Oxygen-glucose Deprivation In Neurons Research Articles

Berberine Alleviates Ischemic Brain Injury by Enhancing Autophagic Flux via Facilitation of TFEB Nuclear Translocation.

Berberine has been demonstrated to alleviate cerebral ischemia/reperfusion injury, but its neuroprotective mechanism has yet to be understood. Studies have indicated that ischemic neuronal damage was frequently driven by autophagic/lysosomal dysfunction, which could be restored by boosting transcription factor EB (TFEB) nuclear translocation. Therefore, this study investigated the pharmacological effects of berberine on TFEB-regulated autophagic/lysosomal signaling in neurons after cerebral stroke. A rat model of ischemic stroke and a neuronal ischemia model in HT22 cells were prepared using middle cerebral artery occlusion (MCAO) and oxygen-glucose deprivation (OGD), respectively. Berberine was pre-administered at a dose of 100[Formula: see text]mg/kg/d for three days in rats and 90[Formula: see text][Formula: see text]M in HT22 neurons for 12[Formula: see text]h. 24[Formula: see text]h after MCAO and 2[Formula: see text]h after OGD, the penumbral tissues and OGD neurons were obtained to detect nuclear and cytoplasmic TFEB, and the key proteins in the autophagic/lysosomal pathway were examined using western blot and immunofluorescence, respectively. Meanwhile, neuron survival, infarct volume, and neurological deficits were assessed to evaluate the therapeutic efficacy. The results showed that berberine prominently facilitated TFEB nuclear translocation, as indicated by increased nuclear expression in penumbral neurons as well as in OGD HT22 cells. Consequently, both autophagic activity and lysosomal capacity were simultaneously augmented to alleviate the ischemic injury. However, berberine-conferred neuroprotection could be greatly counteracted by lysosomal inhibitor Bafilomycin A1 (Baf-A1). Meanwhile, autophagy inhibitor 3-Methyladenine (3-MA) also slightly neutralized the pharmacological effect of berberine on ameliorating autophagic/lysosomal dysfunction. Our study suggests that berberine-induced neuroprotection against ischemic stroke is elicited by enhancing autophagic flux via facilitation of TFEB nuclear translocation in neurons.

The neuroprotective mechanisms of naringenin: Inhibition of apoptosis through the PI3K/AKT pathway after hypoxic-ischemic brain damage

Ethnopharmacological relevanceNaringenin (NGN) is a widely distributed flavonoid with potent antioxidant and neuroprotective properties. Neuroprotective agents play a crucial role in the treatment of hypoxic-ischemic encephalopathy (HIE). It has shown potential therapeutic effects for neurological disorders. However, its efficacy on HIE is yet to be investigated. Aim of the studyThis study aims to investigate the potential neuroprotective effect of naringenin and its underlying molecular mechanisms in reducing oxidative stress, apoptosis, and improving brain outcomes following HIE. Additionally, the study aims to identify the potential targets, mechanisms, and functions of naringenin using network pharmacology analysis. Materials and methodsNeonatal mice were exposed to the hypoxic-ischemic brain damage (HIBD) model to determine brain water content, and brain tissue was subjected to hematoxylin and eosin (HE), immunohistochemistry (IHC), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and Nissl staining to investigate its neuroprotective effects. Furthermore, the neonatal mouse primary neuron oxygen-glucose deprivation (OGD) model to measure reactive oxygen species (ROS) production in vitro. The protein levels were characterized by Western Blot, and mRNA levels were evaluated by a real-time quantitative PCR detecting system (qPCR). Transmission electron microscopy (TEM) and mitochondrial fluorescent staining were used to observe mitochondrial morphology. Neuronal nuclei (NeuN) and microtubule-associated protein 2 (MAP2) were detected by Immunofluorescence (IF). Finally, network pharmacology was employed to determine the common target of naringenin and HIE. The core genes were obtained via protein-protein interaction networks (PPI) analysis and molecular docking was examined, and the mechanism of action was explored through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Additionally, small interfering RNA (siRNA) was constructed for verification. ResultsNaringenin has a neuroprotective effect in HIBD by modulating Vegfa expression and activating the PI3K/AKT pathway to inhibit apoptosis. Furthermore, molecular docking results suggest that Vegfa is a potential binding target of naringenin, and silencing Vegfa partially reverses the pharmacological effects of NGN. ConclusionOur findings suggest that naringenin demonstrates potential clinical application for treating HIE as a novel neuroprotective agent.

