RESUMEN

This study investigates the molecular mechanisms behind ischaemia/reperfusion (I/R) injury in the brain, focusing on neuronal apoptosis. It scrutinizes the role of the Jun proto-oncogene in apoptosis, involvement of SOCS1 in neural precursor cell accumulation in ischaemic regions, and the upregulation of C-EBPß in the hippocampus following I/R. Key to the study is understanding how Jun controls C-EBPß degradation via SOCS1, potentially offering new clinical treatment avenues for I/R. Techniques such as mRNA sequencing, KEGG enrichment analysis and protein-protein interaction (PPI) in mouse models have indicated involvement of Jun (AP-1) in I/R-induced cerebral damage. The study employs middle cerebral artery occlusion in different mouse models and oxygen-glucose deprivation/reoxygenation in cortical neurons to examine the impacts of Jun and SOCS1 manipulation on cerebral I/R injury and neuronal damage. The findings reveal that I/R reduces Jun expression in the brain, but its restoration lessens cerebral I/R injury and neuron death. Jun activates SOCS1 transcriptionally, leading to C-EBPß degradation, thereby diminishing cerebral I/R injury through the SOCS1/C-EBPß pathway. These insights provide a deeper understanding of post-I/R cerebral injury mechanisms and suggest new therapeutic targets for cerebral I/R injury. KEY POINTS: Jun and SOCS1 are poorly expressed, and C-EBPß is highly expressed in ischaemia/reperfusion mouse brain tissues. Jun transcriptionally activates SOCS1. SOCS1 promotes the ubiquitination-dependent C-EBPß protein degradation. Jun blunts oxygen-glucose deprivation/reoxygenation-induced neuron apoptosis and alleviates neuronal injury. This study provides a theoretical basis for the management of post-I/R brain injury.

RESUMEN

OBJECTIVE: To investigate the cerebroprotective effects of leptin in vitro and in vivo via the Janus kinase-2 (JAK2)/transcription factor signal transducer and activators of transcription-3 (STAT3) pathway and leptin receptors (LEPR). METHODS: The study used the cellular oxygen-glucose deprivation (OGD) model in PC12 cells and the middle cerebral artery occlusion (MCAO) rat model of cerebral ischaemia-reperfusion injury (CIRI) to assess changes in gene expression and protein levels following leptin pretreatment. The methylated DNA immunoprecipitation (MeDIP) assay measured DNA methylation levels. RESULTS: The optimal leptin concentration for exerting neuroprotective effects against ischaemia-reperfusion injury in PC12 cells was 200 ng/ml in vitro, but excessive leptin diminished this effect. Leptin pretreatment in the MCAO rat model demonstrated a similar effect to previously reported leptin administration post-CIRI. In addition to regulating the expression of inflammation-related cytokines, Western blot analysis showed that leptin pretreatment upregulated BCL-2 and downregulated caspase 3 levels. The MeDIP analysis demonstrated that DNA methylation regulated LEPR gene expression in the MCAO rat model when leptin pretreatment was used. CONCLUSION: Exogenous leptin might bind to extra-activated LEPR by reducing the methylation level of the LEPR gene promoter region, which leads to an increase in phosphorylated JAK2/STAT3 and apoptotic signalling pathways.

Asunto(s)

Metilación de ADN , Janus Quinasa 2 , Leptina , Ratas Sprague-Dawley , Receptores de Leptina , Daño por Reperfusión , Factor de Transcripción STAT3 , Transducción de Señal , Animales , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Janus Quinasa 2/metabolismo , Ratas , Factor de Transcripción STAT3/metabolismo , Transducción de Señal/efectos de los fármacos , Receptores de Leptina/metabolismo , Receptores de Leptina/genética , Masculino , Leptina/metabolismo , Células PC12 , Infarto de la Arteria Cerebral Media/metabolismo , Infarto de la Arteria Cerebral Media/patología , Modelos Animales de Enfermedad , Fármacos Neuroprotectores/farmacología , Apoptosis/efectos de los fármacos , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Caspasa 3/metabolismo

