Cerebral Ischemia Research Articles

VEGFD/VEGFR3 signaling contributes to the dysfunction of the astrocyte IL-3/microglia IL-3Rα cross-talk and drives neuroinflammation in mouse ischemic stroke.

Astrocyte-derived IL-3 activates the corresponding receptor IL-3Rα in microglia. This cross-talk between astrocytes and microglia ameliorates the pathology of Alzheimer's disease in mice. In this study we investigated the role of IL-3/IL-3Rα cross-talk and its regulatory mechanisms in ischemic stroke. Ischemic stroke was induced in mice by intraluminal occlusion of the right middle cerebral artery (MCA) for 60 min followed by reperfusion (I/R). Human astrocytes or microglia subjected to oxygen-glucose deprivation and reoxygenation (OGD/Re) were used as in vitro models of brain ischemia. We showed that both I/R and OGD/Re significantly induced decreases in astrocytic IL-3 and microglial IL-3Rα protein levels, accompanied by pro-inflammatory activation of A1-type astrocytes and M1-type microglia. Importantly, astrocyte-derived VEGFD acting on VEGFR3 of astrocytes and microglia contributed to the cross-talk dysfunction and pro-inflammatory activation of the two glial cells, thereby mediating neuronal cell damage. By using metabolomics and multiple biochemical approaches, we demonstrated that IL-3 supplementation to microglia reversed OGD/Re-induced lipid metabolic reprogramming evidenced by upregulated expression of CPT1A, a rate-limiting enzyme for the mitochondrial β-oxidation, and increased levels of glycerophospholipids, the major components of cellular membranes, causing reduced accumulation of lipid droplets, thus reduced pro-inflammatory activation and necrosis, as well as increased phagocytosis of microglia. Notably, exogenous IL-3 and the VEGFR antagonist axitinib reestablished the cross-talk of IL-3/IL-3Rα, improving microglial lipid metabolic levels via upregulation of CPT1A, restoring microglial phagocytotic function and attenuating microglial pro-inflammatory activation, ultimately contributing to brain recovery from I/R insult. Our results demonstrate that VEGFD/VEGFR3 signaling contributes to the dysfunction of the astrocyte IL-3/microglia IL-3Rα cross-talk and drives pro-inflammatory activation, causing lipid metabolic reprogramming of microglia. These insights suggest VEGFR3 antagonism or restoring IL-3 levels as a potential therapeutic strategy for ischemic stroke.

Global research trends and prospects on immune-related therapy in ischemic stroke: a bibliometric analysis.

Following ischemic stroke, non-neuronal cells within the nervous system play a crucial role in maintaining neurovascular unit functions, regulating metabolic and inflammatory processes of the nervous system. Investigating the functions and regulation of these cells, particularly immune cells, deepens our understanding of the complex mechanisms of neuroinflammation and immune modulation after ischemic stroke and provides new perspectives and methods for immune-related therapy. The annual distribution, journals, authors, countries, institutions, and keywords of articles published between 2015 and 2024 were visualized and analyzed using CiteSpace and other bibliometric tools. A total of 1,089 relevant articles or reviews were included, demonstrating an overall upward trend; The terms "cerebral ischemia," "immune response," "brain ischemia," "cerebral inflammation," "neurovascular unit," and "immune infiltration," etc. are hot keywords in this field. In recent years, research on immune-related therapy for ischemic stroke has focused on mechanisms of occurrence, protection and repair of the blood-brain barrier (BBB) by non-neuronal cells, and regulation of immunosuppression and inflammation. Among these, reducing BBB disruption to minimize secondary brain damage has become a hotspot. At the same time, the complex roles of immune responses have attracted attention, particularly the balance between regulatory T cells and Th17 cells in regulating neuroinflammation and promoting neurological function recovery, which is crucial to reduce secondary neuronal damage and improve prognosis, potentially establishing a pivotal frontier in this domain of investigation.

Ultrasound assessment of endothelial dysfunction in Egyptian migraine patients

BackgroundIt becomes clearer that migraine is associated with vascular risks; however, preclinical vascular involvement is not sufficiently addressed. Evidences point that migraine attacks affect vascular endothelium. The aim of this study was to investigate endothelial dysfunction in migraineurs through assessment of carotid intima-media thickness (CIMT) and flow-mediated dilatation (FMD) of the brachial artery and their correlation with clinical characteristics of migraine, headache severity, and brain magnetic resonance imaging (MRI) findings.ResultsA statistically significant difference was found between migraineurs and controls where carotid IMT was significantly higher and FMD of the brachial artery was significantly lower in migraineurs compared to controls. Carotid IMT was significantly higher and FMD was significantly lower in chronic migraine compared to episodic migraine patients. Mean IMT values were significantly higher in patients receiving ergots and in patients with subcortical white matter lesions in brain MRI. Mean FMD values were significantly lower in patients receiving ergots. There was a significant negative correlation between FMD and carotid IMT, age of the patients, disease duration, duration of headache attacks, headache frequency, and migraine disability assessment questionnaire (MIDAS) score. There were significant positive correlations between carotid IMT and age of patients, disease duration, headache frequency, MIDAS score, and number of MRI white matter lesions. For diagnosing endothelial dysfunction in migraineurs, the sensitivity and specificity of IMT were 72.5 and 70%, respectively, with a cut-off value of 0.575 mm and that of FMD were 82.5 and 90%, respectively, with a cut-off value of 20.55%.ConclusionMigraine coincides with endothelial dysfunction which promotes atherogenesis and increased risk of cerebral ischemia. FMD could be used as a potential biomarker for endothelial dysfunction in migraine. The affection of IMT and FMD is more in patients receiving ergots which may influence the selection of treatment in migraineurs in the future.