AQP4 knockout promotes neurite outgrowth via upregulating GAP43 expression in infant rats with hypoxic-ischemic brain injury.

Neonatal hypoxic-ischemic encephalopathy (NHIE) induces severe cerebral damage and neurological dysfunction, with seldom effective therapy. Aquaporin-4 (AQP4)isinvolvedin aggravating brain damage induced by NHIE. This study aimed to investigate the role of AQP4 underlying the pathogenesis of NHIE. Neonatal Sprague-Dawley rats were used to establish neonatal hypoxic-ischemic (HI) models, and the expression of AQP4 in the cortex, hippocampus, and lung tissues was detected by real-time quantitative polymerase chain reaction as well as Western blot. Primary cortical neurons were cultured for the oxygen-glucose deprivation (OGD) model, and siRNA was used to silence the expression of AQP4. Immunostaining of Tuj1 was performed to observe the axonal growth. CRISPER/Cas9 technology was used to knock out AQP4. The results demonstrated thatAQP4 was upregulated in the cortex, hippocampus, and lung tissues in neonatal rats with HI andOGD neurons. Besides, silencing AQP4 promoted axonal growth of OGD neurons, and AQP4 knockout notably improved long-term neurobehavioral impairment. Furthermore, GAP43 was found closely correlated with AQP4 via GeneMANIA prediction. Significant downregulation of GAP43 was induced in OGD neurons, while AQP4 knockout markedly upregulated its expression in rats. This indicated that the depletion of AQP4 may enhance axonal regeneration and promote the long-term neurobehavioral recovery associated with the upregulation of GAP43 expression.

Scutellarin protects cortical neurons against neonatal hypoxic-ischemic encephalopathy injury via upregulation of vascular endothelial growth factor.

Neonatal hypoxic-ischemic encephalopathy (NHIE) causes devastating cerebral damage and neurological deficits that seldom have effective therapies. This study aimed to explore the mechanisms underlying the therapeutic efficacy of Scutellarin in NHIE. NHIE models were successfully established. Zea-longa score and triphenyte-trazoliumchloride (TTC) staining demonstrated that hypoxia and ischemia (HI) insult induced prominent neurological dysfunctions and brain infarction. Protein microarray was applied to detect the differentially expressed genes in the cortex, hippocampus, and lung tissues of HI rats, which revealed the downregulation of vascular endothelial growth factor (VEGF) in these tissues. Additionally, double immunostaining uncovered VEGF expression was localized in the neurons. Besides, VEGF was decreasingly expressed in oxygen-glucose deprivation (OGD) neurons, which was intriguingly reversed by Scutellarin treatment. Moreover, VEGF silencing increased OGD-induced neuronal apoptosis and attenuated neurite outgrowth, which was enhanced by Scutellarin administration. GeneMANIA predicted a close correlation of VEGF with caspase 3, caspase 7, and interleukin(IL)-1β, and qRT-PCR revealed that Scutellarin treatment depressed the expression levels of them elevated in OGD neurons, but the Scutellarin-depressed levels of these factors were prominently increased after VEGF silencing. Our findings suggested that Scutellarin exerted neuroprotective effects in NHIE potentially through mediating VEGF-targeted inactivation of caspase 3, caspase 7, and IL-1β.

Beta-Boswellic Acid Protects Against Cerebral Ischemia/Reperfusion Injury via the Protein Kinase C Epsilon/Nuclear Factor Erythroid 2-like 2/Heme Oxygenase-1 Pathway.

Ischemic strokes are associated with a high rate of disability and death globally. Cerebral ischemia/reperfusion (I/R) injury is a type of brain damage associated with oxidative stress after an ischemic stroke. Beta-boswellic acid (β-BA) reportedly exerts antioxidant and neuroprotective effects, but its role in cerebral I/R injury is unclear. The aim of this research was to investigate the neuroprotective effects, as well as the mechanisms of β-BA in cerebral I/R injury. In vivo experiments were conducted using a rat middle cerebral artery occlusion and reperfusion (MCAO/R) model, and in vitro experiments were performed using a rat neuronal oxygen-glucose deprivation and reoxygenation (OGD/R) model. Triphenyltetrazolium chloride staining, neurological function scores, terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling, hematoxylin and eosin staining, and antioxidant levels in the brain were used to assess the effects of β-BA. Flow cytometry was used to detect reactive oxygen species and apoptotic cells. Western blotting and immunofluorescence staining were used to measure protein levels. The results showed that β-BA markedly improved neurological deficits and decreased infarct volume and necrotic neurons in rats. The in vitro results showed that β-BA protected neurons against OGD/R-induced injury. Additionally, β-BA significantly increased the phosphorylation of protein kinase C epsilon (PRKCE) at S729, the translocation of nuclear factor erythroid 2-like 2 (NFE2L2), and expression of heme oxygenase-1 (HMOX1). This study demonstrates that β-BA exerts neuroprotective effects against cerebral I/R via the activation of the PRKCE/NFE2L2/HMOX1 pathway and is a potential therapeutic candidate for ischemic stroke.

