Inhibition Of Neuronal Apoptosis Research Articles

Peroxiredoxin 6 secreted by Schwann-like cells protects neuron against ischemic stroke in rats via PTEN/PI3K/AKT pathway

Schwann cells can promote the survival of damaged neurons and axon regeneration by secreting or releasing some proteins and factors which may provide effective strategies to the remedy for ischemic stroke. The models of middle cerebral artery occlusion and oxygen-glucose deprivation (OGD) were established. Peroxiredoxin 6 (PRDX6) was found in Schwann-like cell conditioned medium (SCLC-CM) by mass spectrometry. The rehabilitative performance of SCLC-CM on focal cerebral ischemia of rats and on OGD-induced PC12 cells were assessed. SCLC-CM significantly improved neurological recovery, reducing the infarct volume of rats after stroke. PRDX6 could significantly inhibit neuron apoptosis in the OGD injury by mediating oxidative stress and activating the PTEN/PI3K/AKT pathway. In conclusion, PRDX6 secreted by Schwann-like cell protects neuron against focal cerebral ischemia, SCLC-CM might be a new effective early intervention for ischemic stroke.

Canopy Fibroblast Growth Factor Signaling Regulator 2 (CNPY2) Inhibits Neuron Apoptosis in Parkinson's Disease via the AKT/GSK3β Pathway.

Parkinson's disease (PD) is a neurodegenerative disorder caused by the progressive loss of dopaminergic neurons. Canopy fibroblast growth factor signaling regulator 2 (CNPY2) is down-regulated in this disease, but its functions are unknown. This study investigates the effects and regulation of CNPY2 in the apoptosis of neurons in PD. We established a PD model in vivo by a five consecutive days-injection of 1-methyl-4- phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) to mice. In vitro, the human SH-SY5Y neuroblastoma cells, after differentiation, were treated with 1-Methyl-4-phenylpyridinium iodide (MPP+) for modeling. The cells were transfected with a recombinant vector overexpressing CNPY2 followed by MPP+ treatment. Expression of CNPY2 and proteins related to apoptosis was detected by real-time PCR, western blot, or immunofluorescence staining. The ROS level and mitochondrial membrane potential were determined by flow cytometry. Cell viability and apoptosis were measured by MTT assay and TUNEL staining. CNPY2 level was down-regulated both in the brain and retina of PD mice and also inhibited in neurons by MPP+ in vitro. Overexpression of CNPY2 repressed the level of Bax and cleaved caspase-3, enhanced Bcl-2 level, and promoted neurite length under MPP+ treatment. CNPY2 overexpression reduced the accumulation of ROS and mitochondria dysfunction in neurons. The AKT/ GSK3β signaling pathway was activated by overexpressed CNPY2 to inhibit MPP+-induced neuronal apoptosis, which was confirmed using an AKT inhibitor MK-2206 2HCl. CNPY2 alleviates oxidative stress, mitochondria dysfunction, and apoptosis of neurons induced by MPP+ by activating the AKT/ GSK3β signaling pathway.

Rhamnazin Ameliorates Traumatic Brain Injury in Mice via Reduction in Apoptosis, Oxidative Stress, and Inflammation

Background: Traumatic brain injury (TBI) is posing serious health challenges for people across the globe due to high morbidity and mortality. However, none of the agents prevents or limits the damage caused by TBI because of its multifactorial etiology. Thus, the discovery of novel agents which can act via several pathways could serve the purpose and afford favorable consequence against TBI. Therefore, in the present article, we intended to investigate the protective effect of rhamnazin (RMZ), a dimethoxyflavone against experimentally induced TBI in mice. Methods: The effect of RMZ was investigated on cerebral edema and grip test score after induction of experimental brain injury in rats. The effect of RMZ was also investigated on neuronal degeneration in brain tissues of the experimental mice via Nissl staining and flow cytometry analysis. The expression of Bax and Bcl-2 was also quantified using Western blot analysis. The level of inflammatory cytokines (TNF-α and IL-1β) and oxidative stress markers (malondialdehyde, superoxide dismutase, and glutathione peroxidase) was also determined using enzyme-linked immunosorbent assay. Results: RMZ showed a significant reduction in edema and improved grip strength. It also prevented neuronal degeneration via inhibition of neuronal apoptosis as shown by flow cytometry analysis. RMZ showed an antiapoptotic effect via reduction of Bax and increased the expression of Bcl-2 in Western blot analysis. It also showed to inhibit oxidative stress and inflammation compared to the TBI group. Conclusion: Collectively, our study is first to demonstrate the protective effect of RMZ against experimentally induced TBI in rats.