RESUMEN

Reperfusion injury, which is distinct from ischaemic injury, occurs when blood flow is restored in previously ischaemic brain tissue, further compromising neurons and other cells and worsening the injury. There is currently a lack of pharmaceutical agents and therapeutic interventions that specifically mitigate cerebral ischaemia/reperfusion (I/R) injury. Ginsenoside Rg1 (Rg1), a protopanaxatriol-type saponin isolated from Panax ginseng C. A. Meyer, has been found to protect against cerebral I/R injury, but its intricate protective mechanisms remain to be elucidated. Numerous studies have shown that autophagy plays a crucial role in protecting brain tissue during the I/R process and is emerging as a promising therapeutic strategy for effective treatment. In this study, we investigated whether Rg1 protected against I/R damage in vitro and in vivo by regulating autophagy. Both MCAO and OGD/R models were established. SK-N-AS and SH-SY5Y cells were subjected to OGD followed by reperfusion with Rg1 (4-32 µM). MCAO mice were injected with Rg1 (30 mg·kg-1·d-1. i.p.) for 3 days before and on the day of surgery. Rg1 treatment significantly mitigated ischaemia/reperfusion injury both in vitro and in vivo. Furthermore, we demonstrated that the induction of autophagy contributed to I/R injury, which was effectively inhibited by Rg1 in both in vitro and in vivo models of cerebral I/R injury. Rg1 inhibited autophagy through multiple steps, including impeding autophagy initiation, inducing lysosomal dysfunction and inhibiting cathepsin enzyme activities. We revealed that mTOR activation was pivotal in mediating the inhibitory effect of Rg1 on autophagy. Treatment with Torin-1, an autophagy inducer and mTOR-specific inhibitor, significantly reversed the impact of Rg1 on autophagy, decreasing its protective efficacy against I/R injury both in vitro and in vivo. In conclusion, our results suggest that Rg1 may serve as a promising drug candidate against cerebral I/R injury by inhibiting autophagy through activation of mTOR signalling.

RESUMEN

Dual-specificity phosphatase 12 (DUSP12) is abnormally expressed under various pathological conditions and plays a crucial role in the pathological progression of disorders. However, the role of DUSP12 in cerebral ischaemia/reperfusion injury has not yet been investigated. This study explored the possible link between DUSP12 and cerebral ischaemia/reperfusion injury using an oxygen-glucose deprivation/reoxygenation (OGD/R) model. Marked decreases in DUSP12 levels have been observed in cultured neurons exposed to OGD/R. DUSP12-overexpressed neurons were resistant to OGD/R-induced apoptosis and inflammation, whereas DUSP12-deficient neurons were vulnerable to OGD/R-evoked injuries. Further investigation revealed that DUSP12 overexpression or deficiency affects the phosphorylation of apoptosis signal-regulating kinase 1 (ASK1), c-Jun NH2-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK) in neurons under OGD/R conditions. Moreover, blockade of ASK1 diminished the regulatory effect of DUSP12 deficiency on JNK and p38 MAPK activation. In addition, DUSP12-deficiency-elicited effects exacerbating neuronal OGD/R injury were reversed by ASK1 blockade. In summary, DUSP12 protects against neuronal OGD/R injury by reducing apoptosis and inflammation through inactivation of the ASK1-JNK/p38 MAPK pathway. These findings imply a neuroprotective function for DUSP12 in cerebral ischaemia/reperfusion injury.

Asunto(s)

Apoptosis , Fosfatasas de Especificidad Dual , Glucosa , Inflamación , MAP Quinasa Quinasa Quinasa 5 , Neuronas , Oxígeno , Daño por Reperfusión , Proteínas Quinasas p38 Activadas por Mitógenos , Animales , Ratones , Células Cultivadas , Fosfatasas de Especificidad Dual/metabolismo , Fosfatasas de Especificidad Dual/genética , Glucosa/metabolismo , Inflamación/metabolismo , Inflamación/patología , MAP Quinasa Quinasa Quinasa 5/metabolismo , Sistema de Señalización de MAP Quinasas , Neuronas/metabolismo , Neuronas/patología , Oxígeno/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Transducción de Señal , Proteína Quinasa 14 Activada por Mitógenos

RESUMEN

OBJECTIVE: To explore the effect of acupuncture treatment on cerebral ischaemia-reperfusion injury (CIRI) and reveal the underlying mechanism of the effect based on nuclear receptor coactivator 4 (NCOA4) mediated ferritinophagy. METHODS: Sprague-Dawley male rats were divided into four groups: the sham group, model group, acupuncture group, and sham acupuncture group. After 2 h of middle cerebral artery occlusion (MCAO), reperfusion was performed for 24 h to induce CIRI. The rats were treated with acupuncture at the Neiguan (PC6) and Shuigou (GV26) acupoints. Their neurological function was evaluated by taking their Bederson scores at 2 h after ischaemia and 24 h after reperfusion. Triphenyltetrazolium chloride staining was applied to assess the cerebral infarct volume at 24 h after reperfusion. The malondialdehyde (MDA) and ferrous iron (Fe2+) levels were observed after 24 h of reperfusion using an assay kit. Western blotting was performed to detect the expression of NCOA4 and ferritin heavy chain 1 (FTH1) at 24 h after reperfusion. Moreover, the colocalization of ferritin with neurons, NCOA4 with microtubule-associated protein 1 light chain 3 (LC3), and NCOA4 with ferritin was visualized using immunofluorescence staining. RESULTS: Acupuncture significantly improved neurological function and decreased cerebral infarct volume in the acupuncture group. Following CIRI, the expression of NCOA4, LC3 and FTH1 was increased, which enhanced ferritinophagy and induced an inappropriate accumulation of Fe2+ and MDA in the ischaemic brain. However, acupuncture dramatically downregulated the expression of NCOA4, LC3 and FTH1, inhibited the overactivation of ferritinophagy, and decreased the levels of MDA and Fe2+. CONCLUSIONS: Acupuncture can inhibit NCOA4-mediated ferritinophagy and protect neurons against CIRI in a rat model.

