RESUMEN
In the realm of this study, obtaining a comprehensive understanding of ischemic brain injury and its molecular foundations is of paramount importance. Our study delved into single-cell data analysis, with a specific focus on sub-celltypes and differentially expressed genes in the aftermath of ischemic injury. Notably, we observed a significant enrichment of the "ATP METABOLIC PROCESS" and "ATP HYDROLYSIS ACTIVITY" pathways, featuring pivotal genes such as Pbx3, Dguok, and Kif21b. A remarkable finding was the consistent upregulation of genes like Fabp7 and Bcl11a within the MCAO group, highlighting their crucial roles in regulating the pathway of mitochondrial ATP synthesis coupled proton transport. Furthermore, our network analysis unveiled pathways like "Neuron differentiation" and "T cell differentiation" as central in the regulatory processes of sub-celltypes. These findings provide valuable insights into the intricate molecular responses and regulatory mechanisms that govern brain injury. The shared differentially expressed genes among sub-celltypes emphasize their significance in orchestrating responses post-ischemic injury. Our research, viewed from the perspective of a medical researcher, contributes to the evolving understanding of the molecular landscape underlying ischemic brain injury, potentially paving the way for targeted therapeutic strategies and improved patient outcomes.
Asunto(s)
Adenosina Trifosfato , Infarto de la Arteria Cerebral Media , Cinesinas , Mitocondrias , Células Precursoras de Oligodendrocitos , Transducción de Señal , Animales , Transducción de Señal/genética , Infarto de la Arteria Cerebral Media/patología , Infarto de la Arteria Cerebral Media/metabolismo , Mitocondrias/metabolismo , Adenosina Trifosfato/metabolismo , Adenosina Trifosfato/biosíntesis , Cinesinas/genética , Cinesinas/metabolismo , Células Precursoras de Oligodendrocitos/metabolismo , Humanos , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Masculino , Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Ratas , Proteínas Proto-OncogénicasRESUMEN
Ischaemic stroke is a common condition that can lead to cerebral ischaemia-reperfusion injury. Phillygenin (PHI), a natural bioactive compound derived from Forsythia suspensa, has been shown to play a crucial role in regulating inflammation across various diseases. However, its specific regulatory effects in ischaemic stroke progression remain unclear. In this study, we established a middle cerebral artery occlusion (MCAO) rat model. Treatment with PHI (50 or 100 mg/kg) significantly reduced cerebral infarction in MCAO rats. PHI treatment also mitigated the increased inflammatory response observed in these rats. Additionally, PHI suppressed microglial activation by reducing iNOS expression, a marker of M1-type polarization of microglia, and attenuated increased brain tissue apoptosis in MCAO rats. Furthermore, PHI's anti-inflammatory effects in MCAO rats were abrogated upon co-administration with GW9662, a peroxisome proliferator-activated receptor γ (PPARγ) inhibitor. In summary, PHI attenuated microglial activation and apoptosis in cerebral ischaemia-reperfusion injury through PPARγ activation, suggesting its potential as a therapeutic agent for mitigating cerebral ischaemia-reperfusion injury.
Asunto(s)
Apoptosis , Infarto de la Arteria Cerebral Media , Microglía , PPAR gamma , Ratas Sprague-Dawley , Daño por Reperfusión , Animales , PPAR gamma/metabolismo , Apoptosis/efectos de los fármacos , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Microglía/efectos de los fármacos , Microglía/metabolismo , Microglía/patología , Ratas , Masculino , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/patología , Infarto de la Arteria Cerebral Media/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , LignanosRESUMEN
BACKGROUND: Ischemic stroke leads a primary cause of mortality in human diseases, with a high disability rate worldwide. This study aims to investigate the function of ß-1,4-galactosyltransferase 1 (B4galt1) in mouse brain ischemia/reperfusion (I/R) injury. METHODS: Recombinant human B4galt1 (rh-B4galt1) was intranasally administered to the mice model of middle cerebral artery occlusion (MCAO)/reperfusion. In this study, the impact of rh-B4galt1 on cerebral injury assessed using multiple methods, including the neurological disability status scale, 2,3,5-triphenyltetrazolium chloride (TTC), Nissl and TUNEL staining. This study utilized laser speckle Doppler flowmeter to monitor the cerebral blood flow. Western blotting was performed to assess the protein expression levels, and fluorescence-labeled dihydroethidium method was performed to determine the superoxide anion generation. Assay kits were used for the measurement of iron, malondialdehyde (MDA) and glutathione (GSH) levels. RESULTS: We demonstrated that rh-B4galt1 markedly improved neurological function, reduced cerebral infarct volume and preserved the completeness of blood-brain barrier (BBB) for preventing damage. These findings further illustrated that rh-B4galt1 alleviated oxidative stress, lipid peroxidation, as well as iron deposition induced by I/R. The vital role of ferroptosis was proved in brain injury. Furthermore, the rh-B4galt1 could increase the levels of TAZ, Nrf2 and HO-1 after I/R. And TAZ-siRNA and ML385 reversed the neuroprotective effects of rh-B4galt1. CONCLUSIONS: The results indicated that rh-B4galt1 implements neuroprotective effects by modulating ferroptosis, primarily via upregulating TAZ/Nrf2/HO-1 pathway. Thus, B4galt1 could be seen as a promising novel objective for ischemic stroke therapy.