LncRNA-GAS5 promotes spinal cord repair and the inhibition of neuronal apoptosis via the transplantation of 3D printed scaffold loaded with induced pluripotent stem cell-derived neural stem cells.

BackgroundStem cell transplantation has been increasingly used for spinal cord repair, and some achievements have been made. However, limited stem cell sources as well as immune rejection and ethical issues have restricted its wide application. Therefore, to achieve further breakthroughs regarding the application of stem cell transplantation to treat spinal cord injury (SCI), it is important to develop a stem cell line that can effectively avoid immune rejection and ethical issues.MethodsUrine cells (UCs) were induced to differentiate into induced pluripotent stem cells (iPSCs), which then further differentiated into neural stem cells (NSCs). Relevant tests were performed, and three-dimensional (3D) printed scaffolds were prepared. Thirty C57BL/6 mice were divided into 5 groups based on a random number table: a sham group, an SCI group, an SCI + control group, an SCI + siNC group, and an SCI + siGAS5 group (n=6). The latter 4 groups replicated SCI models. Mice in the SCI + control group were transplanted with 3D scaffolds loaded with iPSC-derived NSCs (iPSd-NSCs). Mice in the SCI + siNC group and the SCI + siGAS5 group were transplanted with scaffolds loaded with iPSd-NSCs-siNC and 3D scaffolds loaded with iPSd-NSCs-siGAS5, respectively. Mice in the other groups were injected with the same amount of normal saline. Hematoxylin-eosin staining was used to observe the histopathology of the injured spinal cord, the Basso-Mouse Scale was used to assess the motor function of the hind limbs of the mice, and Western blot was used to detect the expression of apoptosis-related proteins after SCI.ResultsiPSd-NSCs were successfully induced and differentiated, and 3D printed heparin sulfate-collagen scaffolds were prepared, inside which a 3D loose porous structure was shown by electron microscopy. Morphological observations showed that iPSd-NSC transplantation improved SCI in mice, while GAS5 silencing inhibited the reparative effect of iPSd-NSC transplantation on SCI in mice. Western blot results indicated that iPSd-NSC transplantation significantly increased the expression level of B cell lymphoma/leukemia-2 (Bcl-2) (P<0.01) but decreased the expression levels of Bcl-2 associated X protein, cytochrome C, and cleaved caspase-3 (P<0.001).ConclusionsThe overexpression of lncRNA-GAS5 can promote spinal cord repair and inhibit neural apoptosis via the transplantation of 3D printed scaffolds loaded with iPSd-NSCs.

Maternal Subclinical Hypothyroidism in Rats Impairs Spatial Learning and Memory in Offspring by Disrupting Balance of the TrkA/p75NTR Signal Pathway

Maternal subclinical hypothyroidism (SCH) during pregnancy can adversely affect the neurodevelopment of the offspring. The balance of nerve growth factor (NGF)-related tropomyosin receptor kinase A/p75 neurotrophin receptor (TrkA/p75NTR) signaling in the hippocampus is important in brain development, and whether it affects cognitive function in maternal SCH’s offspring is not clear. In this study, we found that compared with the control (CON) group, expression of proliferation-related proteins [NGF, p-TrkA, phospho-extracellular signal-regulated kinase 1/2 (p-ERK1/2) and phospho-cAMP response element-binding protein (p-CREB)] decreased in the hippocampus of the offspring in the SCH group, overt hypothyroidism (OHT) group, and the group with levothyroxine (L-T4) treatment for SCH from gestational day 17 (E17). In contrast, expression of apoptosis-related proteins [pro-NGF, p75NTR, phospho-C-Jun N-terminal kinase (p-JNK), p53, Bax and cleaved caspase-3] was increased. The two groups with treatment with L-T4 for SCH from E10 and E13, respectively, showed no significant difference compared with the CON group. L-T4 treatment enhanced relative expression of NGF by increasing NGF/proNGF ratio in offspring from maternal SCH rats. In conclusion, L-T4 treatment for SCH from early pregnancy dramatically ameliorated cognitive impairment via TrkA/p75NTR signaling, which involved activation of the neuronal proliferation and inhibition of neuronal apoptosis in SCH rats’ offspring.

BAPTA, a calcium chelator, neuroprotects injured neurons in vitro and promotes motor recovery after spinal cord transection in vivo.

