Animal Models Of Ischemic Stroke Research Articles

Tetramethylpyrazine Analogue T-006 Protects Neuronal and Endothelial Cells Against Oxidative Stress via PI3K/AKT/mTOR and Nrf2 Signaling.

T-006, a novel neuroprotective derivative of tetramethylpyrazine (TMP), exhibits multifunctional neuroprotective properties. T-006 has been shown to improve neurological and behavioral functions in animal models of ischemic stroke and neurodegenerative diseases. The present study aims to further elucidate the mechanisms underlying the protective effects of T-006 against oxidative injuries induced by glutamate or hypoxia. Mouse hippocampal HT22 cells were used to evaluate the neuroprotective effects of T-006 against glutamate-induced injuries, while mouse brain endothelial bEnd.3 cells were used to evaluate the cerebrovascular protective effects of T-006 against oxygen-glucose deprivation followed by reperfusion (OGD/R)-induced injuries. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry were used to measure cell viability and oxidative stress. Western blot and immunofluorescence analyses of protein expression were used to study cell signaling pathways. T-006 exhibited significant protective effects in both oxidative injury models. In HT22 cells, T-006 reduced cell death and enhanced antioxidant capacity by upregulating mTOR and nuclear factor erythroid 2-related factor 2/Heme oxygenase-1 (Nrf2/HO-1) signaling. Similarly, in bEnd.3 cells, T-006 reduced oxidative injuries and preserved tight junction integrity through Nrf2/HO-1 upregulation. These effects were inhibited by LY294002, a Phosphoinositide 3-kinase (PI3K) inhibitor. T-006 may exert its neuroprotective and cerebrovascular protective effects via the regulation of PI3K/AKT-mediated pathways, which facilitate downstream mTOR and Nrf2 signaling, leading to improved cell survival and antioxidant defenses.

Antioxidant Effect of Naringin Demonstrated Through a Bayes' Theorem Driven Multidisciplinary Approach Reveals its Prophylactic Potential as a Dietary Supplement for Ischemic Stroke.

Naringin (NAR), a flavanone glycoside, occurs widely in citrus fruits, vegetables, and alcoholic beverages. Despite evidence of the neuroprotective effects of NAR on animal models of ischemic stroke, brain cell-type-specific data about the antioxidant efficacy of NAR and possible protein targets of such beneficial effects are limited. Here, we demonstrate the brain cell type-specific prophylactic role of NAR, an FDA-listed food additive, in an in vitro oxygen-glucose deprivation (OGD) model of cerebral ischemia using MTT and DCFDA assays. Using Bayes' theorem-based predictive model, we first ranked the top-10 protein targets (ALDH2, ACAT1, CTSB, FASN, LDHA, PTGS1, CTSD, LGALS1, TARDBP, and CDK1) from a curated list of 289 NAR-interacting proteins in neurons that might be mediating its antioxidant effect in the OGD model. When preincubated with NAR for 2days, N2a and CTX-TNA2 cells could withstand up to 8h of OGD without a noticeable decrease in cell viability. This cerebroprotective effect is partly mediated by reducing intracellular ROS production in the above two brain cell types. The antioxidant effect of NAR was comparable with the equimolar (50µM) concentration of clinically used ROS-scavenger and neuroprotective edaravone. Molecular docking of NAR with the top-10 protein targets from Bayes' analysis showed the lowest binding energy for CDK1 (- 8.8kcal/M). Molecular dynamics simulation analysis showed that NAR acts by inhibiting CDK1 by stably occupying its ATP-binding cavity. Considering diet has been listed as a risk factor for stroke, NAR may be explored as a component of functional food for stroke or related neurological disorders.

MGlu4R, mGlu7R, and mGlu8R allosteric modulation for treating acute and chronic neurodegenerative disorders.