Asunto(s)

Terapia por Acupuntura , Isquemia Encefálica , Daño por Reperfusión , Ratas , Masculino , Animales , Ratas Sprague-Dawley , Isquemia Encefálica/genética , Isquemia Encefálica/terapia , Isquemia Encefálica/metabolismo , Infarto Cerebral , Daño por Reperfusión/genética , Daño por Reperfusión/terapia , Daño por Reperfusión/metabolismo , Ferritinas/genética , Coactivadores de Receptor Nuclear/metabolismo

RESUMEN

BACKGROUND: The gut-brain axis (GBA) plays a central role in cerebral ischaemia-reperfusion injury (CIRI). Rhubarb, known for its purgative properties, has demonstrated protective effects against CIRI. However, it remains unclear whether this protective effect is achieved through the regulation of the GBA. AIM: This study aims to investigate the mechanism by which rhubarb extract improves CIRI by modulating the GBA pathway. METHODS: We identified the active components of rhubarb extract using LC-MS/MS. The model of middle cerebral artery occlusion (MCAO) was established to evaluate the effect of rhubarb extract. We conducted 16S rDNA sequencing and untargeted metabolomics to analyze intestinal contents. Additionally, we employed HE staining, TUNEL staining, western blot, and ELISA to assess intestinal barrier integrity. We measured the levels of inflammatory cytokines in serum via ELISA. We also examined blood-brain barrier (BBB) integrity using Evans blue (EB) penetration, transmission electron microscopy (TEM), western blot, and ELISA. Neurological function scores and TTC staining were utilized to evaluate neurological outcomes. RESULTS: We identified twenty-six active components in rhubarb. Rhubarb extract enhanced α-diversity, reduced the abundance of Enterobacteriaceae, and partially rectified metabolic disorders in CIRI rats. It also ameliorated pathological changes, increased the expressions of ZO-1, Occludin, and Claudin 1 in the colon, and reduced levels of LPS and d-lac in serum. Furthermore, it lowered the levels of IL-1ß, IL-6, IL-10, IL-17, and TNF-α in serum. Rhubarb extract mitigated BBB dysfunction, as evidenced by reduced EB penetration and improved hippocampal microstructure. It upregulated the expressions of ZO-1, Occludin, Claudin 1, while downregulating the expressions of TLR4, MyD88, and NF-κB. Similarly, rhubarb extract decreased the levels of IL-1ß, IL-6, and TNF-α in the hippocampus. Ultimately, it reduced neurological function scores and cerebral infarct volume. CONCLUSION: Rhubarb effectively treats CIRI, potentially by inhibiting harmful bacteria, correcting metabolic disorders, repairing intestinal barrier function, alleviating BBB dysfunction, and ultimately improving neurological outcomes.

Asunto(s)

Isquemia Encefálica , Enfermedades Metabólicas , Fármacos Neuroprotectores , Daño por Reperfusión , Rheum , Ratas , Animales , Neuroprotección , Rheum/metabolismo , Ocludina/metabolismo , Interleucina-6 , Factor de Necrosis Tumoral alfa/genética , Eje Cerebro-Intestino , Cromatografía Liquida , Claudina-1 , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Espectrometría de Masas en Tándem , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/patología , Azul de Evans/uso terapéutico , Daño por Reperfusión/metabolismo , Enfermedades Metabólicas/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/tratamiento farmacológico