AimDespite animal evidence of a role of calcium in the pathogenesis of spinal cord injury, several studies conducted in the past found calcium blockade ineffective. However, those studies involved oral or parenteral administration of Ca++ antagonists. We hypothesized that Ca++ blockade might be effective with local/immediate application (LIA) at the time of neural injury.MethodsIn this study, we assessed the effects of LIA of BAPTA (1,2‐bis (o‐aminophenoxy) ethane‐N, N, N′, N'‐tetraacetic acid), a cell‐permeable highly selective Ca++ chelator, after spinal cord transection (SCT) in mice over 4 weeks. Effects of BAPTA were assessed behaviorally and with immunohistochemistry. Concurrently, BAPTA was submitted for the first time to multimodality assessment in an in vitro model of neural damage as a possible spinal neuroprotectant.ResultsWe demonstrate that BAPTA alleviates neuronal apoptosis caused by physical damage by inhibition of neuronal apoptosis and reactive oxygen species (ROS) generation. This translates to enhanced preservation of electrophysiological function and superior behavioral recovery.ConclusionThis study shows for the first time that local/immediate application of Ca++ chelator BAPTA is strongly neuroprotective after severe spinal cord injury.

Xuesaitong injection (lyophilized) combined with aspirin and clopidogrel protect against focal cerebral ischemic/reperfusion injury in rats by suppressing oxidative stress and inflammation and regulating the NOX2/IL-6/STAT3 pathway.

Combination of aspirin (ASA) and clopidogrel (CLP) [dual antiplatelet therapy (DAPT)] has been limited in reducing early recurrent stroke events. Xuesaitong injection (lyophilized) (XST) made of total saponins from P. notoginseng, which significantly improves cerebral circulation and has been widely used in clinical applications for decades to treat and prevent ischemic stroke. Here, we confirmed the protective effect and mechanism of XST combined with DAPT (XST+ASA+CLP) on cerebral ischemia/reperfusion injury, exploring their better pharmacological action for clinical patients. Sprague-Dawley rats (SD rats) (n=9 in each group) were randomly assigned to three groups and pretreated with XST, ASA+CLP, or XST+ASA+CLP for 7 days. Then rats were subjected to 2 h of middle cerebral artery occlusion (MCAO) followed by reperfusion for 24 h. Therapeutic effect of XST+ASA+CLP was measured by infarct volume, neurological behavior and regional cerebral blood flow (rCBF). Inhibition of neuronal apoptosis and glial cells was determined by immunofluorescent staining. We studied the protein levels of neurotrophic factors, neuroplasticity-related factors, oxidative stress indicators and inflammatory factors by ELISA assay. XST+ASA+CLP group showed significant reduction in infarct volumes and neurological deficit scores. XST+ASA+CLP group also had higher levels in rCBF and synaptic growth, and showed remarkable inhibition of microglia and astrocytes activation and the neuronal apoptosis. In addition, XST+ASA+CLP group had lower levels of NADPH, protein carbonyl, 4-hydroxynonenal (4-HNE), 8-hydroxydeoxyguanosine (8-OHdG) and several inflammatory cytokines. Moreover, XST+ASA+CLP group also had lower levels of NOX2, inducible nitric oxide synthase (iNOS), interleukin (IL)-6, and p-STAT3/STAT3. These results demonstrate that a combination of XST, ASA, and CLP effectively protected rats against middle cerebral artery occlusion/reperfusion (MCAO/R) injury by suppressing the NOX2/IL-6/ STAT3 pathway. These novel findings provide theoretical basis and experimental evidence for the rationality of clinical combined use of drugs in the treatment of ischemic stroke.

Role of melatonin in the dynamics of acute spinal cord injury in rats.

Melatonin is well‐documented to have the ability of reducing nerve inflammation and scavenging free radicals. However, the therapeutic effect of melatonin on spinal cord injury has not been fully described. In this study, we assessed the effect of melatonin on T9 spinal cord injury established by Allen method in rats. Melatonin deficiency significantly delayed the recovery of sensory and motor functions in SCI rats. Treatment with melatonin significantly alleviated neuronal apoptosis and accelerated the recovery of spinal cord function. These results suggest that melatonin is effective to ameliorate spinal cord injury through inhibition of neuronal apoptosis and promotion of neuronal repair.

Neuroprotective effects of zonisamide on cerebral ischemia injury via inhibition of neuronal apoptosis

It is known that neuronal apoptosis contributes to pathology of cerebral ischemia injury. Zonisamide (ZNS) has shown anti-apoptosis effects in recent studies. The present study investigated whether the anti-apoptotic effect can account for the neuroprotective action of ZNS on cerebral ischemia. Neuronal cells were maintained under oxygen-glucose deprivation conditions to simulate cerebral ischemia and treated with ZNS simultaneously. The apoptosis of the cells and expression of apoptosis-related proteins were investigated by flow cytometry and western blot analysis, respectively. A cerebral ischemia mouse model was created via middle cerebral artery occlusion, and the mice were treated with ZNS. Neurological deficit scores and infarct volumes of the cerebral ischemia mice were measured. The apoptosis status of the neuronal cells was evaluated by TUNEL staining. In vitro, the ZNS treatment inhibited both the apoptosis of the neuronal cells and apoptosis-related protein expression (caspase-3, caspase-8, and calpain-1) induced by the oxygen-glucose deprivation. The anti-apoptosis effect of ZNS could occur through the blocking of reactive oxygen species. Moreover, ZNS treatment significantly ameliorated neurological deficits and reduced infarct volumes in the cerebral ischemia mice model. In this study, ZNS exerted neuroprotective effects by inhibition of apoptosis in neuronal cells in cerebral ischemia. Therefore, ZNS might be a promising therapy for cerebral ischemia.