Neuroprotection, defined as safeguarding neurons from damage and death by inhibiting diverse pathological mechanisms, continues to be a promising approach for managing a range of central nervous system (CNS) disorders, including acute conditions such as ischemic stroke and traumatic brain injury (TBI) and chronic neurodegenerative diseases like Parkinson's disease (PD), Alzheimer's disease (AD), and multiple sclerosis (MS). These pathophysiological conditions involve excessive glutamatergic (Glu) transmission activity, which can lead to excitotoxicity. Inhibiting this excessive Glu transmission has been proposed as a potential therapeutic strategy for treating the CNS disorders mentioned. In particular, ligands of G protein-coupled receptors (GPCRs), including metabotropic glutamatergic receptors (mGluRs), have been recognized as promising options for inhibiting excessive Glu transmission. This review discusses the complex interactions of mGlu receptors with their subtypes, including the formation of homo- and heterodimers, which may vary in function and pharmacology depending on their protomer composition. Understanding these intricate details of mGlu receptor structure and function enhances researchers' ability to develop targeted pharmacological interventions, potentially offering new therapeutic avenues for neurological and psychiatric disorders. This review also summarizes the current knowledge of the neuroprotective potential of ligands targeting group III mGluRs in preclinical cellular (in vitro) and animal (in vivo) models of ischemic stroke, TBI, PD, AD, and MS. In recent years, experiments have shown that compounds, especially those activating mGlu4 or mGlu7 receptors, exhibit protective effects in experimental ischemia models. The discovery of allosteric ligands for specific mGluR subtypes has led to reports suggesting that group III mGluRs may be promising targets for neuroprotective therapy in PD (mGlu4R), TBI (mGlu7R), and MS (mGlu8R).

Histomorphometric analysis of the soleus muscle in different phases of ischemic stroke in an animal model

Objective: To analyze the muscle mass and cross-sectional area of the muscular fiber of the hemiparetic soleus in an animal model, in order to identify the muscular adaptations that occur in ischemic stroke. Method: Twelve Rattus norvegicus were divided into 2 groups: stroke group (n=6) and control group (n=6). Each group was subdivided into two subgroups, with evaluations at 7 days (CG7 and SG7) and 21 days (CG21 and SG21) after the accident. Their soleus muscles were removed for muscle mass analysis and cross-sectional area of the muscular fibers (CSAMF) measurement. The adopted statistical significance was 5%. Result: Significant differences in the muscle mass were observed between CG7 (0.120 ± 0.005 g) vs. SG7 (0.100 ± 0.004 g; p=0.035), and between CG21 (0.130 ± 0.010 g) vs. SG21 (0.078 ± 0.006; p=0.012). Significant differences in the relative muscle mass were observed between CG7 (0.044 ± 0.002 g) vs SG7 (0.039 ± 0.003; p=0.025), and CG21 (0.044 ± 0.003) vs. SG21 (0.028 ± 0.002; p=0.011). The CSAMF showed significant differences between CG7 (2,322 μm2 [2312-2453]) vs. SG7 (2,056 μm2 [2,022-2,135]; p=0.012), and CG21 (2,667 μm2 [2,692-2,845]) vs. SG21 (2,050 μm2 [2,034-2,161]; p=0.006). Conclusion: In this study in animal models of ischemic stroke, there was a significant loss of muscle mass, and this loss was accentuated in the longer term of the injury, highlighting the importance of future research on types of muscle fibers and applicability in human patients.

Panax notoginseng saponins promotes angiogenesis after cerebral ischemia-reperfusion injury

BackgroundIschemic stroke is a devastating disease that can result in permanent disability and death, and angiogenesis plays a critical role in the recovery and survival of patients and animal models of ischemic stroke. Panax notoginseng has been used as a key herb in the treatment of stroke diseases due to its effect in promoting blood circulation and removing blood stasis. However, the role of Panax notoginseng saponins, in promoting angiogenesis is unclear. PurposeThis study is aimed to investigate the effect of Xueshuantong (XST) injection, composed of Panax notoginseng saponins in post-stroke revascularization. MethodIn the present study, a middle cerebral artery occlusion/reperfusion model was established in Sprague-Dawley rats, with XST and the positive drug Dl-3-n-butylphthalide (NBP) administered via intraperitoneal injection to observe vascular changes after stroke. The protective and pro-angiogenic effects of XST after stroke were demonstrated by Triphenyltetrazolium chloride staining and optical coherence tomography angiography. Subsequently, network pharmacology and molecular docking techniques, as well as in vitro experimental validation, were used to further analyze the potential mechanism by which XST promotes angiogenesis. ResultsThe results showed that XST could reduce the cerebral infarction region in rats. And the neovascularization in the ischemic area of the rat brain significantly increased after 7 or 14 days of XST administration. Furthermore, XST could activate the vascular endothelial growth factor A (VEGFA)/vascular endothelial growth factor receptor 2 (VEGFR2), and hypoxia-inducible factor 1 (HIF-1) signaling pathways. ConclusionXST may promote post-stroke angiogenesis by affecting the HIF1-α/VEGFA/VEGFR2 signaling pathways.

