The role of Wnt/β-catenin pathway in the protection process by dexmedetomidine against cerebral ischemia/reperfusion injury in rats.

To assess the role of glycogen synthase kinase-3β (GSK3β) and β-catenin in the protection of ischemic injury by dexmedetomidine (Dex). Adult male Sprague-Dawley rats were subjected to (middle cerebral artery occlusion, MCAO) for 2 h followed by reperfusion and Dex was administered 30min before MCAO. The neurological deficit score, cerebral infarct size and neuron survival were evaluated at 24 h after reperfusion. The expression of pAKT, pGSK3β and β-catenin in the ischemic penumbra was assayed by Western blot at 2 h after reperfusion. We found that the Dex-induced increment of neuron survival in the ischemic penumbra was diminished by the PI3K inhibitor LY294002 and the β-catenin inhibitor XAV939, respectively. The increasing expression of pAKT, pGSK3β and β-catenin induced by Dex was markedly inhibited by LY294002. And the increasing expression of β-catenin in nuclei induced by Dex was markedly inhibited by XAV939. At the same time, the GSK3β inhibitor SB216763 also caused an increment of neuron survival and an increasing expression of pGSK3β and β-catenin in the ischemic penumbra. Our data suggested that treatment with Dex reduced cerebral injury in rats exposed to cerebral ischemia-reperfusion (I/R) by the activation of the PI3K/AKT/GSK3β pathways as well the activation of downstream Wnt/β-catenin pathway. And the Wnt/β-catenin pathway may play an important role in the protection against cerebral ischemia/reperfusion injury in rats.

Li P ，Zhang Y ，Liu H 《-》

Both PI3K/Akt and ERK1/2 pathways participate in the protection by dexmedetomidine against transient focal cerebral ischemia/reperfusion injury in rats.

Dexmedetomidine (Dex) has been demonstrated to provide neuroprotection against ischemia/reperfusion (I/R) injury. However, the exact mechanism of this protection remains unknown. Here, we explored the neuroprotective effect of Dex in rats exposed to cerebral I/R-induced by middle cerebral artery occlusion (MCAO) and the role of phosphatidylinositol 3-kinase (PI3K)/Akt, extracellular signal-regulated kinase 1/2 (ERK1/2), and glycogen synthase kinase-3β (GSK-3β) in this protective action. Adult male Sprague-Dawley rats were subjected to MCAO for 90 min followed by reperfusion for 24h and Dex (15 μg/kg, i.v.) was administered immediately after the onset of MCAO. The neurological deficit score, cerebral infarct volume, brain edema, and neuron survival were evaluated at 24h of reperfusion. The effect of Dex on p-Akt, p-ERK1/2 and p-GSK-3β expression in the ischemic hemisphere was assayed by Western blot. Treatment of rats exposed to I/R with Dex caused not only marked reduction in the neurological deficit score, cerebral infarct volume, and brain edema (P <0.01 vs. I/R alone), but also a decrease in neuron death in hippocampal CA1 and cortex (P<0.01 vs. I/R alone). The Dex-induced increment of neuron survival in the ischemic CA1 and cortex was diminished by the PI3K inhibitor LY294002 and the MEK inhibitor U0126. The increasing expressions of p-Akt and p-ERK1/2 induced by Dex in the ischemic hemisphere were markedly inhibited by LY294002 (or wortmannin) and U0126 (or PD98059), respectively. The up-regulation of p-GSK-3β by Dex in the ischemic hemisphere was significantly decreased by both LY294002 (or wortmannin) and U0126 (or PD98059). Our data demonstrated that treatment with Dex reduced cerebral injury in rats exposed to transient focal I/R, and this was mediated by the activation of the PI3K/Akt and ERK1/2 pathways as well the phosphorylation of downstream GSK-3β.

Zhu YM ，Wang CC ，Chen L ，Qian LB ，Ma LL ，Yu J ，Zhu MH ，Wen CY ，Yu LN ，Yan M ... -  《-》

Protective effect of dexmedetomidine against diabetic hyperglycemia-exacerbated cerebral ischemia/reperfusion injury: An in vivo and in vitro study.