Atorvastatin inhibits neuronal apoptosis via activating cAMP/PKA/p-CREB/BDNF pathway in hypoxic-ischemic neonatal rats.

Neuronal apoptosis is one of the main pathological processes of hypoxic-ischemic brain damage (HIBD) and is involved in the development of hypoxic-ischemic encephalopathy (HIE) in neonates. Atorvastatin has been found to have neuroprotective effects in some nervous system diseases, but its role in regulating the pathogenesis of neonatal HIBD remains elusive. Thus, this study aimed to explore the effects and related mechanisms of atorvastatin on the regulation of neuronal apoptosis after HIBD in newborn rats. The rat HIBD model and the neuronal oxygen glucose deprivation (OGD) model were established routinely. Atorvastatin, cAMP inhibitor (SQ22536), and BDNF inhibitor (ANA-12) were used to treat HIBD rats and OGD neurons. Cerebral infarction, learning and memory ability, cAMP/PKA/p-CREB/BDNF signaling molecules, and apoptosis-related indicators (TUNEL, cleaved caspase-3, and Bax/Bcl2) were then examined. In vivo, atorvastatin reduced cerebral infarction, improved learning and memory ability, decreased the number of TUNEL-positive neurons, inhibited the expression of cleaved caspase-3 and Bax/Bcl2, and activated the cAMP/PKA/p-CREB/BDNF pathway in the cerebral cortex after HIBD. In vitro, atorvastatin also decreased the apoptosis-related indicators and activated the cAMP/PKA/p-CREB/BDNF pathway in neurons after OGD. Furthermore, inhibition of cAMP or BDNF attenuated the effect of atorvastatin on the reduction of neuronal apoptosis, suggesting that atorvastatin inhibits HIBD-induced neuronal apoptosis and alleviates brain injury in neonatal rats mainly by activating the cAMP/PKA/p-CREB/BDNF pathway. In conclusion, atorvastatin may be developed as a potential drug for the treatment of neonatal HIE.

Exosomes Secreted by Hypoxia-Pre-conditioned Adipose-Derived Mesenchymal Stem Cells Reduce Neuronal Apoptosis in Rats with Spinal Cord Injury.

Neuronal death is the main cause of nerve function impairment after spinal cord injury (SCI). Exosome-based therapy has become a novel strategy for tissue injury repair. We designed a method to treat SCI using exosomes secreted by adipose tissue-derived stromal cells (ADSCs) under hypoxic conditions. We established a neuronal oxygen-glucose deprivation and reperfusion (OGD/R) model in vitro to simulate the hypoxic environment after SCI. We observed that exosomes derived from hypoxia-conditioned ADSCs (Hypo-exo) significantly reduced neuronal apoptosis after OGD. By establishing a rat SCI model, we found that Hypo-exo can significantly reduce the formation of cavities in the injured area and improve the functional recovery of the hindlimbs of rats after injury. To explore the molecular mechanism, we conducted microRNA sequencing analysis of exosomes. Through real-time polymerase chain reaction, dual luciferase reporter assays and signaling pathway chip analysis, we determined that miR-499a-5p regulates the JNK3/c-jun-apoptotic signaling pathway by targeting JNK3. Further, we verified the expression of the key proteins in the JNK3/c-jun-apoptotic signaling pathway by immunofluorescence and Western blotting. These results support the hypothesis that Hypo-exo can reduce neuronal apoptosis after SCI and may provide new methods to treat SCI.

Down-Regulated microRNA-192-5p Protects Against Hypoxic-Ischemic Brain Damage via Regulation of YAP1-Mediated Hippo Signaling Pathway.