RESUMEN

Cerebral ischemia-reperfusion injury （CIRI） has a very high incidence， disability， and mortality rates， which seriously affects human life and health. In recent years， modern medicine has made some progress in the diagnosis and treatment of CIRI， but there are still problems such as difficulties in postoperative rehabilitation and adverse drug reactions， and new therapeutic drugs for CIRI are urgently needed. As an important class of active ingredients in traditional Chinese medicine， flavonoids can play antioxidant， apoptosis inhibition， anti-inflammatory， and other pharmacological effects to improve brain tissue damage， which is important for improving the quality of life of CIRI patients and slowing down the aging of the social population. Numerous studies have found that flavonoids in traditional Chinese medicine can regulate cell surface receptors Toll-like receptor 4/nuclear factor-kappaB （TLR4/NF-κB）， phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt）， adenylate-activated protein kinase/mammalian target of rapamycin protein （AMPK/mTOR）， Ras homologous gene family member A/Rho-associated coiled-coil protein kinase （RhoA/ROCK）， nuclear factor E2-associated factor 2/Kelch-like epoxychloropropane-associated protein-1/haemoglobin oxygenase 1 （Nrf2/Keap1/ HO-1）， Notch， and other signaling pathways， so as to regulate the transcription and expression of related proteins after CIRI， alleviate brain tissue injury， and improve CIRI. This paper analyzed the relevant literature in China and abroad in recent years， reviewed the mechanism of action and related pathways of flavonoids in traditional Chinese medicine to improve CIRI， and explored the new therapeutic direction of CIRI at the metabolic level， with a view to providing a basis for the further development and application of flavonoids in traditional Chinese medicine.

RESUMEN

Ischaemic stroke is a common cerebrovascular disease. Long non-coding RNA (lncRNA) of small nucleolar RNA host gene (SNHG15) has been supposedly performed a regulatory role in many diseases. Nonetheless, the function of SNHG15 in cerebral ischaemia-reperfusion injury has not been clarified. The OGD/R of Neuro2A cells simulated the ischaemic and reperfused states of the brain. Neuro2a cell line with stable transfection of plasmid with silent expression of SNHG15 was constructed. Neuro2a cell lines transfected with miR-153-3p mimic (miR-153-3p-mimics) and miR-153-3p inhibitor (miR-153-3p-inhibition) were constructed. Expression of SNHG15, mi R-200a, FOXO3 and ATG7 in mouse brain tissue and N2a cells was identified by qRT-PCR. Western blot (WB) analysis of mouse brain tissue and Neuro2a cells revealed the presence of the proteins ATG5, Cle-caspase-3, Bax, Bcl-2, LC3 II/I and P62 (WB). The representation and distribution of LC3B were observed by immunofluorescence. The death of cells was measured using a technique called flow cytometry (FACS). SNHG15 was highly expressed in cerebral ischaemia-reperfusion injury model. Down-regulation of SNHG15 lead to lower apoptosis rate and decreased autophagy. Dual luciferase assay and co-immunoprecipitation (CoIP) found lncRNA SNHG15/miR-153-3p/ATG5. Compared to cells transfected with NC suppression, cells transfected with miR-153-3p-inhibition had substantially greater overexpression of LC 3 II/I, ATG5, cle-Caspase-3, and Bax, as determined by a recovery experiment, the apoptosis rate was elevated, yet both P62 and Bcl-2 were significantly lower and LC3+ puncta per cells were significantly increased. Co-transfection of miR-153-3p-inhibition and sh-SNHG15 could reverse these results. LncRNA SNHG15 regulated autophagy and prevented cerebral ischaemia-reperfusion injury through mediating the miR-153-3p/ATG5 axis.

