Brain Cell Apoptosis Research Articles

ADAR1 exacerbates ischemic brain injury via astrocyte-mediated neuron apoptosis

Astrocytes affect stroke outcomes by acquiring functionally dominant phenotypes. Understanding molecular mechanisms dictating astrocyte functional status after brain ischemia/reperfusion may reveal new therapeutic strategies. Adenosine deaminase acting on RNA (ADAR1), an RNA editing enzyme, is not normally expressed in astrocytes, but highly induced in astrocytes in ischemic stroke lesions. The expression of ADAR1 steeply increased from day 1 to day 7 after middle cerebral artery occlusion (MCAO) for 1 h followed by reperfusion. ADAR1 deficiency markedly ameliorated the volume of the cerebral infarction and neurological deficits as shown by the rotarod and cylinder tests, which was due to the reduction of the numbers of activated astrocytes and microglia. Surprisingly, ADAR1 was mainly expressed in astrocytes while only marginally in microglia. In primary cultured astrocytes, ADAR1 promoted astrocyte proliferation via phosphatidylinositol 3-kinase (PI3K)/Akt pathway. Furthermore, ADAR1 deficiency inhibited brain cell apoptosis in mice with MCAO as well as in activated astrocyte-conditioned medium-induced neurons in vitro. It appeared that ADAR1 induces neuron apoptosis by secretion of IL-1β, IL-6 and TNF-α from astrocytes through the production of reactive oxygen species. These results indicated that ADAR1 is a novel regulator promoting the proliferation of the activated astrocytes following ischemic stroke, which produce various inflammatory cytokines, leading to neuron apoptosis and worsened ischemic stroke outcome.

Neurotoxicity and the potential molecular mechanisms of mono-2-ethylhexyl phthalic acid (MEHP) in zebrafish

Mono-2-ethylhexyl phthalic acid (MEHP) is the most toxic metabolite of plasticizer di-2-ethylhexyl phthalic acid (DEHP), and there is limited information available on the effects of MEHP on neurotoxicity. This study aims to examine the neurotoxicity of MEHP and preliminarily explore its potential molecular mechanisms. We found that MEHP impeded the growth of zebrafish embryos and the neurodevelopmental-related gene expression at environmentally relevant concentrations. MEHP exposure also induces oxidative stress response and brain cell apoptosis accompanied by a decrease in acetylcholinesterase (AChE) activity in zebrafish larvae. RNA-Seq and bioinformatics analysis showed that MEHP treatment altered the nervous system, neurogenic diseases, and visual perception pathways. The locomotor activity in dark-to-light cycles and phototaxis test confirmed the abnormal neural behavior of zebrafish larvae. Besides, the immune system has produced a large number of differentially expressed genes related to neural regulation. Inflammatory factor IL1β and IL-17 signaling pathways highly respond to MEHP, indicating that inflammation caused by immune system imbalance is a potential mechanism of MEHP-induced neurotoxicity. This study expands the understanding of the toxicity and molecular mechanisms of MEHP, providing a new perspective for in-depth neurotoxicity exploration of similar compounds.

Piceatannol improved cerebral blood flow and attenuated JNK3 and mitochondrial apoptotic pathway in a global ischemic model to produce neuroprotection.

Cerebral ischemia is one of the leading causes of death and disability worldwide. The only FDA-approved treatment is recanalization with systemic tissue plasminogen activators like alteplase, although reperfusion caused by recanalization can result in neuroinflammation, which can cause brain cell apoptosis. Therefore, after an ischemic/reperfusion injury, interventions are needed to minimize the neuroinflammatory cascade. In the present study, piceatannol (PCT) was studied for its neuroprotective efficacy in a rat model of global ischemic injury by attenuating c-Jun N-terminal kinase 3 (JNK3) downstream signaling. PCT is a resveratrol analog and a polyphenolic stilbenoid naturally occurring in passion fruit and grapes. The neuroprotective efficacy of PCT (1, 5, 10 mg/kg) in ischemic conditions was assessed through pre- and post-treatment. Cerebral blood flow (CBF) and tests for functional recovery were assessed. Protein and gene expression were done for JNK3 and other inflammatory markers. A docking study was performed to identify the amino acid interaction. The results showed that PCT improved motor and memory function as measured by a functional recovery test believed to be due to an increase in cerebral blood flow. Also, the caspase signaling which promotes apoptosis was found to be down-regulated; however, nitric oxide synthase expression was up-regulated, which could explain the enhanced cerebral blood flow (CBF). According to our findings, PCT impeded c-Jun N-terminal kinase 3 (JNK3) signaling by suppressing phosphorylation and disrupting the mitochondrial apoptotic pathway, which resulted in the neuroprotective effect. Molecular docking analysis was performed to investigate the atomic-level interaction of JNK3 and PCT, which reveals that Met149, Leu206, and Lys93 amino acid residues are critical for the interaction of PCT and JNK3. According to our current research, JNK3 downstream signaling and the mitochondrial apoptosis pathway are both inhibited by PCT, which results in neuroprotection under conditions of global brain ischemia. Piceatannol attenuated JNK3 phosphorylation during the ischemic condition and prevented neuronal apoptosis.

