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Background: The cornu ammonis 1 (CA1) region of the hippocampus is a sensitive area that is susceptible to injury caused by cerebral ischemia. High-mobility group box 1 (HMGB1) and phosphorylated c-Jun N-terminal kinase (p-JNK) play important roles in mediating cerebral ischemic injury., Objective: To elucidate the mechanism through which electroacupuncture (EA) via the Baihui (GV20) and Zusanli (ST36) acupoints protects neurons., Methods: A rat model of permanent middle cerebral artery occlusion (pMCAO) was established. Sprague-Dawley rats were divided into four groups: sham-operated control, pMCAO control, EA, and sham-EA (SEA). In the EA and SEA groups, the GV20 and ST36 acupoints were selected for treatment. However, the SEA group was treated only by superficial pricking of the skin at the two acupoints without the application of electricity. Neurological function was assessed using the neurological deficit function score, and neuronal damage was detected through Nissl staining. HMGB1 and p-JNK expression was evaluated using immunohistochemical staining and western blot assays., Results: The behavioural experiments showed that the EA treatment improved the neurological deficits in the pMCAO rats. The Nissl staining results revealed that EA reduced neural tissue damage. The immunohistochemical staining and western blot results showed that EA inhibited HMGB1 and p-JNK overexpression. By contrast, none of these EA effects were observed in the SEA group., Conclusion: EA may reduce ischemia-induced neuronal damage in the hippocampal CA1 region by inhibiting the overexpression of both HMGB1 and p-JNK.

Introduction: Cerebral ischemic strokes cause brain damage, primarily through inflammatory factors. One of the regions most affected by middle cerebral artery occlusion (MCAO) is the hippocampus, specifically the CA1 area, which is highly susceptible to ischemia. Previous studies have demonstrated the anti-inflammatory properties of quercetin. Therefore, this study aimed to investigate the neuroprotective effects of quercetin on hippocampal CA1 neurons following MCAO., Materials and Methods: Fifty-six male Albino Wistar rats were divided into seven groups (intact, sham, stroke, vehicle, and three quercetin-treated groups receiving 5, 10, and 20 mg/kg, respectively), each containing 8 rats. Various assessments, including brain water content, the rotarod test, the Bederson neurological score, the Morris water maze (MWM) test, the shuttle box test, histopathological evaluations, and measurements of interleukin-10 (IL-10) and interleukin-1β (IL-1β) levels, were conducted across the groups., Results: Compared with control rats, 5 and 10 mg/kg quercetin-treated rats presented significant improvements in brain water content, neurological function, and motor function and improved performance in the MWM and shuttle box tests. Histopathological analyses revealed better preservation of CA1 neurons in these groups. Additionally, IL-10 levels significantly increased, whereas IL-1β levels significantly decreased. However, the group receiving 20 mg/kg quercetin showed no statistically significant changes in the parameters assessed (P > 0.05)., Conclusion: Quercetin may help prevent or ameliorate brain injuries caused by acute stroke, suggesting its neuroprotective effects. The reduction in IL-1β and increase in IL-10 may play key roles in quercetin's protective mechanism., Competing Interests: Declarations. Ethics approval and consent to participate: This study was conducted without commercial input or involvement in the design, implementation, analysis, or reporting. This study was approved by the Research Ethics Committee of Laboratory Animals, Mazandaran University of Medical Sciences (Ethics Approval Code: IR.MAZUMS.AEC.1401.046). All procedures performed in this study were in accordance with the ARRIVE guidelines and the ethical standards of the Institutional Research Ethics Committee of Mazandaran University of Medical Sciences and in accordance with the Guidance on the Operation of the Animals (Scientific Procedures) Act 1986 and associated guidelines. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Post-stroke depression (PSD) is a significant complication in stroke patients, increases long-term mortality, and exaggerates ischemia-induced brain injury. However, the underlying molecular mechanisms and effective therapeutic targets related to PSD have remained elusive. Here, we employed an animal behavioral model of PSD by combining the use of middle cerebral artery occlusion (MCAO) followed by spatial restraint stress to study the molecular underpinnings and potential therapies of PSD. Interestingly, we found that sub-chronic application of gastrodin (Gas), a traditional Chinese medicinal herb Gastrodia elata extraction, relieved depression-related behavioral deficits, increased the impaired expression of synaptic transmission-associated proteins, and restored the altered spine density in hippocampal CA1 of PSD animals. Furthermore, our results indicated that the anti-PSD effect of Gas was dependent on membrane cannabinoid-1 receptor (CB1R) expression. The contents of phosphorated protein kinase A (p-PKA) and phosphorated Ras homolog gene family member A (p(ser188)-RhoA) were decreased in the hippocampus of PSD-mice, which was reversed by Gas treatment, and CB1R depletion caused a diminished efficacy of Gas on p-PKA and p-RhoA expression. In addition, the anti-PSD effect of Gas was partially blocked by PKA inhibition or RhoA activation, indicating that the anti-PSD effect of Gas is associated with the CB1R-mediated PKA/RhoA signaling pathway. Together, our findings revealed that Gas treatment possesses protective effects against the post-stroke depressive-like state; the CB1R-involved PKA/RhoA signaling pathway is critical in mediating Gas's anti-PSD potency, suggesting that Gas application may be beneficial in the prevention and adjunctive treatment of PSD., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Ischemic stroke is a devastating disease in which mitochondrial damage or dysfunction substantially contributes to brain injury. Mitochondrial uncoupling protein-2 (UCP2) is a member of the UCP family, which regulates production of mitochondrial superoxide anion. UCP2 is reported to be neuroprotective for ischemic stroke-induced brain injury. However, the molecular mechanisms of UCP2 in ischemic stroke remain incompletely understood. In this study, we investigated whether and how UCP2 modulates neuroinflammation and regulates neuronal ferroptosis following ischemic stroke in vitro and in vivo. Wild-type (WT) and UCP2 knockout (Ucp2 -/- ) mice were subjected to middle cerebral artery occlusion (MCAO). BV2 cells (mouse microglial cell line) and HT-22 cells (mouse hippocampal neuronal cell line) were transfected with small interfering (si)-RNA or overexpression plasmids to knockdown or overexpress UCP2 levels. Cells were then exposed to oxygen-glucose deprivation and reoxygenation (OGD/RX) to simulate hypoxic injury in vitro. We found that UCP2 expression was markedly reduced in a time-dependent manner in both in vitro and in vivo ischemic stroke models. In addition, UCP2 was mainly expressed in neurons. UCP2 deficiency significantly enlarged infarct volumes, aggravated neurological deficit scores, and exacerbated cerebral edema in mice after MCAO. In vitro knockdown of Ucp2 and in vivo genetic depletion of Ucp2 (Ucp2 -/- mice) increased neuronal ferroptosis-related indicators, including Fe 2+ , malondialdehyde, glutathione, and lipid peroxidation. Overexpression of UCP2 in neuronal cells resulted in reduced ferroptosis. Moreover, knockdown of UCP2 exacerbated neuroinflammation in BV2 microglia and mouse ischemic stroke models, suggesting that endogenous UCP2 inhibits neuroinflammation following ischemic stroke. Upregulation of UCP2 expression in microglia appeared to decrease the release of pro-inflammatory factors and increase the levels of anti-inflammatory factors. Further investigation showed that UCP2 deletion inhibited expression of AMPKα/NRF1 pathway-related proteins, including p-AMPKα, t-AMPKα, NRF1, and TFAM. Thus, UCP2 protects the brain from ischemia-induced ferroptosis by activating AMPKα/NRF1 signaling. Activation of UCP2 represents an attractive strategy for the prevention and treatment of ischemic stroke., Competing Interests: Declarations. Ethics Approval: The study was approved by the Ethics Committee of Renmin Hospital of Wuhan University and performed in compliance with the ARRIVE guidelines. Consent for Publication: Not applicable. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Experimental rat models of stroke and hemorrhage are important tools to investigate cerebrovascular disease pathophysiology mechanisms, yet how significant patterns of functional impairment induced in various models of stroke are related to changes in connectivity at the level of neuronal populations and mesoscopic parcellations of rat brains remain unresolved. To address this gap in knowledge, we employed two middle cerebral artery occlusion models and one intracerebral hemorrhage model with variant extent and location of neuronal dysfunction. Motor and spatial memory function was assessed and the level of hippocampal activation via Fos immunohistochemistry. Contribution of connectivity change to functional impairment was analyzed for connection similarities, graph distances and spatial distances as well as the importance of regions in terms of network architecture based on the neuroVIISAS rat connectome. We found that functional impairment correlated with not only the extent but also the locations of the injury among the models. In addition, via coactivation analysis in dynamic rat brain models, we found that lesioned regions led to stronger coactivations with motor function and spatial learning regions than with other unaffected regions of the connectome. Dynamic modeling with the weighted bilateral connectome detected changes in signal propagation in the remote hippocampus in all 3 stroke types, predicting the extent of hippocampal hypoactivation and impairment in spatial learning and memory function. Our study provides a comprehensive analytical framework in predictive identification of remote regions not directly altered by stroke events and their functional implication., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Objective: Cerebral ischemia-reperfusion injury (CIRI) is a major obstacle to neurological recovery after clinical treatment of ischemic stroke. The aim of this study was to investigate the molecular mechanism of Nek6 alleviating CIRI through autophagy after cerebral ischemia., Materials and Methods: A mouse model of CIRI was constructed by middle cerebral artery occlusion (MCAO). TUNEL staining was used to observe the apoptosis of neuronal cells. The oxygen glucose deprivation/reoxygenation (OGD/R) model was established by hypoxia and reoxygenation. The cell apoptosis and activity was detected. Western blot was performed to detect the expression of autophagy-related proteins, protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and adenosine 5'-monophosphate-activated protein kinase (AMPK)/mTOR signaling pathway-related proteins. Cellular autophagy flux was observed by fluorometric method. NIMA-related kinase 6 (Nek6) mRNA stability was detected by actinomycin D treatment. Methylation RNA immunoprecipitation technique was used to detect Nek6 methylation level., Results: Nek6 expression was increased in both MCAO and OGD/R models. Overexpression of Nek6 in OGD/R inhibited apoptosis, decreased LC3II and Beclin-1 expression, increased p62 expression, and occurred lysosome dysfunction. Interference with Nek6 has opposite results. Nek6 overexpression promoted p-Akt and p-mTOR protein expressions, inhibited p-AMPK and p-UNC-51-like kinase 1 protein expressions and cell apoptosis, while LY294002, Rapamycin or RSVA405 treatment reversed this effect. Abnormal methyltransferase·like protein 3 (METTL3) expression in CIRI enhanced m6A modification and promoted Nek6 expression level., Conclusion: This study confirmed that Nek6 regulates autophagy and alleviates CIRI through the mTOR signaling pathway, which provides a novel therapeutic strategy for patients with ischemic stroke in the future., Competing Interests: Declarations. Ethics approval and consent to participate: All animal experiments are approved by the Ethics committee of The First Affiliated Hospital of Zhengzhou University. Consent for publication: Not applicable. Competing interests: All authors declare that they have no conflict of interest., (© 2024. The Author(s).)

