiogenesis by down-regulating proangiogenic proteins, and therefore it should be considered as a potential anticancer drug targeting tumor-derived angiogenesis.

Cytochalasin H, an Active Anti-angiogenic Constituent of the Ethanol Extract of <i>Gleditsia sinensis</i> Thorns

“…19) In the present study, we attempted to identify the active constituents of EEGS that display these cytostatic and anti-angiogenic properties. Unexpectedly, activityguided fractionation and spectral studies identified cytochalasin H, a fungal secondary metabolite, as an active constituent of G. sinensis thorns.…”

“…19) EEGS efficiently inhibited cellular mobility and 3-dimensional vascular tube formation by HUVEC primary cells, which led to the suppression of vessel formation in the matrigels that had been premixed with pro-angiogenic proteins and implanted in nude mice. In the present study, we tested the in vitro anti-angiogenic potential of G. sinensis thorns which had been purchased from 5 different herbal plant suppliers.…”

“…Statistical analyses revealed that the tube length and branch number were significantly reduced by the treatment with 125 nmol/L of cytochalasin H. In our previous study, proteomic and immunoassay studies revealed that both the intracellular and extracellular expression of the pro-angiogenic protein EDN1 was reduced by treatment with EEGS in a dosedependent manner. 19) In an effort to verify whether cytochalasin H, an active anti-angiogenic constituent isolated from the EEGS, could also down-regulate EDN1 expression, the extracellular levels of EDN1 that had been released by HUVEC were quantified using an ELISA. As shown in Fig.…”

“…19) In brief, angioreactors that had been pre-filled with growth factor-reduced BME with combinations of VEGF, FGF2, and increasing concentrations of the drug were subcutaneously implanted into the dorsal flanks of 6 week old C57BL/6 nude mice. After 12 d, the implanted angioreactors were harvested, and the degree of new vessel formation was determined by FITC-lectin mediated quantification of the vascular endothelial cells that had infiltrated into the angioreactors.…”

“…17,18) The results of our previous study demonstrated that the ethanol extract of G. sinensis thorns (EEGS) efficiently inhibits the angiogenic activity of HUVEC and reduces pro-angiogenic factor-induced new vessel formation in a matrigel plug assay. 19) To date, there have been no scientific reports identifying the active anti-angiogenic constituents of G. sinensis thorns. In the present study, we isolated and identified cytochalasin H, as an active anti-angiogenic constituent of the extract of G. sinensis thorns, using activity-guided fractionation.…”