Polygala tenuifolia root extract attenuates ischemic stroke by inhibiting HMGB1 trigger neuroinflammation

Polygala tenuifolia Willd., a famous traditional Chinese medicine, has been widely applied to treat central nervous system diseases. In this study, P. tenuifolia root extract exhibited a moderate anti-ischemic effect on in-vitro oxygen-glucose deprivation/reperfusion (OGD/R) model. In transient middle cerebral artery occlusion (tMCAO) rats, P. tenuifolia root extract significantly attenuated brain infarction and neurological deficits in a dose-dependent manner. Compared with the sham group, the release of damage-associated molecular patterns (DAMPs)-HMGB1 in the ischemic brain was significantly higher, which was inhibited by P. tenuifolia root extract. To further explore such neuroprotective effects whether associated with aseptic inflammation, HMGB1-activated BV2 microglial cells model was established. The extract of P. tenuifolia was found to inhibit the downstream inflammatory response driven by HMGB1, with an IC50 value of 49.46 μg/mL. In addition, the extract was also found to be able to directly interact with HMGB1 in the surface plasmon resonance (SPR) experiment. Phytochemical studies showed that the extract of P. tenuifolia root contains a large number of terpenoids, oligosaccharides and phenolic compounds, which likely contribute to the above observed biological activities. Our results not only provide some data support for the clinical application of P. tenuifolia against cerebral ischemia, but also clarify the potential target of P tenuifolia's anti-inflammatory properties.

Proposed Hypothesis of TWEAK/Fn14 Receptor Modulation in Autism Spectrum Disorder.

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder with a complex, multiple etiology that is marked by impaired social interaction, communication, and repetitive behaviour. There is presently no pharmaceutical treatment for the core symptoms of ASD, even though the prevalence of ASD is increasing worldwide. Treatment of autism spectrum disorder involves the interaction of numerous signalling pathways, such as the Wnt/beta-catenin pathway, probiotics and kynurenine pathway, PPAR pathway, PI3K-AKT-mTOR pathway, Hedgehog signaling pathway, etc. The scientific literature has revealed TWEAK/Fn14 to not be explored in the autism spectrum disorder. In vitro and in vivo, TWEAK can control a wide range of cellular responses. Recent research has revealed that TWEAK and Fn14 are expressed in the Central Nervous System (CNS) and upregulated in perivascular endothelial cells, astrocytes, neurons, and microglia in response to various stimuli, including cerebral ischemia. This upregulation is followed by cell death and an increase in Blood-brain Barrier (BBB) permeability. The study has revealed that Aurintricarboxylic Acid (ATA) acts as an agent that suppresses TWEAK/Fn14 signaling. Similarly, from the discussion, it has been emphasized that the proposed molecular TWEAK/Fn14 signalling pathway can be considered as a therapeutic approach in the management of autism spectrum disorder.

The impact of intravenous agitated saline contrast echocardiography on subclinical neuronal injury determined by neuron-specific enolase

Abstract Introduction Patent foramen ovale (PFO) is a congenital defect in interatrial septum with 25-30% prevelance. Diagnostic methods include transesophageal echocardiography (TEE), magnetic resonance imaging (MRI), and computed tomography (CT), but transthoracic echocardiography (TTE) is the primary diagnostic method. TTE involves the injection of agitated saline intravenously, and the passage of microbubbles from the right atrium to the left atrium is monitored. The reliability of this method has been questioned in various case series, with a low incidence (0.062%) of generally transient, mild clinical cerebrovascular events reported. Silent cerebral ischemia (SSI) is asymptomatic, radiographically identified acute neuronal damage. SSI is associated with an increased risk of stroke, dementia, and cognitive impairment. Neuron-specific enolase (NSE) with high specificity for neural cells can be used to detect SSI. While symptomatic patients have been evaluated in reported case series, there is no study determining the relationship and reliability of intravenous agitated saline echocardiography (SCE) with SSI. This study aims to determine whether TTE, performed on patients with PFO without neurological history and a healthy control group, can cause SSI by evaluating NSE with SCE. Materials and methods 52 patients undergoing SCE for color flow through interatrial septum or prominent interatrial septal aneurysm and are diagnosed with PFO were included. Additionally, 51 control patients without detected PFO or ASA were also included. Serum samples were taken from the patients before and 12 hours after the SCE test, and NSE levels were measured by the ELISA method. Results The mean baseline NSE level was 12.7 ng/mL with a median of 5.3 ng/mL in the PFO group, and in the control group, the mean was 9.1 ng/mL with a median of 6.4 ng/mL. At 12 hours, the mean NSE levels in the PFO group were 9.8 ng/mL with a median of 6.6 ng/mL, and in the control group, the mean was 12.7 ng/mL with a median of 5.8 ng/mL. The increase in NSE levels after SCE, was not statistically significant when comparing the PFO and control groups (p=0.77). No relationship was found between age, presence of interatrial septal aneurysm, presence of mitral annulus calcification, the number of bubbles passed in the SCE test, and the basal and 12-hour change in NSE levels (p=0.926). NSE level changes were found to be correlated between groups (p<0.001). Conclusion This study is the first to evaluate whether SCE causes subclinical neuronal damage. Our study suggests that when evaluated with NSE, the SCE method is safe and does not cause subclinical neuronal damage in patients without underlying neuronal susceptibility.results