Objective: Ferroptosis is an iron-dependent form of programmed cell death associated with lipid peroxidation. Though diabetes worsens cerebral injury and clinical outcomes in stroke, it is poorly understood whether ferroptosis contributes to diabetes-exacerbated stroke. This study aimed to identify ferroptosis-associated differentially expressed genes in ischemic stroke under diabetic condition and then explore their roles using comprehensive bioinformatics analyses., Methods: Type 1 diabetes (T1D) model was established in male mice at 8-10 weeks of age by one intraperitoneal injection of streptozotocin (110 mg/kg). Ischemic stroke was induced by a transient 45-minute middle cerebral artery occlusion and evaluated three days thereafter. Ischemic brain cortex was dissected 24 h after the reperfusion and subjected to bulk tissue RNA sequencing followed by bioinformatics analysis and verification of key findings via quantitative real-time PCR., Results: Enlarged infarct size was seen in diabetic, as compared with non-diabetic mice, in conjunction with worsened neurological behaviors. Both body and spleen weights were reduced in diabetic as compared with non-diabetic mice. There was a trend for reduced survival rate in diabetic mice following the stroke. In RNA sequencing analysis, we identified 1299 differentially expressed genes in ischemic brain between diabetic and non-diabetic mice, with upregulation and downregulation for 732 and 567 genes, respectively. Among these genes, 27 genes were associated with ferroptosis. Further analysis reveals that solute carrier family 25 member 28(SLC25A28) and sterol carrier protein 2(SCP2) were the top genes associated with ferroptosis in diabetic mice following ischemic stroke. In several bioinformatics analyses, we found SLC25A28, one of the top ferroptosis-related genes, is involved in several metabolic and regulatory pathways as well as the regulatory complexity of microRNAs and circular RNAs, which demonstrates the potential role of SLC25A28 in diabetes-exacerbated stroke. Drug network analysis suggests SLC25A28 as a potential therapeutic target for ameliorating ischemic injury in diabetes., Conclusions: Our bulk RNA sequencing and bioinformatics analyses show that altered ferroptosis signaling pathway was associated with the exacerbation of experimental stroke injury under diabetic condition. Especially, additional investigation into the mechanisms of SLC25A28 and SCP2 in diabetes-exacerbated stroke will be explored in the future study., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Introduction: This study aimed to explore the characteristics of post-stroke sleep dysfunction and verify their association with gut dysbiosis and the related amino acid metabolism disorders. This was achieved by using fecal microbiota transplantation (FMT) in a non-human primate stroke model., Methods: Twenty adult male cynomolgus monkeys were divided into the sham (n = 4), middle cerebral artery occlusion (MCAO, n = 5), MCAO + FMT (n = 3), and donor (n = 8) groups. The MCAO+FMT group received FMT at post-MCAO week 4. Sleep parameters, gut microbiota, gamma-aminobutyric acid (GABA), and glutamine (Gln) in the cerebrospinal fluid (CSF) were measured at baseline and postoperative weeks 4, 8, and 12., Results: At postoperative weeks 4, 8, and 12, the MCAO group showed decreased sleep efficiency, measured as the percentage of sleep during the whole night (82.3 ± 3.2 % vs 91.3 ± 2.5 %, 79.0 ± 3.75 % vs 90.8 ± 3.2 %, and 69.5 ± 4.8 % vs 90.5 ± 2.7 %; all P < 0.05), lower relative abundance of Lactobacillus (all P < 0.05), and reduced GABA concentrations in the CSF (317.3 ± 30.6 nmol/L vs 437.7 ± 25.6 nmol/L, 303.1 ± 48.9 nmol/L vs 4 40.9 ± 37.8 nmol/L, and 337.9 ± 49.4 nmol/L vs 457.4 ± 39.2 nmol/L; all P < 0.05) compared with the sham group. Sleep efficiency at post-FMT weeks 4 and 8 (84.7 ± 1.1 % vs 79.0 ± 3.75 %, and 84.1 ± 2.0 % vs 69.5 ± 4.8 %; both P < 0.05) and GABA concentration in the CSF at post-FMT week 4 (403.1 ± 25.4 nmol/L vs 303.1 ± 48.9 nmol/L, P < 0.05) was higher in the MCAO+FMT group than in the MCAO group., Conclusions: Post-stroke sleep dysfunction in monkeys is characterized by impaired sleep coherence, associated with decreased levels of probiotics such as Lactobacillus, GABA, and Gln in the CSF and can be ameliorated using FMT., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Jinsheng Zeng reports financial support was provided by National Natural Science Foundation of China. Fubing Ouyang reports financial support was provided by National Natural Science Foundation of China. Yicong Chen reports was provided by the Basic and Applied Basic Research Foundation Natural Science Foundation of Guangdong Province. Fubing Ouyang reports financial support was provided by the Medical Scientific Research Foundation of Guangdong Province. Jinsheng Zeng reports financial support was provided by Sun Yat-sen University Clinical Research 5010 Program. Yicong Chen reports financial support was provided by the Kelin Star Talent Support Program of the First Affiliated Hospital, Sun Yat-sen University. Jinsheng Zeng reports financial support was provided by Guangdong Provincial Key Laboratory of Diagnosis and Treatment of Major Neurological Diseases. Jinsheng Zeng reports was provided by Guangdong Province International Cooperation Base for Early Intervention and Functional Rehabilitation of Neurological Diseases. Jinsheng Zeng reports financial support was provided by Guangdong Provincial Engineering Center for Major Neurological Disease Treatment. Jinsheng Zeng reports financial support was provided by Guangdong Provincial Translational Medicine Innovation Platform for Diagnosis and Treatment of Major Neurological Disease and Guangzhou Clinical Research and Translational Center for Major Neurological Diseases. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

In recent years, the mechanism of acupuncture in the treatment of post-stroke cognitive impairment (PSCI) has not been fully elucidated. The balance between mitochondrial fission and fusion is important for PSCI. Our previous research demonstrated that electroacupuncture can improve learning and memory in middle cerebral artery ischemia reperfusion (MCAO/R) rats. However, the specific mechanism by which electroacupuncture improves learning and memory in MCAO/R rats by regulating mitochondrial fission and fusion needs to be further investigated. The MCAO/R rats was developed using the line-bolt method. The rats were randomly divided into sham-operated (Sham), model (MCAO/R), electroacupuncture (MCAO/R + EA) and sham-electroacupuncture (MCAO/R + sham EA) groups. Investigating the effects of EA on the expression of Sirtuin1 (SIRT1), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), Optic atrophy 1R + (OPA1) and Dynamin-related protein 1 (DRP1) in hippocampal neurons and on the morphology and function of hippocampal neurons and mitochondria. EA was able to reduce neurologic deficit scores and cerebral infarct volume and improve new object discrimination in MCAO/R rats, but there were no significant changes in these indices in the sham-electroacupuncture group. Moreover, EA increased the expression of SIRT1, PGC-1α, and OPA1 in hippocampal tissues, inhibited the expression of DRP1, attenuated neuronal and mitochondrial damage, and reduced mitochondrial fragmentation. The mechanism by which EA improves learning memory deficits in MCAO/R rats may be related to the inhibition of SIRT1/PGC-1α expression, the enhancement of mitochondrial fusion and the obstruction of its fission, and the reduction of hippocampal neuronal damage., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Purpose: Neurological impairments are the leading cause of post-stroke mortality, while stroke-related cardiovascular diseases rank second in significance. This study investigates the potential protective effects of MitoTEMPO (2,2,6,6-tetramethyl-4-[[2-(triphenylphosphonio) acetyl] amino]-1-piperidinyloxy, monochloride, monohydrate), a mitochondria-specific antioxidant, against cardiac and neurological complications following stroke. The objective is to assess whether MitoTEMPO can be utilized as a protective agent for individuals with a high risk of stroke., Materials and Methods: Seventeen-week-old male Wistar Albino rats were randomly assigned to three groups: SHAM, ischemia-reperfusion and MitoTEMPO + ischemia-reperfusion (MitoTEMPO injection 0.7 mg/kg/day for 14 days). The SHAM group underwent a sham operation, while the ischemia-reperfusion group underwent 1-h middle cerebral artery occlusion followed by three days of reperfusion. Afterwards, noninvasive thoracic electrical bioimpedance and electrocardiography measurements were taken, and sample collection was performed for histological and biochemical examinations., Results: Our thoracic electrical bioimpedance and electrocardiography findings demonstrated that MitoTEMPO exhibited a protective effect on most parameters affected by ischemia-reperfusion compared to the SHAM group. Furthermore, our biochemical and histological data revealed a significant protective effect of MitoTEMPO against oxidative damage., Conclusions: The findings suggest that both ischemia-reperfusion-induced cardiovascular abnormalities and the protective effect of MitoTEMPO may involve G-protein coupled receptor-mediated signaling mechanisms. This study was conducted with limitations including a single gender, a uniform age group, a specific stroke model limited to middle cerebral artery, and pre-scheduled only one ischemia-reperfusion period. In future studies, addressing these limitations may enable the implementation of preventive measures for individuals at high risk of stroke.