Hypoxic-ischemic brain damage (HIBD) is a familiar neurological disorder. Emerging reports manifest that microRNAs (miRs) are related to the progression of HIBD. The goal of this study is to explore the mechanism of miR-192-5p in HIBD via regulation of Yes-associated protein 1 (YAP1)-mediated Hippo signaling pathway. The miR-192-5p, YAP1, and Hippo pathway-related factors Phospho (p)-Triaminoguanidinium azide (TAZ) in hippocampal tissues and neurons were detected. The regulatory relationship between miR-192-5p and YAP1 was verified. Neonatal hypoxic ischemia and oxygen-glucose deprivation (OGD) were used to simulate HIBD in vivo and in vitro. The neurobehavioral impairment, neuronal damage and vascular endothelial growth factor (VEGF) expression of neonatal rats in each group were detected. The viability, apoptosis and VEGF expression of hippocampal neurons in each group were also examined. MiR-192-5p expression was elevated while YAP1 expression was reduced in hippocampal tissues of HIBD rats in vivo and OGD neurons in vitro. MiR-192-5p had a targeting relation with YAP1. Suppressed miR-192-5p or overexpressed YAP1 in HIBD rats alleviated neurobehavioral impairment and neuronal damage, and decreased the expression levels of p-TAZ and VEGF expression in vivo. Reduced miR-192-5p or augmented YAP1 decelerated the neuron apoptosis, decreased the p-TAZ level and VEGF level and promoted cell viability of OGD hippocampal neurons in vitro. The study highlights that inhibited miR-192-5p protects against HIBD via regulation of YAP1 and Hippo signaling pathway, which is beneficial for HIBD treatment.

Oxygen-glucose deprivation in neurons: implications for cell transplantation therapies

Cell replacement therapies hold the potential to restore neuronal networks compromised by neurodegenerative diseases (such as Parkinson's disease or Huntington's disease), or focal tissue damage (via a stroke or spinal cord injury). Despite some promising results achieved to date, transplanted cells typically exhibit poor survival in the central nervous system, thus limiting therapeutic efficacy of the graft. Although cell death post-transplantation is likely to be multifactorial in causality, growing evidence suggests that the lack of vascularisation at the graft site, and the resulting ischemic host environment, may play a fundamental role in the fate of grafted cells. Herein, we summarise data showing how the deprivation of either oxygen, glucose, or both in combination, impacts the survival of neurons and review strategies which may improve graft survival in the central nervous system.

Exosomes from Human Umbilical Vein Endothelial Cells Ameliorate Ischemic Injuries by Suppressing the RNA Component of Mitochondrial RNA-processing Endoribonuclease via the Induction of miR-206/miR-1-3p Levels

Exosomes might mediate the effects of remote ischemic post-conditioning (RIPostC) treatment on vital organs. The present study aimed to explore the role of RNA component of mitochondrial RNA-processing endoribonuclease (RMRP) in the effects of human umbilical vein endothelial cell (HUVEC)-derived exosomes on ischemic injuries in vitro and in vivo. HUVECs were subjected to oxygen-glucose deprivation (OGD) treatment and exosomes were collected OGD-treated human neural cells were incubated with HUVEC-derived exosomes. Changes in cell viability, apoptosis, and RMRP-mediated PI3K/Akt/mTOR pathway activity were detected. The role of RMRP inhibition in the anti-OGD effects of exosomes was further determined by upregulating RMRP expression in human neural cells. The potential RMRP inhibitory factors in exosomes were explored using microarray detection. The effects of exosomes were validated with MCAO mouse models. In OGD neurons incubated with the exosomes, cell viability was improved and cell apoptosis was suppressed. At molecular level, exosomes on downregulated RMRP, p-PI3K, p-Akt, and p-mTOR, while induced eNOS. After the overexpression of RMRP, the cell protective effects of exosomes were counteracted, which was associated with the re-activation of PI3K/Akt/mTOR pathway. Based on the detection of microarray, the induced levels of miR-206 and miR-1-3p by OGD in HVUECs contributed to the RMPR inhibition. Additionally, injection of exosomes restricted infarction area and suppressed RMRP in MCAO mice. Collectively, exosomes from OGD HUVECs could protect neurons against ischemia-induced injuries, and the effects were associated with the suppression of RMRP in neurons via distance transfer of miR-206 and miR-1-3p.

IL-27 Protects the Brain from Ischemia-Reperfusion Injury via the gp130/STAT3 Signaling Pathway.