Activation of Nurr1 with Amodiaquine Protected Neuron and Alleviated Neuroinflammation after Subarachnoid Hemorrhage in Rats

Background. Nurr1, a member of the nuclear receptor 4A family (NR4A), played a role in neuron protection, anti‐inflammation, and antioxidative stress in multidiseases. We explored the role of Nurr1 on subarachnoid hemorrhage (SAH) progression and investigated the feasibility of its agonist (amodiaquine, AQ) as a treatment for SAH. Methods. SAH rat models were constructed by the endovascular perforation technique. AQ was administered intraperitoneally at 2 hours after SAH induction. SAH grade, mortality, weight loss, neurological performance tests, brain water content, western blot, immunofluorescence, Nissl staining, and qPCR were assessed post‐SAH. In vitro, hemin was introduced into HT22 cells to develop a model of SAH. Results. Stimulation of Nurr1 with AQ improved the outcomes and attenuated brain edema. Nurr1 was mainly expressed in neuron, and administration of AQ alleviated neuron injury in vivo and enhanced the neuron viability and inhibited neuron apoptosis and necrosis in vitro. Besides, AQ reduced the amount of IL‐1β+Iba‐1+ cells and inhibited the mRNA level of proinflammatory cytokines (IL‐1β and TNF‐α) and the M1‐like phenotype markers (CD68 and CD86). AQ inhibited the expression of MMP9 in HT22 cells. Furthermore, AQ reduced the expression of nuclear NF‐κB and Nurr1 while increased cytoplasmic Nurr1 in vivo and in vitro. Conclusion. Pharmacological activation of Nurr1 with AQ alleviated the neuron injury and neuroinflammation. The mechanism of antineuroinflammation may be associated with the Nurr1/NF‐κB/MMP9 pathway in the neuron. The data supported that AQ might be a promising treatment strategy for SAH.

A Network Pharmacology-Based Analysis of the Protective Mechanism of Miao Medicine Xuemaitong Capsule Against Secondary Brain Damage in the Ischemic Area Surrounding Intracerebral Hemorrhage.

Intracerebral hemorrhage (ICH) is a devastating disease with the high mortality. The poor outcome of ICH is partially due to a combination of various secondary insults, including in the ischemic area. Xuemaitong capsule (XMT), a kind of traditional Chinese medicine, has been applied to clinic practice. The purpose of this study is to explore the mechanism of XMT in alleviating secondary damage in the ischemic area after ICH. We screened XMT target, compound components, and ICH-related targets using network pharmacology, cluster analysis, and enrichment analysis. We found that the tumor necrosis factor (TNF) signaling pathway might be the key signaling pathway for XMT treatment of ICH. An ICH rat model was established, as demonstrated by poor neurologic score. In the ICH rats, Western blot analysis and immunofluorescence indicated the upregulated expression of TNF receptor 1 (TNFR1), mitogen-activated protein kinase (MAPK), nuclear factor-κB (NF-κB), and caspase-3 (CASP3). Importantly, administration of XMT alleviated inflammation, edema, and increased perfusion in the ischemic area, whereas the expression of TNFR1, MAPK, NF-κB, and CASP3 was decreased. Furthermore, Fluoro-Jade B and terminal deoxynucleotidyl transferase-mediated digoxigenin-deoxyuridine nick-end labeling staining revealed that XMT application also inhibited apoptosis and degradation of ischemic area neurons. In conclusion, this evidence elucidates that XMT alleviates neuron apoptosis, ischemic area inflammation, edema, and perfusion through the TNFR1-mediated CASP3/NF-κB/MAPK axis. SIGNIFICANCE STATEMENT: Tumor necrosis factor (TNF) is the key signaling pathway of Xuemaitong (XMT) to intervention during intracerebral hemorrhage. Fourteen key targets [intercellular adhesion molecule 1, interleukin (IL) 6, TNF, C-C motif chemokine ligand 2, prostaglandin-endoperoxide synthase 2, v-rel reticuloendotheliosis viral oncogene homolog A, matrix metalloproteinase 9, endothelin-1 (EDN1), mitogen-activated protein kinase (MAPK) 1, fos proto-oncogene protein, caspase-3 (CASP3), jun proto-oncogene, IL1B, MAPK8] are retrieved from the data base. XMT can inhibit neuron apoptosis in the ischemic area via regulating TNF receptor 1 (TNFR1)/CASP3. XMT alleviates inflammation and edema through regulating TNFR1/nuclear factor-κB and TNFR1/MAPK signaling pathways. XMT alleviates hypoperfusion in the cerebral ischemic area through mediating TNFR1/MAPK/EDN1.