Ischemic stroke is followed by an increased susceptibility to bacterial infections, which exacerbate histological stroke outcome, neurological deficits and memory impairment due to increased neuroinflammation and neurotransmitter dysfunction. Pharmacological activation of nicotinic acetylcholine receptors was suggested to mitigate brain inflammatory responses in ischemic stroke. The functional responses associated with nicotinic acetylcholine receptor activation were unknown. In this study, male NMRI mice subjected to transient intraluminal middle cerebral artery occlusion (MCAO) were intraperitoneally exposed to vehicle treatment or Escherichia coli lipopolysaccharide (LPS; 4 mg/kg)-induced sepsis-like state 24 h post-MCAO, followed by intraperitoneal administration of vehicle or nicotine (0.5 mg/kg) 30 min later. Over 96 h, rectal temperature, neurological deficits, spontaneous locomotor activity, working memory, ischemic injury, synaptic plasticity, and brain inflammatory responses were evaluated by temperature measurement, behavioral analysis, infarct volumetry, electrophysiological recordings, and polymerase-chain reaction analysis. LPS-induced sepsis induced hypothermia, increased general and focal neurological deficits, reduced spontaneous exploration behavior, reduced working memory, and increased infarct volume post-MCAO. Additional treatment with nicotine attenuated LPS-induced hypothermia, reduced neurological deficits, restored exploration behavior, restored working memory, and reduced infarct volume. Local field potential recordings revealed that LPS-induced sepsis decreased long-term potentiation (LTP) in the dentate gyrus post-MCAO, whereas concomitant nicotine exposure restored LTP in the contralateral dentate gyrus. LPS-induced sepsis increased microglial/ macrophage Iba-1 mRNA and astrocytic GFAP mRNA levels post-MCAO, whereas add-on nicotine treatment reduced astrocytic GFAP mRNA. Taken together, these findings indicate that acute nicotine exposure enhances functional stroke recovery. Future studies will have to evaluate the effects of (1) chronic nicotine exposure, a clinically relevant vascular risk factor, and (2) the cessation of nicotine exposure, which is widely recommended post-stroke, but might have detrimental effects in the early stroke recovery phase., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Systemic inflammatory stimulus is a risk factor for the incidence of ischemic stroke and contributes to poorer clinical outcomes. Solute carrier 15A3 (SLC15A3) is a peptide/histidine transporter that is implicated in regulating inflammatory responses. However, whether SLC15A3 affects the progression of ischemic stroke associated with systemic inflammation is unclear. The transient middle cerebral artery occlusion (tMCAO) mice with LPS administration (LPS/tMCAO) were prepared as an in vivo model, and LPS-induced BV2 cells under oxygen-glucose deprivation (OGD) exposure were utilized as an in vitro model. We found that SLC15A3 was highly expressed in the ischemic penumbra of LPS/tMCAO mice, and its inhibition reduced infarct area, attenuated neurological deficit, recovered motor function, and mitigated apoptotic neurons. Knockdown of SLC15A3 suppressed the proinflammatory M1-type markers and promoted the levels of M2-associated genes. The in vitro results confirmed that SLC15A3 overexpression promoted microglia polarizing towards M1 subtypes, while SLC15A3 inhibition exerted an opposite effect. In addition, we demonstrated that the p65 signaling pathway and HIF1α were activated by LPS/OGD. Luciferase reporter assay showed that inhibiting p65 using its specific inhibitor BAY 11-7082 or silencing HIF1α using siRNAs reduced the transcriptional activity of SLC15A3 in LPS/OGD-induced BV2 cells. Results in NIH 3T3 cells also confirmed that p65 and HIF1α directly bound to the SLC15A3 promoter to activate SLC15A3 transcription. In conclusion, this work shows that SLC15A3, transcriptionally activated by p65 and HIF1α, contributes to poor outcomes in ischemic stroke associated with systemic inflammation by promoting microglial cells polarizing towards M1 types., Competing Interests: Declarations. Ethics statement: The animal experiments were performed complied with the Guide for the Care and Use of Laboratory Animals, and the ethics were approved by the ethical committee of Jilin University (No. 202002031). Consent to participate: Not applicable. Consent to publish: Not applicable. Competing interests: The authors declare no conflicts of interest., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: Epileptic seizures occur frequently after stroke due to changes in brain function and structure, and up to around 10% of stroke patients experience stroke recurrence in the first year. We aimed to establish the risk of acute symptomatic seizures in patients with recurrent stroke., Methods: Retrospective cohort study including consecutive admissions to a Stroke Unit due to acute ischemic stroke, during a 5-year period. Additional inclusion of patients admitted to two centers in different countries to corroborate findings (confirmatory cohort). We aimed to compare acute symptomatic seizure incidence in patients with and without previous stroke. Patients with history of epilepsy were excluded. Logistic regression modeling was performed to identify predictors in middle cerebral artery (MCA) stroke., Results: We included 1473 patients (1085 with MCA stroke), of which 117 had a recurrent ischemic stroke (84 with MCA stroke). Patients with recurrent stroke had a seizure risk during hospital stay similar to that of patients with a first-ever stroke (5.1% vs. 4.5%, OR 1.15, 95% CI .48-2.71, p = .75). Risk of acute symptomatic seizures was also similar (5.0% vs. 4.1, OR 1.22, 95% CI .29-5.27, p = .78). Older age, female sex, and hemorrhagic transformation were predictors of seizures in patients with a first MCA ischemic stroke, but not in recurrent stroke patients. Electrographic characteristics were similar between the two groups in patients who had an electroencephalogram (46 with first stroke, 5 with recurrent stroke). The low rate of seizures (1.5%) in the confirmatory cohort (n = 198) precluded full comparison with the initial cohort. Nevertheless, the rate of seizures was not higher in stroke recurrence., Significance: History of previous stroke was not associated with an increased risk of acute symptomatic seizures during hospital stay. Larger, prospective studies, with prospective electrophysiological evaluation, are needed to explore the impact of stroke recurrence on seizure risk., (© 2024 International League Against Epilepsy.)

Stroke induces cardiac dysfunction, which increases poststroke mortality and morbidity. An imbalance in the autonomic nervous system resulting from brain injury may serve as the underlying mechanism. The present study investigated whether transcutaneous auricular vagus nerve stimulation (taVNS) attenuates poststroke cardiac dysfunction by activating the parasympathetic nervous system. Adult male mice were subjected to transient middle cerebral artery occlusion (MCAO) and reperfusion surgery. The mice in the treatment group received repeated taVNS starting 60 min after the onset of cerebral ischemia. To assess whether the effects of taVNS were associated with parasympathetic activity, the MCAO mice in the atropine group received intraperitoneal injections of atropine to inhibit parasympathetic activity prior to taVNS. taVNS significantly increased the left ventricular ejection fraction (EF), attenuated myocardial apoptosis, reduced myocardial hypertrophy, and reduced fibrosis following stroke. The beneficial effects of taVNS on cardiac dysfunction were abolished by atropine administration. Heart rate variability (HRV) analysis and western blot analysis revealed that taVNS increased parasympathetic activity but decreased sympathetic dominance in mice with MCAO. Furthermore, the cardioprotective effects of taVNS were associated with muscarinic acetylcholine receptor activation, PI3K-Akt pathway modulation, and eNOS regulation in the heart. Therefore, taVNS alleviates cardiac dysfunction after stroke and is associated with activation of the parasympathetic nervous system., Competing Interests: Declaration of competing interest We declare that all authors have no financial or personal relationships with other people or organizations that can influence our work. There is no professional or other personal interest of any nature or kind in any product, service or company., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Electroacupuncture (EA) has been shown to ameliorate brain injury and protect against intestinal injury after ischemic stroke. These protective effects are closely associated with the enhancement of regulatory T (Treg) cell numbers and function in the intestine, as well as the inhibition of intestinal γδ T cell production and their migration to the brain. This study aimed to elucidate the potential mechanism by which EA regulates intestinal Treg cell differentiation after stroke. Sprague-Dawley rats were divided into three groups: the sham group, the middle cerebral artery occlusion (MCAO) group, and the MCAO plus EA (MEA) group. The MCAO model was generated by occluding the middle cerebral artery. EA was applied to Baihui (GV20) acupoint once daily. Samples were collected 3 days after reperfusion. Our results showed that EA reduced the inflammatory response in the brain and intestine after ischemic stroke. EA treatment increased the percentage of Treg cells in the small intestine of rats. EA increased the levels of SCFAs, while also inhibiting histone deacetylase activity (HDAC). Additionally, acetylated Foxp3 protein in the small intestine was increased after EA treatment. These results suggest that EA at GV20 alleviates brain and intestinal inflammatory injury in stroke rats, potentially through the enhancement of SCFA-mediated Foxp3 acetylation in Treg cells., Competing Interests: Declarations. Ethics Approval: This study was performed in line with the Guide for the Care and Use of Laboratory Animals published by the National Institutes of Health. Approval was granted by the Institutional Animal Care and Use Committee of the Nanjing University of Chinese Medicine (202103A003). Consent to Participate: Not applicable. Consent for Publication: Not applicable. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

People with type 1 diabetes (T1D) have a significantly elevated risk of stroke, but the mechanism through which T1D worsens ischemic stroke remains unclear. This study was aimed at investigating the roles of T1D-associated changes in the gut microbiota in aggravating ischemic stroke and the underlying mechanism. Fecal 16SrRNA sequencing indicated that T1D mice and mice with transplantation of T1D mouse gut microbiota had lower relative abundance of butyric acid producers, f&#95;Erysipelotrichaceae and g&#95;Allobaculum, and lower content of butyric acid in feces. After middle cerebral artery occlusion (MCAO), these mice had poorer neurological outcomes and more severe inflammation, but higher expression of myeloid differentiation factor 88 (MyD88) in the ischemic penumbra; moreover, the microglia were inclined to polarize toward the pro-inflammatory type. Administration of butyrate to T1D mice in the drinking water alleviated the neurological damage after MCAO. Butyrate influenced the response and polarization of BV2 and decreased the production of inflammatory cytokines via MyD88 after oxygen-glucose deprivation/reoxygenation. Knocking down MyD88 in the brain alleviated neurological outcomes and decreased the concentrations of inflammatory cytokines in the brain after stroke in mice with transplantation of T1D mouse gut microbiota. Poor neurological outcomes and aggravated inflammatory responses of T1D mice after ischemic stroke may be partly due to differences in microglial polarization mediated by the gut microbiota-butyrate-MyD88 pathway. These findings provide new ideas and potential intervention targets for alleviating neurological damage after ischemic stroke in T1D., Competing Interests: Declarations. Ethics Approval: All experimental procedures were approved by the Ethics Committee of the Second Affiliated Hospital of Harbin Medical University (animal ethics approval number: SYDW2019-105) and were conducted in accordance with the Guidelines for the Care and Use of Laboratory Animals of Harbin Medical University. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Levodopa (L-DOPA) treatment is a clinically effective strategy for improving motor function in patients with ischemic stroke. However, the mechanisms by which modulating the dopamine system relieves the pathology of the ischemic brain remain unclear. Emerging evidence from an experimental mouse model of ischemic stroke, established by middle cerebral artery occlusion (MCAO), suggested that L-DOPA has the potential to modulate the inflammatory and immune response that occurs during a stroke. Here, we aimed to demonstrate the therapeutic effect of L-DOPA in regulating the systemic immune response and improving functional deficits in mice with ischemia. Transient MCAO led to progressive degeneration of nigrostriatal dopamine neurons and significant rotational behavior in mice. Exogenous L-DOPA treatment attenuated the striatonigral degeneration and reversed motor behavioral impairment. Notably, treatment with L-DOPA significantly increased IL-13 but reduced IFN-γ in infarct lesions. To investigate the role of IL-13 in motor behavior, we stereotaxically injected anti-IL-13 antibodies into the infarct area of the mouse brain one week after MCAO, followed by L-DOPA treatment. The intervention reduced dopamine, IL-13, and IL-10 levels and exacerbated motor function. IL-13 is potentially expressed on CD4 T cells, while IL-10 is mainly expressed on microglia rather than astrocytes. Finally, IL-13 activates the phagocytosis of microglia, which may contribute to neuroprotection by eliminating degenerating neurons. Our study provides evidence that the L-DOPA-activated dopamine system modulates peripheral immune cells, resulting in the expression of anti-inflammatory and neuroprotective cytokines in mice with ischemic stroke., Competing Interests: Competing interests: The authors declare no competing interests. Ethics approval: All studies were conducted following the guidelines of the Institutional Animal Care and Use Committee of Inha University in Incheon (INHA170908-513-5; INHA180420-558-4; INHA200518-701-7; INHA230525-875-2; INHA240723-939)., (© 2024. The Author(s).)