RESUMEN

ETHNOPHARMACOLOGICAL RELEVANCE: The combination of Alisma and Atractylodes (AA), a classical traditional Chinese herbal decoction, may protect against cerebral ischaemia/reperfusion injury (CIRI). However, the underlying mechanism has not been characterized. Intriguingly, exosomal microRNAs (miRNAs) have been recognized as vital factors in the pharmacology of Chinese herbal decoctions. AIM OF THE STUDY: The aim of the present study was to assess whether the neuroprotective effect of AA was dependent on the efficient transfer of miRNAs via exosomes in the brain. MATERIALS AND METHODS: Bilateral common carotid artery ligation (BCAL) was used to induce transient global cerebral ischaemia/reperfusion (GCI/R) in C57BL/6 mice treated with/without AA. Neurological deficits were assessed with the modified neurological severity score (mNSS) and Morris water maze (MWM) test. Western blot (WB) analysis was used to detect the expression of sirtuin 1 (SIRT1) in the cerebral cortex. The inflammatory state was quantitatively evaluated by measuring the expression of phospho-Nuclear factor kappa B (p-NF-κB), Interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) using WB analysis and glial fibrillary acidic protein (GFAP) immunohistochemical staining. The protein expression of zonula occluden-1 (ZO-1), occludin, caudin-5 and CD31 was examined by immunohistochemical staining to determine blood‒brain barrier (BBB) permeability. Exosomes were extracted from the brain interstitial space by ultracentrifugation and identified by transmission electron microscopy (TEM), WB analysis and nanoparticle tracking analysis (NTA). The origin of exosomes was clarified by measuring the specific mRNAs within exosomes via Real Time Quantitative PCR (RT‒qPCR). Differential miRNAs in exosomes were identified using microarray screening and were validated by RT‒qPCR. Exosomes were labelled with fluorescent dye (PKH26) and incubated with bEnd.3 cells, the supernatant was collected, IL-1ß/TNF-α expression was measured using enzyme-linked immunosorbent assay (ELISA), total RNA was extracted, and miR-200a-3p/141-3p expression was examined by RT‒qPCR. In addition, the levels of miR-200a-3p/141-3p in oxygen glucose deprivation/reoxygenation (OGD/R)-induced bEnd.3 cells were quantified. The direct interaction between miR-200a-3p/141-3p and the SIRT1 3' untranslated region (3'UTR) was measured by determining SIRT1 expression in bEnd.3 cells transfected with the miR-200a-3p/141-3p mimic/inhibitor. RESULTS: Severe neurological deficits and memory loss caused by GCI/R in mice was markedly ameliorated by AA treatment, particularly in the AA medium-dose group. Moreover, AA-treated GCI/R-induced mice showed significant increases in SIRT1, ZO-1, occludin, caudin-5, and CD31 expression levels and decreases in p-NF-κB, IL-1ß, TNF-α, and GFAP expression levels compared with those in untreated GCI/R-induced mice. Furthermore, we found that miR-200a-3p/141-3p was enriched in astrocyte-derived exosomes from GCI/R-induced mice and could be inhibited by treatment with a medium dose of AA. The exosomes mediated the transfer of miR-200a-3p/141-3p into bEnd.3 cells, promoted IL-1ß and TNF-α release and downregulated the expression of SIRT1. No significant changes in the levels of miR-200a-3p/141-3p were observed in OGD/R-induced bEnd.3 cells. The miR-200a-3p/141-3p mimic/inhibitor decreased/increased SIRT1 expression in bEnd.3 cells, respectively. CONCLUSION: Our findings demonstrated that AA attenuated inflammation-mediated CIRI by inhibiting astrocyte-derived exosomal miR-200a-3p/141-3p by targeting the SIRT1 gene, which provided further evidence and identified a novel regulatory mechanism for the neuroprotective effects of AA.

Asunto(s)

Alisma , Atractylodes , Isquemia Encefálica , MicroARNs , Daño por Reperfusión , Ratones , Animales , Sirtuina 1/genética , Alisma/genética , Alisma/metabolismo , FN-kappa B , Factor de Necrosis Tumoral alfa/farmacología , Células Endoteliales/metabolismo , Astrocitos/metabolismo , Ocludina , Ratones Endogámicos C57BL , MicroARNs/metabolismo , Isquemia Encefálica/metabolismo , Daño por Reperfusión/metabolismo , Apoptosis

RESUMEN

As an endogenous gas signalling molecule, hydrogen sulfide (H2S) is frequently present in a variety of mammals and plays a significant role in the cardiovascular and nervous systems. Reactive oxygen species (ROS) are produced in large quantities as a result of cerebral ischaemia-reperfusion, which is a very serious class of cerebrovascular diseases. ROS cause oxidative stress and induce specific gene expression that results in apoptosis. H2S reduces cerebral ischaemia-reperfusion-induced secondary injury via anti-oxidative stress injury, suppression of the inflammatory response, inhibition of apoptosis, attenuation of cerebrovascular endothelial cell injury, modulation of autophagy, and antagonism of P2X7 receptors, and it plays an important biological role in other cerebral ischaemic injury events. Despite the many limitations of the hydrogen sulfide therapy delivery strategy and the difficulty in controlling the ideal concentration, relevant experimental evidence demonstrating that H2S plays an excellent neuroprotective role in cerebral ischaemia-reperfusion injury (CIRI). This paper examines the synthesis and metabolism of the gas molecule H2S in the brain as well as the molecular mechanisms of H2S donors in cerebral ischaemia-reperfusion injury and possibly other unknown biological functions. With the active development in this field, it is expected that this review will assist researchers in their search for the potential value of hydrogen sulfide and provide new ideas for preclinical trials of exogenous H2S.