Therapeutic effect of metformin on inflammation and apoptosis after spinal cord injury in rats through the Wnt/β-catenin signaling pathway

ObjectiveTo verify the effect of metformin on spinal cord injury (SCI) through Wnt/β-catenin signaling pathway. BackgroundSCI is a serious traumatic disease of the central nervous system. Wnt/β-catenin signaling pathway plays important roles in SCI. Metformin has been reported to exert neuroprotective effects in the central nervous system. Whether metformin could improve SCI through Wnt/β-catenin signaling pathway remains unclear. MethodsRats were divided into sham group, SCI group, SCI + metformin group, metformin + XAV939 group (XAV939 is an effective inhibitor of the Wnt/β-catenin signaling pathway), and methylprednisolone group. BBB scores were used to detect motor function recovery at different time points (0, 1, 3, 7, 14, 21, and 28 days) in SCI rats. Western blot analysis, immunofluorescence, TUNEL, HE and Nissl staining were used to observe the morphological characteristics of spinal cord tissue and the expression of inflammation and apoptosis in spinal cord neurons. ResultsMetformin(50 mg/kg) promoted motor functional recovery in rats after SCI, increased the expressions of β-catenin and brain derived neurotrophic factor (BDNF), inhibited neuron apoptosis and inflammatory response, and improved the recovery of pathological morphology at the injury site by activating the Wnt/β-catenin signaling pathway. ConclusionWe found a possible mechanism that metformin could reduce inflammation and apoptosis, and promote functional recovery of SCI rats through activating Wnt/β-catenin signaling pathway.

MiR-27a promotion resulting from silencing of HDAC3 facilitates the recovery of spinal cord injury by inhibiting PAK6 expression in rats

AimsSpinal cord injury (SCI) is one of the most devastating diseases that challenges neurology and medicine, leading to paraplegia or quadriplegia worldwide. Neuroprotection conferred by histone deacetylase (HDAC) inhibitors against various insults and deficits in the central nervous system has been reported previously. Herein, we set out to ascertain whether HDAC3 inhibition exerts neuroprotective effects against SCI. Main methodsA modified Allen's weight-drop method was performed to induce experimental SCI in rats. Basso-Beattie-Bresnahan (BBB) scores were used to assess locomotor function. Flow cytometric analysis of AnnexinV-FITC/PI double staining, TUNEL staining, and immunoblotting analysis of apoptosis-related proteins were performed to determine apoptosis in H2O2-induced cell injury of primary rat neurons. Key findingsUpregulated HDAC3 and downregulated miR-27a were observed in spinal cord tissues of SCI rats and H2O2-injured neurons. HDAC3 knockdown by its specific shRNA restored the locomotor function of SCI rats and prevented rat neurons from H2O2-induced apoptosis through promotion of miR-27a. miR-27a targeted PAK6 (encoding P21-activated kinase 6) and inhibited its expression. The effects of HDAC3 knockdown on the locomotor function of SCI rats and H2O2-induced apoptosis of rat neurons were lost upon further PAK6 overexpression. SignificanceThe present study uncovers that silencing HDAC3 inhibited PAK6 expression by upregulating miR-27a, eventually inhibiting neuron apoptosis and promoting the recovery of SCI, which might provide a novel therapeutic target for SCI.

MiR-410-3p overexpression ameliorates neurological deficits in rats with hypoxic-ischemic brain damage