Objective: To explore the mechanism of electroacupuncture pretreatment (EP) for relieving post-stroke spasticity in rats., Methods: Eighteen rats were randomized equally into sham-operated group, middle cerebral artery occlusion (MCAO) group, and MCAO+EP group. In MCAO+EP group, the rats received electroacupuncture at the acupoints Qubin and Baihui for 3 consecutive days prior to MCAO. Neurological deficits and cognitive function of the rats were evaluated, and pathologies in the hippocampus were examined using HE, Nissl, and TUNEL staining. The expressions of IL-4, IL-6, TNF-α, and TMAO in the brain tissues were detected with ELISA, and the mRNA and protein expression levels of NF-κB p65, NLRP3, caspase-3, and caspase-9 were determined with qRT-PCR, Western blotting, and immunohistochemistry., Results: The rats receiving MCAO had significantly increased neurological deficit scores and showed increased muscle tension, number of apoptotic neurons, and expressions of IL-6, TNF-α, NF-κB p65, NLRP3, caspase-3 and caspase-9 in the hippocampus and significantly reduced length of time for new object recognition. Microscopically, the cells in the hippocampus of the MCAO rats showed uneven and loosened arrangement and unclear cell boundaries. In contrast, the rats in I/R+EP group showed significantly lowered neurological deficit scores and dystonia rating scores, reduced cell apoptosis, lowered hippocampal expressions of IL-6, TNF-α, caspase-3, caspase-9, and NF-κB p65, increased time for new object recognition, tightly arranged and uniformly stained hippocampal cells with clear boundaries, with also an increased number of active neurons and enhanced expression of IL-4 in the hippocampus., Conclusion: EP alleviates post-stroke spasticity in rats by inhibiting inflammatory responses and hippocampal neuronal apoptosis mediated by the NF-κB/NLRP3 signaling pathway.

Background: Post-stroke cognitive impairment (PSCI) severely reduces quality of life of patients with stroke. This study aimed to assess the effects of electroacupuncture (EA) on PSCI and the role of the mTOR/NLRP3-mediated autophagy-inflammatory pathway in this process., Methods: The rat focal cerebral ischemia model was established using middle cerebral artery occlusion (MCAO). Following successful induction of the model, EA was applied to the bilateral Fengchi, Fengfu, and Dazhui acupoints, and brain tissue samples were collected on day 15. Cognitive function was assessed using the Morris water maze test. Cerebral infarct volume was quantified by Triphenyltetrazolium chloride (TTC) staining. Hematoxylin-eosin and TUNEL staining were performed to evaluate pathological changes and apoptosis rates. Apoptosis-, inflammation-, and autophagy-related biomarkers were measured, and autophagosomes were visualized using transmission electron microscopy., Results: MCAO rats exhibited slower weight gain, reduced mobility, increased infarct size, pathological damage, and apoptosis, confirming successful establishment of the MCAO rat model. Following EA treatment, MCAO rats displayed faster weight gain, improved mobility, and shorter escape latency. EA also reduced the area of cerebral infarction and alleviated pathological damage and apoptosis in MCAO rats. Furthermore, EA downregulated IL-1β, IL-18, NLRP3, and LC3 II/LC3 I expression and upregulated p62, mTOR, and Beclin-1 expression in MCAO rats. EA treatment also decreased the number of autophagosomes in these rats., Conclusions: EA effectively mitigates post-stroke cognitive impairment by reducing apoptosis, inflammation, and autophagy through the regulation of the mTOR/NLRP3-mediated autophagy-inflammatory pathway, offering valuable therapeutic insights for stroke rehabilitation., (© 2024. The Author(s).)

Endoplasmic reticulum stress (ERS) and ferroptosis are linked to cerebral ischemia reperfusion injury (CIRI). The neuroprotective properties of 1α, 25-dihydroxyvitamin D3 (VitD3 or 1,25-D3) have been well established; however, the mechanism by which VitD3 treats CIRI through ERS and ferroptosis has not been examined. Hence, we developed middle cerebral artery occlusion/reperfusion (MCAO/R) model in SD rats to ascertain if VitD3 preconditioning mediates ERS and ferroptosis involving of p53 signaling. In this study, we observed that VitD3 can reduce infarction volume and cerebral edema, which leads to the improvement of nerve function. HE, Nissl and Tunel staining showed that VitD3 treatment significantly improved the morphology of neuronal cells and reduced their death. The expression and activation of Vitamin D receptor (VDR), PKR-like ER kinase (PERK), C/EBP-homologous protein (CHOP), p53, nuclear factor erythroid 2-related factor 2 (Nrf2), glutathione peroxidase 4 (GPX4) and reactive oxygen species (ROS) in the ischemic penumbral area were detected by real-time qPCR, Western-blotting and Elisa. The results showed that after VitD3 treatment, VDR increased, ERS-related indices (PERK, CHOP) significantly decreased and ferroptosis-related indices (Nrf2, GPX4) increased. As a VDRs antagonist, pyridoxal-5-phosphate (P5P) can partially block the neuroprotective effects of VitD3. Therefore, CIRI can induce ERS and ferroptosis in the ischemic penumbra area and VitD3 may ameliorate nerve damage in CIRI rats by up-regulating VDR, alleviating p53-associated ERS and ferroptosis., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Vasogenic brain edema, a potentially life-threatening consequence following an acute ischemic stroke, is a major clinical problem. This research aims to explore the therapeutic benefits of nimodipine, a calcium channel blocker, in mitigating vasogenic cerebral edema and preserving blood-brain barrier (BBB) function in an ischemic stroke rat model. In this research, animals underwent the induction of ischemic stroke via a 60-min blockage of the middle cerebral artery and treated with a nonhypotensive dose of nimodipine (1 mg/kg/day) for a duration of five days. The wet/dry method was employed to identify cerebral edema, and the Evans blue dye extravasation technique was used to assess the permeability of the BBB. Furthermore, immunofluorescence staining was utilized to assess the protein expression levels of matrix metalloproteinase-9 (MMP-9) and intercellular adhesion molecule-1 (ICAM-1). The study also examined mitochondrial function by evaluating mitochondrial swelling, succinate dehydrogenase (SDH) activity, the collapse of mitochondrial membrane potential (MMP), and the generation of reactive oxygen species (ROS). Post-stroke administration of nimodipine led to a significant decrease in cerebral edema and maintained the integrity of the BBB. The protective effects observed were associated with a reduction in cell apoptosis as well as decreased expression of MMP-9 and ICAM-1. Furthermore, nimodipine was observed to reduce mitochondrial swelling and ROS levels while simultaneously restoring MMP and SDH activity. These results suggest that nimodipine may reduce cerebral edema and BBB breakdown caused by ischemia/reperfusion. This effect is potentially mediated through the reduction of MMP-9 and ICAM-1 levels and the enhancement of mitochondrial function., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Following ischemic stroke, substance P (SP)-mediated neurogenic inflammation is associated with profound blood-brain barrier (BBB) dysfunction, cerebral edema, and elevated intracranial pressure (ICP). SP elicits its effects by binding the neurokinin 1 tachykinin receptor (NK1-R), with administration of an NK1-R antagonist shown to ameliorate BBB dysfunction and cerebral edema in rodent and permanent ovine stroke models. Given the importance of reperfusion in clinical stroke, this study examined the efficacy of NK1-R antagonist treatment in reducing cerebral edema and ICP in an ovine model of transient middle cerebral artery occlusion (tMCAo). Anesthetized sheep ( n =  24) were subject to 2-hours tMCAo and randomized ( n =  6/group) to receive early NK1-R treatment (days 1-3 post-stroke), delayed NK1-R treatment (day 5 post-stroke), or saline vehicle. At 6-days post-stroke animals were re-anaesthetized and ICP measured, followed by MRI to evaluate infarction, edema and BBB dysfunction. Following both early and delayed NK1-R antagonist administration, ICP was significantly reduced on day 6 compared to vehicle animals (p < 0.05), accompanied by a reduction in cerebral edema, midline shift and BBB dysfunction (p < 0.05). This study demonstrates that NK1-R antagonist treatment is an effective novel therapy for cerebral edema and elevated ICP following stroke in an ovine model, warranting future clinical evaluation., Competing Interests: Declaration of conflicting interestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: The NK1-R antagonist drug (EU-C-001) was kindly donated by PresSura Neuro, however, they had no input in the experimental design, conduct of the study, nor data analysis.

Calpain and PARP-NF-κB signaling are reported to participate in the ischemic brain injury. In this study, it was investigated whether calpain was contributed to the neurovascular unit (NVU) damage through up-regulating PARP-NF-κB signaling during experimental ischemic stroke. Male Sprague-Dawley rats were suffered from 90 min of middle cerebral artery occlusion, followed by reperfusion. The NVU damage was evaluated by the permeability of blood-brain barrier (BBB), the degradation of proteins in extracellular matrix and tight junctions, and ultrastructural changes. The inflammatory response was determined by the expression of inflammatory genes driven by PARP-NF-κB signaling and the activities of myeloperoxidase (MPO). Treatment with MDL 28,170, a calpain inhibitor, improved neurological functions, reduced TUNEL staining index, lessened brain swelling, and decreased infarct volume in ischemic rats. Moreover, it reduced the BBB permeability, enhanced the levels of laminin, collagen IV and occludin, and attenuated the ultrastructural damage of NVU in penumbra and core after induction of ischemia. Meanwhile, it enhanced the levels of cytosolic IκBα, lessened the levels of nuclear PARP and NF-κB p65, reduced the levels of ICAM-1, TNF-α, IL-1β, MMP-9, and MMP-2,and suppressed the activities of MPO in penumbra and core. These data showed that calpain inhibition suppressed PARP-NF-κB signaling-mediated inflammatory response, reduced NVU damage, and protected brain against ischemic stroke, suggesting the involvement of calpain in the NVU damage through up-regulating PARP-NF-κB signaling during brain ischemia., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Acid sphingomyelinase (ASM) inhibitors are widely used for the treatment of post-stroke depression. They promote neurological recovery in animal stroke models via neurorestorative effects. In a previous study, we found that antidepressants including amitriptyline, fluoxetine, and desipramine increase cerebral angiogenesis post-ischemia/reperfusion (I/R) in an ASM-dependent way. To elucidate the underlying mechanisms, we investigated the effects of the functional ASM inhibitor amitriptyline in two models of I/R injury, that is, in human cerebral microvascular endothelial hCMEC/D3 cells exposed to oxygen-glucose deprivation and in mice exposed to middle cerebral artery occlusion (MCAO). In addition to our earlier studies, we now show that amitriptyline increased mitochondrial reactive oxygen species (ROS) formation in hCMEC/D3 cells and increased ROS formation in the vascular compartment of MCAO mice. ROS formation was instrumental for amitriptyline's angiogenic effects. ROS formation did not result in excessive endothelial injury. Instead, amitriptyline induced a profound metabolic reprogramming of endothelial cells that comprised reduced endothelial proliferation, reduced mitochondrial energy metabolism, reduced endoplasmic reticulum stress, increased autophagy/mitophagy, stimulation of antioxidant responses and inhibition of apoptotic cell death. Specifically, the antioxidant heme oxygenase-1, which was upregulated by amitriptyline, mediated amitriptyline's angiogenic effects. Thus, heme oxygenase-1 knockdown severely compromised angiogenesis and abolished amitriptyline's angiogenic responses. Our data demonstrate that ASM inhibition reregulates a complex network of metabolic and mitochondrial responses post-I/R that contribute to cerebral angiogenesis without compromising endothelial survival., (© 2024. The Author(s).)