Injectable, Micellar Chitosan Self‐Healing Hydrogel for Asynchronous Dual‐Drug Delivery to Treat Stroke Rats

AbstractStroke is a common disease with high mortality worldwide. The endogenous neural regeneration during the intracerebral hemorrhage (ICH) stroke is restricted by the brain cavity, inflammation, cell apoptosis, and neural scar formation. Biomaterials serving as temporary supporting matrices are highly demanded as injectable implants for brain tissue regeneration. Herein, a chitosan micellar self‐healing hydrogel (CM hydrogel) with comparable modulus (≈150 Pa) to brain, shape adaptability, and proper swelling (≈105%) is developed from phenolic chitosan (PC) and a micellar crosslinker (DPF). Two model drugs are individually packaged in the hydrophilic network and hydrophobic micelle cavities of CM hydrogel, and they feature asynchronous releasing kinetics, including a first‐order rapid release for hydrophilic drug and a zero‐order sustained release for hydrophobic drug. The dual‐drug loaded CM (CMD) hydrogel delivers two clinical drugs corresponding to the anti‐inflammatory and neurogenesis phases of the stroke to ICH rats through brain injection. The rats receiving CMD hydrogel show behavioral improvement (≈84% recovery) and balanced brain midline shift (≈0.98 left/right hemibrain ratio). Immunohistochemistry reveals neurogenesis (doublecortin‐ and nestin‐ positive cells) and evidence of angiogenesis (≈18 µm diameter vessels lined with CD31‐positive cells). The injectable CMD hydrogel offers a novel asynchronous drug delivery platform for treating ICH stroke.

Thiacloprid impairs honeybee worker learning and memory with inducing neuronal apoptosis and downregulating memory-related genes

Neonicotinoids are among the most widely used insecticides in the world and are recognized as a potential cause of pollinator decline. Previous studies have demonstrated that the neonicotinoid thiacloprid has adverse effects on foraging and memory behaviors. However, there is no direct evidence linking thiacloprid-induced neuronal cell damage in the brains of honeybees to learning and memory dysfunction. Adult honeybee (Apis mellifera L.) workers were chronically exposed to sub-lethal concentrations of thiacloprid. We discovered that thiacloprid negatively affected their survival, food consumption, and body weight. In addition, sucrose sensitivity and memory performance were impaired. We evaluated the apoptosis of honeybee brain cells using TUNEL (Terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP-biotin nick-end labeling) and Caspase-3 assays, which revealed that thiacloprid increases the dose-dependent apoptosis of neurons in the mushroom bodies (MB) and antennal lobes (AL). We also determined the abnormal transcripts of multiple genes, including vitellogenin (Vg), immune system genes (apidaecin and catalase), and memory-associated genes (pka, creb, Nmdar1, Dop2, Oa1, Oa-2R, and Oa-3R). These results indicate that exposure to sublethal concentrations of thiacloprid cause abnormal expression of memory-related genes and apoptosis of brain cells in the AL and MB, which may contribute to the memory disorder induced by thiacloprid exposure.

Qilong capsule alleviates ponatinib-induced ischemic stroke in a zebrafish model by regulating coagulation, inflammation and apoptosis

Ethopharmacological relevanceQilong capsule (QLC) is a compound traditional Chinese medicine commonly used to treat ischemic stroke (IS). QLC is made of eight kinds of medicinal materials. It has two kinds of monarch medicine and six kinds of minister medicine. However, the pharmacodynamic mechanism of QLC is still unknown. Aim of the studyThe aim of this paper was to evaluate the pharmacology mechanism of QLC against ischemic stroke through coagulation, inflammation and apoptosis. Materials and methodsWe used an existing zebrafish model to explore the protective mechanism of QLC on IS. We treated normally-developing zebrafish larvae with QLC and ponatinib 2 days post fertilization (dpf), and used the area of cerebral vascular thrombosis, red blood cell staining intensity, and brain cell apoptosis to quantitate QLC efficacy against IS. Evaluation of brain inflammation in zebrafish by observing macrophage aggregation and migration. In addition, we also explored the effect of QLC on zebrafish angiogenesis. Quantitative polymerase chain reaction (qPCR) was used to detect changes in the expression of genes involved in coagulation, inflammation, vascular endothelium, and apoptosis. ResultsWe found that QLC reduced the area affected by ponatinib-induced cerebral vascular embolism, erythrocyte staining intensity, and the number of apoptotic brain cells. For inflammation, QLC can improve the aggregation and migration of macrophages. QLC can significantly promote the formation of blood vessels in zebrafish. qPCR showed that QLC inhibited the expression of genes related to coagulation, inflammation, and apoptosis. ConclusionQilong capsule had a significant protective efficacy in ponatinib-induced IS.