Asunto(s)
Isquemia Encefálica , Ferroptosis , Galactosiltransferasas , Hemo-Oxigenasa 1 , Factor 2 Relacionado con NF-E2 , Transducción de Señal , Animales , Humanos , Masculino , Ratones , Isquemia Encefálica/metabolismo , Isquemia Encefálica/prevención & control , Ferroptosis/efectos de los fármacos , Ferroptosis/fisiología , Galactosiltransferasas/metabolismo , Hemo-Oxigenasa 1/metabolismo , Infarto de la Arteria Cerebral Media , Proteínas de la Membrana , Ratones Endogámicos C57BL , Factor 2 Relacionado con NF-E2/metabolismo , Daño por Reperfusión/prevención & control , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Transducción de Señal/fisiología , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/metabolismoRESUMEN
The therapeutic potential of suppressing polypyrimidine tract-binding protein 1 (Ptbp1) messenger RNA by viral transduction in a post-stroke dementia mouse model has not yet been examined. In this study, 3 days after cerebral ischemia, we injected a viral vector cocktail containing adeno-associated virus (AAV)-pGFAP-mCherry and AAV-pGFAP-CasRx (control vector) or a cocktail of AAV-pGFAP-mCherry and AAV-pGFAP-CasRx-SgRNA-(Ptbp1) (1:5, 1.0 × 1011 viral genomes) into post-stroke mice via the tail vein. We observed new mCherry/NeuN double-positive neuron-like cells in the hippocampus 56 days after cerebral ischemia. A portion of mCherry/GFAP double-positive astrocyte-like glia could have been converted into new mCherry/NeuN double-positive neuron-like cells with morphological changes. The new neuronal cells integrated into the dentate gyrus and recognition memory was significantly ameliorated. These results demonstrated that the in vivo conversion of hippocampal astrocyte-like glia into functional new neurons by the suppression of Ptbp1 might be a therapeutic strategy for post-stroke dementia.
Asunto(s)
Astrocitos , Isquemia Encefálica , Modelos Animales de Enfermedad , Ribonucleoproteínas Nucleares Heterogéneas , Hipocampo , Neurogénesis , Proteína de Unión al Tracto de Polipirimidina , Animales , Proteína de Unión al Tracto de Polipirimidina/metabolismo , Proteína de Unión al Tracto de Polipirimidina/genética , Astrocitos/metabolismo , Hipocampo/metabolismo , Hipocampo/patología , Ratones , Isquemia Encefálica/metabolismo , Isquemia Encefálica/terapia , Ribonucleoproteínas Nucleares Heterogéneas/metabolismo , Ribonucleoproteínas Nucleares Heterogéneas/genética , Masculino , Neuronas/metabolismo , Memoria , Ratones Endogámicos C57BL , Vectores Genéticos/genética , Vectores Genéticos/administración & dosificaciónRESUMEN
Glutaminyl cyclase (QC) and its isoenzyme (isoQC) catalyze the formation of N-terminal pyroglutamate (pGlu) from glutamine on a number of neuropeptides, peptide hormones and chemokines. Chemokines of the C-C ligand (CCL) motif family are known to contribute to inflammation in neurodegenerative conditions. Here, we used a model of transient focal cerebral ischemia to explore functional, cellular and molecular responses to ischemia in mice lacking genes for QC, isoQC and their substrate CCL2. Mice of the different genotypes were evaluated for functional consequences of stroke, infarct volume, activation of glia cells, and for QC, isoQC and CCL2 expression. The number of QC-immunoreactive, but not of isoQC-immunoreactive, neurons increased robustly in the infarct area at 24 and 72 h after ischemia. In parallel, immunohistochemical signals for the QC substrate CCL2 increased from 24 to 72 h after ischemia induction without differences between genotypes analyzed. The increase in CCL2 was accompanied by morphological activation of Iba1-immunoreactive microglia and recruitment of MHC-II-positive cells at 72 h after ischemia. Among other chemokines quantified in the brain tissue, CCL17 showed higher concentrations at 72 h compared to 24 h after ischemia. Collectively, these data suggest a critical role for QC in inflammatory processes in the stroke-affected brain.