A meta-analysis of animal studies evaluating the effect of hydrogen sulfide on ischemic stroke: is the preclinical evidence sufficient to move forward?

Hydrogen sulfide (H2S) is a gasotransmitter that has been studied for its potential therapeutic effects, including its role in the pathophysiology and treatment of stroke. This systematic review and meta-analysis aimed to determine the sufficiency of overall preclinical evidence to guide the initiation of clinical stroke trials with H2S and provide tailored recommendations for their design. PubMed, Web of Science, Scopus, EMBASE, and MEDLINE were searched for studies evaluating the effect of any H2S donor on in vivo animal models of regional ischemic stroke, and 34 publications were identified. Pooling of the effect sizes using the random-effect model revealed that H2S decreased the infarct area by 34.5% (95% confidence interval (CI) 28.2-40.8%, p < 0.0001), with substantial variability among the studies (I2 = 89.8%). H2S also caused a 37.9% reduction in the neurological deficit score (95% CI 29.0-46.8%, p < 0.0001, I2 = 63.8%) and in the brain water content (3.2%, 95% CI 1.4-4.9%, p = 0.0014, I2 = 94.6%). Overall, the studies had a high risk of bias and low quality of evidence (median quality score 5/15, interquartile range 4-9). The majority of the included studies had a "high" or "unclear" risk of bias, and none of the studies overall had a "low" risk. In conclusion, H2S significantly improves structural and functional outcomes in in vivo animal models of ischemic stroke. However, the level of evidence from preclinical studies is not sufficient to proceed to clinical trials due to the low external validity, high risk of bias, and variable design of existing animal studies.

A Systematic Review and Meta-Analysis of Therapeutic Hypothermia and Pharmacological Cotherapies in Animal Models of Ischemic Stroke.

Therapeutic hypothermia (TH) lessens ischemic brain injury. Cytoprotective agents can augment protection, although it is unclear which combinations are most effective. The objective of this study is to identify which cytoprotective drug works best with delayed TH. Following PRISMA guidelines, a systematic review (PubMed, Web of Science, MEDLINE, Scopus) identified controlled experiments that used an invivo focal ischemic stroke model and evaluated the efficacy of TH (delay of ≥1 hour) coupled with cytoprotective agents. This combination was our main intervention compared with single treatments with TH, drug, or no treatment. Endpoints were brain injury and neurological impairment. The CAMARADES checklist for study quality and the SYRCLE's risk of bias tool gauged study quality. Twenty-five studies were included. Most used young, healthy male rats, with only one using spontaneously hypertensive rats. Two studies used mice models, and six used adult animals. Study quality was moderate (median score = 6), and risk of bias was high. Pharmacological agents provided an additive effect on TH for all outcomes measured. Magnesium coupled with TH had the greatest impact compared with other agent-TH combinations on all outcomes. Longer TH durations improved both behavioral and histological outcomes and had greater cytoprotective efficacy than shorter durations. Anti-inflammatories were the most effective in reducing infarction (standardized mean difference [SMD]: -1.64, confidence interval [CI]: [-2.13, -1.15]), sulfonylureas reduced edema the most (SMD: -2.32, CI: [-3.09, -1.54]), and antiapoptotic agents improved behavioral outcomes the most (normalized mean difference: 52.38, CI: [45.29, 59.46]). Statistically significant heterogeneity was observed (I2 = 82 - 98%, all p < 0.001), indicating that studies wildly differ in their effect size estimates. Our results support the superiority of adding cytoprotective therapies with TH (vs. individual or no therapy). Additional exploratory and confirmatory studies are required to identify and thoroughly assess combination therapies owing to limited work and inconsistent translational quality.

β2-Integrins Regulate Microglial Responses and the Functional Outcome of Hemorrhagic Stroke InVivo.