Asunto(s)

Lesiones Encefálicas , Isquemia Encefálica , Sulfuro de Hidrógeno , Daño por Reperfusión , Animales , Sulfuro de Hidrógeno/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Estrés Oxidativo , Infarto Cerebral/tratamiento farmacológico , Isquemia Encefálica/tratamiento farmacológico , Lesiones Encefálicas/tratamiento farmacológico , Mamíferos

RESUMEN

OBJECTIVES: This study aimed to observe the effect of the combination of astragaloside IV (AST IV) and Panax notoginseng saponins (PNS) on cerebral ischaemia-reperfusion injury (CIRI) and explore the specific mechanism of the nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated combination of AST IV and PNS against CIRI based on ferroptosis and inflammatory response. METHODS: The therapeutic effect and mechanism of AST IV and PNS were evaluated by constructing a Sprague-Dawley rat middle cerebral artery ischaemia-occlusion-reperfusion model. The specific mechanism of the combination of AST IV and PNS against CIRI was revealed through the combined intervention of the Nrf2-specific inhibitor brusatol. KEY FINDINGS: After AST IV and PNS treatment, the cerebral infarction area of the rats was reduced; behavioural performance was improved; Fe2+, malondialdehyde, lipid peroxidation, interleukin-6, interleukin-1ß, tumour necrosis factor-α and myeloperoxidase levels were reduced; and glutathione and glutathione peroxidase 4 levels were increased. In addition, the expression of Nrf2 was significantly increased, the combined treatment was more effective than the single treatment, and the Nrf2 inhibitor brusatol could reverse the effects of the combined intervention of AST IV and PNS. CONCLUSIONS: The findings of this study suggest that combining AST IV and PNS attenuates CIRI by activating Nrf2 to inhibit ferroptosis and inflammatory responses.

Asunto(s)

Ferroptosis , Panax notoginseng , Daño por Reperfusión , Saponinas , Ratas , Animales , Panax notoginseng/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Ratas Sprague-Dawley , Saponinas/farmacología , Saponinas/uso terapéutico , Inflamación/tratamiento farmacológico , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/prevención & control , Daño por Reperfusión/metabolismo

RESUMEN

Cerebral ischaemia-reperfusion injury (CIRI) is a critical component of ischaemic stroke pathogenesis. Ferroptosis contributes to and aggravates CIRI, whereas the P62/Kelch-like ECH-associated protein 1 (Keap1)/NF-E2-related factor 2 (Nrf2) pathway exerts neuroprotective effects. Astragaloside IV (AST IV) is the primary active ingredient of Astragalus, an herb with anti-CIRI properties used in traditional Chinese medicine. However, the mechanism of its anti-CIRI action is unclear. This study examined the mechanisms underlying the anti-CIRI action of AST IV using a combination of in vitro and in vivo approaches. We established an erastin-induced ferroptosis model, oxygen and glucose deprivation/reoxygenation (OGD/R)-induced model in SH-SY5Y cells, and middle cerebral artery occlusion-reperfusion (MCAO/R) model using Sprague-Dawley rats. The extent of cell damage and brain damage in rats, ferroptosis indicator changes, and expression of P62, Keap1, and Nrf2 were investigated. AST IV inhibited erastin-induced ferroptosis, attenuated OGD/R-induced cell damage, and ameliorated sensorimotor dysfunction and injury in the MCAO/R model. Further, AST IV promoted Nrf2 activation, inhibited ferroptosis, and reduced cell damage. Notably, these effects were inhibited by ML385, an Nrf2 inhibitor. AST IV increased the P62 and Nrf2 levels and decreased the Keap1 levels. P62 silencing reduced the effects of AST IV on the P62/Keap1/Nrf2 pathway and ferroptosis. Our findings suggest that AST IV mitigates CIRI by inhibiting ferroptosis via activation of the P62/Keap1/Nrf2 pathway. This study provides an important scientific basis and direction for the application and research of AST IV and provides new potential targets and ideas for the study of the pathological mechanism of CIRI.

Asunto(s)

Isquemia Encefálica , Ferroptosis , Neuroblastoma , Daño por Reperfusión , Accidente Cerebrovascular , Ratas , Humanos , Animales , Factor 2 Relacionado con NF-E2/metabolismo , Ratas Sprague-Dawley , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Transducción de Señal , Oxígeno/metabolismo , Daño por Reperfusión/metabolismo

RESUMEN

Cerebral ischaemia/reperfusion (I/R) injury is caused by blood flow restoration after an ischaemic insult, and effective treatments targeting I/R injury are still insufficient. Oxidative stress plays a critical role in the pathogenesis of cerebral I/R injury. This study investigated whether vitamin D receptor (VDR) could inhibit oxidative stress caused by cerebral I/R injury and explored the detailed mechanism. VDR was highly expressed in brain tissues of mice with cerebral I/R injury. Pretreatment with the active vitamin D calcitriol and synthetic vitamin D analogue paricalcitol (PC) reduced autophagy and apoptosis, improved neurological deficits and decreased infarct size in mice after cerebral I/R. Calcitriol or PC upregulated VDR expression to prevent cerebral I/R injury by affecting oxidative stress. Silencing of VDR reversed the protective effects of calcitriol or PC on brain tissues in mice with cerebral I/R. The bioinformatics analysis revealed that VDR interacted with SMAD family member 3 (SMAD3). It was validated through the chromatin immunoprecipitation assay that SMAD3 can bind to the VDR promoter and VDR can bind to the SMAD3 promoter. Collectively, these findings provide evidence that reciprocal activation between SMAD3 and VDR transcription factors defines vitamin D-mediated oxidative stress to prevent cerebral I/R injury.