Asunto(s)
Aminoaciltransferasas , Isquemia Encefálica , Inflamación , Animales , Aminoaciltransferasas/metabolismo , Aminoaciltransferasas/genética , Ratones , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Isquemia Encefálica/genética , Inflamación/patología , Inflamación/metabolismo , Inflamación/genética , Quimiocina CCL2/metabolismo , Masculino , Ratones Endogámicos C57BL , Microglía/metabolismo , Microglía/patología , Neuronas/metabolismo , Neuronas/patologíaRESUMEN
Matrix metalloproteinases (MMPs) such as MMP-9, 3, and 2 degrade the cellular matrix and are believed to play a crucial role in ischemic stroke. We examined how the duration of ischemia (up to 4 h) and treatment with recombinant tissue plasminogen activator altered the comparative expression of these MMPs in experimental ischemic stroke with reperfusion. Both prolonged ischemia and r-tPA treatment markedly increased MMP-9 expression in the ischemic hemisphere (all p < 0.0001). The duration of ischemia and r-tPA treatment also significantly increased MMP-2 expression (p < 0.01-0.001) in the ischemic hemisphere (p < 0.01) but to a lesser degree than MMP-9. In contrast, MMP-3 expression significantly decreased in the ischemic hemisphere (p < 0.001) with increasing duration of ischemia and r-tPA treatment (p < 0.05-0001). MMP-9 expression was prominent in the vascular compartment and leukocytes. MMP-2 expression was evident in the vascular compartment and MMP-3 in NeuN+ neurons. Prolonging the duration of ischemia (up to 4 h) before reperfusion increased brain hemorrhage, infarction, swelling, and neurologic disability in both saline-treated (control) and r-tPA-treated mice. MMP-9 and MMP-2 expression were significantly positively correlated with, and MMP-3 was significantly negatively correlated with, infarct volume, swelling, and brain hemorrhage. We conclude that in experimental ischemic stroke with reperfusion, the duration of ischemia and r-tPA treatment significantly altered MMP-9, 3, and 2 expression, ischemic brain injury, and neurological disability. Each MMP showed unique patterns of expression that are strongly correlated with the severity of brain infarction, swelling, and hemorrhage. In summary, in experimental ischemic stroke in male mice with reperfusion, the duration of ischemia, and r-tPA treatment significantly altered the immunofluorescent expression of MMP-9, 3, and 2, ischemic brain injury, and neurological disability. In this model, each MMP showed unique patterns of expression that were strongly correlated with the severity of brain infarction, swelling, and hemorrhage.
Asunto(s)
Isquemia Encefálica , Metaloproteinasa 2 de la Matriz , Metaloproteinasa 3 de la Matriz , Metaloproteinasa 9 de la Matriz , Activador de Tejido Plasminógeno , Animales , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones , Masculino , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Accidente Cerebrovascular/tratamiento farmacológico , Accidente Cerebrovascular/metabolismo , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Accidente Cerebrovascular Isquémico/tratamiento farmacológico , Accidente Cerebrovascular Isquémico/metabolismo , Accidente Cerebrovascular Isquémico/patología , Factores de TiempoRESUMEN
Agathisflavone is a flavonoid that exhibits anti-inflammatory and anti-oxidative properties. Here, we investigated the neuroprotective effects of agathisflavone on central nervous system (CNS) neurons and glia in the cerebellar slice ex vivo model of neonatal ischemia. Cerebellar slices from neonatal mice, in which glial fibrillary acidic protein (GFAP) and SOX10 drive expression of enhanced green fluorescent protein (EGFP), were used to identify astrocytes and oligodendrocytes, respectively. Agathisflavone (10 µM) was administered preventively for 60 min before inducing ischemia by oxygen and glucose deprivation (OGD) for 60 min and compared to controls maintained in normal oxygen and glucose (OGN). The density of SOX-10+ oligodendrocyte lineage cells and NG2 immunopositive oligodendrocyte progenitor cells (OPCs) were not altered in OGD, but it resulted in significant oligodendroglial cell atrophy marked by the retraction of their processes, and this was prevented by agathisflavone. OGD caused marked axonal demyelination, determined by myelin basic protein (MBP) and neurofilament (NF70) immunofluorescence, and this was blocked by agathisflavone preventative treatment. OGD also resulted in astrocyte reactivity, exhibited by increased GFAP-EGFP fluorescence and decreased expression of glutamate synthetase (GS), and this was prevented by agathisflavone pretreatment. In addition, agathisflavone protected Purkinje neurons from ischemic damage, assessed by calbindin (CB) immunofluorescence. The results demonstrate that agathisflavone protects neuronal and myelin integrity in ischemia, which is associated with the modulation of glial responses in the face of ischemic damage.
Asunto(s)
Animales Recién Nacidos , Cerebelo , Flavonoides , Fármacos Neuroprotectores , Animales , Fármacos Neuroprotectores/farmacología , Ratones , Cerebelo/metabolismo , Cerebelo/efectos de los fármacos , Flavonoides/farmacología , Oligodendroglía/efectos de los fármacos , Oligodendroglía/metabolismo , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Glucosa/metabolismo , BiflavonoidesRESUMEN
We reported that infiltrated Ly6C+ macrophages express brain-derived neurotrophic factor (BDNF) only at the cerebral cortex infarct in a rat dMCAO model. However, the changein neuron-expressed BDNF, the niche components that induce the Ly6C+ cells to express BDNF, and the cellular sources of these components, remain unclear. In this study, immunofluorescence double staining was performed to label BDNF and Ly6C on brain sections at 3, 24, and 48 h following distal middle cerebral artery occlusion (dMCAO) of male rats, and to stain BDNF with Ly6C, IL-4R, and IL-10R. A neutralizing anti-IL-4 antibody was injected into the infarct, and the IL-4 and BDNF concentrations in the subareas of the infarct were determined using enzyme-linked immunosorbent assay. To find out the cellular sources of IL-4, the markers for microglia, T cells, and neurons were co-stained with IL-4 separately. In certain infarct subareas, the main BDNF-expressing cells shifted quickly from NeuN+ neurons to Ly6C+ cells during 24-48 h post-stroke, and the Ly6C+/BDNF+ cells mostly expressed IL-4 receptor. Following IL-4 neutralizing antibody injection, the BDNF, IL-4 protein levels, and BDNF+/Ly6C+ cells decreased significantly. The main IL-4-expressing cell type in this infarct subarea is not neuron either, but immune cells, including microglia, monocyte, macrophages, and T cells. The neurons, maintained BDNF and IL-4 expression in the peri-infarct area. In conclusion, in a specific cerebral subarea of the rat dMCAO model, IL-4 secreted by immune cells is one of the main inducers for Ly6C+ cells to express BDNF.