Stroke is one of the leading causes of death and long-term disabilities worldwide. In addition to interruption of blood flow, inflammation is widely recognized as an important factor mediating tissue destruction in stroke. Depending on their phenotype, microglia, the main leukocytes in the CNS, are capable of either causing further tissue damage or promoting brain restoration after stroke. β2-integrins are cell adhesion molecules that are constitutively expressed on microglia. The function of β2-integrins has been investigated extensively in animal models of ischemic stroke, but their role in hemorrhagic stroke is currently poorly understood. We show in this study that dysfunction of β2-integrins is associated with improved functional outcome and decreased inflammatory cytokine expression in the brain in a mouse model of hemorrhagic stroke. Furthermore, β2-integrins affect microglial phenotype and cytokine responses invivo. Therefore, our findings suggest that targeting β2-integrins in hemorrhagic stroke may be beneficial.

Efficacy of bumetanide in animal models of ischemic stroke: a systematic review and meta-analysis.

This meta-analysis aimed to describe the efficacy of bumetanide in improving infarct volume, brain edema, and behavioral outcomes in animal models of cerebral ischemia. Embase, PubMed and Web of Science databases were searched from their inception to February 2024 (INPLASY:202430023). Data on the animal species, stroke model, drug dose, time of treatment, method of administration, study quality, and outcomes were extracted and pooled in a meta-analysis. The combined standardized mean difference (SMD) or mean difference (MD) estimates and 95% confidence intervals (CIs) were calculated using random- or fixed-effects models. Thirteen eligible studies involving >200 animals fulfilled the inclusion criteria and were included in this meta-analysis. Meta-analyses demonstrated that bumetanide treatment significantly reduced cerebral infarct volume (SMD: -0.42; 95% CI: -0.75, -0.09; p < 0.01; n = 186 animals) and consistently relieved brain edema (SMD: -1.39; 95% CI: -2.06, -0.72; p < 0.01; n = 64 animals). Subgroup analyses demonstrated that bumetanide treatment reduced infarct volume in transient but not permanent cerebral ischemia models. When administered after the stroke, it was more effective than treatment initiation before the stroke. Eight studies assessed the effect of bumetanide on behavioral function and the results showed that bumetanide treatment significantly improved neurobehavioral deficits (SMD: -2.35; 95% CI: -2.72, -1.97; p < 0.01; n = 250 animals). We conclude that bumetanide appears to be effective in reducing infarct volume and brain edema and improving behavioral recovery in animal models of cerebral ischemia. This mechanism needs to be confirmed through further investigation.

Morphine Induced Neuroprotection in Ischemic Stroke by Activating Autophagy Via mTOR-Independent Activation of the JNK1/2 Pathway.

Morphine (Mor) has exhibited efficacy in safeguarding neurons against ischemic injuries by simulating ischemic/hypoxic preconditioning (I/HPC). Concurrently, autophagy plays a pivotal role in neuronal survival during IPC against ischemic stroke. However, the involvement of autophagy in Mor-induced neuroprotection and the potential mechanisms remain elusive. Our experiments further confirmed the effect of Mor in cellular and animal models of ischemic stroke and explored its potential mechanism. The findings revealed that Mor enhanced cell viability in a dose-dependent manner by augmenting autophagy levels and autophagic flux in neurons subjected to oxygen-glucose deprivation/reoxygenation (OGD/R). Pretreatment of Mor improved neurological outcome and reduced infarct size in mice with middle cerebral artery occlusion/reperfusion (MCAO/R) at 1, 7 and 14 days. Moreover, the use of autophagy inhibitors nullified the protective effects of Mor, leading to reactive oxygen species (ROS) accumulation, increased loss of mitochondrial membrane potential (MMP) and neuronal apoptosis in OGD/R neurons. Results further demonstrated that Mor-induced autophagy activation was regulated by mTOR-independent activation of the c-Jun NH2- terminal kinase (JNK)1/2 Pathway, both in vitro and in vivo. Overall, these findings suggested Mor-induced neuroprotection by activating autophagy, which were regulated by JNK1/2 pathway in ischemic stroke.

A Rat Model of Middle Cerebral Artery Occlusion/Reperfusion without Damaging the Anatomical Structure of Cerebral Vessels.

