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 γ-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.

An unusual type I ribosome-inactivating protein from Agrostemma githago L.

Agrostemma githago L. (corn cockle) is an herbaceous plant mainly growing in Europe. The seeds of the corn cockle are toxic and poisonings were widespread in the past by consuming contaminated flour. The toxic principle of Agrostemma seeds was attributed to triterpenoid secondary metabolites. Indeed, this is in part true. However Agrostemma githago L. is also a producer of ribosome-inactivating proteins (RIPs). RIPs are N-glycosylases that inactivate the ribosomal RNA, a process leading to an irreversible inhibition of protein synthesis and subsequent cell death. A widely known RIP is ricin from Ricinus communis L., which was used as a bioweapon in the past. In this study we isolated agrostin, a 27 kDa RIP from the seeds of Agrostemma githago L., and determined its full sequence. The toxicity of native agrostin was investigated by impedance-based live cell imaging. By RNAseq we identified 7 additional RIPs (agrostins) in the transcriptome of the corn cockle. Agrostin was recombinantly expressed in E. coli and characterized by MALDI-TOF-MS and adenine releasing assay. This study provides for the first time a comprehensive analysis of ribosome-inactivating proteins in the corn cockle and complements the current knowledge about the toxic principles of the plant.