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Isquemia Encefálica , Factor Neurotrófico Derivado del Encéfalo , Interleucina-4 , Macrófagos , Animales , Masculino , Ratas , Isquemia Encefálica/metabolismo , Isquemia Encefálica/inmunología , Isquemia Encefálica/patología , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Modelos Animales de Enfermedad , Infarto de la Arteria Cerebral Media/metabolismo , Infarto de la Arteria Cerebral Media/patología , Interleucina-4/metabolismo , Macrófagos/metabolismo , Macrófagos/inmunología , Neuronas/metabolismo , Neuronas/patología , Ratas Sprague-DawleyRESUMEN
Cerebrovascular disease, one of the high-risk diseases worldwide, is high in morbidity, disability, mortality, and recurrence rates, which brings many harms to human beings such as physical and mental harm, economic losses, and impairment of social relations. Cerebral ischemia-reperfusion injury (CIRI) is one of the most common pathological manifestations, with mild hypothermia therapy being the most commonly used treatment in clinical practice. In this study, the research team established a CIRI animal model and found that the neuronal apoptosis rate was significantly increased, accompanied by significant ferroptosis, increased inflammation and oxidative stress damage in brain tissue, and obviously inhibited SIRT1/AMPK pathway. However, after mild hypothermia treatment, the pathological changes of CIRI rats were significantly reversed, and the SIRT1/AMPK pathway was reactivated. Therefore, mild hypothermia may achieve the purpose of CIRI repair by activating the SIRT1/AMPK signaling pathway, and targeted regulation of the SIRT1/AMPK signaling pathway may be a research direction for optimizing mild hypothermia therapy or developing new treatment plans for CIRI.
Asunto(s)
Proteínas Quinasas Activadas por AMP , Apoptosis , Hipotermia Inducida , Neuronas , Estrés Oxidativo , Daño por Reperfusión , Transducción de Señal , Sirtuina 1 , Sirtuina 1/metabolismo , Daño por Reperfusión/terapia , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Animales , Hipotermia Inducida/métodos , Neuronas/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Masculino , Isquemia Encefálica/terapia , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Ratas Sprague-Dawley , Ratas , Modelos Animales de EnfermedadRESUMEN
Ischemic stroke (IS) is a severe cerebrovascular disease with high disability and mortality rates, where the inflammatory response is crucial to its progression and prognosis. Efferocytosis, the prompt removal of dead cells, can reduce excessive inflammation after IS injury. While electroacupuncture (EA) has been shown to decrease inflammation post-ischemia/reperfusion (I/R), its link to efferocytosis is unclear. Our research identified ATP-binding cassette transporter A1 (Abca1) as a key regulator of the engulfment process of efferocytosis after IS by analyzing public datasets and validating findings in a mouse model, revealing its close ties to IS progression. We demonstrated that EA can reduce neuronal cell death and excessive inflammation caused by I/R. Furthermore, EA treatment increased Abca1 expression, prevented microglia activation, promoted M2 microglia polarization, and enhanced their ability to phagocytose injured neurons in I/R mice. This suggests that EA's modulation of efferocytosis could be a potential mechanism for reducing cerebral I/R injury, making regulators of efferocytosis steps a promising therapeutic target for EA benefits.