Background and Purpose: The pathophysiological features of acute ischemic stroke (AIS) often involve dysfunction of the blood-brain barrier (BBB), characterized by the degradation of tight junction proteins (Tjs) leading to increased permeability. This dysfunction can exacerbate cerebral injury and contribute to severe complications. The permeability of the BBB fluctuates during different stages of AIS and is influenced by various factors. Developing effective therapies to restore BBB function remains a significant challenge in AIS treatment. High levels of vascular endothelial growth factor (VEGF) in the early stages of AIS have been shown to worsen BBB breakdown and stroke progression. Our study aimed to investigate the protective effects of the VEGF receptor inhibitor Axitinib on BBB dysfunction and cerebral ischemia/reperfusion-induced injury., Methods: BEnd3 cell exposed to oxygen-glucose deprivation (OGD) model was constructed to estimate pharmacological activity of Axitinib (400 ng/ml) on anti-apoptosis and pathological barrier function recovery. In vivo, rats were subjected to a 1 h transient middle cerebral artery occlusion and 23 h reperfusion (tMCAO/R) to investigate the permeability of BBB and cerebral tissue damage. Axitinib was administered through the tail vein at the beginning of reperfusion. BBB integrity was assessed by Evans blue leakage and the expression levels of Tjs claudin-5 and occludin., Results: Our research revealed that co-incubation with Axitinib enhanced the cell viability of OGD-insulted bEnd3 cells, decreased LDH leakage rate, and suppressed the expression of apoptosis-related proteins cytochrome C and Bax. Axitinib also mitigated the damage to Tjs and facilitated the restoration of transepithelial electrical resistance in OGD-insulted bEnd.3 cells. In vivo, Axitinib administration reduced intracerebral Evans blue leakage and up-regulated the expression of Tjs in the penumbra brain tissue in tMCAO/R rats. Notably, 10 mg/kg Axitinib exerted a significant anti-ischemic effect by decreasing cerebral infarct volume and brain edema volume, improving neurological function, and reducing pro-inflammatory cytokines IL-6 and TNF-α in the brain., Conclusions: Our study highlights Axitinib as a potent protectant of blood-brain barrier function, capable of promoting pathological blood-brain barrier recovery through VEGF inhibition and increased expression of tight junction proteins in AIS. This suggests that VEGF antagonism within the first 24 h post-stroke could be a novel therapeutic approach to enhance blood-brain barrier function and mitigate ischemia-reperfusion injury., Competing Interests: Declaration of competing interest All the authors have approved the manuscript and agree with submission to Experimental Neurology. There are no conflicts of interest to declare., (Copyright © 2024 Elsevier Inc. All rights reserved.)

This research investigated the preventive effects of myrtenol (MYR), fatty acid nanocarriers (FANC), and myrtenol-loaded FANC (MYR + FANC) on neurological disturbance, stroke volume, the levels of malondialdehyde (MDA), superoxide dismutase (SOD), and tumor necrosis factor-alpha (TNF-α) in the brain with ischemia-reperfusion injuries induced by middle cerebral artery occlusion (MCAO) in rats. Seventy two Wistar male rats were divided into six main groups. The groups were sham, ischemia-reperfusion group (MACO), MACO-MYR (50 mg/kg), MACO-FANC (50 and 100 mg/kg), and MACO-MYR + FANC (50 mg/kg). Stroke volume, neurological deficit scores, and the brain levels of MDA, SOD, and TNF-α were examined with TTC staining, observation, and ELISA, respectively. Pretreatment with MYR, FANC (100 mg/kg), and MYR + FANC reduced the neurological deficit score and cerebral infarction volume. MYR, FANC (100 mg/kg), and MYR + FANC pretreatment increased and decreased brain SOD and MDA levels compared to MACO group, respectively. The TNF-α level decreased in the MYR + FANC group compared to MCAO and MCAO-MYR groups in the brain. The use of FANC (100 mg/kg), MYR, and MYR + FANC has protective effects against oxidative stress and ischemia-reperfusion injury. FANC probably improve the bioavailability of MYR, as MYR+ FANC had more therapeutic effects on the reduction of ischemia-reperfusion injuries, inflammation, and oxidative stress., (© 2024 John Wiley & Sons Ltd.)

Accurate assessment of post-stroke deficits is crucial in translational research. Recent advances in machine learning offer precise quantification of rodent motor behavior post-stroke, yet detecting lesion-specific upper extremity deficits remains unclear. Employing proximal middle cerebral artery occlusion (MCAO) and cortical photothrombosis (PT) in mice, we assessed post-stroke impairments via the Staircase test. Lesion locations were identified using 7 T-MRI. Machine learning was applied to reconstruct forepaw kinematic trajectories and feature analysis was achieved with MouseReach , a new data-processing toolbox. Lesion reconstructions pinpointed ischemic centers in the striatum (MCAO) and sensorimotor cortex (PT). Pellet retrieval alterations were observed, but were unrelated to overall stroke volume. Instead, forepaw slips and relative reaching success correlated with increasing cortical lesion size in both models. Striatal lesion size after MCAO was associated with prolonged reach durations that occurred with delayed symptom onset. Further analysis on the impact of selective serotonin reuptake inhibitors in the PT model revealed no clear treatment effects but replicated strong effect sizes of slips for post-stroke deficit detection. In summary, refined movement analysis unveiled specific deficits in two widely-used mouse stroke models, emphasizing the value of deep behavioral profiling in preclinical stroke research to enhance model validity for clinical translation., Competing Interests: Declaration of conflicting interestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: ME reports grants from Bayer and fees paid to the Charité from Abbot, Amgen, AstraZeneca, Bayer, Boehringer Ingelheim, BMS, Daiichi Sankyo, Sanofi, Novartis, Pfizer, all outside the submitted work. All other authors report no conflict of interest.

Background: Stroke is a type of acute brain damage that can lead to a series of serious public health challenges. Demonstrating the molecular mechanism of stroke-related neural cell degeneration could help identify a more efficient treatment for stroke patients. Further elucidation of factors that regulate microglia and nuclear factor (erythroid-derived 2)-like 1 (Nrf1) may lead to a promising strategy for treating neuroinflammation after ischaemic stroke. In this study, we investigated the possible role of pterostilbene (PTS) in Nrf1 regulation in cell and animal models of ischaemia stroke., Methods: We administered PTS, ITSA1 (an HDAC activator) and RGFP966 (a selective HDAC3 inhibitor) in a mouse model of middle cerebral artery occlusion-reperfusion (MCAO/R) and a model of microglial oxygen‒glucose deprivation/reperfusion (OGD/R). The brain infarct size, neuroinflammation and microglial availability were also determined. Dual-luciferase reporter, Nrf1 protein stability and co-immunoprecipitation assays were conducted to analyse histone deacetylase 3 (HDAC3)/Nrf1-regulated Nrf1 in an OGD/R-induced microglial injury model., Results: We found that PTS decreased HDAC3 expression and activity, increased Nrf1 acetylation in the cell nucleus and inhibited the interaction of Nrf1 with p65 and p65 accumulation, which reduced infarct volume and neuroinflammation (iNOS/Arg1, TNF-α and IL-1β levels) after ischaemic stroke. Furthermore, the CSF1R inhibitor PLX5622 induced elimination of microglia and attenuated the therapeutic effect of PTS following MCAO/R. In the OGD/R model, PTS relieved OGD/R-induced microglial injury and TNF-α and IL-1β release, which were dependent on Nrf1 acetylation through the upregulation of HDAC3/Nrf1 signalling in microglia. However, the K105R or/and K139R mutants of Nrf1 counteracted the impact of PTS in the OGD/R-induced microglial injury model, which indicates that PTS treatment might be a promising strategy for ischaemia stroke therapy., Conclusion: The HDAC3/Nrf1 pathway regulates the stability and function of Nrf1 in microglial activation and neuroinflammation, which may depend on the acetylation of the lysine 105 and 139 residues in Nrf1. This mechanism was first identified as a potential regulatory mechanism of PTS-based neuroprotection in our research, which may provide new insight into further translational applications of natural products such as PTS., (© 2024. The Author(s).)