The effects of extremely low-frequency magnetic field exposure on apoptosis, neurodegeneration and trace element levels in the rat brain

Aim: The aim of this study was to investigate the effects of 1mT, 1.5 mT, and 2 mT extremely low-frequency magnetic fields, which were within the limits for public environmental and occupational magnetic field exposure guidelines, on apoptosis, neurodegeneration and trace elements in rat brain cells. Material and Method: A total of 35 adult male Wistar rats were allocated into four main groups: Group 1 (n=8) was healthy controls; Group 2 (n=9) was exposed to 1 mT extremely low-frequency magnetic field; Group 3 (n=9) was exposed to 1.5 mT extremely low-frequency magnetic field and Group 4 (n=9) was exposed to 2 mT extremely low-frequency magnetic field. All the rats in the exposure groups were exposed to 50 Hz extremely low-frequency magnetic field for 4 hours per day, 5 days per week for 30 days in the Helmholtz coils. After the exposure, rats were sacrificed and rat brains were evaluated for histopathological and immunohistochemical changes as well as about the trace element levels in the brain. Results: Different levels of exposure to extremely low-frequency magnetic field doses caused increases in Ca levels and increased apoptosis in the rat brain. As the applied extremely low-frequency magnetic field levels increased, so did the apoptosis and Ca levels in the brain tissues. Conclusion: Extremely low-frequency magnetic field exposure caused neurodegeneration in rat brain tissue, increased apoptosis, and increased Ca concentration. These changes may cause various biological damage, especially cancer in healthy tissues and measures should be taken to minimize extremely low-frequency magnetic field exposure in daily life in terms of protecting public health.

Protective effects of histone deacetylase 6 specific inhibitor tubastatin A on subarachnoid hemorrhage in rats and the underlying mechanisms.

Subarachnoid hemorrhage (SAH) is a serious cerebrovascular disease. Early brain injury (EBI) and cerebral vasospasm are the main reasons for poor prognosis of SAH patients. The specific inhibitor of histone deacetylase 6 (HDAC6), tubastatin A (TubA), has been proved to have a definite neuroprotective effect on a variety of animal models of acute and chronic central nervous system diseases. However, the neuroprotective effect of TubA on SAH remains unclear. This study aims to investigate the expression and localization of HDAC6 in the early stage of SAH, and to evaluate the protective effects of TubA on EBI and cerebral vasospasm after SAH and the underlying mechanisms. Adult male SD rats were treated with modified internal carotid artery puncture to establish SAH model. In the first part of the experiment, rats were randomly divided into 6 groups: a sham group, a SAH-3 h group, a SAH-6 h group, a SAH-12 h group, a SAH-24 h group, and a SAH-48 h group. At 3, 6, 12, and 24 h after SAH modeling, the injured cerebral cortex of rats in each group was taken for Western blotting to detect the expression of HDAC6. In addition, the distribution of HDAC6 in the cerebral cortex of the injured side was measured by immunofluorescence double staining in SAH-24 h group rats. In the second part, rats were randomly divided into 4 groups: a sham group, a SAH group, a SAH+TubAL group (giving 25 mg/kg TubA), and a SAH+TubAH group (giving 40 mg/kg TubA). At 24 h after modeling, the injured cerebral cortex tissue was taken for Western blotting to detect the expression levels of HDAC6, endothelial nitric oxide synthase (eNOS), and inducible nitric oxide synthase (iNOS), terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining to detect apoptosis, and hematoxylin and eosin (HE) staining to detect the diameter of middle cerebral artery. The protein expression of HDAC6 began to increase at 6 h after SAH (P<0.05), peaked at 24 h (P<0.001), and decreased at 48 h, but there was still a difference compared with the sham group (P<0.05). HDAC6 is mainly expressed in the cytoplasm of the neurons. Compared with the sham group, the neurological score was decreased significantly and brain water content was increased significantly in the SAH group (both P<0.01). Compared with the SAH group, the neurological score was increased significantly and brain water content was decreased significantly in the SAH+TubAH group (both P<0.05), while the improvement of the above indexes was not significant in the SAH+TubAL group (both P>0.05). Compared with the sham group, the expression of eNOS was significantly decreased (P<0.01) and the expressions of iNOS and HDAC6 were significantly increased (P<0.05 and P<0.01, respectively) in the SAH group. Compared with the SAH group, the expression of eNOS was significantly increased, and iNOS and HDAC6 were significantly decreased in the SAH+TubA group (all P<0.05). Compared with the SAH group, the number of TUNEL positive cells was significantly decreased and the diameter of middle cerebral artery was significantly increased in the SAH+TubA group (both P<0.05) . HDAC6 is mainly expressed in neurons and is up-regulated in the cerebral cortex at the early stage of SAH. TubA has protective effects on EBI and cerebral vasospasm in SAH rats by reducing brain edema and cell apoptosis in the early stage of SAH. In addition, its effect of reducing cerebral vasospasm may be related to regulating the expression of eNOS and iNOS.