Ischemic stroke stands as the primary cause of long-term disability and mortality among adults worldwide. Animal models of ischemic stroke have significantly contributed to our understanding of its pathological mechanisms and the development of potential treatments. Presently, there are two common methods involving filament (endovascular suture) techniques to induce animal models of cerebral ischemia. However, these methods have inherent limitations, such as reduced blood perfusion to the brain, damage to the external carotid artery system, impaired food and/or water intake, and sensory dysfunction of the face. This article introduces a new method for inducing a rat ischemic stroke model without compromising the cerebral vascular anatomy. In this study, the common carotid artery (CCA) of Sprague-Dawley rats was exposed, and an incision was made. A filament was then inserted through the incision into the internal carotid artery to occlude the middle cerebral artery. After 1.5 h of induced ischemia, the occluding filament was fully removed from both the internal carotid artery and the CCA. The incision in the CCA was subsequently sutured using 11-0 microsurgical sutures under a microscope (magnification 4x). Through the utilization of microsurgical techniques to repair the CCA, this study successfully developed a unique method to induce an ischemic stroke model in rats while preserving the anatomical integrity of cerebral blood vessels.

MiR-122 overexpression alleviates oxygen-glucose deprivation-induced neuronal injury by targeting sPLA2-IIA.

Ischemic stroke (IS) is a neurological disease with significant disability and mortality. MicroRNAs were proven to be associated with cerebral ischemia. Previous studies have demonstrated miR-122 downregulation in both animal models of IS and the blood of IS patients. Nonetheless, the role and mechanism of miR-122-5p in IS remain unclear. We established primary human and mouse astrocytes, along with HT22 mouse hippocampal neuronal cells, through oxygen-glucose deprivation/reoxygenation (OGD/R) treatment. To assess the impact of miR-122, we employed CCK8 assays, flow cytometry, RT-qPCR, western blotting, and ELISA to evaluate cell viability, apoptosis, reactive oxygen species (ROS) generation, and cytokine expression. A dual-luciferase reporter gene assay was employed to investigate the interaction between miR-122 and sPLA2-IIA. Overexpression of miR-122 resulted in decreased apoptosis, reduced cleaved caspase-3 expression, and increased cell viability in astrocytes and HT22 cells subjected to OGD/R. RT-qPCR and ELISA analyses demonstrated a decrease in mRNA and cytokine levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-α in both astrocytes and HT22 cells following miR-122 overexpression. Moreover, miR-122 overexpression reversed OGD/R-induced ROS levels and 8-OHdG formation in astrocytes. Additionally, miR-122 overexpression decreased the mRNA and protein expression of inducible nitric oxide synthase (iNOS). Furthermore, we found that miR-122 attaches to the 3'-UTR of sPLA2-IIA, thereby downregulate its expression. Our study demonstrates that miR-122-mediated inhibition of sPLA2-IIA attenuates OGD/R-induced neuronal injury by suppressing apoptosis, alleviating post-ischemic inflammation, and reducing ROS production. Thus, the miR-122/sPLA2-IIA axis may represent a promising target for IS treatment.

The Effects of IRL-1620 in Post-ischemic Brain Injury: A Systematic Review and Meta-analysis of Experimental Studies.

Sovateltide (IRL-1620), an endothelin B receptor agonist, has previously demonstrated neuroprotective and neuroregenerative effects in animal models of acute ischemic stroke. Recently, clinical trials indicated that it could also be effective in humans with stroke. Here, we systematically investigate whether IRL-1620 may be used for the treatment of ischemia-induced brain injury. A systematic review was performed following the Preferred Reporting Items for Systematic reviews and Meta-Analyses guidelines. MEDLINE (PubMed) and Scopus databases were searched for eligible studies up to December 2022. The databases ClinicalTrials.gov and Pharmazz Inc. were screened for unpublished or ongoing trials. Only studies in English were evaluated for eligibility. Meta-analysis of the included studies was also conducted. Finally, seven studies were included in the review, all in animal rat models because of scarcity of clinical trials. Six studies, all in middle cerebral artery occlusion (MCAO) models, were selected for meta-analysis. In the two studies assessing mortality, no deaths were reported in the IRL-1620 group 24h after MCAO, whereas the vehicle group had almost a five times higher mortality risk (risk ratio 5.3, 95% confidence interval 0.7-40.1, I2 = 0%). In all five studies evaluating outcome on day 7 after MCAO, IRL-1620 was associated with statistically significantly lower neurological deficit and improved motor performance compared with the vehicle. Infract volume, differentiation potential of neuronal progenitor cells, and mitochondrial fate also improved with IRL-1620 administration. According to the above, in animal MCAO models, IRL-1620 enhanced neurogenesis and neuroprotection and improved outcome. Future studies are needed to expand our understanding of its effects in human study participants with acute ischemic stroke as well as in other common causes of cerebral ischemia including cardiac arrest.