The foam cell-derived exosomal miRNA Novel-3 drives neuroinflammation and ferroptosis during ischemic stroke.

Large artery atherosclerosis (LAA) is a prevalent cause of acute ischemic stroke (AIS). Understanding the mechanisms linking atherosclerosis to stroke is essential for developing appropriate intervention strategies. Here, we found that the exosomal miRNA Novel-3 is selectively upregulated in the plasma of patients with LAA-AIS. Notably, Novel-3 was predominantly expressed in macrophage-derived foam cells, and its expression correlated with atherosclerotic plaque vulnerability in patients undergoing carotid endarterectomy. Exploring the function of Novel-3 in a mouse model of cerebral ischemia, we found that Novel-3 exacerbated ischemic injury and targeted microglia and macrophages expressing ionized calcium-binding adapter molecule 1 in peri-infarct regions. Mechanistically, Novel-3 increased ferroptosis and neuroinflammation by interacting with striatin (STRN) and downregulating the phosphoinositide 3-kinase-AKT-mechanistic target of rapamycin signaling pathway. Blocking Novel-3 activity or overexpressing STRN provided neuroprotection under ischemic conditions. Our findings suggest that exosomal Novel-3, which is primarily derived from macrophage-derived foam cells, targets microglia and macrophages in the brain to induce neuroinflammation and could serve as a potential therapeutic target for patients with stroke who have atherosclerosis.

Cardiac autonomic changes after stroke

Abstract Introduction There is a highly complex and clinically important relationship between the cardiac and the nervous system, called the heart-brain axis [1]. Therefore, acute brain damage may lead to alterations in cardiac function due to changes in the autonomic nervous system[2,3]. Novel computerised biosignal analysis techniques and advanced biosignal assessment could provide a valuable tool to sensitively analyse cardiac autonomic function (CAF). CAF can be characterized by two complementary digital biomarkers, periodic repolariation dynamics (PRD) and deceleration capacity (DC). The aim of this study was to determine CAF by means of PRD and DC in patients with acute ischemic stroke (AIS) or transient ischemic attack (TIA) and to analyze them according to their relationship with specific cardiac (high-sensitive Troponin T (hsTnT) and N-terminal pro-B-type natriuretic peptide (NT-pro BNP)) and stroke (location, type, severity) characteristics. Methoden / Methods Between February 22, 2021, and May 5, 2023, 282 patients after an AIS or TIA were enrolled in the study. During the inpatient stay, all relevant parameters were recorded and stored in a database. To assess CAF, a 30-minute biosignal measurement was performed with a high resolution (1kHz) electrocardiogram (ECG). Cardiac autonomic biomarkers, PRD and DC, were calculated semiautomatically afterwards from patients. Results Fifteen ECGs were excluded due to poor recording quality. Consequently, data from 261 AIS and 21 TIA patients were analyzed. A total of 105 patients (37.2 %) had pathological PRD (≥ 5.75 deg2) and/or DC (≤ 2.5 ms), 78 patients (27.7 %) had pathological PRD, and 57 patients (20.2 %) had pathological DC. The median PRD was 3.13 deg2 and the median DC was 5.30 ms (see Figure). There were no significant differences in the distribution of PRD and DC between male and female patients (PRD p = 0.38, DC p = 0.30), and between AIS and TIA patients (PRD p = 0.13, DC p = 0.66). Patients with middle cerebral artery (MCA) infarction had significantly higher median PRD values (3.56 deg2 [IQR 1.51 - 6.59 ] vs. 2.81 deg2 [IQR 1.15 - 5.99], p = 0.047) and lower median DC values (4.66 ms [IQR 2.65 – 7.76] vs. 6.11 ms [IQR 3.15 – 10.61], p = 0.011) than patients with stroke located in another cerebral regions (see Figure). Patients with pathological PRD or DC had significantly higher hsTnT (13.50 ng/l [IQR 9.7 – 24.1] vs. 11.05 ng/l [ IQR 7.13 – 16.23], p < 0.001) and NT-pro BNP (269.00 ng/l [IQR 117.50 – 687.00] vs. 125.50 ng/l [IQR 57.00 – 288.25], p < 0.001) values. Conclusions The study showed that a considerable proportion, more than one third of all patients, had evidence of cardiac autonomic dysfunction after acute cerebral ischaemia. Of particular interest, PRD was higher and DC was lower in patients with MCA ischaemia, reflecting a potentially higher cardiac risk profile. Cardiac autonomic dysfunction was also associated with increased hsTnT and NT-proBNP levels.