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Transportador 1 de Casete de Unión a ATP , Electroacupuntura , Inflamación , Ratones Endogámicos C57BL , Microglía , Fagocitosis , Daño por Reperfusión , Animales , Microglía/metabolismo , Microglía/patología , Electroacupuntura/métodos , Transportador 1 de Casete de Unión a ATP/metabolismo , Daño por Reperfusión/patología , Daño por Reperfusión/terapia , Daño por Reperfusión/metabolismo , Inflamación/patología , Masculino , Isquemia Encefálica/patología , Isquemia Encefálica/metabolismo , Isquemia Encefálica/terapia , Ratones , Neuronas/metabolismo , Neuronas/patología , Modelos Animales de Enfermedad , EferocitosisRESUMEN
BACKGROUND: Cerebral ischemia-reperfusion injury (CIRI) is a prevalent neurological disorder, characterized by the oxidative stress and inflammatory response induced during the ischemia-reperfusion process, leading to significant damage to brain cells. Ginsenoside Rb1, a natural medicinal ingredient, possesses potential neuroprotective effects. This study aims to investigate the mechanism of action of ginsenoside Rb1 in CIRI and its protective effects on brain injury. METHODS: We utilized a mouse CIRI model and randomly divided the mice into control group, CIRI group, and ginsenoside Rb1 treatment group. The effects of Rb1 on brain tissue damage, apoptosis, expression of inflammatory factors, and pyroptotic cell numbers in CIRI mice were observed through triphenyl tetrazolium chloride (TTC) staining, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, real-time reverse transcription polymerase chain reaction (qRT-PCR), and electron microscopy. In a cell model, the regulatory effect of Rb1 on oxygen-glucose deprivation/reperfusion (OGD/R)-induced HT22 cell pyroptosis via the nuclear respiratoty factor 2/tumor necrosis factor-α (TNF-α)-induced Protein 3 (TNFAIP3, aka A20)/eukaryotic translation elongation factor 1A2 (Nrf2/A20/eEF1A2) axis was detected using Western blot and TUNEL staining. Additionally, the impact of Nrf2 inhibitor ML385 and eEF1A2 overexpression on the neuroprotective effect of Rb1 was assessed. Using the comprehensive experimental methods mentioned above, the neuroprotective mechanism of Rb1 in CIRI was thoroughly evaluated. RESULTS: Our findings demonstrate that treatment with ginsenoside Rb1 alleviated behavioral deficits induced by CIRI and reduced pathological damage in brain tissue. Furthermore, ginsenoside Rb1 treatment notably decreased oxidative stress and the inflammatory response induced by CIRI, leading to lower levels of inflammatory factors (p < 0.05). Further experimental results indicated that ginsenoside Rb1 promoted antioxidant and anti-inflammatory responses by regulating the activity of the Nrf2/A20/eEF1A2 axis. Additionally, ginsenoside Rb1 inhibited the activation of the NOD-like receptor thermal protein domain associated protein 3 (NLRP3) inflammasome, thereby reducing the release of inflammatory factors and the occurrence of cell apoptosis. CONCLUSION: Our study results suggest that ginsenoside Rb1 exerts neuroprotective effects and alleviates brain injury induced by CIRI by regulating the Nrf2/A20/eEF1A2 axis and inhibiting the activation of the NLRP3 inflammasome. These findings provide new treatment insights for CIRI and support ginsenoside Rb1's development as a therapeutic drug. However, despite the promising nature of our findings, further research is required to validate these discoveries and explore the feasibility and safety of ginsenoside Rb1 in clinical applications. We hope that our study can provide new directions and strategies for the treatment and prevention of CIRI, contributing to the development of neuroprotective drugs.
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Ginsenósidos , Factor 2 Relacionado con NF-E2 , Daño por Reperfusión , Ginsenósidos/farmacología , Ginsenósidos/uso terapéutico , Animales , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Daño por Reperfusión/tratamiento farmacológico , Factor 2 Relacionado con NF-E2/metabolismo , Ratones , Masculino , Transducción de Señal/efectos de los fármacos , Modelos Animales de Enfermedad , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Ratones Endogámicos C57BL , Isquemia Encefálica/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/patología , Estrés Oxidativo/efectos de los fármacos , Apoptosis/efectos de los fármacos , Línea CelularRESUMEN
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 oxygenâglucose 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-1ß 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-1ß 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.
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Histona Desacetilasas , Accidente Cerebrovascular Isquémico , Ratones Endogámicos C57BL , Microglía , Enfermedades Neuroinflamatorias , Estilbenos , Animales , Histona Desacetilasas/metabolismo , Microglía/metabolismo , Microglía/efectos de los fármacos , Ratones , Accidente Cerebrovascular Isquémico/tratamiento farmacológico , Accidente Cerebrovascular Isquémico/metabolismo , Masculino , Enfermedades Neuroinflamatorias/tratamiento farmacológico , Enfermedades Neuroinflamatorias/metabolismo , Estilbenos/farmacología , Estilbenos/uso terapéutico , Modelos Animales de Enfermedad , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/complicaciones , Transducción de Señal/efectos de los fármacos , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismoRESUMEN
OBJECTIVE: This study aimed to further elucidate the mechanism of ginsenoside Rg1 in the treatment of cerebral ischemia-reperfusion. METHODS: In this study, we observed the apoptosis of RM cells (microglia) after oxygen-glucose deprivation/reoxygenation (OGD/R) modeling before and after Rg1 administration, changes in mitochondrial membrane potential, changes in the content of Reactive oxygen species (ROS) and inflammatory vesicles NLR Family Pyrin Domain Containing 3 (NLRP3), and the expression levels of autophagy-related proteins, inflammatory factors, and apoptosis proteins. We further examined the pathomorphological changes in brain tissue, neuronal damage, changes in mitochondrial morphology and mitochondrial structure, and the autophagy-related proteins, inflammatory factors, and apoptosis proteins expression levels in CI/RI rats before and after administration of Rg1 in vivo experiments. RESULTS: In vitro experiments showed that Rg1 induced mitochondrial autophagy, decreased mitochondrial membrane potential, and reduced ROS content thereby inhibiting NLRP3 activation, decreasing secretion of inflammatory factors and RM cell apoptosis by regulating the PTEN induced putative kinase 1(Pink1) /Parkin signaling pathway. In vivo experiments showed that Rg1 induced mitochondrial autophagy, inhibited NLRP3 activation, improved inflammatory response, and reduced apoptosis by regulating the Pink1/Parkin signaling pathway, and Rg1 significantly reduced the area of cerebral infarcts, improved the pathological state of brain tissue, and attenuated the neuronal damage, thus improving cerebral ischemia/reperfusion injury in rats. CONCLUSION: Our results suggest that ginsenoside Rg1 can ameliorate cerebral ischemia-reperfusion injury by modulating Pink1/ Parkin-mediated mitochondrial autophagy in microglia and inhibiting microglial NLRP3 activation.