Neonatal hypoxic-ischemic encephalopathy (HIE) is major cause of neonatal death or long-term neurodevelopmental disabilities, which becomes a major practical problem currently in clinic. Whereas, its pathophysiology and underlying molecular mechanism is not clear. MicroRNAs are involved in the normal growth and development of neuronal cells. Herein, the objective of this research was to examine the roles of miR-410-3p in neurological deficits, neuronal injury and neuron apoptosis after hypoxic-ischemic and to explore its associated mechanisms. We established the hypoxic-ischemic brain damage (HIBD) model and oxygen glucose deprivation (OGD) model. Zea-longa score and TTC staining were used to detect the acute cerebral dysfunction after HIBD. QPCR verification exhibited notable downregulation of miR-410-3p expression at 24 h in rats after HIBD as well as that in PC12, SY5Y cells and primary cortical neurons post OGD. To further determine the function of miR-410-3p, lentivirus-mediated overexpression virus was applied in vivo and in vitro. Behavioral tests, including Morris water maze, open field test, Y maze test, neurological severity score and rotating rod test, were performed to evaluate long-term behavioral changes of rats at 1 month post HIBD. The results showed that the number of cells together with the axonal length were reduced post OGD. While the increase of cells number and the axonal length was measured after upregulating miR-410-3p. Meanwhile, miR-410-3p overexpression inhibited neuron apoptosis and enhanced neuronal survival. In addition, long-term motor and cognitive functions were remarkably recovered in HIBD rats with miR-410-3p overexpression. Together, miR-410-3p exerts a critical role in protecting neuronal growth as well as promoting motor and cognitive function recovery in neonatal rats subjected to HIBD. The current study therefore provides critical insights to develop the activator of miR-410-3p for the clinical treatment of HIBD in future clinic trial.

Electroacupuncture Inhibits Neuronal Autophagy and Apoptosis via the PI3K/AKT Pathway Following Ischemic Stroke.

Electroacupuncture (EA) is a safe and effective therapy for ischemic stroke in both clinical and laboratory settings. However, the underlying mechanism behind EA treatment for stroke remains unclear. Here, we aimed to evaluate whether EA treatment at the acupoints of Zusanli (ST36) and Quchi (LI11) exerted a neuroprotective effect on ischemic stroke rats by modulating autophagy and apoptosis via the PI3K/AKT/mTOR signaling pathway. EA was performed at 24 h following brain ischemia/reperfusion (I/R) for 30 min per day for 3 days. Our results indicated that EA treatment significantly decreased neurological deficits and cerebral infarct volume in ischemic stroke rats. Also, EA intervention markedly reduced neuronal apoptosis by suppressing the activation of cleaved caspase-3 (CCAS3) at 72 h following I/R, as shown by a Western blot analysis. Furthermore, EA treatment after ischemic stroke suppressed the ischemia activated expression level of LC3II/I and Atg7 and increased the ischemia inhibited expression level of PI3K, phosphorylation of mTOR, phosphorylation of AKT, P62 and LAMP1, hence mediating the autophagy level of the neurocyte, which was reversed by the PI3K inhibitor Dactolisib. In summary, our results indicate that the protective effects of EA treatment at points of Quchi (LI11) and Zusanli (ST36) in rats following cerebral I/R injury was associated with the inhibition of neuronal apoptosis and autophagy via activating the PI3K/AKT/mTOR signaling pathway.

Suppression of lncRNA Xist Improves Sevoflurane-Induced Social and Emotional Impairment by Activating miR-98-5p/EDEM1 Signaling Axis in Neonatal Mice

Numerous animal studies have suggested that sevoflurane can induce neuronal cell death and damage social and emotional behaviors and endoplasmic reticulum stress is a crucial underlying mechanism. The goal of this study was to investigate the role and the mechanisms of long noncoding RNA X-inactive specific transcript (XIST) in sevoflurane-induced social and emotional impairment and neuron apoptosis in neonatal mice. The performance in social and emotional tests and the expression levels of lncRNA-XIST after sevoflurane exposure in neonatal mice and hippocampal neuronal cells were measured. Also, the effects of the suppression of lncRNA-XIST on neuron apoptosis and endoplasmic reticulum stress were determined. Subsequently, the association between lncRNA-XIST and miR-98-5p as well as EDEM1 in hippocampal neuronal cells was explored. Our results showed that social and emotional behaviors were impaired in neonatal mice and lncRNA-XIST and EDEM1were increasingly expressed in hippocampal neuronal cells after sevoflurane exposure and suppression of lncRNA-XIST decreased sevoflurane-induced neuron apoptosis and endoplasmic reticulum stress in vitro. Further in vitro studies showed that overexpression of miR-98-5p decreased apoptosis and endoplasmic reticulum stress while lncRNA Xist negative regulated miR-98-5p expression and the effects of lncRNA Xist on apoptosis and endoplasmic reticulum stress through sponging miR-98-5p. Moreover, suppression of lncRNA-XIST improved sevoflurane-induced social and emotional impairment and alleviated sevoflurane-induced apoptosis and endoplasmic reticulum stress accompanied in neonatal mice. Our findings reveal that knockdown of lncRNA-XIST may contribute to ameliorating sevoflurane-induced social and emotional impairment via inhibiting neuron apoptosis and endoplasmic reticulum stress, which may be achieved by regulating miR-98-5p/EDEM1 axis. Funding Statement: This work was supported by the National Natural Science Foundation of China (81400929, 81471240, 81641042 and 81603545), the Bureau of Chinese Medicine, Zhejiang, China (2018ZB065) and the Innovative Talents Project of Zhejiang province (2016). Declaration of Interests: There is no conflict of interest in the authors. Ethics Approval Statement: The study protocol was approved by the Zhejiang University Institutional review board of Animal studies (ZJU2019-16778). The Guidance Recommendations for Experimental Animal Care and Use of the Ministry of Science and Technology of the People’s Republic of China supervised all experimental procedures.