Illustrate the metabolic regulatory mechanism of Taohong Siwu decoction in ischemic stroke by mass spectrometry imaging.

Metabolic dysregulation in the ischemic region has been increasingly recognized as a contributing factor to ischemic stroke pathogenesis. Taohong Siwu decoction (THSWD), a traditional Chinese medicine preparation used to enhance blood circulation, is frequently employed in treating ischemic stroke. However, the metabolic regulatory mechanism underlying the therapeutic effects of THSWD in ischemic stroke remains largely unexplored. In this study, we employed desorption electrospray ionization mass spectrometry imaging (DESI-MSI) to investigate the metabolic changes in the brain tissue of ischemic stroke rat model. Our investigation revealed that 30 metabolites exhibited significant dysregulation in the ischemic brain regions, specifically the cortex and striatum, following ischemic injury. Following the treatment of THSWD, almost all the dysregulated metabolites got different degrees of callback. Further pathway analysis indicated that THSWD might exert its therapeutic effects by restoring energy metabolism, improving neurotransmitter metabolism, recovering polyamine metabolism, and so on. DESI-MSI offers a favorable methodology for investigating the alterations in the spatial distribution and level within the ischemic brain region following treatment with THSWD in ischemic stroke. These findings provide a novel perspective on the underlying mechanisms of the efficacy of THSWD in ischemic stroke treatment.

Glial 'omics in ischemia: Acute stroke and chronic cerebral small vessel disease.

Vascular injury and pathologies underlie common diseases including ischemic stroke and cerebral small vessel disease (CSVD). Prior work has identified a key role for glial cells, including microglia, in the multifaceted and temporally evolving neuroimmune response to both stroke and CSVD. Transcriptional profiling has led to important advances including identification of distinct gene expression signatures in ischemia-exposed, flow cytometrically sorted microglia and more recently single cell RNA sequencing-identified microglial subpopulations or clusters. There is a reassuring degree of overlap in the results from these two distinct methodologies with both identifying a proliferative and a separate type I interferon responsive microglial element. Similar patterns were later seen using multimodal and spatial transcriptomal profiling in ischemia-exposed microglia and astrocytes. Methodological advances including enrichment of specific neuroanatomic/functional regions (such as the neurovascular unit) prior to single cell RNA sequencing has led to identification of novel cellular subtypes and generation of new credible hypotheses as to cellular function based on the enhanced cell sub-type specific gene expression patterns. A ribosomal tagging strategy focusing on the cellular translatome analyses carried out in the acute phases post stroke has revealed distinct inflammation-regulating roles for microglia and astrocytes in this setting. Early spatial transcriptomics experiments using cerebral ischemia models have identified regionally distinct microglial cell clusters in ischemic core versus penumbra. There is great potential for combination of these methods for multi-omics approaches to further elucidate glial responses in the context of both acute ischemic stroke and chronic CSVD.

Dual-tDCS Ameliorates Cerebral Injury and Promotes Motor Function Recovery via cGAS-STING Signaling Pathway in a Rat Model of Ischemic Stroke.

Ischemic stroke is one of the leading causes of death and disability. Dual transcranial direct current stimulation (dual-tDCS) is a promising intervention to treat ischemic stroke, but its efficacy and underlying mechanism remain to be verified. Cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway has recently emerged as a key mediator in cerebral injury. However, little is known about the effect of cGAS-STING on neuronal damage in ischemic stroke, and it remains to be studied whether the cGAS-STING pathway is involved in tDCS intervention for ischemic stroke. Therefore, we aimed to investigate whether dual-tDCS can alleviate ischemic brain injury in a rat model of ischemic stroke and if so, whether via cGAS-STING pathway. Middle cerebral artery occlusion (MCAO) was employed to induce a rat model of ischemic stroke. Male SD rats weighing 250-280g were randomly assigned to the Sham, MCAO, Dual-tDCS, Dual-tDCS + RU.521, and Dual-tDCS + 2'3'-cGAMP groups, with 10 rats in each group completing the experiment. Behavioral, morphological, MRI, and molecular biological methods were performed. We found that the cGAS-STING pathway was activated and expressed in neurons after MCAO. Dual-tDCS improved motor function and infarct volume, inhibited neuronal apoptosis, promoted the expression of neurotrophins (BDNF and NGF), CD31, and VEGF, and suppressed inflammation reaction after MCAO via the cGAS-STING pathway. Taken together, dual-tDCS may improve MCAO-induced brain injury and promote the recovery of motor function, resulting from the inhibition of neuronal apoptosis and inflammation reaction, as well as promotion of the expression of nerve plasticity- and angiogenesis-related proteins, via cGAS-STING pathway.