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Autofagia , Ginsenósidos , Microglía , Mitocondrias , Proteína con Dominio Pirina 3 de la Familia NLR , Proteínas Quinasas , Daño por Reperfusión , Ubiquitina-Proteína Ligasas , Ginsenósidos/farmacología , Animales , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Autofagia/efectos de los fármacos , Microglía/efectos de los fármacos , Microglía/metabolismo , Proteínas Quinasas/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Ratas , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Masculino , Ratas Sprague-Dawley , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Apoptosis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Fármacos Neuroprotectores/farmacologíaRESUMEN
Air pollution has recently emerged as a significant risk factor for ischemic stroke. Although there is a robust association between higher concentrations of ambient particulate matter (PM2.5) and increased incidence and mortality rates of ischemic stroke, the precise mechanisms underlying PM2.5-induced ischemic stroke remain to be fully elucidated. The purpose of this study was to examine the synergistic effect of PM2.5 and hypoxic stress using in vivo and in vitro ischemic stroke models. Intravenously administered PM2.5 exacerbated the ischemic brain damage induced by middle cerebral artery occlusion (MCAo) in Sprague Dawley rats. Alterations in autophagy flux and decreased levels of tight junction proteins were observed in the brain of PM2.5-administered rats after MCAo. The underlying mechanism of PM2.5-induced potentiation of ischemic brain damage was investigated in neurons, perivascular macrophages, and brain endothelial cells, which are the major components of the integrated neurovascular unit. Co-treatment with PM2.5 and oxygen-glucose deprivation (OGD) amplified the effects of OGD on the reduction of viability in primary neurons, immortalized murine hippocampal neuron (HT-22), and brain endothelial cells (bEND.3). After co-treatment with PM2.5 and OGD, the Akt/ß-catenin and autophagy flux were significantly inhibited in HT-22 cells. Notably, the protein levels of metalloproteinase-9 and cystatin C were elevated in the conditioned media of murine macrophages (RAW264.7) exposed to PM2.5, and tight junction protein expression was significantly decreased after OGD exposure in bEND.3 cells pretreated with the conditioned media. Our findings suggest that perivascular macrophages may mediate PM2.5-induced brain endothelial dysfunction following ischemia and that PM2.5 can exacerbate ischemia-induced neurovascular damage.
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Contaminantes Atmosféricos , Autofagia , Glucosa , Material Particulado , Proteínas Proto-Oncogénicas c-akt , beta Catenina , Animales , Masculino , Ratones , Ratas , Contaminantes Atmosféricos/toxicidad , Autofagia/efectos de los fármacos , beta Catenina/metabolismo , Isquemia Encefálica/metabolismo , Células Endoteliales/efectos de los fármacos , Glucosa/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Oxígeno/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacosRESUMEN
BACKGROUND: G protein-coupled receptor 68 (GPR68), an orphan receptor, has emerged as a promising therapeutic target for mitigating neuronal inflammation and oxidative damage. This study explores the protective mechanisms of GPR68 in cerebral ischemia-reperfusion injury (CIRI). METHODS: An in vivo middle cerebral artery occlusion/reperfusion (MCAO/R) mouse model was established. Mice received intraperitoneal injections of Ogerin, a selective GPR68 agonist. In vitro, GPR68 was overexpressed in SH-SY5Y and HMC3 cells, and the effects of oxygen-glucose deprivation/reperfusion (OGD/R) on cell viability were assessed using real-time quantitative polymerase chain reaction (RT-qPCR), enzyme-linked immunosorbent assay (ELISA), and flow cytometry. RESULTS: The expression of GPR68 was suppressed in cells subjected to OGD/R treatment, whereas its upregulation conferred protection to SH-SY5Y and HMC3 cells. In vivo, levels of GPR68 were reduced in brain tissues affected by MCAO/R, correlating with oxidative stress, inflammation, and neurological damage. Treatment with a GPR68 agonist decreased brain infarction, apoptosis, and dysregulated gene expression induced by MCAO/R. Mechanistically, GPR68 agonist treatment may inhibit the activation of the NF-κB/Hif-1α pathway, thereby reducing oxidative and inflammatory responses and enhancing protection against CIRI. CONCLUSIONS: This study confirms that the GPR68/NF-κB/Hif-1α axis modulates apoptosis, inflammation, and oxidative stress in CIRI, indicating that GPR68 is a potential therapeutic target for CIRI.
