Background. Ischemic stroke is a severe acute cerebrovascular disease which can be improved with neuroprotective therapies at an early stage. However, due to the lack of effective neuroprotective drugs, most stroke patients have varying degrees of long-term disability. In the present study, we investigated the role of exosomes derived from CXCR4-overexpressing BMSCs in restoring vascular function and neural repair after ischemic cerebral infarction. Methods. BMSCs were transfected with lentivirus encoded by CXCR4 (BMSCCXCR4). Exosomes derived from BMSCCXCR4 (ExoCXCR4) were isolated and characterized by transmission electron microscopy and dynamic light scattering. Western blot and qPCR were used to analyze the expression of CXCR4 in BMSCs and exosomes. The acute middle cerebral artery occlusion (MCAO) model was prepared, ExoCXCR4 were injected into the rats, and behavioral changes were analyzed. The role of ExoCXCR4 in promoting the proliferation and tube formation for angiogenesis and protecting brain endothelial cells was determined in vitro. Results. Compared with the control groups, the ExoCXCR4 group showed a significantly lower mNSS score at 7 d, 14 d, and 21 d after ischemia/reperfusion ( P < 0.05 ). The bEnd.3 cells in the ExoCXCR4 group have stronger proliferation ability than other groups ( P < 0.05 ), while the CXCR4 inhibitor can reduce this effect. Exosomes control (ExoCon) can significantly promote the migration of bEnd.3 cells ( P < 0.05 ), while there was no significant difference between the ExoCXCR4 and ExoCon groups ( P > 0.05 ). ExoCXCR4 can further promote the proliferation and tube formation for the angiogenesis of the endothelium compared with ExoCon group ( P < 0.05 ). In addition, cobalt chloride (COCl2) can increase the expression of β-catenin and Wnt-3, while ExoCon can reduce the expression of these proteins ( P < 0.05 ). ExoCXCR4 can further attenuate the activation of Wnt-3a/β-catenin pathway ( P < 0.05 ). Conclusions. In ischemia/reperfusion injury, ExoCXCR4 promoted the proliferation and tube formation of microvascular endothelial cells and play an antiapoptotic role via the Wnt-3a/β-catenin pathway.

No abstract

Exosomes derived from the cerebral endothelial cells play essential roles in protecting neurons from hypoxia injury, but little is known regarding the biological effects and mechanisms of exosomes on brain plasticity. In this study, exosomes were isolated from rodent cerebral endothelial cells (bEnd.3 cells) by ultracentrifugation, either endothelial cell-derived exosomes (EC-Exo) or PBS was injected intraventricularly 2 h after the middle cerebral artery occlusion/reperfusion (MCAO/R) model surgery in the Exo group and control group, respectively. Sham group rats received the same surgical but not ischemic procedure. We evaluated the motor function of rats after MCAO/R, and the foot-fault rate of the Exo group was significantly lower than that of the control group within 23 days (p < 0.05); the Catwalk analysis also showed gait difference between two groups (p < 0.05). On day 28 after MCAO/R, we euthanized the rats, removed the motor cortex from the brain, and then sequenced the genes by using GO and KEGG to find transcriptome analysis of biological terms and functional annotations: The pathway enrichment revealed that the function of synaptic transmission, regulation of synaptic plasticity, and regulation of synaptic vesicle cycle was significantly enriched with the Exo group than control group. Furthermore, the upregulation of synapsin-I expression in the motor cortex (p < 0.05) as well as the increase of the length of the dendrites were found in the Exo group (p < 0.05) than the control group. We determined the content of exosome microRNA levels, and microRNA-126-3p was the highest (TPM) by transcriptome analysis. Moreover, the microRNA-126-3p protected PC12 cells from apoptosis and increased neurite outgrowth, illustrating the mechanism of how exosomes play a role in altering brain plasticity. This study demonstrated that EC-Exo promoted functional motor recovery in the MCAO/R model, exosomes were critical for the reconstruction of synaptic function in ischemic brain injury, and microRNA-126-3p from EC-Exo could serve as a treatment for nerve damage.

Neuropathic pain is one of the important challenges in the clinic. Although a lot of research has been done on neuropathic pain (NP), the molecular mechanism is still elusive. We aimed to investigate whether the Wnt/β-catenin pathway was involved in NP caused by sustaining dorsal root ganglion (DRG) compression with the chronic compression of dorsal root ganglion model (CCD). Our RNA sequencing results showed that several genes related to the Wnt pathway have changed in DRG and spinal cord dorsal horn (SCDH) after CCD surgery. Therefore, we detected the activation of the Wnt/β-catenin pathway in DRG and SCDH and found active β-catenin significantly upregulated in DRG and SCDH 1 day after CCD surgery and peaked on days 7-14. Immunofluorescence results also confirmed nuclear translocalization of active β-catenin in DRG and SCDH. Additionally, rats had obvious mechanical induced pain after CCD surgery and the pain was significantly alleviated after the application of the Wnt/β-catenin pathway inhibitor XAV939. Furthermore, we found that the levels of proinflammatory factors tumor necrosis factor-α (TNF-α) and interleukin-18 (IL-18) were significantly elevated in CCD rat serum, while the levels of them were correspondingly decreased after the Wnt/β-catenin pathway being inhibited. The results of Spearman correlation coefficient analysis showed that the levels of TNF-α and IL-18 were negatively correlated with the mechanical withdrawal thresholds (MWT) after CCD surgery. Collectively, our findings suggest that the Wnt/β-catenin pathway plays a critical role in the pathogenesis of NP and may be an effective target for the treatment of NP.