Edaravone dexborneol promotes M2 microglia polarization against lipopolysaccharide-induced inflammation via suppressing TLR4/MyD88/NF-κB pathway.

Edaravone dexborneol (ED) is a novel neuroprotective compound that consists of two active ingredients, edaravone and ( +)-borneol in a 4:1 ratio, which has been shown the anti-inflammatory properties in animal models of ischemic stroke, cerebral hemorrhage, and autoimmune encephalomyelitis. However, the effect of ED on the polarization of microglia in neuroinflammation has not been elucidated. This study was to investigate the effects of ED on the polarization of microglia induced by lipopolysaccharide (LPS) and potential mechanisms. BV-2 microglial cells were incubated with ED (100, 200, and 400 µM) for 2h, followed by lipopolysaccharide (LPS, 1µg/ml) for 12h. The researchers used the Griess method, western blot, immunocytochemistry, and subcellular fractionation to assess the effects and potential mechanisms of ED on neuroinflammatory reactions. The expression of ROS and the activities of antioxidant enzymes (SOD, GPx, and CAT) in LPS-induced BV-2 cells were also measured using the DCFH-DA fluorescent probe and colorimetric methods, respectively. It was observed that ED significantly declined the levels of TLR4/NF-κB pathway-associated proteins (TLR4, MyD88, p65, p-p65, IκBα, p-IκBα, IKKβ, p-IKKβ) and therefore inhibited LPS-induced production of NO, IL-1β, and TNF-α. Moreover, ED markedly downregulated the M1 marker (iNOS) and upregulated the M2 marker (Arginase-1, Ym-1). In addition, ED also reduced ROS generation and enhanced GPx activity. ED induced the polarization of LPS-stimulated microglia from M1 to M2 against inflammation by negatively regulating the TLR4/MyD88/NF-κB signaling pathway. Additionally, ED performed antioxidative function by depleting the intracellular excessive ROS caused by LPS through the enhancement of the enzymatic activity of GPx. ED may be a potential agent to attenuate neuroinflammation via regulating the polarization of microglia.

Epigallocatechin gallate improves neuronal damage in animal model of ischemic stroke and glutamate-exposed neurons via modulation of hippocalcin expression.

Epigallocatechin gallate (EGCG) is a polyphenolic component of green tea that has anti-oxidative and anti-inflammatory effects in neurons. Ischemic stroke is a major neurological disease that causes irreversible brain disorders. It increases the intracellular calcium concentration and induces apoptosis. The regulation of intracellular calcium concentration is important to maintain the function of the nervous system. Hippocalcin is a neuronal calcium sensor protein that controls intracellular calcium concentration. We investigated whether EGCG treatment regulates the expression of hippocalcin in stroke animal model and glutamate-induced neuronal damage. We performed middle cerebral artery occlusion (MCAO) to induce cerebral ischemia. EGCG (50 mg/kg) or phosphate buffered saline was injected into the abdominal cavity just before MCAO surgery. The neurobehavioral tests were performed 24 h after MCAO surgery and cerebral cortex tissue was collected. MCAO damage induced severe neurobehavioral disorders, increased infarct volume, and decreased the expression of hippocalcin in the cerebral cortex. However, EGCG treatment improved these deficits and alleviated the decrease in hippocalcin expression in cerebral cortex. In addition, EGCG dose-dependently alleviated neuronal cell death and intracellular calcium overload in glutamate-exposed neurons. Glutamate exposure reduced hippocalcin expression, decreased Bcl-2 expression, and increased Bax expression. However, EGCG treatment mitigated these changes caused by glutamate toxicity. EGCG also attenuated the increase in caspase-3 and cleaved caspase-3 expressions caused by glutamate exposure. The effect of EGCG was more pronounced in non-transfected cells than in hippocalcin siRNA-transfected cells. These findings demonstrate that EGCG protects neurons against glutamate toxicity through the regulation of Bcl-2 family proteins and caspase-3. It is known that hippocalcin exerts anti-apoptotic effect through the modulation of apoptotic pathway. Thus, we can suggest evidence that EGCG has a neuroprotective effect by regulating hippocalcin expression in ischemic brain damage and glutamate-exposed cells.

