Astragaloside IV (AS-IV), an active component isolated from Astragalus membranaceus, suppresses the activation of ERK1/2 and JNK, and downregulates matrix metalloproteases (MMP)-2, (MMP)-9 in MDA-MB-231 breast cancer cells. Astragaloside IV is a bioactive saponin first isolated from the dried plant roots of the genus Astragalus, which is used in traditional Chinese medicine.1 It dose-dependently inhibits human adenovirus type 3 (HAdV-3) in A549 cells (IC50 = 23 μM; LC50 = 865 μM).It inhibits replication of HAdV-3 and decreases HAdV-3-induced apoptosis. It has diverse protective effects for the cardiovascular, immune, digestive, and nervous systems. In particular, it reduces myocardial infarct size in dogs when administered prior to coronary ligation and reduces reperfusion arrhythmias in isolated rat hearts.

MDA-MB-231 cells treated as indicated or tumor tissues are harvested and lysed in Mg2+ lysis buffer containing 50 mM Tris (pH 7.5), 10 mM MgCl2, 0.5 M NaCl, and protease inhibitor cocktail. Equal amounts of lysates are incubated with PAK-PBD beads at 4°C for 1 h. PAK-PBD beads are pelleted by centrifugation and washed with ish buffer containing 25 mM Tris (pH 7.5), 30 mM MgCl2, 40 mM NaCl. Active Rac1 is detected by western blotting.

CCK-8 assay is adopted to determine cell viability. cultured NSCLC cells are seeded into 96-well plates at the density of 4×104 (cells/well). Then 10 μL?well CCK8 solution is added and incubated in dark at 37°C for another 2 h. The absorbance is determined with the wavelength of 490 nm.

Transient cerebral ischemia and reperfusion is prepared by BCCAO. Mice are randomly divided into the Sham, Model, Astragaloside IV (10 mg/kg) and Astragaloside IV (20 mg/kg) treatment groups. The Astragaloside IV treatment groups are intragastrically administered 7 days before the surgery and terminated on the day of sacrifice. On the day of the surgery, Astragaloside IV is administrated 2 h prior to ischemia. The Sham-operated and Model groups are treated with distilled water. After the mice are anesthetized with an intraperitoneal injection of chloral hydrate (350 mg/kg), the bilateral common carotid arteries are exposed and carefully separated with a small ventral neck incision and occluded twice (20 min each) with ligated surgical silk as described previously with minor modifications. There is a 10 min reperfusion period between the two occlusion periods (ischemia 20 min ? reperfusion 10 min ? ischemia 20 min). Sham-operated mice are subjected to the same surgical operation without the surgical silk ligation. Mouse body temperature is maintained at 37±0.5°C during the surgery with heating equipment until recovery from the anesthesia.

DMSO: 100 mg/mL (127.39 mM), Sonication is recommended.