The occurrence of ischemia-reperfusion (I/R) injury leads to dysfunction as well as high rates of morbidity and mortality in stroke, and new effective therapeutic strategies for I/R are still needed. We investigated the effect of IL-27 on I/R injury-induced neurological function impairment, cerebral infarction volume and variation in levels of inflammatory factors in mice with middle cerebral artery occlusion (MCAO), as well as concentration of LDH and neuronal apoptosis in a neuron oxygen-glucose deprivation and reperfusion (OGD/R) model mediated by gp130/STAT3 signaling in vitro. Our results indicated that IL-27 could bind to its receptor of gp130 to attenuate the I/R injury-induced impairment function and cerebral infarction volume, and decrease inflammatory cytokines TNF-α, IL-1β and MCP-1 but increase anti-inflammatory factors IL-10 and TGF-β in vivo, while inhibiting LDH leakage and neuronal apoptosis through activation of STAT3 to antagonize I/R induction. Our results suggest that IL-27 may protect the brain from I/R injury through the gp130/STAT3 signaling pathway.

Neuroprotective Effects of Oxymatrine on PI3K/Akt/mTOR Pathway After Hypoxic-Ischemic Brain Damage in Neonatal Rats.

Oxymatrine (OMT), a quinolizidine alkaloid extracted from traditional Chinese herb Sophora flavescens Ait, has drawn attention because of its beneficial bioactivities against hypoxic–ischemic brain damage (HIBD). However, the underlying molecular mechanism remains unclear. In this study, we determined the in vivo and in vitro effects of OMT on seven-day old Sprague–Dawley rats with HIBD and in a rat model of primary hippocampal neuron oxygen glucose deprivation reoxygenation (OGD/R). This study was aimed to evaluate whether OMT exerted neuroprotective effects mediated by the (phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin) PI3K/Akt/mTOR pathway after HIBD. Experimental results showed that the alkaloid significantly improved the early neurofunctional development, brain water content, abnormal pathological changes, and necrosis of neurons after HIBD. Moreover, OMT enhanced the cell viability and stabilized the mitochondrial permeability transition pore in the primary hippocampal neurons after OGD/R. OMT significantly decreased the autophagosome generation, elevated the expression of PI3K, Akt, and mTOR, and simultaneously reversed the mRNA expression of microtubule-associated protein 1-light chain 3 (LC3), Beclin-1, and sequestosomel (P62) induced by hypoxia and ischemia. However, these protective effects against HIBD could be suppressed when rapamycin, a specific inhibitor of mTOR, was included. Hence, the OMT exerted neuroprotective effects against HIBD by attenuating excessive autophagy by mediating the PI3K/Akt/mTOR pathway.

Neuroprotective Effects Against Cerebral Ischemic Injury Exerted by Dexmedetomidine via the HDAC5/NPAS4/MDM2/PSD-95 Axis.

Numerous evidences have highlighted the efficient role of dexmedetomidine (DEX) in multi-organ protection. In the present study, the neuroprotective role of DEX on cerebral ischemic injury and the underlining signaling mechanisms were explored. In order to simulate cerebral ischemic injury, we performed middle cerebral artery occlusion in mice and oxygen-glucose deprivation in neurons. Immunohistochemistry, Western blot analysis, and RT-qPCR were used to examine expression of HDAC5, NPAS4, MDM2, and PSD-95 in hippocampus tissues of MCAO mice and OGD-treated neurons. MCAO mice received treatment with DEX and sh-PSD-95, followed by neurological function evaluation, behavioral test, infarct volume detection by TTC staining, and apoptosis by TUNEL staining. Additionally, gain- and loss-of-function approaches were conducted in OGD-treated neuron after DEX treatment. Cell viability and apoptosis were assessed with the application of CCK-8 and flow cytometry. The interaction between MDM2 and PSD-95 was evaluated using Co-IP assay, followed by ubiquitination of PSD-95 detection. As per the results, HDAC5 and MDM2 were abundantly expressed, while NPAS4 and PSD-95 were poorly expressed in hippocampus tissues of MCAO mice and OGD-treated neurons. DEX elevated viability, and reduced LDH leakage rate and apoptosis rate of OGD-treated neurons, which was reversed following the overexpression of HDAC5. Moreover, HDAC5 augmented MDM2 expression via NPAS4 inhibition. MDM2 induced PSD-95 ubiquitination and degradation. In MCAO mice, DEX improved neurological function and behaviors and decreased infarct volume and apoptosis, which was negated as a result of PSD-95 silencing. DEX plays a neuroprotective role against cerebral ischemic injury by disrupting MDM2-induced PSD-95 ubiquitination and degradation via HDAC5 and NPAS4.

