Apelin-13 Protects against Ischemic Blood-Brain Barrier Damage through the Effects of Aquaporin-4.

Apelin-13 has been found to have protective effects on many neurological diseases, including cerebral ischemia. However, whether Apelin-13 acts on blood-brain barrier (BBB) disruption following cerebral ischemia is largely unknown. Aquaporin-4 (AQP4) has a close link with BBB due to the high concentration in astrocyte foot processes and regulation of astrocytes function. Here, we aimed to test Apelin-13's effects on ischemic BBB injury and examine whether the effects were dependent on AQP4. We detected the expression of AQP4 induced by Apelin-13 injection at 1, 3, and 7 days after middle cerebral artery occlusion. Meanwhile, we examined the effects of Apelin-13 on neurological function, infarct volume, and BBB disruption owing to cerebral ischemia in wild type mice, and tested whether such effects were AQP4 dependent by using AQP4 knock-out mice. Furthermore, we assessed the possible signal transduction pathways activated by Apelin-13 to regulate AQP4 expression via astrocyte cultures. It was found that Apelin-13 highly increased AQP4 expression as well as reduced neurological scores and infarct volume. Importantly, Apelin-13 played a role of BBB protection in both types of mice by reducing BBB permeability, increased vascular endothelial growth factor, upregulated endothelial nitric oxide synthase, and downregulated inducible NOS. In morphology, we demonstrated Apelin-13 suppressed tight junction opening and endothelial cell swelling via electron microscopy detection. Meanwhile, Apelin-13 also alleviated apoptosis of astrocytes and promoted angiogenesis. Interestingly, effects of AQP4 on neurological function and infarct volume varied with time course, while AQP4 elicited protective effects on BBB at all time points. Statistical analysis of 2-way analysis of variance with replication indicated that AQP4 was required for these effects. In addition, Apelin-13 upregulated phosphorylation of extracellular signal-regulated kinase (ERK) and Akt as well as AQP4 protein in cultured astrocytes. The latter was inhibited by ERK and phosphatidylinositol 3'-kinase (PI3K) inhibitors. Our data suggest that Apelin-13 protects BBB from disruption after cerebral ischemia both morphologically and functionally, which is highly associated with the increased levels of AQP4, possibly through the activation of ERK and PI3K/Akt pathways. This study provides double targets to protection of ischemic BBB damage, which can present new insights to drugs development.

Chu H ，Yang X ，Huang C ，Gao Z ，Tang Y ，Dong Q ... -  《-》

Protection of Vascular Endothelial Growth Factor to Brain Edema Following Intracerebral Hemorrhage and Its Involved Mechanisms: Effect of Aquaporin-4.

Chu H ，Tang Y ，Dong Q 《PLoS One》

Apelin-13 protects neurovascular unit against ischemic injuries through the effects of vascular endothelial growth factor.

Apelin-13 has protective effects on many neurological diseases, including cerebral ischemia. Here, we aimed to test Apelin-13's effects on ischemic neurovascular unit (NVU) injuries and investigate whether the effects were dependent on vascular endothelial growth factor (VEGF). We detected the expression of VEGF and its receptors (VEGFRs) induced by Apelin-13 injection at 1d, 3d, 7d and 14d after middle cerebral artery occlusion (MCAO). Meanwhile, we examined the effects of Apelin-13 on NVU in both in vivo and in vitro experiments as well as whether the effects were VEGF dependent by using VEGF antibody. We also assessed the related signal transduction pathways via multiple inhibitors. We demonstrated Apelin-13 highly increased VEGF and VEGFR-2 expression, not VEGFR-1. Importantly, Apelin-13 led to neurological functions improvement by associating with promotion of angiogenesis as well as reduction of neuronal death and astrocyte activation, which was markedly blocked by VEGF antibody. In cell cultures, Apelin-13 protected neurons, astrocytes and endothelial cells against oxygen-glucose deprivation (OGD) injuries. Moreover, the effect of Apelin-13 to up-regulate VEGF was suppressed by extracellular signal-regulated kinase (ERK) inhibitor U0126 and phosphatidylinositol 3'-kinase (PI3K) inhibitor LY294002. Our data suggest protective effects of Apelin-13 on ischemic NVU injuries are highly associated with the increase of VEGF binding to VEGFR-2, possibly acting through activation of ERK and PI3K/Akt pathways.

Huang C ，Dai C ，Gong K ，Zuo H ，Chu H ... -  《-》

Protection of granulocyte-colony stimulating factor to hemorrhagic brain injuries and its involved mechanisms: effects of vascular endothelial growth factor and aquaporin-4.

Chu H ，Tang Y ，Dong Q 《-》

Protective Effect of 4-Methoxy Benzyl Alcohol on the Blood-Brain Barrier after Cerebral Ischemia Reperfusion Injury.

Damage of the blood-brain barrier (BBB) during the process of cerebral ischemic injury is a key factor that influences the therapeutic efficacy to the cerebral ischemic injury. This work was designed to investigate the mechanisms underlying the protective effects of 4-methoxy benzyl alcohol (4-MA) on the BBB by developing a cerebral ischemia/reperfusion model of rats (MCAO/R). The MCAO/R was developed through a thread embolism method. The neurologic scales, the brain infarct rate, and the Evans blue (EB) contents of the brains were detected. Meanwhile, the release of nitric oxide (NO) and activities of NO synthase (NOS) in brain tissues were measured. Western blotting analyses were also used to assess the protein expressions of aquaporin-4 (AQP-4), occludin, and claudin-5 in brain tissue. After rats were pretreated with different concentrations of 4-MA, the neurologic scores, the infarct rate, and the EB contents in the brain tissues were significantly decreased. The release of NO and the activities of neuronal NOS and inducible NOS were notably inhibited. Furthermore, the protein expression of AQP-4 was markedly decreased, whereas the protein expressions of claudin-5 and occludin were significantly increased. In conclusion, the 4-MA decreases the permeability of BBB when focal cerebral ischemia occurs. The inhibition of the NOS pathways, the attenuation of the protein expression of AQP-4, and the enhancement of the expressions of the tight junction proteins might contribute to the protective effects of 4-MA on the BBB.

He F ，Duan X ，Dai R ，Li Y ，Lin Q ... -  《-》