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@#BACKGROUND: Small extracellular vesicles (sEVs) from bone marrow mesenchymal stem cells (BMSCs) have shown therapeutic potential for cerebral ischemic diseases. However, the mechanisms by which BMSC-derived sEVs (BMSC-sEVs) protect neurons against cerebral ischemia/reperfusion (I/R) injury remain unclear. In this study, we explored the neuroprotective effects of BMSC-sEVs in the primary culture of rat cortical neurons exposed to oxygen-glucose deprivation and reperfusion (OGD/R) injury. METHODS: The primary cortical neuron OGD/R model was established to simulate the process of cerebral I/R in vitro. Based on this model, we examined whether the mechanism through which BMSC-sEVs could rescue OGD/R-induced neuronal injury. RESULTS: BMSC-sEVs (20 μg/mL, 40 μg/mL) significantly decreased the reactive oxygen species (ROS) productions, and increased the activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx). Additionally, BMSC-sEVs prevented OGD/R-induced neuronal apoptosis in vivo, as indicated by increased cell viability, reduced lactate dehydrogenase (LDH) leakage, decreased terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining-positive cells, down-regulated cleaved caspase-3, and up-regulated Bcl-2/Bax ratio. Furthermore, Western blot and flow cytometry analysis indicated that BMSC-sEV treatment decreased the expression of phosphorylated calcium/calmodulin-dependent kinase II (p-CaMK II)/CaMK II, suppressed the increase of intracellular calcium concentration ([Ca2+]i) caused by OGD/R in neurons. CONCLUSIONS: These results demonstrate that BMSC-sEVs have significant neuroprotective effects against OGD/R-induced cell injury by suppressing oxidative stress and apoptosis, and Ca2+/CaMK II signaling pathways may be involved in this process.
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Aim To investigate the protective effects of ginkgolide K (GK) on neurovascular unit injured by ischemic stroke and the potential mechanism associated with hypoxia-inducible factor-1α (HIF-1α) pathway. Methods The BV-2 cells and EA. hy926 cells suffered from oxygen-glucose deprivation and reperfusion (OGD/R) were applied to mimic the injury of neurovascular unit induced by cerebral ischemia in vitro. After 4 h OGD insult, BV-2 cells and EA. hy926 cells received reperfusion and treated with GK. The levels of inflammatory cytokines in the supernatant of BV-2 cells were detected, while the protective effects of GK on EA. hy926 cells were also evaluated after GK administration for 24 h. The p-Akt and p-Erk expressions were examined by Western blot after 1 h of GK treatment, while HIF-1α was detected after 6 h of GK treatment. In addition, PI3K inhibitor LY294002 was applied to further verify the potential mechanisms underlying the beneficial effects of GK. The expressions of p-Akt after 1 h of GK treatment, and the protein levels of HIF-1α pathway after 6 h of GK treatment were also analyzed by Western blot. Results GK significantly inhibited the levels of TNF-α, IL-6 and IL-1β in supernatant of BV-2 cells injured by OGD/R, through increasing p-Akt and decreasing p-Erk expressions, and then affecting HIF-1α pathway. In addition, LY294002 reduced the regulatory effect of GK. Furthermore, GK significantly improved viability and inhibited the release of LDH in supernatant of EA. hy926 cells suffered from OGD/R, and up-regulated the expressions of p-Akt, HIF-1α, HO-1 and VEGF, while cleaved caspase-3/9 was inhibited. Conclusions GK exerts multi-effects on reducing neurovascular unit injury induced by ischemic stroke, and the potential mechanism might be associated with the different regulatory effects of HIF-1α in different cells.