Dexmedetomidine (Dex) has been noted to have neuroprotective effect against cerebral ischemia-reperfusion (I/R) injury. However, the effect of Dex in diabetic hyperglycemia-exacerbated cerebral I/R injury and its underlying mechanism remain unclear. The infarct volume and brain edema were evaluated by 2,3,5-triphenyltetrazolium chloride staining and standard wet-dry method. Modified neurological severity score was utilized to assess the neurological deficits. The oxidative stress and inflammation were evaluated by detecting reactive oxygen species (ROS), malondialdehyde (MDA), tumor necrosis factor (TNF)-α and interleukin (IL)-1β. Terminal deoxynucleotidyl transferase dUTP nick end labeling assay and cell count kit-8 were applied to measure cell apoptosis and viability. Dex treatment reduced infarct volume, decreased brain water content and improved neurological deficit in middle cerebral artery occlusion/reperfusion (MCAO/R) mice. Dex treatment reduced the levels of ROS, MDA, TNF-α and IL-1β in the entire middle cerebral artery territory of diabetic mice subjected to MCAO/R, as well as in primary culture of mouse hippocampal neurons stimulated with 50 mM glucose and oxygen glucose deprivation/reperfusion. Dex treatment inhibited neuronal apoptosis induced by diabetic hyperglycemia-exacerbated cerebral I/R injury. Dex upregulated nuclear factor of activated T-cells 5 (NFAT5) and Sirtuin 1 (SIRT1) expression, induced NF-E2-related factor 2 (Nrf2) translocation from cytoplasm to nucleus and inhibited the acetylation of Nrf2. However, these changes triggered by Dex treatment were abrogated by NFAT5 knockdown. Dex protects against diabetic hyperglycemia-exacerbated cerebral I/R injury through attenuation of oxidative stress, inflammation and apoptosis. The underlying mechanism is at least the NFAT5/SIRT1/Nrf2 signaling pathway dependent.

Chen L ，Cao J ，Cao D ，Wang M ，Xiang H ，Yang Y ，Ying T ，Cong H ... -  《-》

Wnt/β-catenin signaling pathway contributes to isoflurane postconditioning against cerebral ischemia-reperfusion injury and is possibly related to the transforming growth factorβ1/Smad3 signaling pathway.

The Wnt/β-catenin signaling pathway plays an important role in ischemia-reperfusion(I/R) injury, and the transforming growth factor(TGF)-β/Smad signaling pathway participates in the neuroprotection effect induced by isoflurane(ISO) postconditioning. In this study, we aimed to explore the role of the Wnt/|[beta]|-catenin β-catenin signaling pathway in the neuroprotection effect induced by ISO postconditioning, and investigate the interaction of Wnt/β-catenin and TGF-β/Smad signaling pathway in this neuroprotection effect. Cerebral I/R injury was established in Sprague-Dawley rats by using the middle cerebral artery occlusion (MCAO) model for 90 min followed by 24 h reperfusion. Postconditioning by inhalation of ISO was performed for 60 min after ischemia at the onset of reperfusion. Neurological deficit scoring, 2,3,5-triphenyl tetrazolium chloride staining and Nissl staining were adopted to evaluate brain injury. Apoptosis of the hippocampus and cortex neurons was detected by TUNEL staining. The expression levels of Wnt3a, GSK-3β, β-catenin, Cyclin D1, VEGF, Caspase 3, TGF-β1, Smad3 and p-Smad3 were determined by immunofluorescence (IF) staining, quantitative real-time polymerase chain reaction (qRT-PCR), and Western blot. Various targeted inhibitors were administered via intraperitoneal injection or lateral ventricle injection. In the cortex region, the neurological deficit score, infarct volumes and neuron apoptosis increased, and the expression level of the Wnt3a, GSK-3β, β-catenin, VEGF and Cyclin D1 decreased in the MCAO group compared with the Sham group. In the MCAO + ISO group, the neurological deficit score, infarct volumes and neuron apoptosis reduced significantly, the expression levels of Wnt3a, β-catenin, VEGF and Cyclin D1 increased, while the expression level of GSK-3β and Caspase 3 decreased relative to MCAO group. When Wnt inhibitor(DKK-1) was given in advance followed by ISO postconditioning, the neurological deficit score, infarct volumes, neuron apoptosis and the expression level of GSK-3β and Caspase 3 increased. qRT-PCR and IF showed similar changes in the protein levels of all groups. However, the expression level of β-catenin in nuclear and cytoplasm both decreased significantly after pre-injection with the TGF-β1 inhibitor(LY2157299) and Smad3 inhibitor(SIS3), whereas the expression levels of TGF-β1, Smad3 and p-Smad3 were almost unchanged. The expression levels of all the related proteins and morphological changes in the hippocampus region were consistent with that of the cortex. ISO postconditioning can reduce cerebral I/R injury by activating the Wnt/β-catenin signaling pathway and may be related to the TGF-β/Smad3 signaling pathway.