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Subunidad alfa del Factor 1 Inducible por Hipoxia , FN-kappa B , Fármacos Neuroprotectores , Receptores Acoplados a Proteínas G , Daño por Reperfusión , Animales , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Daño por Reperfusión/metabolismo , Daño por Reperfusión/tratamiento farmacológico , Ratones , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , FN-kappa B/metabolismo , Fármacos Neuroprotectores/farmacología , Masculino , Infarto de la Arteria Cerebral Media/metabolismo , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Humanos , Transducción de Señal/efectos de los fármacos , Isquemia Encefálica/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Ratones Endogámicos C57BL , Estrés Oxidativo/efectos de los fármacos , Modelos Animales de Enfermedad , Línea Celular TumoralRESUMEN
Pre-clinical trials have demonstrated the neuroprotective effects of transplanted human neural stem cells (hNSCs) during the post-ischemic phase. However, the exact neuroprotective mechanism remains unclear. Tunneling nanotubes (TNTs) are long plasma membrane bridges that physically connect distant cells, enabling the intercellular transfer of mitochondria and contributing to post-ischemic repair processes. Whether hNSCs communicate through TNTs and their role in post-ischemic neuroprotection remains unknown. In this study, non-immortalized hNSC lines derived from fetal human brain tissues were examined to explore these possibilities and assess the post-ischemic neuroprotection potential of these hNSCs. Using Tau-STED super-resolution confocal microscopy, live cell time-lapse fluorescence microscopy, electron microscopy, and direct or non-contact homotypic co-cultures, we demonstrated that hNSCs generate nestin-positive TNTs in both 3D neurospheres and 2D cultures, through which they transfer functional mitochondria. Co-culturing hNSCs with differentiated SH-SY5Y (dSH-SY5Y) revealed heterotypic TNTs allowing mitochondrial transfer from hNSCs to dSH-SY5Y. To investigate the role of heterotypic TNTs in post-ischemic neuroprotection, dSH-SY5Y were subjected to oxygen-glucose deprivation (OGD) followed by reoxygenation (OGD/R) with or without hNSCs in direct or non-contact co-cultures. Compared to normoxia, OGD/R dSH-SY5Y became apoptotic with impaired electrical activity. When OGD/R dSH-SY5Y were co-cultured in direct contact with hNSCs, heterotypic TNTs enabled the transfer of functional mitochondria from hNSCs to OGD/R dSH-SY5Y, rescuing them from apoptosis and restoring the bioelectrical profile toward normoxic dSH-SY5Y. This complete neuroprotection did not occur in the non-contact co-culture. In summary, our data reveal the presence of a functional TNTs network containing nestin within hNSCs, demonstrate the involvement of TNTs in post-ischemic neuroprotection mediated by hNSCs, and highlight the strong efficacy of our hNSC lines in post-ischemic neuroprotection. Human neural stem cells (hNSCs) communicate with each other and rescue ischemic neurons through nestin-positive tunneling nanotubes (TNTs). A Functional mitochondria are exchanged via TNTs between hNSCs. B hNSCs transfer functional mitochondria to ischemic neurons through TNTs, rescuing neurons from ischemia/reperfusion ROS-dependent apoptosis.
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Comunicación Celular , Técnicas de Cocultivo , Mitocondrias , Células-Madre Neurales , Neuronas , Humanos , Células-Madre Neurales/metabolismo , Células-Madre Neurales/citología , Neuronas/metabolismo , Mitocondrias/metabolismo , Encéfalo/metabolismo , Encéfalo/embriología , Diferenciación Celular , Nanotubos/química , Feto , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Estructuras de la Membrana CelularRESUMEN
Ischemic stroke is a serious condition that results in high rates of illness and death. Anaerobic glycolysis becomes the primary means of providing energy to the brain during periods of low oxygen levels, such as in the aftermath of an ischemic stroke. This process is essential for maintaining vital brain functions and has significant implications for recovery following a stroke. Energy supply by anaerobic glycolysis and acidosis caused by lactic acid accumulation are important pathological processes after ischemic stroke. Numerous natural products regulate glucose and lactate, which in turn modulate anaerobic glycolysis. This article focuses on the relationship between anaerobic glycolysis and ischemic stroke, as well as the associated signaling pathways and natural products that play a therapeutic role. These natural products, which can regulate anaerobic glycolysis, will provide new avenues and perspectives for the treatment of ischemic stroke in the future.