MELLT3 protects against cerebral ischemia-reperfusion (I/R) injury through up-regulation of m6A modification

Ischemic cerebrovascular disease is a leading cause of death globally and is often exacerbated by cerebral ischemic/reperfusion injury (CIRI). The exact mechanisms underlying I/R injury are unclear. In this study, we aimed to determine the role of m6A-modified methylase complex methyltransferase-like 3 (METTL3) in cerebral ischemia-reperfusion (I/R) injury. We found that m6A and METTL3 levels increased in OGD/RX-induced mouse astrocytes-cerebellar (MA-C) and the brain of middle cerebral artery occlusion (MCAO) model mice. METTL3 siRNA treatment reduced OGD-RX-induced MAC cell viability and proliferation, which increased with METTL3 over-expression. Flow cytometry analysis showed that silencing METTL3 significantly enhanced OGD/RX-induced MAC apoptosis, which was significantly reduced with METTL3 up-regulation. In an MCAO model, METTL3 overexpression significantly reduced cerebral infarction area and decreased brain cell apoptosis, indicating that METTL3 OE treatment could ameliorate brain edema and injury. Thus, METTL3 could be used as a target to treat I/R injury.

Salvianolic acids for injection alleviates cerebral ischemia-induced neurodegeneration by inhibiting endoplasmic reticulum stress and neuroinflammation

BackgroundCerebral ischemia is one of the leading causes of neurological deficit, dementia, and disability globally. Salvianolic acids for injection (SAFI), a multi-component drug derived from Salvia miltiorrhiza L., has been clinically used in China to treat ischemic stroke. PurposeThis study was designed to investigate the mechanisms of SAFI in alleviating cerebral ischemia-induced neurodegeneration, especially its regulatory effects on excessive ER stress. Basic proceduresThe rat acute cerebral ischemic injury model was established by a 1.5 h occlusion of the middle cerebral artery followed by 24 h of reperfusion. The neurological deficits, neurobehavior, cerebral infarct volume and edema of rats were examined. Hematoxylin and eosin staining, transmission electron microscopy analysis and TUNEL staining were performed to study the brain injury and cell apoptosis. Western blot and immunofluorescence assay were carried out to study the effects and mechanism of SAFI on ischemic injury. Main findingsResults showed that SAFI significantly inhibited the acute neurodegeneration, reduced brain infarct volumes and decreased brain water content of rats. SAFI also downregulated the relative protein levels of Bax/Bcl-2, Bim, Caspase-12, and reduced the number of TUNEL-positive cells in brain tissue, indicating the anti-apoptotic effects of SAFI. Both of immunofluorescence assay and western blot results showed that SAFI downregulated the expression of endoplasmic reticulum (ER) stress maker GRP78. SAFI dramatically inhibited the ER stress-related PERK/eIF2α/ATF4/CHOP and IRE1α/TRAF2/ASK1/JNK signaling pathways. SAFI also alleviated NF-κB triggered inflammatory response, and inhibited the production of IL-6 and IL-β. ConclusionsOur studies indicated that inhibition of excessive ER stress contributes to the protective effects of SAFI against cerebral ischemia-induced neurodegeneration and neuroinflammation.

Sub-chronic toxicity of broflanilide on the nervous system of zebrafish (Danio rerio)

ABSTRACT The new insecticide broflanilide, which possesses high insecticidal activity against agricultural pests, acts on the γ-aminobutyric acid receptor in the insect nervous system. At present, few studies assessed its effect on the nervous system of zebrafish, especially its sub-chronic toxicity on the zebrafish brain. In this study, the sub-chronic toxicity of broflanilide on the zebrafish brain was assessed. After 21 days of exposure at 36.3 μg/L, broflanilide caused oxidative damage to the zebrafish brain, increased the levels of reactive oxygen species and malondialdehyde, and inhibited the activities of superoxide dismutase, catalase, and glutathione peroxidase. Overall, these changes result in brain cell apoptosis, and inhibited growth and development. Moreover, broflanilide affects the release of acetylcholinesterase, γ-aminobutyric acid, 5-hydroxy tryptamine, and dopamine as well as the expression of related genes in the brain, leading to abnormal zebrafish behaviour. These results corroborate the sub-chronic toxicity of broflanilide on zebrafish brain, which helps the further understanding of the potential environmental risk of broflanilide.