Remote liver ischaemic preconditioning protects rat brain against cerebral ischaemia-reperfusion injury by activation of an AKT-dependent pathway.

What is the central question of this study? Can remote liver ischaemic preconditioning (RLIPC) protect rat brain against cerebral ischaemia-reperfusion injury? What is the main finding and its importance? Pretreatment with RLIPC reduced cerebral infarct volume, improved neurological outcomes and inhibited neuron apoptosis. RLIPC led to increased phosphorylation of AKT, while inhibition of AKT abolished the effects of RLIPC. Our data suggest that liver ischaemic preconditioning exerts a strong neuroprotective effect against cerebral ischaemia-reperfusion injury by activating an AKT-dependent pathway. Remote limb ischaemic preconditioning has been shown to have beneficial effects in protecting brains against ischaemia-reperfusion (I/R) injury. However, little is known regarding the effect of remote liver ischaemic conditioning (RLIPC). We therefore investigated the effect of RLIPC on brain tissues suffering from I/R injury. Rats were randomly assigned to a sham group, a control group or a RLIPC group. Rats in all groups except for the sham group received middle cerebral artery occlusion (MCAO) for 1h, followed by 48h of reperfusion. For the RLIPC rats, four cycles of 5min of liver ischaemia (portal vein, hepatic arterial and venous trunk occlusion) with 5min intermittent reperfusion were carried out before cerebral ischaemia. Infarct volume was assessed after 48h of reperfusion. Blood samples were taken for serum lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) tests. Morphological changes of cortical tissue and cellular apoptosis were determined. Right cortex tissues were taken for western blotting measurements. Our data demonstrate that RLIPC reduced cerebral I/R injury, decreased the volume of the MCAO-evoked infarct region, decreased serum levels of LDH and CK-MB, and reduced neurological deficits and apoptosis after I/R injury. Moreover, rats receiving RLIPC showed increased cortical AKT phosphorylation, but protein phosphorylation level was unchanged in the survivor activating factor enhancement (SAFE) signalling pathway. Accordingly, inhibition of AKT with wortmannin abolished the neuroprotective action of liver preconditioning. Our study showed for the first time that liver ischaemic preconditioning effectively protects brain against cerebral I/R injury by activating an AKT-dependent pathway.

Treatment of Brain Edema by Wogonoside Is Associated with Inhibition of Neuronal Apoptosis and SIRT1 Activation in Rats.

BackgroundBrain edema and neuronal apoptosis are closely associated with loss of neurological function and death in rats with subarachnoid hemorrhage (SAH). The present study investigated the effect of wogonoside on brain edema induced by SAH in rats and studied the mechanism involved.Material/MethodsThe rats were intra-gastrically administered 10, 20, 50, 100, 150 and 200 mg/kg doses of wogonoside 24 h prior to SAH induction. Western blotting was used to assess levels of pro-apoptotic protein, SIRT1, ZO-1, and p53 protein expression. Apoptotic nuclei were detected using immunofluorescence and TUNEL staining.ResultsWogonoside treatment significantly suppressed edema formation in SAH-induced rats. Pre-treatment with wogonoside exhibited an inhibitory effect on SAH-induced extravascular Evans blue staining in rats. The expression of ZO-1, Occludin, and Claudin-5 proteins was increased by wogonoside in the SAH-induced rats. The inhibitory effect of SAH was completely reversed in the rats treated with the 200 mg/kg dose of wogonoside. The expression of SIRT1 protein was upregulated, and p53 and AC-p53 were downregulated by wogonoside in SAH rats. Wogonoside treatment significantly reduced SAH-mediated promotion of Bax, Puma, Noxa, Bid, and cleaved Caspase-3 expression. In the SAH-induced rats, pre-treatment with wogonoside reduced the TUNEL-positive cell count.ConclusionsThe present study demonstrated that wogonoside prevents brain edema development and apoptosis of neurons in rats by promoting SIRT1 expression and suppression of p53 activation. Therefore, wogonoside has therapeutic potential for the treatment of edema and needs to be investigated further to completely define the mechanism involved.

