Anti-angiogenic effects of aqueous extract from Agrostemma githago L. seed in human umbilical vein endothelial cells via regulating Notch/VEGF, MMP2/9, ANG2, and VEGFR2

Abnormal angiogenesis contributes to the pathogenesis of various diseases. The medicinal usage of Agrostemma githago L. seed (A. githago herein) has been stated in traditional medicine. This study aims to investigate the anti-angiogenic potential of aqueous extract of A. githago. In order to test the effect of A. githago extract, its impact on HUVECs, T98G, and HGF2PI2 cells was assessed by looking at cellular viability, changes in the distribution of cells in different phases of the cell cycle, induction of oxidative stress, and apoptosis. In addition, the release of VEGF, ANG2, and MMP2/9 factors, along with the expressions of the critical Notch signaling pathway players and VEGF receptors (VEGFR), was measured. Furthermore, a gamma-secretase inhibitor (LY411575) was applied to determine whether Notch inhibition restores A. githago effects. As a further characterization, total phenolic and flavonoid contents of A. githago were estimated, and five triterpene saponin compounds were identified using LC-ESI-MS. In response to A. githago extract, a reduction in total cell viability, along with the induction of ROS and apoptosis, was detected. Exposure to the A. githago extract could modulate the release of VEGF and ANG2 from T98G and HUVECs, respectively. In addition, A. githago reduced the release of MMP2/9. Furthermore, Notch1, DLL4, and HEY2 transcripts and protein expressions were up-regulated, while VEGFR2 was down-regulated in treated HUVEC cells. Treatment with the A. githago extract resulted in a dose-dependent inhibition of AKT phosphorylation. Inhibition of Notch signaling retrieved the viability loss, reduced intracellular ROS, and alleviated the impaired tube formation in A. githago-treated HUVECs. Overall, these data underscore the anti-angiogenic potential of A. githago via inducing apoptosis, modifying the expression levels of VEGF/VEGFR2, and impacting the release of MMP2/9 and ANG2, effects that are most probably modulated through the Notch/VEGF signaling axis.