Effect of electroacupuncture at "Baihui" (GV20) and "Zusanli" (ST36) on angiogenesis in the brain of middle cerebral artery occlusion rats based on HIF/VEGF/Notch signaling pathway

To observe the effect of electroacupuncture (EA) on hypoxia-inducible factor (HIF)/vascular endothelial growth factor (VEGF)/Notch signaling pathway and related factors in rats with cerebral ischemia (CI), so as to explore its regulatory mechanism underlying improvement of CI by promoting cerebral microangiogenesis. Fifteen male SD rats were randomly selected as the sham surgery (sham) group, while other 85 rats were used to prepare CI model according to the modified Zea-Longa method. The successful CI model rats (n=60) were randomly allocated to model, EA, EA+inhibitor, and inhibitor groups, with 15 rats in each group. EA stimulation (1 Hz/20 Hz, 1-2 mA) was applied to "Baihui"(GV20) and right "Zusanli"(ST36) for 30 min, once a day for 7 days. Rats of the 2 inhibitor groups received intraperitoneal injection of YC-1 (HIF-1α inhibitor, 2.5 mg/kg), once a day for 7 days. The modified neurological severity scale (mNSS, including the motor ［muscular state, abnormal movement］, sensory ［visual, tactile and proprioceptive］, balance and reflex functions, 0-18 points) was used to assess the rats' neurological deficit state. The percentage of cerebral infarct volume was measured after 2, 3, 5-triphenyl tetrazolium chloride (TTC) staining, and pathological changes of the ischemic penumbra cortex were observed after H.E. staining. The immunoactivity of CD34 was determined using immunohistochemistry, followed by calculating the microvascular density (MVD) in the ischemic penumbra cortex, and the expression levels of HIF-1α, VEGF, vascular endothelial growth factor receptor 2 (VEGFR2), Notch1, and Delta like 4 ligand (DLL4) mRNAs and proteins in the ischemic penumbra of brain tissue were detected using real-time PCR and Western blot, respectively. In contrast to the sham group, the mNSS score, percentage of cerebral infarct volume, MVD, and expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 proteins and mRNAs were all significantly increased in the model group (P<0.01). In comparison with the model group, the mNSS score and percentage of cerebral infarct volume were significantly decreased in the EA group (P<0.01), and increased in the inhibitor group (P<0.01), and the MVD, and expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 proteins and mRNAs were further strikingly increased in the EA group (P<0.01), but obviously decreased in the inhibitor group (P<0.05, P<0.01). Comparison between EA and EA+inhibitor groups showed that the effects of EA were basically eliminated in lowering mNSS score and percentage of cerebral infarct volume (P<0.01), and in up-regulating MVD, and expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 mRNAs and proteins (P<0.01). The levels of mNSS score and percentage of cerebral infarct volume were markedly higher (P<0.01) in the inhibitor group than those in the EA+inhibitor group, while the MVD, expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 mRNAs and proteins were significantly lower in the inhibitor group than in the EA+inhibitor group (P<0.01), suggesting an elimination of EA after administration of HIF-1α inhibitor. H.E. staining showed loosened structure and disordered arrangement of neurons, swollen and vacuolized cells with ruptured nuclei, swollen and deformed microvessels in the ischemic penumbra of the brain tissue in the model group, which was improved in the EA group, including reduction of cellular swelling degree and vacuol-like changes, relatively intact of nuclei, and increase of new capillaries. EA of GV20 and ST36 can improve neurological deficit and reduce cerebral infarct volume in CI rats, which may be associated with its functions in promoting angiogenesis in ischemic penumbra area and in up-regulating the activities of HIF/VEGF/Notch signaling pathway and related factors.

METTL14 Promotes Ischemic Stroke-induced Brain Injury by Stabilizing HDAC3 Expression in an m6A-IGF2BP3 Mechanism.

N6-methyladenosine (m6A) modification is the most abundant post-transcriptional modification of mRNAs and has been identified to play critical roles in ischemic stroke (IS). Herein, this study aimed to investigate the function and mechanism of Methyltransferase-like 14 (METTL14) methylase in cerebral IS. Murine BV-2 microglial cell OGD/R models and rat middle cerebral artery occlusion (MCAO) models were established to mimic IS-induced neuronal damage in vitro and brain injury in vivo. Levels of METTL14, Histone Deacetylase 3 (HDAC3) and cGAS-STING axis-related proteins were detected using qRT-PCR or western blotting. Cell proliferation and inflammation were assessed by CCK-8 assay, EdU assay and ELISA. Flow cytometry detected microglia polarization. Cell pyroptosis was analyzed by detecting related-protein markers by western blotting. The m6A modification was determined by methylated RNA immunoprecipitation assay. Brain injury was analyzed by evaluating infarct volume and neurologic score. METTL14 levels were higher in OGD/R-induced microglial cells, primary microglia and infarct brain tissues of rat MCAO models. Functionally, METTL14 silencing reversed OGD/R-induced proliferation inhibition, inflammation and pyroptosis in microglial cells and primary microglia in vitro, and ameliorated cerebral ischemic injury in rat MCAO models. Mechanistically, METTL14 induced HDAC3 m6A modification in an IGF2BP3-dependent manner, and could activate cGAS-STING pathway through HDAC3. Moreover, HDAC3 overexpression reversed the neuroprotective effects of METTL14 silencing. METTL14 silencing reversed ischemic stroke-induced brain injury by inducing HDAC3 m6A modification in an IGF2BP3-dependent mechanism, recommending a novel insight for ameliorating cerebral ischemic stroke.