PI3K/Akt and ERK1/2 pathways are responsible for sodium butyrateinduced inhibition of neuronal apoptosis in rats with cerebral infarction.

The aim of this study was to elucidate the neuronal protection effect of sodium butyrate (NaB) on neuronal apoptosis in rats with cerebral infarction (CI), and the involvement of the phosphatidilinositol 3-kinase/protein kinase B (PI3K/Akt) and extracellular-signal-regulated kinase 1/2 (ERK1/2) pathways. MI model in rats was performed by middle cerebral artery occlusion (MCAO). Three hours after reperfusion, gastric administration of 5 or 10 mg/kg NaB was performed. Neurological deficit score, infarct size and brain edema were evaluated in rats after 24 h of reperfusion. Enzyme-linked immunosorbent assay (ELISA) was conducted to determine contents of oxidative stress factors. Lactate dehydrogenase (LDH) activity, cell viability and apoptosis in extracted neurons were determined. Moreover, expression levels of Bcl-2, Bax, Akt and ERK1/2 were examined. NaB treatment markedly reduced infarct size and brain edema content in CI rats, and NaB treatment improved viability, decreased LDH activity and reversed contents of malondialdehyde (MDA) and superoxide dismutase (SOD) in a dose-dependent manner. In addition, NaB treatment dose-dependently reduced apoptotic rate and Bax level, as well as enhanced Bcl-2 level. Protein levels of Akt and ERK1/2 were markedly upregulated in NaB-treated neurons. NaB treatment alleviates neuronal apoptosis via the PI3K/Akt and ERK1/2 pathways in CI rats, thus protecting the deterioration of CI.

Effect and mechanism of DL-3-n-butylphthalide on emotional memory in mice with chronic alcoholism

Objective To investigate the improvement and possible mechanism of dl-3-n-butylphthalide (NBP) on the emotional memory of chronic alcoholism model mice. Methods Twenty-four adult male C57/BL6 mice were randomly divided into control group (CON, n=8), model group (AT, n=8), treatment group (AT+ NBP, n=8). Mice in AT group and AT+ NBP group were administrated with alcohol to establish chronic alcoholism model. In the AT+ NBP group, the mice was administrated with NBP (40 mg/kg) by gavage once a day for 14 days during the alcohol modeling period. Ang the mice in AT group and CON group was given the same dose of corn oil by gavage.Open field test was used to evaluate anxiety-like behavior, tail suspension test to evaluate depression-like behavior, Morris water maze and new object recognition to evaluate memory ability, and TUNEL staining to evaluate the number of neuron apoptosis. The primary cultured neurons were interfered by alcohol and NBP at the cell level, and the calcium concentration in the neurons was detected by fluorescence calcium imaging. Descriptive analysis, t-test and one-way ANOVA were processed by SPSS 17.0. Results The results of open field test showed that the exploration time of AT+ NBP group was longer than that of AT group ((50.68±7.82)s, (38.50±13.93)s; t=-2.16, P<0.05)). In the spatial memory test, the target quadrant exploration time of AT+ NBP group was longer than that of AT group ((28.02±7.13)s, (20.98±5.58)s; t=-2.20, P<0.05). In short memory test, the cognitive coefficient RI (0.83±0.08) of AT+ NBP group was higher than that of AT group (0.68±0.10) (t=-3.13, P<0.05). Compared with CON group, the number of neuron apoptosis in prefrontal cortex in AT group increased ((17.33±2.51), (115.67±6.50); t=-24.41, P<0.001), and that in AT+ NBP group decreased compared with AT group((45.00±5.57)) (t=14.29, P<0.001). The number of apoptosis neurons in the dentate gyrus of the hippocampus in AT group (13.75±4.79) was also less than that in the AT+ NBP group (5.75±3.30) (t=2.75, P<0.05). Calcium concentration in nerve cells was detected that the three concentrations of alcohol (100 mmol/L, 200 mmol/L and 300 mmol/L) led to a significant increase in the RFU within the nerve cells (△F/F) ((1.43±0.32), (2.31±1.39), (1.21±0.73); t=-7.67, -2.85, -2.86, all P<0.05). In comparison, the changes of RFU in the three groups treated with NBP treatment were relatively stable ((0.04±0.01), (-0.03±0.01), (-0.04±0.02); t=7.96, 2.96, 2.92, all P< 0.05). Conclusion The 3-n-Butylphthalide can improve the learning and memory ability of chronic alcoholism model mice, which may be related with the inhibition of neuron apoptosis and the influence of intracellular calcium homeostasis. Key words: DL-3-n-Butylphthalide; Alcoholism; Anxiety; Memory; Neuronal apoptosis; Calcium homeostasis