Neuroprotective effects of microRNA-140-5p on ischemic stroke in mice via regulation of the TLR4/NF-κB axis

Background and aimIschemic stroke is one of the main causes of death worldwide and permanent global disability. On the basis of existing literature data, the study was carried out in an effort to explore how miR-140-5p affects ischemic stroke and whether the mechanism relates to toll-like receptor-4 (TLR4) and nuclear factor-kappa B (NF-κB). MethodsFirstly, middle cerebral artery occlusion (MCAO) was performed to establish mouse models of ischemic stroke in vivo, while primary neurons were exposed to oxygen-glucose deprivation (OGD) to set up an ischemic stroke model in vitro. RT-qPCR was then applied to detect the miR-140-5p expression patterns, whereas Western blot was adopted to detect the expression patterns of TLR4, NF-κB, and apoptosis-related factors. In addition, based gain-function of experiments using miR-140-5p mimic and TLR4 over-expression plasmid, neurological function score, TTC staining, TUNEL staining, as well as flow cytometry were carried out to evaluate the effects of miR-140-5p and TLR4 on MCAO mice and OGD neurons. Moreover, dual-luciferase reporter assay was applied to validate the targeting relationship between miR-140-5p and TLR4. ResultsInitial findings revealed that miR-140-5p was poorly-expressed, while TLR4 was highly-expressed in ischemic stroke. It was verified that miR-140-5p targeted TLR4 and downregulated its expression. MiR-140-5p over-expression was observed to inhibit the apoptosis of neurons under OGD exposure and restrain the progression of ischemic stroke, while TLR4 over-expression promoted the apoptosis and disease progression. Besides, miR-140-5p over-expression led to a decrease in NF-κB protein levels, which were increased by TLR4 over-expression. ConclusionIn conclusion, our data indicates that miR-140-5p over-expression may be instrumental for the therapeutic targeting of ischemic stroke by alleviating neuron injury with the involvement of the TLR4/NF-κB axis.

Agrin Involvement in Synaptogenesis Induced by Exercise in a Rat Model of Experimental Stroke

Background Agrin is a proteoglycan that aggregates nicotinic acetylcholine receptors (AChRs) on neuromuscular junctions and takes part in synaptogenesis in the development of the central nervous system. However, its effects on neural repair and synaptogenesis after stroke are still unclear. Objective This study aimed to investigate the effects of agrin on neural repair and synaptogenesis after stroke and the effects of exercise on this process in vivo and in vitro. Methods Exercise with gradually increased intensity was initiated at 1 day after middle cerebral artery occlusion (MCAO) for a maximum of 14 days. Neurological deficit scores and foot fault tests were used to assess the behavioral recovery. Western blotting, immunofluorescence, and electron microscopic images were used to detect the expression of agrin, synaptogenesis-related proteins, and synaptic density in vivo. In vitro, the ischemic neuron model was established via oxygen-glucose deprivation (OGD). The lentivirus overexpressed agrin and CREB inhibitor were used to investigate the mechanism by which agrin promoted synaptogenesis. Results Exercise promoted behavioral recovery and this beneficial role was linked to the upregulated expression of agrin and increased synaptic density. Overexpressed agrin promoted synaptogenesis in OGD neuron, CREB inhibitor downregulated the expression of agrin and hampered synaptogenesis in cultured neurons. Conclusions These results indicated that exercise poststroke improved the recovery of behavioral function after stroke. Synaptogenesis was an important and beneficial factor, and agrin played a critical role in this process and could be a potential therapeutic target for the treatment of stroke and other nervous system diseases.