Analysis of nerve excitability in the dentate gyrus of the hippocampus in cerebral ischaemia-reperfusion mice

Ischemic stroke often leads to cognitive dysfunction, which delays the recovery process of patients. However, its pathogenesis is not yet clear. In this study, the cerebral ischemia-reperfusion model was built as the experimental object, and the hippocampal dentate gyrus (DG) was the target brain area. TTC staining was used to evaluate the degree of cerebral infarction, and nerve cell membrane potentials and local field potentials (LFPs) signals were collected to explore the mechanism of cognitive impairment in ischemia-reperfusion mice. The results showed that the infarcted area on the right side of the brain of the mice in the model group was white. The resting membrane potential, the number of action potential discharges, the post-hyperpolarization potential and the maximum ascending slope of the hippocampal DG nerve cells in the model mice were significantly lower than those in the control group ( P < 0.01); the peak time, half-wave width, threshold and maximum descending slope of the action potential were significantly higher than those in the control group ( P < 0.01). The time-frequency energy values of LFPs signals in the θ and γ bands of mice in the ischemia and reperfusion groups were significantly reduced ( P < 0.01), and the time-frequency energy values in the reperfusion group were increased compared with the ischemia group ( P < 0.01). The signal complexity of LFPs in the ischemia and reperfusion group was significantly reduced ( P < 0.05), and the signal complexity in the reperfusion group was increased compared with the ischemia group ( P < 0.05). In summary, cerebral ischemia-reperfusion reduced the excitability of nerve cells in the DG area of the mouse hippocampus; cerebral ischemia reduced the discharge activity and signal complexity of nerve cells, and the electrophysiological indicators recovered after reperfusion, but it failed to reach the healthy state during the experiment period.

Construction of HEK293T cell line stably expressing TRPM2 channel based on PiggyBac transposition system and its application in drug screening for cerebral ischemia and other diseases

To establish a cell line stably expressing the transient receptor potential melastatin 2 (TRPM2) channel for screening TRPM2 inhibitors based on PiggyBac transposition system. A plasmid PiggyBac-human TRPM2 (pPB-hTRPM2) eukaryotic expression vector was constructed using PiggyBac transposition system. The plasmid and a helper plasmid were co-transfected into HEK293T cells to express TRPM2, which was identified by fluorescence and patch-clamp assays. The high throughput screening performance was assessed with the Z ´ factor. Calcium imaging and patch clamp techniques were employed to assess the initial activity of eleven compound molecules, confirming the inhibitory effects of the primary molecules on TRPM2. The protective effect of the screened compounds on damaged cells was validated using the oxygen-glucose deprivation/reperfusion (OGD/R) injury model and CCK-8 kit. The level of cellular reactive oxygen species (ROS) was detected by flow cytometry. The neuroprotective effects of the compounds were evaluated using a transient middle cerebral artery occlusion (tMCAO) mouse model. The HEK293T cells transfected with pPB-hTRPM2-EGFP showed high TRPM2 expression. Puromycin-resistant cells, selected through screening, exhibited robust fluorescence. Whole-cell patch results revealed that induced cells displayed classical TRPM2 current characteristics comparable to the control group, showing no significant differences (P>0.05). With a Z ´ factor of 0.5416 in calcium imaging, the model demonstrated suitability for high-throughput screening of TRPM2 inhibitors. Calcium imaging and electrophysiological experiments indicated that compound 6 significantly inhibited the TRPM2 channel. Further experiments showed that 1.0 μmol/L of compound 6 enhanced cell viability (P<0.05) and reduced the level of ROS (P<0.05) of SH-SY5Y under OGD/R injury. 0.3 and 1.0 mg/kg of compound 6 reduced the cerebral infarction volume in tMCAO mice (both P<0.05). A stable TRPM2 gene expressing cell line has been successfully established using PiggyBac gene editing in this study. TRPM2 channel inhibitors were screened through calcium imaging and patch clamp techniques, and an inhibitor compound 6 was identified. This compound can alleviate cell damage after OGD/R by reducing cellular ROS levels and has a protective effect against cerebral ischemia-reperfusion injury in mice.