Introduction: Mangiferin is a Chinese herbal extract with multiple biological activities. Mangiferin can penetrate the blood‒brain barrier and has potential in the treatment of nervous system diseases. These findings suggest that mangiferin protects the neurological function in ischemic stroke rats by targeting multiple signaling pathways. However, little is known about the effect and mechanism of mangiferin in alleviating poststroke cognitive impairment., Methods: Cerebral ischemia/reperfusion (I/R) rats were generated via middle cerebral artery occlusion. Laser speckle imaging was used to monitor the cerebral blood flow. The I/R rats were intraperitoneally (i.p.) injected with 40 mg/kg mangiferin for 7 consecutive days. Neurological scoring, and TTC staining were performed to evaluate neurological function. Behavioral experiments, including the open field test, elevated plus maze, sucrose preference test, and novel object recognition test, were performed to evaluate cognitive function. Metabolomic data from brain tissue with multivariate statistics were analyzed by gas chromatography‒mass spectrometry and liquid chromatography‒mass spectrometry., Results: Mangiferin markedly decreased neurological scores, and reduced infarct areas. Mangiferin significantly attenuated anxiety-like and depression-like behaviors and enhanced learning and memory in I/R rats. According to the metabolomics results, 13 metabolites were identified to be potentially regulated by mangiferin, and the differentially abundant metabolites were mainly involved in lipid metabolism., Conclusions: Mangiferin protected neurological function and relieved poststroke cognitive impairment by improving lipid metabolism abnormalities in I/R rats., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Stroke is a globally dangerous disease capable of causing irreversible neuronal damage with limited therapeutic options. Meldonium, an inhibitor of carnitine-dependent metabolism, is considered an anti-ischemic drug. However, the mechanisms through which meldonium improves ischemic injury and its potential to protect neurons remain largely unknown., Methods: A rat model with middle cerebral artery occlusion (MCAO) was used to investigate meldonium's neuroprotective efficacy in vivo. Infarct volume, neurological deficit score, histopathology, neuronal apoptosis, motor function, morphological alteration and antioxidant capacity were explored via 2,3,5-Triphenyltetrazolium chloride staining, Longa scoring method, hematoxylin and eosin staining, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay, rotarod test, transmission electron microscopy and Oxidative stress index related kit. A primary rat hippocampal neuron model subjected to oxygen-glucose deprivation reperfusion was used to study meldonium's protective ability in vitro. Neuronal viability, mitochondrial membrane potential, mitochondrial morphology, respiratory function, ATP production, and its potential mechanism were assayed by MTT cell proliferation and cytotoxicity assay kit, cell-permeant MitoTracker ® probes, mitochondrial stress, real-time ATP rate and western blotting., Results: Meldonium markedly reduced the infarct size, improved neurological function and motor ability, and inhibited neuronal apoptosis in vivo. Meldonium enhanced the morphology, antioxidant capacity, and ATP production of mitochondria and inhibited the opening of the mitochondrial permeability transition pore in the cerebral cortex and hippocampus during cerebral ischemia-reperfusion injury (CIRI) in rats. Additionally, meldonium improved the damaged fusion process and respiratory function of neuronal mitochondria in vitro. Further investigation revealed that meldonium activated the Akt/GSK-3β signaling pathway to inhibit mitochondria-dependent neuronal apoptosis., Conclusion: Our study demonstrated that meldonium shows a neuroprotective function during CIRI by preserving the mitochondrial function, thus prevented neurons from apoptosis., (© 2024. The Author(s).)

Background: Clinically, ischemic reperfusion injury is the main cause of stroke injury. This study aimed to assess the effectiveness of fingolimod in suppressing inflammation caused by ischemic brain injury and explore its pharmacological mechanisms., Methods: In total, 75 male Sprague-Dawley rats were randomly and equally assigned to five distinct groups: sham, middle cerebral artery occlusion/reperfusion (MCAO/R) surgery, fingolimod low-dose (F-L), fingolimod medium-dose (F-M), and fingolimod high-dose (F-H). Neurobehavioral tests, 2,3,5-triphenyltetrazolium chloride staining, and the brain tissue drying-wet method were conducted to evaluate neurological impairment, cerebral infarction size, and brain water content. Enzyme-linked immunosorbent assay was employed to quantify pro-inflammatory cytokines interleukin (IL)-1β, IL-6, and tumor necrosis factor-alpha (TNF-α) protein levels. Western blotting and immunohistochemical staining were performed to assess high mobility group box 1 (HMGB1), toll-like receptor 4 (TLR4), and nuclear factor kappa-B p65 (NF-κBp65) levels., Results: Rats in the F-L, F-M, and F-H groups exhibited lower Longa scores, reduced infarction volumes, and decreased brain edema than those in the MCAO/R group. Additionally, the F-L, F-M, and F-H groups exhibited lower serum levels of IL-1β, IL-6, and TNF-α than those of the MCAO/R group. Additionally, F-L, F-M, and F-H treatments resulted in decreased HMGB1, TLR4, and NF-κBp65 protein expression levels in the hippocampus of MCAO/R rats., Conclusions: Fingolimod was found to reduce ischemic brain injury in a dose-dependent manner. Moreover, it was also found to alleviate inflammation following ischemic brain injury via the HMGB1/TLR4/NF‑κB signaling pathway., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). Published by IMR Press.)

The goal of this study is to investigate the role of microbiota-gut-brain axis involved in the protective effect of pair-housing on post-stroke depression (PSD). PSD model was induced by occluding the middle cerebral artery (MCAO) plus restraint stress for four weeks. At three days after MCAO, the mice were restrained 2 h per day. For pair-housing (PH), each mouse was pair housed with a healthy isosexual cohabitor for four weeks. While in the other PH group, their drinking water was replaced with antibiotic water. On day 35 to day 40, anxiety- and depression-like behaviors (sucrose consumption, open field test, forced swim test, and tail-suspension test) were conducted. Results showed pair-housed mice had better performance on anxiety- and depression-like behaviors than the PSD mice, and the richness and diversity of intestinal flora were also improved. However, drinking antibiotic water reversed the effects of pair-housing. Furthermore, pair-housing had an obvious improvement in gut barrier disorder and inflammation caused by PSD. Particularly, they showed significant decreases in CD8 lymphocytes and mRNA levels of pro-inflammatory cytokines (TNF-a, IL-1β and IL-6), while IL-10 mRNA was upregulated. In addition, pair-housing significantly reduced activated microglia and increased Nissl's body in the hippocampus of PSD mice. However, all these improvements were worse in the pair-housed mice administrated with antibiotic water. We conclude that pair-housing significantly improves PSD in association with enhanced functions of microbiota-gut-brain axis, and homeostasis of gut microbiota is indispensable for the protective effect of pair-housing on PSD., Competing Interests: Declaration of competing interest All the authors declared no potential conflicts of interest with respect to the study, authorship, and/or publication of this article., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective: To explore the neuroprotective effects and mechanism of Tanreqing Injection (TRQ) on treating ischemic stroke based on network pharmacology and in vivo experimental validation., Methods: The chemical compounds of TRQ were retrieved based on published data, with targets retrieved from PubChem, Therapeutic Target Database and DrugBank. Network visualization and analysis were performed using Cytoscape, with protein-protein interaction networks derived from the STRING database. Enrichment analysis was performed using Kyoto Encyclopedia of Genes Genomes pathway and Gene Ontology analysis. In in vivo experiments, the middle cerebral artery occlusion (MCAO) model was used. Infarct volume was determined by 2,3,5-triphenyltetrazolium hydrochloride staining and protein expressions were analyzed by Western blot. Molecular docking was performed to predict ligand-receptor interactions., Results: We screened 81 chemical compounds in TRQ and retrieved their therapeutic targets. Of the targets, 116 were therapeutic targets for stroke. The enrichment analysis showed that the apelin signaling pathway was a key pathway for ischemic stroke. Furthermore, in in vivo experiment we found that administering with intraperitoneal injection of 2.5 mL/kg TRQ every 6 h could significantly reduce the infarct volume of MCAO rats (P<0.05). In addition, protein levels of the apelin receptor (APJ)/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway were increased by TRQ (P<0.05). In addition, 41 chemical compounds in TRQ could bind to APJ., Conclusions: The neuroprotective effect of TRQ may be related to the APJ/PI3K/AKT signaling pathway. However, further studies are needed to confirm the findings., (© 2024. The Chinese Journal of Integrated Traditional and Western Medicine Press and Springer-Verlag GmbH Germany, part of Springer Nature.)

Introduction: Calcified arterial cerebral embolism is a rare occurrence among large and medium vessel occlusions causing ischemic stroke and its diagnosis and treatment is a challenge. The sources of calcified embolism might be a calcific atheroma from the aortic arch and carotid artery, but also heart valve disease has been reported in the literature. Calcified embolism is frequently simultaneous on multiple vascular territories. The prognosis of patients is usually poor, including patients treated by using endovascular thrombectomy (EVT) and this diagnosis could be easily missed in the acute phase. In addition, the optimal secondary prevention has not been yet fully stated., Methods: We are presenting two cases of acute stroke due to calcified embolism in the middle cerebral artery (MCA) coming from a complicated carotid atheroma, non-stenosing in the first case (a 49 years old man) and stenosing in the second case (a 71 years old man) without clinical indications to intravenous thrombolysis and/or EVT, extensively investigated in the acute phase and followed-up for over 12 months with a favorable clinical course and the persisting steno-occlusion in the involved MCA. In both cases, antiplatelet treatment and targeting of vascular risk factors were done without recurrences in the follow-up period., Discussion: Cerebral calcified embolism has been reported in 5.9% of cases of acute ischemic stroke in a single center series and only in 1.2% of a large retrospective cohort of EVT-treated patients. In both series the prognosis was poor and only one third of EVT-treated patients had functional independence at 3-months follow-up. The natural history of these subtype of ischemic stroke is relatively poorly understood and both etiological diagnosis and treatment have not yet defined. It is possible that some cases might be underdiagnosed and underreported., Conclusions: Calcified cerebral embolism is a rare cause of stroke, but it is largely underreported and both acute phase and secondary preventive treatment have to be defined., (© 2024. Fondazione Società Italiana di Neurologia.)