LXA4 protects against hypoxic-ischemic damage in neonatal rats by reducing the inflammatory response via the IκB/NF-κB pathway

Hypoxia and the resultant decreases in cerebral blood flow in the perinatal period can lead to neonatal hypoxic-ischemic (HI) brain injury, which can, in turn, cause severe disability or even death. However, the efficacy of current treatment strategies remains limited. Several studies have demonstrated that lipoxin A4 (LXA4), as one of the earliest types of endogenous lipid mediators, can inhibit the accumulation of neutrophils, arrest inflammation, and promote the resolution of inflammation. However, research on LXA4 in the nervous system has rarely been carried out. In the present study, we sought to investigate the protective effect of LXA4 on HI brain damage in neonatal rats, as well as the underlying mechanisms. Through experiments conducted using an HI animal model, we found that the LXA4 intervention promoted the recovery of neuronal function and tissue structure following brain injury while maintaining the integrity of the blood–brain barrier in addition to reducing cerebral edema, infarct volume, and inflammatory responses. Our results suggest that LXA4 interfered with neuronal oxygen-glucose deprivation insults, reduced the expression of inflammatory factors, inhibited apoptosis, and promoted neuronal survival in vitro. Finally, the LXA4 intervention attenuated HI-induced activation of inhibitor kappa B (IκB) and degradation of nuclear factor-κB (NF-κB). In conclusion, our data suggest that LXA4 exerts a neuroprotective effect against neonatal HI brain damage through the IκB/NF-κB pathway. Our findings will help inform future studies regarding the effects of LXA4 on neuroinflammation, blood–brain barrier integrity, and neuronal apoptosis.

Cocaine- and amphetamine-regulated transcript protects synaptic structures in neurons after ischemic cerebral injury

Cocaine-regulated and amphetamine-regulated transcript (CART) is a neuropeptide with reported neuroprotective effects in ischemic cerebral injury. However, its mechanism has not yet been elucidated. Herein, we investigated the role and mechanism of CART in synaptic plasticity in neurons after ischemic cerebral stroke. We found that the survival rate of the oxygen-glucose deprivation (OGD) neurons was increased after CART treatment. Moreover, CART treatment significantly attenuated ischemia-induced neuronal synaptic damage and increased synaptophysin expression. In addition, the number of presynaptic vesicles was increased and the postsynaptic density (PSD) was thickened after CART treatment. Mechanistically, CART treatment enhanced the expression of Arc mRNA in a cAMP response element binding protein (CREB) dependent manner in OGD neurons, and blockade of CREB by KG-501 eliminated the protective effect of CART. Collectively, CART protected the synaptic structure in neurons after ischemic cerebral injury by increasing the Arc expression via upregulating p-CREB.

Melatonin protects against ischemic stroke by modulating microglia/macrophage polarization toward anti-inflammatory phenotype through STAT3 pathway.

AimsMicroglia and infiltrated macrophages play important roles in inflammatory processes after ischemic stroke. Modulating microglia/macrophage polarization from pro‐inflammatory phenotype to anti‐inflammatory state has been suggested as a potential therapeutic approach in the treatment of ischemic stroke. Melatonin has been shown to be neuroprotective in experimental stroke models. However, the effect of melatonin on microglia polarization after stroke and underlying mechanisms remain unknown.MethodsIn vivo, cerebral ischemia was induced by distal middle cerebral artery occlusion (dMCAO) in C57BL/6J mice. Melatonin was injected intraperitoneally (20 mg/kg) at 0 and 24 hours after ischemia. In vitro, the microglial cell line BV2 was stimulated to the pro‐inflammatory state with conditioned media (CM) collected from oxygen‐glucose deprivation (OGD) challenged neuronal cell line Neuro‐2a (N2a). Real‐time PCR was utilized to detect the mRNA expression of microglia phenotype markers. Activation of signal transducer and activator of transcription 3 (STAT3) pathway was determined by Western blot of phosphorylated STAT3 (pSTAT3). A neuron‐microglia co‐culture system was used to determine whether melatonin can inhibit the neurotoxic effect of pro‐inflammatory microglia to post‐OGD neurons.ResultsMelatonin treatment reduced brain infarct and improved neurological functions 3 days after dMCAO, which was accompanied by decreased expression of pro‐inflammatory markers and increased expression of anti‐inflammatory markers in the ischemic brain. In vitro studies confirmed that melatonin directly inhibited the pro‐inflammatory responses in BV2 cells upon exposure to OGD neuron CM. The microglia possessing pro‐inflammatory phenotype exacerbated post‐OGD N2a cells death, whereas melatonin reduced such neurotoxic effect. Further, melatonin enhanced the otherwise inhibited pSTAT3 expression in BV2 cells treated with OGD neuron CM. STAT3 blockade significantly reduced the effect of melatonin on microglial phenotype shift. Conclusion: Melatonin treatment ameliorates brain damage at least partially through shifting microglia phenotype from pro‐inflammatory to anti‐inflammatory polarity in a STAT3‐dependent manner.