Asunto(s)

Isquemia Encefálica , Daño por Reperfusión , Animales , Ratones , Receptores de Calcitriol/genética , Receptores de Calcitriol/metabolismo , Calcitriol/farmacología , Calcitriol/uso terapéutico , Vitamina D/farmacología , Vitamina D/uso terapéutico , Estrés Oxidativo , Daño por Reperfusión/tratamiento farmacológico , Isquemia Encefálica/tratamiento farmacológico

RESUMEN

BACKGROUND: Previous data have reported that Sentrin/SUMO-specific protease 6 (SENP6) is involved in ischaemic brain injury and induces neuronal apoptosis after cerebral ischaemia, but the role of SENP6 in microglia-induced neuroinflammation and its underlying mechanism remain poorly understood. This research systematically explored the function and potential mechanism of SENP6 in microglia-induced neuroinflammation after ischaemic stroke. RESULTS: We first identified an increased protein level of SENP6 in microglia after cerebral ischaemia. Then, we demonstrated that SENP6 promoted detrimental microglial phenotype polarization. Specifically, SENP6-mediated de-SUMOylation of ANXA1 targeted the IκB kinase (IKK) complex and selectively inhibited the autophagic degradation of IKKα in an NBR1-dependent manner, activating the NF-κB pathway and enhancing proinflammatory cytokine expression. In addition, downregulation of SENP6 in microglia effectively reduced cocultured neuronal damage induced by ischaemic stroke. More importantly, we employed an AAV-based technique to specifically knockdown SENP6 in microglia/macrophages, and in vivo experiments showed that SENP6 inhibition in microglia/macrophages notably lessened brain ischaemic infarct size, decreased neurological deficit scores, and ameliorated motor and cognitive function in mice subjected to cerebral ischaemia surgery. CONCLUSION: We demonstrated a previously unidentified mechanism by which SENP6-mediated ANXA1 de-SUMOylation regulates microglial polarization and our results strongly indicated that in microglia, inhibition of SENP6 may be a crucial beneficial therapeutic strategy for ischaemic stroke.

RESUMEN

BACKGROUND: Cerebral ischaemia-reperfusion injury (CIRI) is a common disease characterized by severe attacks and a high disabling rate worldwide. Oxidative stress injury has been proposed as a major risk factor for CIRI. Ginkgo biloba extract (GBE) has been shown to elicit vascular protective effects, the main components of which are Ginkgo flavonoids (GF) and ginkgolides (GL). Our previous study showed that GF and GL played a central role in protecting CIRI, but the mechanism remains unclear. This study aimed to further reveal the protective effect mechanism of GF and GL in rats with CIRI. METHODS: The antioxidant activity in vitro was assessed by the DPPH method. The model used in this study was established by middle cerebral artery occlusion (MCAO) and reperfusion; the level of CIRI was assessed by nerve function score and TTC staining; we measured the oxidative stress indices in the brain cortex, including LDH, GSH-Px, and the protein contents of Akt, p-Akt, Nrf2, and HO-1; HPLC-MS was used to detect drug concentrations in rat plasma at different times after administration of GF and GL; and the pharmacokinetic parameters of each component were calculated by Drug and Statistic Version 3.2.6 (DAS 3.2.6) software and SPSS 17.0. RESULTS: Regarding the DPPH free radical scavenging ability, GF performed better free radical scavenging ability than GL. In terms of the nerve function score and TTC staining, there were no statistically significant differences among the GF, GL and combined groups; however, there were significant differences in reducing the activity of LDH and increasing the activity of GSH-Px in the three administration groups. For the expression of Akt, p-Akt, Nrf2, and HO-1, the combined group had a significant effect compared with that in the GF or GL group. In addition, there was a significant multicomponent interaction in vivo in the combined group compared with the GF or GL group. CONCLUSION: After GF and GL were used in combination, the effect of anti-CIRI was more pronounced. This result indicated that GF and GL might improve CIRI by activating the PI3K/Akt/Nrf2 signalling pathway and promoting multicomponent interactions in vivo.