Chronic vascular pathogenesis results in the reduced serum Metrnl levels in ischemic stroke patients.

Metrnl is a secreted protein involved in neurite outgrowth, insulin sensitivity, immunoinflammatory responses, blood lipids and endothelial protection. In this study, we investigated the role of Metrnl in ischemic stroke. Fifty-eight ischemic stroke patients (28 inpatient patients within 2 weeks of onset and 30 emergency patients within 24 h of onset) and 20 healthy controls were enrolled. Serum Metrnl was measured by enzyme-linked immunosorbent assay. We showed that serum Metrnl levels were significantly reduced in both inpatient and emergency patient groups compared with the controls. Different pathological causes for ischemic stroke such as large artery atherosclerosis and small artery occlusion exhibited similar reduced serum Metrnl levels. Transient ischemic attack caused by large artery atherosclerosis without brain infarction also had lower serum Metrnl levels. Metrnl was correlated with some metabolic, inflammatory and clotting parameters. Reduced serum Metrnl was associated with the severity of intracranial arterial stenosis and the presence of ischemic stroke. In order to elucidate the mechanisms underlying the reduced serum Metrnl levels, we established animal models of ischemic stroke in normal mice, atherosclerotic apolipoprotein E-knockout mice and Metrnl-knockout mice by middle cerebral artery occlusion (MCAO) using intraluminal filament or electrocoagulation. We demonstrated that serum Metrnl levels were significantly lower in atherosclerosis mice than normal mice, whereas acute ischemic stroke injury in normal mice and atherosclerosis mice did not alter serum Metrnl levels. Metrnl knockout did not affect acute ischemic stroke injury and death. We conclude that reduced serum Metrnl levels are attributed to the chronic vascular pathogenesis before the onset of ischemic stroke. Metrnl is a potential target for prevention of ischemic stroke.

Retinoic acid alleviates the reduction of Akt and Bad phosphorylation and regulates Bcl-2 family protein interactions in animal models of ischemic stroke.

Ischemic stroke causes a lack of oxygen and glucose supply to brain, eventually leads to severe neurological disorders. Retinoic acid is a major metabolic product of vitamin A and has various biological effects. The PI3K-Akt signaling pathway is an important survival pathway in brain. Phosphorylated Akt is important in regulating survival and apoptosis. We examined whether retinoic acid has neuroprotective effects in stroke model by regulating Akt and its downstream protein, Bad. Moreover, we investigated the relationship between retinoic acid and Bcl-2 family protein interactions. Animals were intraperitoneally administered vehicle or retinoic acid (5 mg/kg) for four days before surgery and ischemic stroke was induced by middle cerebral artery occlusion (MCAO) surgery. Neurobehavioral tests were performed 24 h after MCAO and cerebral cortical tissues were collected. Cresyl violet staining and TUNEL histochemistry were performed, Western blot and immunoprecipitation analysis were performed to elucidate the expression of various proteins. Retinoic acid reduced neurological deficits and histopathological changes, decreased the number of TUNEL-positive cells, and alleviated reduction of phospho-PDK1, phospho-Akt, and phospho-Bad expression caused by MCAO damage. Immunoprecipitation analysis showed that MCAO damage reduced the interaction between phospho-Bad and 14-3-3, which was attenuated by retinoic acid. Furthermore, retinoic acid mitigated the increase in Bcl-2/Bad and Bcl-xL/Bad binding levels and the reduction in Bcl-2/Bax and Bcl-xL/Bax binding levels caused by MCAO damage. Retinoic acid alleviated MCAO-induced increase of caspase-3 and cleaved caspase-3 expression. We demonstrate that retinoic acid prevented apoptosis against cerebral ischemia through phosphorylation of Akt and Bad, maintenance of phospho-Bad and 14-3-3 binding, and regulation of Bcl-2 family protein interactions. .