Role of caspase-8 in stem cell transplantation alleviates rat cerebral cortex apoptosis after cardiopulmonary resuscitation

To explore the effects and the possible mechanism of bone marrow mesenchymal stem cell (BMMSC) transplantation on apoptosis in rats cerebral cortex after cardiac arrest/cardiopulmonary resuscitation (CA/CPR). The BMMSC of 2 Sprague-Dawley (SD) rats aged 4-5weeks was extracted, and the 3rd passage was used in experimental study. Eighteen Sprague-Dawley (SD) rats were divided into sham group, model group (CA/CPR group) and intervention group (BMMSC group) according to random number table method, with 6 rats in each group. CPR was performed 6 minutes after asphyxia induced CA. In sham group, CA was not induced except performing general surgical procedure. At 1 hour after return of spontaneous circulation (ROSC), 0.5 mL phosphate buffered saline (PBS) was injected through tail vein in CA/CPR group. 2×109/L green fluorescence protein (GFP)-labeled BMMSC was injected through tail vein 1 hour after ROSC in BMMSC group. Neurological deficit score (NDS) were assessed in every group at 72 hours after CPR. Serum S100 calcium binding protein B (S100B) levels were assayed by enzyme linked immunosorbent assay (ELISA). Distribution of BMMSC in brain was observed under a fluorescent microscope. Apoptosis rate in cerebral cortex was assayed by TdT-mediated dUTP nick-end labeling (TUNEL). Western blotting was performed to measure the expression levels of active aspartic acid specific cysteine proteinase (caspase-8 and caspase-9) in cerebral cortex. At 3 days after CPR, compared with sham group, the apoptosis of cerebral cortex cells was increased and brain damage was obvious, NDS score was decreased significantly (56.6±5.5 vs. 80.0±0.0, P < 0.05), and serum S100B was increased markedly (ng/L: 45.1±4.7 vs. 19.1±1.4, P < 0.05), apoptosis rate of cerebral cortex cells increased significantly [(52.9±11.8)% vs. (10.1±1.5)%, P < 0.05], the level of active caspase-8 expression in cerebral cortex was significantly higher (caspase-8/GAPDH: 0.689±0.047 vs. 0.330±0.108, P < 0.05), and there was no significant difference in active caspase-9 protein expression (caspase-9/GAPDH: 0.428±0.014 vs. 0.426±0.021, P > 0.05) in CA/CPR group. After BMMSC transplantation, GFP-labeled BMMSC were primarily detected in cerebral cortex, compared with CA/CPR group, the apoptosis of cerebral cortex cells and brain injury were significantly improved in BMMSC group, NDS score increased significantly (70.6±2.1 vs. 56.6±5.5, P < 0.05), serum S100B levels in BMMSC group were lower (ng/L: 32.0±3.2 vs. 45.1±4.7, P < 0.05), apoptosis rate of cerebral cortex cells decreased significantly [(31.1±3.4)% vs. (52.9±11.8)%, P < 0.05], and the active caspase-8 expression in cerebral cortex in BMMSC group was significantly decreased (caspase-8/GAPDH: 0.427±0.067 vs. 0.689±0.047, P < 0.05). The active caspase-9 expression in cerebral cortex in BMMSC group and CA/CPR group were not significantly different (caspase-9/GAPDH: 0.431±0.022 vs. 0.428±0.014, P > 0.05). BMMSC transplantation can alleviate rat brain damage after CA/CPR possibly by inhibiting the death receptor mediated apoptotic pathway to inhibit the apoptosis of brain cells.

Oral GSH Exerts a Therapeutic Effect on Experimental Salmonella Meningitis by Protecting BBB Integrity and Inhibiting Salmonella-induced Apoptosis.

Bacterial meningitis (BM) is the main cause of the central nervous system (CNS) infection and continues to be an important cause of mortality and morbidity. Glutathione (GSH), an endogenous tripeptide antioxidant, has been proved to exert crucial role in reducing superoxide radicals, hydroxyl radicals and peroxynitrites. The purpose of this study is to expand the application scope of GSH via exploring its therapeutic effect on BM caused by Salmonella typhimurium SL1344 and then provide a novel approach for the treatment of BM. The results suggested that intragastric administration of GSH could significantly increase median survival and improve experimental autoimmune encephalomyelitis score of BM model mice. However, exogenous GSH did not affect the adhesion, invasion and cytotoxicity of SL1344 to C6, BV2 and primary microglia. Due to the contradiction between the therapeutic and bactericidal effects of GSH, the effect of GSH on blood-brain barrier (BBB) was investigated to explore its action target for the treatment of meningitis. GSH was found to repair the damage of BBB and then prevent the leakage of SL1344 from the brain to the blood circulation. The repaired BBB could also effectively reduce the entry of macrophages and neutrophils into the brain, and significantly reverse the microglia activation induced by SL1344. More importantly, exogenous GSH was proved to reduce mouse brain cell apoptosis by inhibiting the activation of caspase-8 followed by caspase-3, and reversing the up-regulation of ICAD and PARP-1 caused by SL1344.