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Productos Biológicos , Glucólisis , Accidente Cerebrovascular Isquémico , Transducción de Señal , Humanos , Glucólisis/fisiología , Glucólisis/efectos de los fármacos , Accidente Cerebrovascular Isquémico/metabolismo , Accidente Cerebrovascular Isquémico/tratamiento farmacológico , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Productos Biológicos/farmacología , Productos Biológicos/uso terapéutico , Animales , Anaerobiosis/fisiología , Isquemia Encefálica/metabolismo , Isquemia Encefálica/tratamiento farmacológicoRESUMEN
Stroke is the second most common cause of death and one of the most common causes of disability worldwide. The intestine is home to several microorganisms that fulfill essential functions for the natural and physiological functioning of the human body. There is an interaction between the central nervous system (CNS) and the gastrointestinal system that enables bidirectional communication between them, the so-called gut-brain axis. Based on the gut-brain axis, there is evidence of a link between the gut microbiota and the regulation of microglial functions through glial activation. This interaction is partly due to the immunological properties of the microbiota and its connection with the CNS, such that metabolites produced by the microbiota can cross the gut barrier, enter the bloodstream and reach the CNS and significantly affect microglia, astrocytes and other cells of the immune system. Studies addressing the effects of short-chain fatty acids (SCFAs) on glial function and the BBB in ischemic stroke are still scarce. Therefore, this review aims to stimulate the investigation of these associations, as well as to generate new studies on this topic that can clarify the role of SCFAs after stroke in a more robust manner.
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Barrera Hematoencefálica , Ácidos Grasos Volátiles , Microbioma Gastrointestinal , Accidente Cerebrovascular Isquémico , Neuroglía , Humanos , Barrera Hematoencefálica/metabolismo , Ácidos Grasos Volátiles/metabolismo , Ácidos Grasos Volátiles/farmacología , Accidente Cerebrovascular Isquémico/metabolismo , Accidente Cerebrovascular Isquémico/fisiopatología , Animales , Neuroglía/metabolismo , Eje Cerebro-Intestino/fisiología , Isquemia Encefálica/metabolismoRESUMEN
Ischemic stroke remains a major cause of disability and mortality. Nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy is involved in cerebral ischemic injury. Additionally, lactylation regulates the progression of ischemia injury. This study aimed to investigate the impact of NCOA4 on ferritinophagy and glycolysis of hippocampal neuron cells and its lactylation modification. Middle cerebral artery occlusion (MCAO) mouse and oxygen-glucose deprivation (OGD)-treated HT22 cell models were generated. Ferritinophagy was evaluated via detecting ferrous iron (Fe 2+ ), glutathione, malondialdehyde, and protein levels. Glycolysis was assessed by examining the glucose consumption, lactate production, and extracellular acidification rate. The lactylation was evaluated using immunoprecipitation and immunoblotting. Brain injury in vivo was analyzed by measuring brain infarct and neurological function. The results showed that NCOA4 expression was increased in the blood of patients with acute ischemia stroke, the peri-infarct region of the brain in MCAO mice (increased percentage: 142.11%) and OGD-treated cells (increased percentage: 114.70%). Knockdown of NCOA4 inhibited ferritinophagy and glycolysis of HT22 cells induced by OGD. Moreover, OGD promoted the lactylation of NCOA4 at lysine (K)450 sites, which enhanced NCOA4 protein stability. Additionally, interfering with NCOA4 attenuated brain infarction and neurological dysfunction in MCAO mice. Lactylation of NCOA4 at K450 sites promotes ferritinophagy and glycolysis of hippocampal neuron cells, thereby accelerating cerebral ischemic injury. These findings suggest a novel pathogenesis of ischemic stroke.
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Ferritinas , Glucólisis , Infarto de la Arteria Cerebral Media , Neuronas , Coactivadores de Receptor Nuclear , Animales , Neuronas/metabolismo , Glucólisis/fisiología , Ratones , Coactivadores de Receptor Nuclear/metabolismo , Ferritinas/metabolismo , Masculino , Infarto de la Arteria Cerebral Media/metabolismo , Isquemia Encefálica/metabolismo , Humanos , Ratones Endogámicos C57BL , Autofagia/fisiología , Hipocampo/metabolismo , Glucosa/deficiencia , Glucosa/metabolismoRESUMEN
Vascular dementia (VaD) is a prevalent form of dementia resulting from chronic cerebral hypoperfusion (CCH). However, the pathogenic mechanisms of VaD and corresponding therapeutic strategies are not well understood. Sirtuin 6 (SIRT6) has been implicated in various biological processes, including cellular metabolism, DNA repair, redox homeostasis, and aging. Nevertheless, its functional relevance in VaD remains unexplored. In this study, we utilized a bilateral common carotid artery stenosis (BCAS) mouse model of VaD to investigate the role of SIRT6. We detected a significant decrease in neuronal SIRT6 protein expression following CCH. Intriguingly, neuron-specific ablation of Sirt6 in mice exacerbated neuronal damage and cognitive deficits after CCH. Conversely, treatment with MDL-800, an agonist of SIRT6, effectively mitigated neuronal loss and facilitated neurological recovery. Mechanistically, SIRT6 inhibited excessive mitochondrial fission by suppressing the CCH-induced STAT5-PGAM5-Drp1 signaling cascade. Additionally, the gene expression of monocyte SIRT6 in patients with asymptomatic carotid stenosis showed a correlation with cognitive outcomes, suggesting translational implications in human subjects. Our findings provide the first evidence that SIRT6 prevents cognitive impairment induced by CCH, and mechanistically, this protection is achieved through the remodeling of mitochondrial dynamics in a STAT5-PGAM5-Drp1-dependent manner.