Environmental Enrichment for Stroke and Traumatic Brain Injury: Mechanisms and Translational Implications.

Acquired brain injuries, such as ischemic stroke, intracerebral hemorrhage (ICH), and traumatic brain injury (TBI), can cause severe neurologic damage and even death. Unfortunately, currently, there are no effective and safe treatments to reduce the high disability and mortality rates associated with these brain injuries. However, environmental enrichment (EE) is an emerging approach to treating and rehabilitating acquired brain injuries by promoting motor, sensory, and social stimulation. Multiple preclinical studies have shown that EE benefits functional recovery, including improved motor and cognitive function and psychological benefits mediated by complex protective signaling pathways. This article provides an overview of the enriched environment protocols used in animal models of ischemic stroke, ICH, and TBI, as well as relevant clinical studies, with a particular focus on ischemic stroke. Additionally, we explored studies of animals with stroke and TBI exposed to EE alone or in combination with multiple drugs and other rehabilitation modalities. Finally, we discuss the potential clinical applications of EE in future brain rehabilitation therapy and the molecular and cellular changes caused by EE in rodents with stroke or TBI. This article aims to advance preclinical and clinical research on EE rehabilitation therapy for acquired brain injury. © 2024 American Physiological Society. Compr Physiol 14:5291-5323, 2024.

Expression of Na+/Ca2+ exchangers was enhanced following pre-treatment of olive leaf extract and olive oil in animal model of ischemic stroke

Background Neuroprotective role of olive and its natural products can introduce them as alternative candidates for the management of neurodegenerative diseases including stroke. The present study was designed to evaluate whether pretreatment of olive oil and leaf extract can attenuate the most important destructive processes in cerebral ischemia called excitotoxicity. Material and Methods The male rats were categorized into control, virgin olive oil (OVV), MCAO, MCAO + OVV (with doses of 0.25, 0.50 and 0.75 ml/kg as treatment groups), olive leaf extract, MCAO + olive leaf extract (with doses 50, 75 and 100 mg/kg as treatment groups) groups. Rats of treatment groups received gastric gavage with olive oil or leaf extract for 30 consecutive days. After pretreatment, the intraluminal filament technique was used to block middle cerebral artery (MCA) transiently. Neurological deficits, infarct volume and expression of Na+/Ca2+ exchangers (NCX1, NCX2 and NCX3) proteins were measured. Results The results revealed that olive oil at doses of 0.50 and 0.75 ml/kg reduced the infarction and neurological score and upregulated NCXs expression in rat brain. In addition, olive leaf extract at doses of 75 and 100 mg/kg attenuated the infarction and neurological score and enhanced NCXs expression in rat brain. Conclusion These findings support the view that olive oil and leaf extract play the neuroprotective role in cerebral ischemia due to the upregulation of NCXs protein expression.

PCSK9 inhibitor protects against ischemic cerebral injury by attenuating inflammation via the GPNMB/CD44 pathway

BackgroundIschemic stroke is the second leading cause of death worldwide, and neuroinflammation has been recognized as a critical player in its progression. Meanwhile, proprotein convertase subtilisin/kexin type 9 inhibitor (PCSK9i) has been demonstrated to inhibit inflammatory response. However, the effects of PCSK9i on ischemic stroke remain unclear and require further investigation. MethodsTemporary middle cerebral artery occlusion (tMCAO) was performed to establish animal models of ischemic stroke in C57BL/6 mice. The PCSK9i were administered subcutaneously after 2 h tMCAO. Neurological function and cerebral infarct volume were measured by mNSS and TTC staining, respectively. RNA-seq was performed to investigate the changes in mechanistic pathways. Western blotting and immunofluorescence were applied to detect expression of GPNMB, CD44, IL-6, and iNOS. ResultsTreatment with PCSK9i significantly improved neurological deficits and reduced the volume of cerebral infarction. PCSK9i suppressed neuroinflammation by activating the GPNMB/CD44 signaling pathway, further exerting their protective effects. ConclusionTaken together, treatment with PCSK9i is an effective way to prevent ischemic stroke-induced brain injury.