Metabolomic analysis and pharmacological validation of the cerebral protective effect of 3,4‑dihydroxybenzaldehyde on cerebral ischemia‑reperfusion injury.

3,4‑Dihydroxybenzaldehyde (DBD), one of the active components of Gastrodia elata, has a cerebral protective effect and can effectively combat cerebral ischemia/reperfusion (I/R) injury in rats. However, the metabolite profiles and underlying mechanisms associated with DBD remain unclear. To explore the level of energy metabolism and pharmacological targets in brain tissue following DBD treatment of stroke. The right middle cerebral artery of the rats was occluded for 2h and reperfused for 24h to simulate brain I/R injury. Pharmacological results showed that DBD reduced cerebral infarct volume, improved neurological function and increased adenosine triphosphate (ATP) content. Mitochondria are the primary sites for ATP generation and cellular energy supply and decreasing mitochondrial dysfunction can alleviate the energy expenditure of ischemic stroke. Through further experiments, it was found that mitochondrial damage was recovered following DBD treatment, which was manifested by the improvement of mitochondrial morphology under an electron microscope and the reduction of oxidative stress damage. The metabolomics of middle cerebral artery occlusion/reperfusion (MCAO/R) rat brain tissue was studied by the liquid chromatography‑tandem mass spectrometry metabolomics method. Significantly different metabolites were screened and the pathways involved included amino sugar and nucleotide sugar metabolism and pentose phosphate pathway. Finally, the present study performed targeted metabolic profiling and validated potential therapeutic targets. Uridine diphosphate N‑acetylglucosamine (UDP‑GlcNAc) levels were decreased in the MCAO/R group but significantly increased in the DBD group. TdT‑mediated dUTP nick end labeling (TUNEL) staining, hematoxylin and eosin staining and western blotting showed that brain cell apoptosis was inhibited and neuronal morphology improved. Among them, the regulatory enzyme O‑GlcNAc transferase (OGT) of UDP‑GlcNAc appeared to be significantly increased and neuronal apoptosis was inhibited following DBD treatment, which was verified by western blotting. Therefore, by analyzing mitochondrial dysfunction following I/R and the characterization of potential markers in mitochondrial energy metabolism, it was shown that OGT could inhibit neuronal apoptosis as a potential therapeutic target for brain I/R injury.

Proteasome activator 28A: A clinical biomarker and pharmaceutical target in acute cerebral infarction therapy

Purpose: To determine the dynamic changes in serum levels of PA28α in patients with acute cerebral infarction (ACI), and to investigate its correlation with infarct size and neurological deficit of the disease. &#x0D; Methods: A total of 100 ACI patients and 100 healthy volunteers were recruited from The First Affiliated Hospital of Xinxiang Medical University as case and control groups, respectively. Their serum levels of PA28α were determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The potential of PA28α in predicting the incidence of ACI was assessed by plotting ROC curves. Multivariate logistic regression analysis was conducted to investigate the risk factors of ACI. In addition, an ACI model in rats was established, and ACI rats were classified into 1, 3, 5, 7 and 14 day subgroups based on the duration post-ACI. Rats in the sham group served as control. &#x0D; Results: Serum level of PA28α was significantly higher in ACI patients than in controls. Moreover, the serum level of PA28α at admission was positively correlated to the NIHSS score and infarct volume of ACI patients. The level of PA28α in ACI rats gradually increased post-ACI, reaching a peak on day 7. The number of apoptotic brain cells in ACI rats gradually decreased after ACI. In addition, PA28α level was negatively correlated to the number of apoptotic brain cells in ACI rats (R2 = 0.5148, p &lt; 0.001). &#x0D; Conclusion: The serum level of PA28α is elevated in ACI patients, and is positively correlated to infarct volume and neurological deficit of the disease. The dynamic change in brain cell apoptosis post-ACI is negatively correlated to the serum level of PA28α. These findings may provide theoretical basis for the diagnosis and treatment of ACI.