Zhang G ，Ge M ，Han Z ，Wang S ，Yin J ，Peng L ，Xu F ，Zhang Q ，Dai Z ，Xie L ，Li Y ，Si J ，Ma K ... -  《-》

PI3K/Akt pathway contributes to neuroprotective effect of Tongxinluo against focal cerebral ischemia and reperfusion injury in rats.

Tongxinluo (TXL), a compound prescription, is formulated according to the collateral disease doctrine of traditional Chinese medicine, and is widely used for the treatment of cardio-cerebrovascular diseases in China. We aimed to investigate the neuroprotective effect of TXL on focal cerebral ischemia and reperfusion injury in rats by attenuating its brain damage and neuronal apoptosis, and to assess the potential role of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway in this protection. Adult Male Sprague-Dawley rats (n=120) were randomly divided into 5 groups: sham, cerebral ischemia and reperfusion (I/R), cerebral ischemia and reperfusion plus TXL (1.6g/kg/day) (TXL1.6), TXL1.6 plus LY294002 and dimethyl sulfoxide (DMSO) (TXL1.6+LY294002), TXL1.6 plus DMSO (TXL1.6+vehicle). Prior to the grouping, TXL1.6 was selected to be the optimal dose of TXL by evaluating the neurological deficits score of five group rats (Sham, I/R, TXL0.4, TXL0.8 and TXL1.6, n=30) at 0, 1, 3, 5, and 7 days after reperfusion. Rats, being subjected to middle cerebral artery occlusion (MCAO) for 90min followed by 24h reperfusion, were the cerebral ischemia/reperfusion models. At 24h after reperfusion, cerebral infarct area was measured via tetrazolium staining and neuronal damage was showed by Nissl staining. The double staining of Terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick end labeling (TUNEL) staining and immunofluorescence labeling with NeuN, was performed to evaluate neuronal apoptosis. Proteins involved in PI3K/Akt pathway were detected by Western blot. The results showed that TXL markedly improved neurological function, reduced cerebral infarct area, decreased neuronal damage, and significantly attenuated neuronal apoptosis, while these effects were eliminated by inhibition of PI3K/Akt with LY294002. We also found that TXL up-regulated the expression levels of p-PDK1, p-Akt, p-c-Raf, p-BAD and down-regulated Cleaved caspase 3 expression notably, which were partially reversed by LY294002. Additionally, the increment of p-PTEN level on which LY294002 had little effect was also detected in response to TXL treatment. These findings demonstrated that TXL provided neuroprotection against cerebral ischemia/reperfusion injury and neuronal apoptosis, and this effect was mediated partly by activation of the PI3K/Akt pathway.

Yu ZH ，Cai M ，Xiang J ，Zhang ZN ，Zhang JS ，Song XL ，Zhang W ，Bao J ，Li WW ，Cai DF ... -  《-》