Effect of "Xingnao Kaiqiao" needling on expression of ferroptosis-related factors in rats with cerebral ischemia-reperfusion injury

To observe the effect of "Xingnao Kaiqiao" needling on the expression of ferroptosis-related proteins in neurons of rats with cerebral ischemia-reperfusion injury (CIRI), so as to explore its mechanism underlying improvement of CIRI. Male Wistar rats were randomly divided into sham operation, model, acupuncture and deferoxamine (DFO) groups, with 18 rats in each group. The CIRI model was established by occlusion of the middle cerebral artery. In the acupuncture group, "Xingnao Kaiqiao" needling was applied to "Shuigou" (GV26), "Neiguan" (PC6) and "Sanyinjiao"(SP6) for 20 min with electroacupuncture (2 Hz/15 Hz, 1 mA) at PC6 and SP6, twice daily for continuous 3 days. Rats of the DFO group received intraperitoneal injection of iron chelator DFO (0.1 g/kg, once daily). The severity of neurological impairment (neurological deficit score, 0-5 points, the lower the score, the severer is the neurological impairment) was evaluated by using Zausinger 6-poins scaling method. The cerebral infarct volume was measured after 2, 3, 5-triphenyltetrazolium chloride (TTC) staining, and the histopathological changes of the ischemic brain tissue were observed after H.E. staining. The mitochondrial structure of the hippocampal neurons on the ischemic side of the brain was observed by using transmission electron microscope. The levels of iron deposition rate (%) in the ischemic penumbra of the brain tissue and hippocampus were observed after Prussian blue staining, and the reactive oxygen species (ROS) content of the cerebral ischemic penumbra was assayed using flow cytometry, and the content of glutathione (GSH) content in the ischemic penumbra was detected by using microplate method. The real-time quantitative PCR was used to detect the expression of glutathione peroxidase 4 (GPX4), divalent metal transporter 1 (DMT1), transferrin (TF), transferrin receptor 1 (TFR1), and ferroportin 1 (FPN1) mRNAs in the ischemic penumbra, and the Western blot was used to detect the expression of GPX4, DMT1, TF, TFR1, FPN1, and ferritin (FER) proteins in the ischemic penumbra. Compared with the sham operation group, the neurological deficit score, GSH content, expression of GPX4 and FPN1 mRNAs and proteins were significantly decreased (P<0.01), while the percentage of cerebral infarct volume, iron deposit rates of the cerebral ischemic penumbra and hippocampus, ROS content, and the expression levels of DMT1, TF, and TFR1 mRNAs and proteins and FER protein were considerably increased (P<0.01) in the model group. In comparison with the model group, the decrease of neurological deficit score, GSH content, expression of GPX4 and FPN1 mRNAs and proteins, and the increase of the percentage of cerebral infarct volume, iron deposit rates of the cerebral ischemic penumbra and hippocampus, ROS content, and the expression levels of DMT1, TF, and TFR1 mRNAs and proteins and FER protein were all reversed in both DFO and acupuncture groups (P<0.01, P<0.05). The effects of acupuncture were significantly superior to those of DFO in lowering the levels of cerebral cortical and hippocampal iron deposit rates, ROS content and in elevating the expression of GPX4 mRNA and protein (P<0.01, P<0.05). H.E. staining showed large necrotic cells, disordered arrangement of cells in the cerebral cortex and hippocampus, with hyperchromic nuclei, vacuole-like changes, widening of cellular space, and cell swelling in the model group, which was relatively milder in the cell damage in both acupuncture and DFO groups. In addition, the ultrastructure of cells in the hippocampus showed irregular cellular nuclear morphology, atrophy of some mitochondria in the cytoplasm, partial mitochondrial membrane rupture and edema, and loosening of the ridge structure in the model group, which was milder in the mitochondrial impairment (including reduced number of mitochondria, broken mitochondrial membrane and reduced ridge structure in fewer cells) in the acupuncture group. The "Xingnao Kaiqiao" needling intervention has a neuroprotective effect in CIRI rats, which may be related to its functions in regulating ferroptosis-related targets and iron metabolism in cerebral ischemic penumbra, reducing oxidative stress injury, and suppressing neuronal ferroptosis.

Xenon and Argon as Neuroprotective Treatments for Perinatal Hypoxic-Ischemic Brain Injury: A Preclinical Systematic Review and Meta-Analysis.

Xenon and argon are currently being evaluated as potential neuroprotective treatments for acquired brain injuries. Xenon has been evaluated clinically as a treatment for brain ischemia with equivocal results in small trials, but argon has not yet undergone clinical evaluation. Several preclinical studies have investigated xenon or argon as treatments in animal models of perinatal hypoxic-ischemic encephalopathy (HIE). A systematic review of MEDLINE and Embase databases was performed. After screening of titles, abstracts, and full text, data were extracted from included studies. A pairwise meta-analysis of neuroprotective efficacy was performed using a random effects model. Heterogeneity was investigated using subgroup analysis, funnel plot asymmetry, and Egger's regression. The protocol was prospectively registered on PROSPERO (CRD42022301986). A total of 21 studies met the inclusion criteria. The data extracted included measurements from 1591 animals, involving models of HIE in mice, rats, and pigs. The meta-analysis found that both xenon and argon had significant (P < .0001) neuroprotective efficacies. The summary estimate for xenon was 39.7% (95% confidence interval [CI], 28.3%-51.1%) and for argon it was 70.3% (95% CI, 59.0%-81.7%). The summary effect for argon was significantly (P < .001) greater than that of xenon. Our results provide evidence supporting further investigation of xenon and argon as neuroprotective treatments for HIE.

