Study On Inducing Moderate Angiogenesis By Xuefu Zhuyu Capsule Through EphB4/EphrinB2

ObjectiveTo study and confirm the angiogenesis effect of Xuefu Zhuyu Capsule (XFZYC) in different conditions on the endothelial cell line HMEC-1. Then at the best angiogenesis condition, to investigate the function of EphB4/EphrinB2in controlling angiogenesis by XFZYC to provide scientific theory basis for its clinical usage.MethodSerum pharmacology technique was adopted. First, human blank serum and XFZYC containing serum (XZC-CS) were prepared. Secondly, Endothelial cells HMEC-1were treated with1.25%,2.5%and5%blank serum or XZC-CS for24h,48h or72h. Then, Thiazolyl blue tetrazolium bromide (MTT), Fluorescence Activating Cell Sorter (FACS), Transwell chamber, adhesion and in vitro tube formation assays were used to observe the effect of XFZYC on cell proliferation, migration, adhesion and tube formation capacities respectively. Real-time PCR and Western blot were used to detect the expressions of EphB4and EphrinB2.Results1. Xuefu Zhuyu Capsule didn’t affect HMEC-1viability.2. Xuefu Zhuyu Capsule didn’t affect HMEC-1number in S phase.3.1.25%XZC-CS significantly improved cell migration at all three time points; but2.5%XZC-CS inhibited cell migration at first24h, then induced cell migration at48h and72h. And5%XZC-CS improved cell migration at24h, and then restained at48h and72h.4.1.25%XZC-CS, inhibit cell adhesion at72h, but2.5%XZC-CS significantly induced cell adhesion at24h and48h, and5%XZC-CS had function of promoting cell adhesion at48h.5. Only2.5%XZC-CS could promote tube formation in vitro at48h, and5%XZC-CS inhibited tube formation in vitro at48h and72h.6. Real-time PCR assay showed that EphB4expression was up-regulatied by2.5%XZC-CS at12h, and then down-regulated at48h. No difference was observed at6h or48h.7. Real-time PCR assay showed that2.5%XZC-CS didn’t affect the expression of EphrinB2.8. Western blot assay observed the expression of EphrinB2was up-regulated by2.5%XZC-CS at24h; but no difference was observed at12h or48h. Conclusion1. The XFZYC induced the best angiogenesis effect at an appropriate condition as2.5%concentration working48h. The celluar mechanism of XFZYC on angiogenesis involved cell migration, adhesion and in vitro tube formation.2. XFZYC inducing angiogenesis in low concentration at a narrow time and inhibiting angiogenesis in high concentration could avoid overgrowth of vessel.3. XFZYC controlling moderate angiogensis closely related with EphB4and EphrinB2.,...