Huanglian Jiedu decoction (HLJD) has been used to treat ischemic stroke in clinic. However, the detailed protective mechanisms of HLJD on ischemic stroke have yet to be elucidated. The aim of this study is to elucidate the underlying pharmacological mechanisms of HLJD based on the inhibition of neuroinflammation and the amelioration of nerve cell damage. A middle cerebral artery occlusion reperfusion (MCAO/R) model was established in rats and received HLJD treatment. Effects of HLJD on neurological function was assessed based on Bederson's score, postural reflex test and asymmetry score. 2, 3, 5-Triphenyltetrazolium chloride (TTC) staining, Hematein and eosin (HE) and Nissl staining were used to observe the pathological changes in brain. Then, transcriptomics was used to screen the differential genes in brain tissue in MCAO/R model rats following HLJD intervention. Subsequently, the effects of HLJD on neutrophil extracellular trap (NET) formation-related neuroinflammation, gamma-aminobutyric acid (GABA)ergic synapse activation, nerve cell damage and proliferation were validated using immunofluorescence, western blot and enzyme-linked immunosorbent assay (ELISA). Our results showed that HLJD intervention reduced the Bederson's score, postural reflex test score and asymmetry score in MCAO/R model rats. Pathological staining indicated that HLJD treatment decreased the cerebral infarction area, mitigated neuronal damage and increased the numbers of Nissl bodies. Transcriptomics suggested that HLJD affected 435 genes in MCAO/R rats. Among them, several genes involving in NET formation and GABAergic synapses pathways were dysregulated. Subsequent experimental validation showed that HLJD reduced the MPO + CitH3 + positive expression area, reduced the protein expression of PAD4, p-P38/P38, p-ERK/ERK and decreased the levels of IL-1β, IL-6 and TNF-α, reversed the increase of Iba1 + TLR4 + , Iba1 + p65 + and Iba1 + NLRP3 + positive expression area in brain. Moreover, HLJD increased GABA levels, elevated the protein expression of GABRG1 and GAT3, decreased the TUNEL positive expression area and increased the Ki67 positive expression area in brain. HLJD intervention exerts a multifaceted positive impact on ischemia-induced cerebral injury in MCAO/R rats. This intervention effectively inhibits neuroinflammation by mitigating NET formation, and concurrently improves nerve cell damage and fosters nerve cell proliferation through activating GABAergic synapses., (© 2024 The Author(s). Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Numerous natural bioactive compounds extracted from Chinese medicines have been proved to be promising and potent agents in the treatment of ischemic stroke. Hydroxysafflor yellow A (HSYA), separated from Carthamus tinctorius, has increasingly attracted attention for its broad spectrum of pharmacological effects, especially of its neuroprotective action. Our previous studies revealed that HSYA plays significant beneficial roles in a dose-dependent manner in rats with focal cerebral ischemia. However, treatment with higher doses of HSYA appeared to bring about adverse reactions in the rats. In present study, we adopted tenuigenin (TEN), extracted from the Polygala tenuifolia root, in combination with HSYA to optimize the therapeutic strategy against ischemic stroke, and further explored the underlying mechanisms of action of the combination in vivo and in vitro. We firstly confirmed the pharmacological efficacies of co-treatment of HSYA and TEN in middle cerebral ischemia occlusion (MCAO) rats and observed the synergistic improvement of infarct volume, cerebral edema, and morphology of neuron cell body. Behavioral experiments indicated that combination of HSYA and TEN could synergistically improve motor and cognitive function in MCAO rats. We also observed increased viability and suppressed cell apoptosis after HSYA and TEN co-treatments in the oxygen-glucose deprivation/reperfusion (OGD/R) SH-SY5Y cells. Furthermore, JAK2/STAT3 and SOCS3 signaling interaction was demonstrated to be a critical responsor to the co-treatment of HSYA and TEN. In the subsequent experiments with silencing SOCS3 in OGD/R-exposed cells, we found that HSYA and TEN might suppress JAK2/STAT3 pathway through different regulatory mechanisms targeting SOCS3-negative feedback signaling. HSYA seemed to impose excessive activation of JAK2/STAT3 to trigger SOCS3-negative feedback signaling, while TEN appeared to provoke SOCS3 inhibitory feedback role directly to further attenuate JAK2-mediated signaling. Collectively, HSYA and TEN might modulate the crosstalk between JAK2/STAT3 and SOCS3 signaling pathways in different manners that eventually contributed to their synergistic therapeutic effects against cerebral ischemic stroke., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Microglia mediated neuroinflammation is one of the major contributors to brain damage in cerebral ischemia reperfusion injury (CI/RI). Recently, RNA modification was found to contribute to the regulation of microglia polarization and the subsequent development of cerebral I/R neuroinflammation. Herein, we investigated the effect and mechanism of m5C RNA modification in the microglia induced CI/RI neuroinflammation. We found that the m5C RNA modification levels decreased in the primary microglia isolated from a mouse model of intraluminal middle cerebral artery occlusion/reperfusion (MCAO/R) and the BV2 microglial cells subjected to oxygen-glucose deprivation and reoxygenation (OGD/R), and this change was accompanied by an increase in the M1/M2 polarization ratio. Furthermore, the expression of m5C demethylase TET1 in microglia increased, which promoted M1 polarization but impeded M2 polarization. Mechanistically, the higher TET1 expression decreased the m5C modification level of RelB and enhanced its mRNA stability, which subsequently increased the M1/M2 polarization ratio. In conclusion, this study provides insight into the role of m5C RNA modification in the pathogenesis of cerebral I/R neuroinflammation and may deepen our understanding on clinical therapy targeting the TET1-RelB axis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective: Epilepsy develops in one third of the patients after perinatal stroke. It is still unclear which vascular syndrome of ischemic stroke carries higher risk of epilepsy. The aim of the current study was to evaluate the risk of epilepsy according to the vascular syndrome of perinatal stroke., Methods: The study included 39 children with perinatal arterial ischemic stroke (13 with anterior or posterior trunk of the distal middle cerebral artery occlusion, 23 with proximal or distal M1 middle cerebral artery occlusion and three with lenticulostriate arteria infarction), and 44 children with presumed perinatal venous infarction. Magnetic resonance imaging obtained at the chronic stage was used to evaluate the vascular syndrome of stroke., Results: The median follow-up time was 15.1 years (95% CI: 12.4-16.5 years), epilepsy developed in 19/83 (22.9%) patients. The cumulative probability to be without epilepsy at 15 years was 75.4% (95% CI: 65.8-86.4). The probability of having epilepsy was higher in the group of proximal or distal M1 artery occlusion compared to patients with periventricular venous infarction (HR 7.2, 95% CI: 2.5-26, p = .0007). Patients with periventricular venous infarction had significantly more often status epilepticus or spike-wave activation in sleep ≥85% of it compared to patients with anterior or posterior trunk of the distal middle cerebral artery occlusion (OR = 81; 95% CI: 1.3-5046, p = .029)., Significance: The emphasis of this study is placed on classifying the vascular syndrome of perinatal stroke and on the targeted follow-up of patients for epilepsy until young adulthood. The risk for having epilepsy after perinatal stroke is the highest in children with proximal or distal M1 middle cerebral artery occlusion. Patients with periventricular venous infarction have a more severe course of epilepsy., (© 2024 The Authors. Epileptic Disorders published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Background: This study aimed to investigate whether moxibustion could affect PI3K/Akt pathway to regulate Transforming acidic coiled-coil containing protein 3 (TACC3) and promote axonal regeneration to improve learning and memory function in middle cerebral artery occlusion (MCAO) rats., Methods: Sixty SD rats were randomly divided into 4 groups: sham-operated control group (SC), model control group (MC), model + moxibustion group (MM), and model + inhibitor + moxibustion group (MIM). The rats in MC, MM, and MIM groups were made into MCAO models, and PI3K inhibitor LY294002 was injected into the rats in MIM group before modeling; while the rats in SC group were only treated with artery separation without monofilament inserting. After that, the rats in MM and MIM groups were intervented with moxibustion. We used the Zea-Longa scale, micro-Magnetic Resonance Imaging (micro-MRI), Morris water maze (MWM), TUNEL, western blot (WB), immunofluorescence and immunohistochemistry to evaluate the neurological deficits, cerebral infarct volume, learning and memory, apoptotic cell percentage in the hippocampal, the expression level of axonal regeneration and PI3K/AKt related proteins, the expression level of TACC3. The detection of 2 h after surgery showed the result before moxibustion and 7 days after the intervention showed the results after moxibustion., Results: After 7 d of intervention, the scores of Zea-Longa and the cerebral infarct volume, the escape latency, the percentage of apoptosis cells of MM group were lower than that of MC and MIM groups; the frequency of rats crossed the previous platform location, PI3K, p-Akt/t-Akt and TACC3, the level of GAP-43 in MM group was more than MC and MIM groups (P < 0.05). While no statistical difference existed between MIM group and MC group (P > 0.05)., Conclusion: Moxibustion can promote axonal regeneration and improve learning and memory of Post-stroke cognitive impairment via activating the PI3K/AKT signaling pathway and TACC3., (Copyright © 2024 IBRO. Published by Elsevier Inc. All rights reserved.)

Aims: Infection is a complication after stroke and outcomes vary by sex. Thus, we investigated if sepsis affects brain from ischemic stroke and sex involvement., Main Methods: Male and female Wistar rats, were submitted to middle cerebral artery occlusion (MCAO) and after 7 days sepsis to cecal ligation and perforation (CLP). Infarct size, neuroinflammation, oxidative stress, and mitochondrial activity were quantified 24 h after CLP in the prefrontal cortex and hippocampus. Survival and neurological score were assessed up to 15 days after MCAO or 8 days after CLP (starting at 2 h after MCAO) and memory at the end., Key Findings: CLP decreased survival, increased neurological impairments in MCAO females. Early, in male sepsis following MCAO led to increased glial activation in the brain structures, and increased TNF-α and IL-1β in the hippocampus. All groups had higher IL-6 in both tissues, but the hippocampus had lower IL-10. CLP potentiated myeloperoxidase (MPO) in the prefrontal cortex of MCAO male and female. In MCAO+CLP, only male increased MPO and nitrite/nitrate in hippocampus. Males in all groups had protein oxidation in the prefrontal cortex, but only MCAO+CLP in the hippocampus. Catalase decreased in the prefrontal cortex and hippocampus of all males and females, and MCAO+CLP only increased this activity in males. Female MCAO+CLP had higher prefrontal cortex complex activity than males. In MCAO+CLP-induced long-term memory impairment only in females., Significance: The parameters evaluated for early sepsis after ischemic stroke show a worse outcome for males, while females are affected during long-term follow-up., Competing Interests: Declaration of competing interest The authors declare no competing interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Recent studies have highlighted the involvement of pyroptosis-mediated cell death and neuroinflammation in ischemic stroke (IS) pathogenesis. DL-3-n-butylphthalide (NBP), a synthesized compound based on an extract from seeds of Apium graveolens, possesses a broad range of biological effects. However, the efficacy and the underlying mechanisms of NBP in IS remain contentious. Herein, we investigated the therapeutic effects of NBP and elucidated its potential mechanisms in neuronal cell pyroptosis and microglia inflammatory responses. Adult male mice underwent permanent distal middle cerebral artery occlusion (dMCAO), followed by daily oral gavage of NBP (80 mg/kg) for 1, 7, or 21 consecutive days. Gene Expression Omnibus (GEO) dataset of IS patients peripheral blood RNA sequencing was analyzed to identify differentially expressed pyroptosis-related genes (PRGs) during the ischemic process. Our results suggested that NBP treatment effectively alleviated brain ischemic damage, resulting in decreased neurological deficit scores, reduced infarct volume, and improved neurological and behavioral functions. RNA sequence data from human unveiled upregulated PRGs in IS. Subsequently, we observed that NBP downregulated pyroptosis-associated markers at days 7 and 21 post-modeling, at both the protein and mRNA levels. Additionally, NBP suppressed the co-localization of pyroptosis markers with neuronal cells to variable degrees and simultaneously mitigated the accumulation of activated microglia. Overall, our data provide novel evidence that NBP treatment significantly attenuates ischemic brain damage and promotes recovery of neurological function in the early and recovery phases after IS, probably by negatively regulating the pyroptosis cell death of neuronal cells and inhibiting toxic neuroinflammation in the central nervous system., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Angiogenesis is crucial in neuroprotection of secondary thalamic injury after cortical infarction. The p75 neurotrophin receptor (p75 NTR ) plays a key role in activating angiogenesis. However, the effects of p75 NTR on angiogenesis in the thalamus after cortical infarction are largely unknown. Herein we investigate whether p75 NTR facilitates angiogenesis to attenuate secondary thalamic damage via activating hypoxia-inducible factor 1α (HIF-1α)/vascular endothelial growth factor (VEGF) pathway mediated by Von Hippel-Lindau (VHL) after distal middle cerebral artery occlusion (dMCAO)., Methods: The male rat model of dMCAO was established. The effects of p75 NTR on the angiogenesis was evaluated using RNA-sequencing, immunohistochemistry, western blot, quantitative real-time polymerase chain reaction, magnetic resonance imaging, behavior tests, viral and pharmacological interventions., Results: We found that the p75 NTR and vessel density were decreased in ipsilateral thalamus after dMCAO. The p75 NTR -VHL interaction was reduced, which promoted the ubiquitination degradation of HIF-1α and reduced VEGF expression after dMCAO. Notably, p75 NTR overexpression restrained the ubiquitination degradation of HIF-1α by inhibiting VHL-HIF-1α interaction, further promoted angiogenesis, increased cerebral blood flow of ipsilateral thalamus and improved neurological function after dMCAO., Conclusion: For the first time, we highlighted that the enhancement of p75 NTR -VHL interaction promoted angiogenesis in attenuating secondary thalamic damage after dMCAO., (© 2024 The Author(s). CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.)