Exercise preconditioning mitigates Ischemia-Reperfusion injury in rats by enhancing mitochondrial respiration

Cerebral ischemia and subsequent reperfusion damage are prevalent in clinical practice, linked to numerous neurodegenerative diseases. Cerebral ischemia deprives brain tissue of essential oxygen and nutrients, disrupting energy metabolism and causing cellular dysfunction. Although reperfusion theoretically aids recovery, it instead initiates complex injury responses such as oxidative stress, apoptosis, and inflammation, worsening brain damage. Recent research suggests that enhancing neuronal energy status by modulating energy metabolism pathways can effectively counter these effects. For instance, boosting mitochondrial function, improving energy provision, and decreasing harmful metabolites can mitigate oxidative stress and cellular injury. This study investigated the protective effects of exercise preconditioning against ischemia–reperfusion injury in rats. It was observed that exercise enhances energy levels and mitochondrial respiration by upregulating the expression of COX4 and NAMPT proteins and activating AMPK and mitochondrial complex V. This process facilitates metabolic reprogramming characterized by the promotion of oxidative phosphorylation (OXPHOS) and the pentose phosphate pathway (PPP), alongside a reduction in glycolysis. Such reprogramming reduces harmful metabolites, mitigating apoptosis and oxidative stress, and is a key factor in alleviating acute ischemic hypoxia-induced brain damage. These findings introduce a novel therapeutic approach for ischemic brain reperfusion injury, underscoring the crucial role of ATP production and metabolic regulation in neuroprotection.

Trajectory of mean platelet volume changes after aneurysmal subarachnoid hemorrhage in patients with or without delayed cerebral ischemia

The morbidity of aneurysmal subarachnoid hemorrhage (aSAH) remains high, particularly because of secondary cerebral lesions that significantly aggravate the primary lesions. The main type of secondary lesions is delayed cerebral ischemia (DCI), in which platelets (PLT) appear to play a key role. Mean platelet volume (MPV) is an indirect marker of platelet activation. We aimed to determine the individual trajectories of MPV over time in patients with and without DCI during the course of aSAH. This is a single-center, retrospective, longitudinal analysis of individual trajectories of MPV over time, in a cohort of aSAH patients included in the Prospective, Observational Registry of Patient with Subarachnoid Hemorrhage in Neurocritical Care Unit (ProReSHA). A mixed-effects linear regression model was used to compare the trajectories of MPV and MPV/PLT ratio between patients who developed a DCI and those who did not. A total of 3634 MPV values were collected in 587 patients. The analysis of MPV as a function of DCI occurrence showed a significant difference in the trajectory over time between patients with DCI and those without, with an estimate of 0.02 (95%CI 0.01, 0.04, p = 0.009). The analysis of the MPV/PLT ratio as a function of DCI occurrence and other covariates showed a significant difference in the trajectory over time only for patients with a modified Fisher score less than 3, with an estimate of -0.59 (95%CI: -0.94, -0.23, p = 0.001). The individual trajectories of MPV over time differ between patients with DCI and those without. However, MPV values vary greatly over time and between patients. Thus it does not appear as a reliable biomarker for stratifying patients based on their specific risk of developing DCI. ClinicalTrials.gov identifier: (NCT02890004), registered in August 2016.

A coloaded liposome in situ gel as a novel therapeutic strategy to treat cerebral ischemia reperfusion injury

BackgroundIschemic stroke has become one of the leading causes of death and disability worldwide in individuals aged 60 and above. However, currently available drugs show limited efficacy. Therefore, research to find more effective and safer therapeutic strategies is an urgent requirement for the treatment of cerebral ischemia reperfusion injury (CIRI). MethodsFirst, the free radical scavenger Edaravone and a Ginseng active ingredient were coloaded into liposomes (aER@Lip), followed by optimization and characterization. Pluronic F127 and F68 at different concentrations were mixed and stored at 4 °C for more than 24 h to obtain gel solutions. Then, aER@Lip was added to the gel solutions to prepare the drug-loaded in situ gel, termed aER@Lip-TSG. ResultsIn vitro experiments showed that aER@Lip-TSG was taken up by cells and had a good protective effect on oxygen-glucose deprivation/reoxygenation in pheochromocytoma 12 cells. In a rat CIRI model, aER@Lip-TSG delivered by intranasal administration not only decreased the apoptosis in brain tissue induced by CIRI, but also decreased the resultant inflammatory response. Moreover, the results suggested that aER@Lip-TSG had good biosafety. ConclusionThis delivery system provides a promising multi-factor combination, synergistic effects, sustained-release capabilities, and is a non-invasive treatment strategy for CIRI. It thus meets the urgent need for effective treatments of central nervous system diseases.