Effects of icariin on alleviating schizophrenia-like symptoms by regulating the miR-144-3p/ATP1B2/mTOR signalling pathway

Schizophrenia is a group of severe mental disorders. Icariin is a main active component of Epimedium, possessing therapeutic effects on various neurodegenerative diseases. The present study investigated whether icariin is effective in alleviating schizophrenia-like symptoms and explored its underlying molecular mechanism. A developmental schizophrenia rat model employing 2-week repeated MK-801 administration was established. Icariin was orally administrated 3 time per day for 2 weeks after the MK-801 administration. Open-field test (OFT), novel object recognition (NOR), rotarod, and Morris water maze (MWM) were performed to examine the therapeutic effects of icariin on behavioural abnormalities. Hematoxylin-eosin (HE) staining on hippocampus slices, and MTT assay and Calcein/PI staining on the SK-N-SH cells treated with MK-801 were carried out to assess the neuroprotective effects of icariin. Furthermore, the regulation of icariin on the miR-144-3p/ATP1B2/mTOR signalling pathway was examined by RT-PCR and Western blots. The results showed that icariin alleviated MK-801-induced anxiety and recognition memory deficits in the OFT and NOR, respectively. Additionally, weakened motor coordination caused by MK-801 was restored by icariin. The MWM test also showed that icariin can improve MK-801-induced impaired spatial memory and swimming ability. Furthermore, brain grey matter atrophy, cytotoxicity, and cell apoptosis caused by MK-801 can be eliminated by icariin. Lastly, icariin can regulate the expression of miR-144-3p and ATP1B2, and enhance the phosphorylation of PI3K, Akt, and mTOR. In conclusion, this study revealed that icariin may have therapeutic effects on schizophrenia-like disorders via regulating the miR-144-3p/ATP1B2/mTOR signalling, suggesting that icariin has potential to become an antipsychotic drug.

Steroid Receptor Coactivator-Interacting Protein (SIP) Alleviates Ischemic Cerebral Infarction Damage Through the NF-κB Pathway.

Ischemic stroke leads to high mortality and disability rates in humans. Cerebral ischemic injury has a severe complex pathophysiological mechanism. The abnormal release of inflammatory cytokines will cause brain tissue damage and destroy the blood-brain barrier integrity, which aggravates the process of brain injury. Therefore, attenuating the level of inflammatory response is critical for the therapy of cerebral ischemia injury. This study examined the rule of SIP treatment to support neuron protective effect after cerebral injury in an animal model of middle cerebral artery occlusion (MCAO). After ischemia/reperfusion, neurological function, neuroglia cells activation, infarction volume, brain water content, brain tissue apoptosis ratio, and inflammatory response were assessed, and quantitative PCR and western blot were also detected, respectively. Treatment of SIP ameliorated neurological dysfunction, brain infarction, brain edema, and brain cell apoptosis after MCAO operation. Overexpression SIP also suppressed pro-inflammatory cytokines release. Furthermore, the protective effect of SIP on brain injury occurs through reduced neuroglia cells activation through downregulation of the NF-κB pathway. In summary, the present work indicated that SIP prevents ischemic cerebral infarction-induced inflammation and apoptosis by blocking inflammasome activation via NF-κB signaling pathway. Those results suggest that SIP treatment is an attractive strategy for prevention of ischemic cerebral infarction.

Dihydromyricetin Attenuates Cerebral Ischemia Reperfusion Injury by Inhibiting SPHK1/mTOR Signaling and Targeting Ferroptosis.

BackgroundDihydromyricetin (DHM) exerts protective effects in various brain diseases. The aim of this research was to investigate the biological role of DHM in cerebral ischemia reperfusion (I/R) injury.MethodsWe generated a rat model of cerebral I/R injury by performing middle cerebral artery occlusion/reperfusion (MCAO/R). The neurological score and brain water content of the experimental rats was then evaluated. The infarct volume and extent of apoptosis in brain tissues was then assessed by 2,3,5-triphenyltetrazolium (TTC) and TdT-mediated dUTP nick end labeling (TUNEL) staining. Hippocampal neuronal cells (HT22) were subjected to oxygen-glucose deprivation/reperfusion (OGD/R) and cell counting kit-8 (CCK-8) assays and flow cytometry were performed to detect cell viability and apoptosis. The levels of lipid reactive oxygen species (ROS) and iron were detected and the expression levels of key proteins were assessed by Western blotting.ResultsDHM obviously reduced neurological deficits, brain water content, infarct volume and cell apoptosis in the brain tissues of MCAO/R rats. DHM repressed ferroptosis and inhibited the sphingosine kinase 1 (SPHK1)/mammalian target of rapamycin (mTOR) pathway in MCAO/R rats. In addition, DHM promoted cell viability and repressed apoptosis in OGD/R-treated HT22 cells. DHM also suppressed the levels of lipid ROS and intracellular iron in OGD/R-treated HT22 cells. The expression levels of glutathione peroxidase 4 (GPX4) was enhanced while the levels of acyl-CoA synthetase long-chain family member 4 (ACSL4) and phosphatidylethanolamine binding protein 1 (PEBP1) were reduced in OGD/R-treated HT22 cells in the presence of DHM. Moreover, the influence conferred by DHM was abrogated by the overexpression of SPHK1 or treatment with MHY1485 (an activator of mTOR).ConclusionThis research demonstrated that DHM repressed ferroptosis by inhibiting the SPHK1/mTOR signaling pathway, thereby alleviating cerebral I/R injury. Our findings suggest that DHM may be a candidate drug for cerebral I/R injury treatment.