Background: The inflammatory response is a key factor in the pathogenesis of cerebral ischemia/reperfusion injury (CIRI), and anti-inflammatory interventions may offer a promising therapeutic strategy. Forsythoside B (FB) is a phenylethanoid glycoside isolated from Forsythiae fructus, which has been reported to have anti-inflammatory effects. However, the mechanism of the neuroprotective effect of FB on CIRI remains unclear., Methods: Adult male Sprague-Dawley rats were subjected to transient middle cerebral artery occlusion/reperfusion (MCAO/R). FB was administered intraperitoneally for 3 days prior to MCAO/R. Cerebral infarct volume and neurological deficit score were used as indices to evaluate MCAO/R injury. The serum levels of inflammatory factors and antioxidant enzymes were measured. The activation of silent information regulator 2 homolog 1 (Sirt1) and the inhibition of the nucleotide-binding oligomerization domain-like receptor with a pyrin domain 3 (NLRP3) pathway were assessed through western blot and immunohistochemistry analysis. Furthermore, the rats were treated with Sirt1 shRNA 3 days before MCAO/R by stereotactical injection into the ipsilateral hemispheric region to assess the impact of Sirt1 knockdown on the protection of FB during MCAO/R., Results: FB reduced cerebral infarct volume and neurological deficit score in MCAO/R rats. FB reduced pathological changes and cell apoptosis in the hippocampal CA1 region and cortex on the ischemic side of rats. FB inhibited the serum levels of inflammatory factors and increased the activities of antioxidant enzymes. Further study showed that FB inhibited the activation of the NLRP3 pathway and induced Sirt1 activation., Conclusion: FB demonstrated neuroprotective and anti-inflammatory effects by inhibiting the NLRP3 pathway through Sirt1 activation in CIRI., Competing Interests: The authors declare that there are no conficts of interest, (Copyright: © 2024 Li et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Purpose: Post-stroke cognitive impairment (PSCI) is a common complication of ischemic stroke episodes. Memory impairment is an important component of the poststroke cognitive syndrome. Microglial activation plays a critical role in stroke-induced neuroinflammation. Previous studies have reported that electroacupuncture (EA) provides neuroprotective effects by reducing the expression levels of the Purinergic receptor P2X ligand-gated ion channel 7 (P2X7) and inhibiting neuroinflammation in rat model of ischemic stroke. Further understanding of the role and connections between P2X7R and microglial activation in EA-induced anti-inflammatory can reveal novel targets for post-stroke memory impairment treatment., Methods: A Middle cerebral artery occlusion and reperfusion (MCAO/R) model was established. We used 2'(3')-O-(4-benzoyl) benzoyl ATP (BzATP) as a P2X7R agonist. Following MCAO/R injury, the rats underwent EA therapy at the Baihui (DU20) and Shenting (DU24) acupoints for seven consecutive days. The Barnes maze test was used to evaluate memory function. Following intervention, a T2 weighted images (T2WI) scan was performed to identify changes in cerebral infarction volume in MCAO/R rats. The levels of Interleukin-1β (IL-1β), Interleukin-6 (IL-6) and Interleukin-4 (IL-4), Interleukin-10 (IL-10) in the peri-infarct hippocampal were examined by ELISA. Immunofluorescence was employed to evaluate Iba-1 + / P2X7R + , Iba-1 + / iNOS + and Iba-1 + / Arg-1 + cell populations in the peri-infarct hippocampal DG area. The protein expression of P2X7R, Nuclear factor E2-related factor 2 (Nrf2), Recombinant nlr family, pyrin domain containing protein 3 (NLRP3), Inducible nitric oxide synthase (iNOS) and Arginase-1 (Arg-1) in the peri-infarct hippocampal were investigated using western blot assays. Besides, we also measured the levels of reactive oxygen species (ROS), superoxide dismutase (SOD) and malondialdehyde (MDA)., Results: We found EA treatment reduced inflammation and oxidative stress, which is consistent with a decrease in P2X7R expression and improved learning and memory functions. In contrast, we found BzATP enhanced inflammation and oxidative stress. Moreover, our results showed EA treatment up-regulated Nrf2, down-regulated NLRP3, and promoted microglia M2 polarization. Finally, EA-mediated positive effects were reversed by intracerebroventricular injection of BzATP, which is consistent with an increase in P2X7R expression., Conclusion: EA ameliorates memory impairment in a rat model of ischemic stroke by reducing inflammation and ROS through the inhibition of P2X7R expression. In turn, this mechanism regulates Nrf2 and NLRP3 expression, suggesting EA is beneficial for ischemic stroke treatment using P2X7R as target., Competing Interests: Declaration of Competing Interest The authors declare the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Currently there is no effective treatment for neonatal stroke, an acute neurologic syndrome with sequelae, due to focal ischemic, thrombotic, or hemorrhagic event occurring in the perinatal period. VCE-004.8, an aminoquinone exhibiting activity on CB2 and PPARγ receptors, is neuroprotective in adult mice models of acute and chronic brain damaging conditions. We hereby aimed to study VCE-004.8 neuroprotection in a rat model of neonatal stroke., Methods: 7-day-old (P7) Wistar rats of both sexes were submitted to Middle Cerebral Artery Occlusion (MCAO), receiving i.p. 30 min after vehicle (MCAO + VEH) or VCE-004.8 5 mg/kg (MCAO + VCE). Non-occluded rats served as controls (SHAM). MCAO consequences were assessed at P14 by MRI, histological (TUNEL staining), biochemical (lactate/n-acetyl aspartate ratio by 1H-NMR spectroscopy) and motor studies (grasp test), and at P37 assessing myelination (MBP signal), hemiparesis and hyperlocomotion. Effects of VCE-004.8 on excitotoxicity (glutamate/n-acetyl aspartate, 1H-NMR), oxidative stress (protein nitrosylation, Oxyblot) and neuroinflammation (Toll-like receptor 4 and TNFa expression, Western blot) were assessed at P14. Therapeutic window was assessed by delaying drug administration for 12 or 18 h., Results: Post-MCAO administration of VCE-004.8 reduced the volume of infarct and histological and biochemical brain damage, reducing hyperlocomotion, restoring motor performance and preserving myelination, in a manner linked to the modulation of excitotoxicity, oxidative stress and neuroinflammation. VCE-004.8 was still effective being administered 12-18 h post-insult., Conclusions: These data suggest that this drug could be effective for the treatment of stroke in newborns., Competing Interests: Declaration of competing interest José Martinez-Orgado had a Research Agreement with Emerald Health Biotechnology España S.L.U. (Córdoba, Spain). The other authors have no relevant financial or non-financial interests to disclose. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Poststroke cognitive impairment (PSCI) is a common complication of stroke, but effective treatments are currently lacking. Repetitive transcranial magnetic stimulation (rTMS) is gradually being applied to treat PSCI, but there is limited evidence of its efficacy. To determine rTMS effects on PSCI, we constructed a transient middle cerebral artery occlusion (tMCAO) rat model. Rats were then grouped by random digital table method: the sham group (n = 10), tMCAO group (n = 10) and rTMS group (n = 10). The shuttle box and Morris water maze (MWM) tests were conducted to detect the cognitive functions of the rats. In addition, synaptic density and synaptic ultrastructural parameters, including the active zone length, synaptic cleft width, and postsynaptic density (PSD) thickness, were quantified and analyzed using an electron microscope. What's more, synaptic associated proteins, including PSD95, SYN, and BDNF were detected by western blot. According to the shuttle box and MWM tests, rTMS improved tMCAO rats' cognitive functions, including spatial learning and memory and decision-making abilities. Electron microscopy revealed that rTMS significantly increased the synaptic density, synaptic active zone length and PSD thickness and decreased the synaptic cleft width. The western blot results showed that the expression of PSD95, SYN, and BDNF was markedly increased after rTMS stimulation. Based on these results, we propose that 20 Hz rTMS can significantly alleviate cognitive impairment after stroke. The underlying mechanism might be modulating the synaptic plasticity and up-regulating the expression PSD95, SYN, and BDNF in the hippocampus., Competing Interests: Declaration of Competing Interest None., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cerebral ischemia-reperfusion injury (CIRI) can induce massive death of ischemic penumbra neurons via oxygen burst, exacerbating brain damage. Parthanatos is a form of caspase-independent cell death involving excessive activation of PARP-1, closely associated with intense oxidative stress following CIRI. 4'-O-methylbavachalcone (MeBavaC), an isoprenylated chalcone component in Fructus Psoraleae, has potential neuroprotective effects. This study primarily investigates whether MeBavaC can act on SIRT3 to alleviate parthanatos of ischemic penumbra neurons induced by CIRI. MeBavaC was oral gavaged to the middle cerebral artery occlusion-reperfusion (MCAO/R) rats after occlusion. The effects of MeBavaC on cerebral injury were detected by the neurological deficit score and cerebral infarct volume. In vitro, PC-12 cells were subjected to oxygen and glucose deprivation/reoxygenation (OGD/R), and assessed cell viability and cell injury. Also, the levels of ROS, mitochondrial membrane potential (MMP), and intracellular Ca 2+ levels were detected to reflect mitochondrial function. We conducted western blotting analyses of proteins involved in parthanatos and related signaling pathways. Finally, the exact mechanism between the neuroprotection of MeBavaC and parthanatos was explored. Our results indicate that MeBavaC reduces the cerebral infarct volume and neurological deficit scores in MCAO/R rats, and inhibits the decreased viability of PC-12 cells induced by OGD/R. MeBavaC also downregulates the expression of parthanatos-related death proteins PARP-1, PAR, and AIF. However, this inhibitory effect is weakened after the use of a SIRT3 inhibitor. In conclusion, the protective effect of MeBavaC against CIRI may be achieved by inhibiting parthanatos of ischemic penumbra neurons through the SIRT3-PARP-1 axis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)