Previous studies had showed that Apelin 13 could protect against apoptosis induced by ischemic/reperfusion (I/R). However, the mechanisms whereby Apelin 13 protected brain I/R remained to be elucidated. The present study was designed to determine whether Apelin 13 provided protection through AMPK/GSK-3β/Nrf2 pathway. In vivo, the I/R model was induced and Apelin 13 was given intracerebroventricularly 15 min before reperfusion. The neurobehavioral scores, infarction volumes, and some cytokines in the brain were measured. For in vitro study, PC12 cells were used. To clarify the mechanisms, proteases inhibitors or siRNA were used. Protein levels were investigated by western blotting. The results showed that Apelin 13 treatment significantly reduced infarct size, improved neurological outcomes, decreased brain edema, and inhibited cell apoptosis, oxidative stress, and neuroinflammation after I/R. Apelin 13 significantly increased the expression of Nrf2 and the phosphorylation levels of AMPK and GSK-3β. Furthermore, in cultured PC12 cells, the same protective effects were also observed. Silencing Nrf2 gene with its siRNA abolished the Apelin 13’s prevention of I/R-induced PC12 cell injury, oxidative stress, and inflammation. Inhibition of AMPK by its siRNA decreased the level of Apelin 13-induced Nrf2 expression and diminished the protective effects of Apelin 13. The interplay relationship between GSK-3β and Nrf2 was also verified with relative overexpression. Using selective inhibitors, we further identified the upstream of AMPK/GSK-3β/Nrf2 is AR/Gα/PLC/IP3/CaMKK. In conclusion, the previous results showed that Apelin 13 protected against I/R-induced ROS-mediated inflammation and oxidative stress through activating the AMPK/GSK-3β pathway by AR/Gα/PLC/IP3/CaMKK signaling, and further upregulated the expression of Nrf2-regulated antioxidant enzymes.

Oxidative stress plays an important role in Parkinson’s disease and other neurodegenerative disorders. Lycium barbarum polysaccharides (LBP), the main active ingredients extracted from the fruits of Lycium barbarum L., have been shown to be a potent antioxidant. In the present study, we investigated the protective effects, and the possible mechanism of action of LBP against 6-hydroxydopamine (6-OHDA)-induced apoptosis in PC12 cells. Our data demonstrated that LBP significantly reversed the 6-OHDA-induced decrease in cell viability, prevented 6-OHDA-induced changes in condensed nuclei and decreased the percentage of apoptotic cells in a dose-dependent manner. Furthermore, LBP also slowed the accumulation of reactive oxygen species (ROS) and nitric oxide (NO), decreased the level of protein-bound 3-nitrotyrosine (3-NT) and intracellular free Ca²⁺, and inhibiting the overexpression of nuclear factor κB (NF-κB), neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS). These results demonstrate that LBP prevents 6-OHDA-induced apoptosis in PC12 cells, at least in part through the ROS-NO pathway.

Buyang Huanwu Decoction (BYHWD) is used to promote blood circulation and is widely used in Chinese clinical practice for the treatment and prevention of ischemic cerebral vascular diseases. However, the mechanism and active compounds of BYHWD used to treat ischemic stroke are not well understood. The current study aimed to identify the potential active components of BYHWD and explore its mechanism using network pharmacology and bioinformatics analyses. The compounds of BYHWD were obtained from public databases. Oral bioavailability and drug‑likeness were screened using the absorption, distribution, metabolism and excretion (ADME) criteria. Components of BYHWD, alongside the candidate targets of each component and the known therapeutic targets of ischemic stroke were collected. A network of target gene compounds and cerebral ischemia compounds was established using network pharmacology data sources. The enrichment of key targets and pathways was analyzed using STRING and DAVID databases. Moreover, three of key targets [IL6, VEGFA and hypoxia‑inducible‑factor‑1α (HIF‑1α)] were verified using western blot analysis. Network analysis determined 102 compounds in seven herbal medicines that were subjected to ADME screening. A total of 42 compounds as well as 79 genes formed the principal pathways associated with ischemic stroke. The 16 key compounds identified were baicalein, beta‑carotene, baicalin, kaempferol, luteolin, quercetin, hydroxysafflor yellow A, isorhamnetin, bifendate, formononetin, calycosin, astragaloside IV, stigmasterol, sitosterol, Z‑ligustilide, and dihydrocapsaicin. The core genes in this network were IL6, TNF, VEGFA, HIF‑1α, MAPK1, MAPK3, JUN, STAT3, IL1B and IL10. Furthermore, the TNF, IL17, apoptosis, PI3K‑Akt, toll‑like receptor, MAPK, NF‑κB and HIF‑1 signaling pathways were identified to be associated with ischemic stroke. Compared with the control group (no treatment), BYHWD significantly inhibited the expression of IL6 and increase the expression of HIF‑1α and VEGFA. Network pharmacology analyses can help to reveal close interactions between multi‑components and multi‑targets and enhance understanding of the potential effects of BYHWD on ischemic stroke.

Effective treatment of ischaemic stroke is required to combat its high prevalence and incidence. Although the combination of Astragalus membranaeus (Fisch.) Bge. (Fabaceae) and Carthamus tinctorius...