Effect of electroacupuncture on behavior and cell apoptosis of brain tissue in traumatic brain injury rats

To observe the effect of electroacupuncture (EA) on behavior, pathomorphology and brain cell apoptosis in traumatic brain injury (TBI) rats, so as to explore its mechanisms underlying treatment of TBI. Male SD rats were randomly divided into control (n=10), sham (n=30), model (n=30) and EA (n=30) groups, the latter three groups were further divided into 3, 7, 14 d subgroups （n=10）. TBI model was established by using a free fall brain injury striking device after exposing the local cranial bone (to induce the left parietal cerebral contusion). Twenty-four hours after modeling, EA (2 Hz, 1 mA) was applied at "Quchi"(LI11), "Neiguan"(PC6), "Zusanli"(ST36) and "Yongquan" (KI1) combined with acupuncture at "Shuigou"(GV26) and "Baihui"(GV20) for 15 min, once a day for 14 consecutive days. After 3, 7, and 14 days treatment, the behavioral function (balance, walking, nerve, and limb retraction) of the rats was evaluated. The histopathological changes of the injured brain tissue were observed by HE staining. TUNEL method was used to detect the apoptosis of cells in the brain injury area. After modeling, the scores of balance and walking in the model group were higher than those in the sham group (P<0.01, P<0.05), the neurological function score and the right limb retraction force were lower than those in the sham group(P<0.01). After 3 days treatment, the neurological function score and the right limb retraction force in the EA group were higher than those in the model group (P<0.05). After 7 and 14 days treatment, the scores of balance and walking function in the EA group were lower than those in the model group (P<0.05, P<0.01), while the scores of nerve function and right limb retraction force were higher (P<0.05, P<0.01). HE staining showed that modeling induced pathological changes such as the inflammatory cell infiltration, interstitial edema, necrosis, nuclear pyknosis and nuclear lysis were relatively milder in the EA group on the 7th and 14th day. Compared with the sham group, the cell apoptosis of brain injury area was higher in the model group (P<0.01); while it was lower on day 7 and 14 in the EA group in comparison with the model group (P<0.01, P<0.05). EA can improve the behavioral function, reduce the apoptosis of brain cells in the injured area, and promote the rehabilitation of craniocerebral injury.

Hyperglycemia aggravates ischemic brain damage via ERK1/2 activated cell autophagy and mitochondrial fission.

BackgroundHyperglycemia is one of the major risk factors for stroke and stroke recurrence, leading to aggravated neuronal damage after cerebral ischemia/reperfusion (I/R). ERK1/2 signaling pathway plays a vital role in cerebral ischemic injury. However, the role of the ERK1/2 pathway in hyperglycemia-aggravated ischemic brain damage is not clear.MethodsStreptozotocin (STZ; 50 mg/kg)-induced diabetes (blood glucose ≥12 mmol/L) or control groups in adult Sprague-Dawley rats were further subdivided into I/R (carotid artery/vein clamping), I/R + PD98059 (I/R plus ERK1/2 inhibitor), and Sham-operated groups (n = 10 each). Neurobehavioral status (Neurological behavior scores) and the volume of the cerebral infarction (TTC staining); brain mitochondrial potential (JCI ratio test) and cell apoptosis (TUNEL assay); RAS protein expression, phosphorylated/total ERK1/2 and Drp-1 (Dynamic-related protein 1) protein levels (Western blotting); mitochondrial fusion-related proteins mitofusin-1/2 (Mfn1/2), optic atrophy (OPA-1) and mitochondrial fission 1 (Fis1), and autophagy-associated proteins Beclin-1, LC3-I/II and P62 (Western blotting and immunohistochemistry) were analyzed.ResultsThe I/R + PD98059 group demonstrated better neurobehavior on the 1st (p < 0.05) and the 3rd day (p < 0.01) than the I/R group. Compared to the Sham group, cerebral ischemia/reperfusion brought about neuronal damage in the I/R group (p <0.01). However, treatment with PD98059 showed an improved situation with faster recovery of mitochondrial potential and less apoptosis of neuronal cells in the I/R + PD98059 group (p < 0.01). The I/R group had a higher-level expression of RAS and phosphorylated ERK1/2 and Drp-1 than the diabetes mellitus (DM) group (p < 0.01). The PD98059 treated group showed decreased expression of p-ERK1/2, p-Drp-1, Fis1, and Beclin-1, LC3-I/II and P62, but increased Mfn1/2 and OPA-1 than the I/R group (p < 0.01).ConclusionHyperglycemia worsens cerebral ischemia/reperfusion-induced neuronal damage via ERK1/2 activated cell autophagy and mitochondrial fission.