Xiaowei Hu

The modulation of microglial polarization from the pro-inflammatory M1 to the anti-inflammatory M2 phenotype shows promise as a therapeutic strategy for ischemic stroke. Quercetin, a natural flavonoid abundant in various plants, possesses anti-inflammatory, anti-apoptotic, and antioxidant properties. Nevertheless, its effect and underlying mechanism on microglia/macrophages M1/M2 polarization in the treatment of cerebral ischemia/reperfusion injury (CI/RI) remain poorly explored. In the current study, we observed that quercetin ameliorated neurological deficits, reduced infarct volume, decreased the number of M1 microglia/macrophages (CD16/32+/Iba1+), and enhanced the number of M2 microglia/macrophages (CD206+/Iba1+) after establishing the CI/RI model in rats. Subsequent in vivo and in vitro experiments indicated that quercetin downregulated M1 markers (CD86, iNOS, TNF-α, IL-1β, and IL-6) and upregulated M2 markers (CD206, Arg-1, IL-10, and TGF-β). Network pharmacology analysis and molecular docking revealed that the PI3K/Akt/NF-κB signaling pathway emerged as the core pathway. Western blot confirmed that quercetin upregulated the phosphorylation of PI3K and Akt, while alleviating the phosphorylation of IκBα and NF-κB both in vivo and in vitro. However, the PI3K inhibitor LY294002 reversed the effects of quercetin on M2 polarization and the expression of key proteins in the PI3K/Akt/NF-κB pathway in primary microglia after oxygen-glucose deprivation/reoxygenation (OGD/R) in vitro. Collectively, our findings demonstrate that quercetin facilitates microglia/macrophages M2 polarization by modulating the PI3K/Akt/NF-κB signaling pathway in the treatment of CI/RI. These findings provide novel insights into the therapeutic mechanisms of quercetin in ischemic stroke.

Exosomes derived from bone mesenchymal stem cells (BMSC-Exos) are one of the main factors responsible for the therapeutic effects of BMSCs. The study aimed to investigate whether BMSC-Exos could promote angiogenesis in ischemic stroke mice via miR-21-5p. In ischemic stroke mice, the therapeutic effects of BMSC-Exos were evaluated by neurological functions and infarct volume. Microvessel density was detected by BrdU/vWF immunofluorescence staining. In in vitro experiments, the proangiogenic effects of BMSC-Exos were assessed via proliferation, migration, and tube formation of human umbilical vein endothelial cells (HUVECs). The miR-21-5p inhibitor was transfected into BMSCs using Lipofectamine 2000. miR-21-5p expression was detected by qRT-PCR. The expression levels of VEGF, VEGFR2, Ang-1, and Tie-2 were determined by Western blot. BMSC-Exos significantly improved neurological functions and reduced infarct volume, upregulated microvessel density, and miR-21-5p expression after cerebral ischemia. In vitro assays revealed that BMSC-Exos enhanced HUVECs functions including proliferation, migration, and tube formation. BMSC-Exos increased the expression levels of VEGF, VEGFR2, Ang-1, and Tie-2. However, the proangiogenic effects of BMSC-Exos on HUVECs were reversed by the miR-21-5p inhibitor. These results suggest that BMSC-Exos could promote angiogenesis via miR-21-5p upregulation, making them an attractive treatment strategy for stroke recovery.

Neuroinflammation manifests following injury to the central nervous system (CNS) and M1/M2 polarization of microglia is closely associated with the development of this neuroinflammation. In this study, multiple databases were used to collect targets regarding luteolin and microglia polarization. After obtaining a common target, a protein-protein interaction (PPI) network was created and further analysis was performed to obtain the core network. Molecular docking of the core network with luteolin after gene enrichment analysis. In vitro experiments were used to examine the polarization of microglia and the expression of related target proteins. A total of 77 common targets were obtained, and the core network obtained by further analysis contained 38 proteins. GO and KEGG analyses revealed that luteolin affects microglia polarization in regulation of inflammatory response as well as the interleukin (IL)-17 and tumor necrosis factor (TNF) signaling pathways. Through in vitro experiments, we confirmed that the use of luteolin reduced the expression of inducible nitric oxide synthase (iNOS), IL-6, TNF-α, p-NFκBIA (p-IκB-α), p-NFκB p65, and MMP9, while upregulating the expression of Arg-1 and IL-10. This study reveals various potential mechanisms by which luteolin induces M2 polarization in microglia to inhibit the neuroinflammatory response.

Bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exos) have been shown to promote angiogenesis after ischemic stroke, in which microRNAs (miRs) are believed to play an important role in exosome-mediated therapeutic effects, though the mechanism is still not clear. In this study, a series of molecular biological and cellular assays, both in vitro and in vivo, were performed to elucidate the role of exosomal miR-486 in angiogenesis following cerebral ischemic and its molecular mechanisms. Our results revealed that BMSC-Exos significantly improved neurological function and increased microvessel density in ischemic stroke rats. In vitro assays showed that BMSC-Exos promoted the proliferation, migration, and tube formation ability of oxygen-glucose deprivation/reoxygenation (OGD/R) injured rat brain microvascular endothelial cells (RBMECs). Importantly, BMSC-Exos increased the expression of miR-486 and phosphorylated protein kinase B (p-Akt) and down-regulated the protein level of phosphatase and tensin homolog (PTEN) in vivo and in vitro. Mechanistic studies demonstrated that transfection with miR-486 mimic enhanced RBMECs angiogenesis and increased p-Akt expression, while inhibited PTEN expression. On the other hand, the miR-486 inhibitor induced an opposite effect, which could be blocked by PTEN siRNA. It was thus concluded that exosomal miR-486 from BMSCs may enhance the functional recovery by promoting angiogenesis following cerebral ischemic injury, which might be related to its regulation of the PTEN/Akt pathway.