Inhibition of PDE4 protects neurons against oxygen-glucose deprivation-induced endoplasmic reticulum stress through activation of the Nrf-2/HO-1 pathway

Inhibition of phosphodiesterase 4 (PDE4) produces neuroprotective effects against cerebral ischemia. However, the involved mechanism remains unclear. Augmentation of endoplasmic reticulum (ER) stress promotes neuronal apoptosis, and excessive oxidative stress is an inducer of ER stress. The present study aimed to determine whether suppression of ER stress is involved in the protective effects of PDE4 inhibition against cerebral ischemia. We found that exposing HT-22 cells to oxygen-glucose deprivation (OGD) significantly activated ER stress, as evidenced by increased expression of the 78-kDa glucose-regulated protein (GRP78), phosphorylated eukaryotic translation-initiation factor 2α (eIF2α), and C/EBP-homologous protein (CHOP). Overexpression of PDE4B increased ER stress, while knocking down PDE4B or treatment with the PDE4 inhibitor, FCPR03, prevented OGD-induced ER stress in HT-22 cells. Furthermore, FCPR03 promoted the translocation of nuclear factor erythroid 2-related factor 2 (Nrf-2) from the cytoplasm to the nucleus. Importantly, the Nrf-2 inhibitor, ML385, blocked the inhibitory role of FCPR03 on OGD-induced ER stress. ML385 also abolished the protective role of FCPR03 in HT-22 cells subjected to OGD. Knocking down heme oxygenase-1 (HO-1), which is a target of Nrf-2, also blocked the protective role of FCPR03, enhanced the level of reactive oxygen species (ROS), and increased ER stress and cell death. We then found that FCPR03 or the antioxidant, N-Acetyl-l-cysteine, reduced oxidative stress in cells exposed to OGD. This effect was accompanied by increased cell viability and decreased ER stress. In primary cultured neurons, we found that FCPR03 reduced OGD-induced production of ROS and phosphorylation of eIF2α. The neuroprotective effect of FCPR03 against OGD in neurons was blocked by ML385. These results demonstrate that inhibition of PDE4 activates Nrf-2/HO-1, attenuates the production of ROS, and thereby attenuates ER stress in neurons exposed to OGD. Additionally, we conclude that FCPR03 may represent a promising therapeutic agent for the treatment of ER stress-related disorders.

Upregulated microRNA-31 inhibits oxidative stress-induced neuronal injury through the JAK/STAT3 pathway by binding to PKD1 in mice with ischemic stroke.

Ischemic stroke (IS), which is characterized by high morbidity, disability, and mortality, is recognized as a major cerebrovascular disease. MicroRNA-31 (miR-31) was reported to participate in the progression of brain disease. The present study was conducted in order to investigate the effect of miR-31 on oxidative stress-induced neuronal injury in IS mice with the involvement of protein kinase D1 (PKD1) and the JAK/STAT3 pathway. C57BL/6J mice were used to establish the middle cerebral artery occlusion (MCAO) model. Astrocytes were transfected with miR-31 mimic, miR-31 inhibitor, si-PKD1, or JAK-STAT3 pathway inhibitor. Following the establishment of an oxygen-glucose deprivation (OGD) model, the astrocytes were cocultured with neuronal OGD. Lower miR-31, higher PKD1 expressions, and activated JAK/STAT3 pathway were found in both the MCAO and OGD models. miR-31 could negatively target PKD1. In an MCAO model, overexpressing miR-31 and silencing PKD1 reduced neuronal injury, cerebral infarct volume, neuron loss, and oxidative stress injury, inhibited the activation of JAK/STAT3 pathway and the expressions of PKD1, interleukin (IL)-1β, IL-6, tumor necrosis factor-α, malondialdehyde, 4-HNE, 8-HOdG, caspase-3, and Bax, but increased the superoxide dismutase content. In the OGD model, overexpression of miR-31 and silencing of PKD1 attenuated oxidative stress-induced neuronal injury, and diminished the lactate dehydrogenase leakage and reactive oxygen species level, accompanied by elevated neuronal viability. These results indicate that miR-31 alleviates inflammatory response as well as an oxidative stress-induced neuronal injury in IS mice by downregulating PKD1 and JAK/STAT3 pathway.