RESUMEN

INTRODUCTION: The study aimed to explore the effects of treatment with black bamboo rhizome extracts on learning and memory and determine the underlying mechanisms in rats with cerebral ischaemia-reperfusion injury. METHODS: Sprague-Dawley rats were randomly divided into the following four groups: control, middle cerebral artery occlusion (MCAO), low-dose drug, and high-dose drug groups. Rats underwent MCAO using a suture method before drug treatment. Then, neurological impairment was assessed using the Longa scoring method, and triphenyl tetrazolium chloride staining was used to analyse the cerebral infarction area. The Elliott formula was used to calculate water content in the brain tissue. A Morris water maze (MWM) was used to assess changes in learning and memory abilities, and Western blotting was used to detect cyclic adenosine phosphate response element-binding protein (CREB) and brain-derived neurotrophic factor (BDNF) expression in the hippocampus of MCAO rats. RESULTS: After treatment with black bamboo rhizome extracts, the neurological dysfunction score was lower in the drug groups than in the MCAO group, and a significant difference was observed between the high-dose drug and MCAO groups (P<0.05). Additionally, the cerebral infarction area was significantly smaller in the drug groups than in the MCAO group (P<0.01), and the effect was more obvious in the high-dose drug group than in the low-dose drug group. There was also a significant difference in water content between the high-dose drug and MCAO groups, and cerebral oedema was significantly reduced in the high-dose drug group (P<0.05). In the MWM, the incubation period was significantly reduced, the number of platform crossings was significantly increased, and the search time was prolonged in the drug groups compared with those in the MCAO group (P<0.05). Moreover, the expression of BDNF and CREB was significantly increased in the drug groups compared to that in the MCAO group, and the increase was more obvious in the high-dose group than in the low-dose group (P<0.05). DISCUSSION: Black bamboo rhizome extracts significantly improved cognitive dysfunction, reduced cerebral oedema, decreased the cerebral infarction area, and improved the neurological function score and learning and memory abilities in rats with cerebral ischaemia-reperfusion injury.

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OBJECTIVES: The study aimed to determine whether dental pulp stem cell-derived exosomes (DPSC-Exos) exert protective effects against cerebral ischaemia-reperfusion (I/R) injury and explore its underlying mechanism. MATERIALS AND METHODS: Exosomes were isolated from the culture medium of human DPSC. Adult male C57BL/6 mice were subjected to 2 hours transient middle cerebral artery occlusion (tMCAO) injury followed by 2 hours reperfusion, after which singular injection of DPSC-Exos via tail vein was administrated. Brain oedema, cerebral infarction and neurological impairment were measured on day 7 after exosomes injection. Then, oxygen-glucose deprivation-reperfusion (OGD/R) induced BV2 cells were studied to analyse the therapeutic effects of DPSC-Exos on I/R injury in vitro. Protein levels of TLR4, MyD88, NF-κB p65, HMGB1, IL-6, IL-1ß and TNF-α were determined by western blot or enzyme-linked immunosorbent assay. The cytoplasmic translocation of HMGB1 was detected by immunofluorescence staining. RESULTS: DPSC-Exos alleviated brain oedema, cerebral infarction and neurological impairment in I/R mice. DPSC-Exos inhibited the I/R-mediated expression of TLR4, MyD88 and NF-κB significantly. DPSC-Exos also reduced the protein expression of IL-6, IL-1ß and TNF-α compared with those of the control both in vitro and in vivo. Meanwhile, DPSC-Exos markedly decreased the HMGB1 cytoplasmic translocation induced by I/R damage. CONCLUSIONS: DPSC-Exos can ameliorate I/R-induced cerebral injury in mice. Its anti-inflammatory mechanism might be related with the inhibition of the HMGB1/TLR4/MyD88/NF-κB pathway.

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Citocinas/metabolismo , Exosomas/trasplante , Daño por Reperfusión/terapia , Animales , Supervivencia Celular , Citoplasma/metabolismo , Pulpa Dental/citología , Pulpa Dental/metabolismo , Modelos Animales de Enfermedad , Exosomas/metabolismo , Proteína HMGB1/metabolismo , Inflamación/metabolismo , Inflamación/terapia , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Microglía/citología , Microglía/metabolismo , Factor 88 de Diferenciación Mieloide/metabolismo , Daño por Reperfusión/metabolismo , Células Madre/citología , Células Madre/metabolismo , Receptor Toll-Like 4/metabolismo , Factor de Transcripción ReIA/metabolismo

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Autophagy is closely associated with cerebral ischaemia/reperfusion injury, but the underlying mechanisms are unknown. We investigated whether Spautin-1 ameliorates cerebral ischaemia/reperfusion injury by inhibiting autophagy and whether its derived pyroptosis is involved in this process. We explored the mechanism of Spautin-1 in cerebral ischaemia/reperfusion. To answer these questions, healthy male Sprague-Dawley rats were exposed to middle cerebral artery occlusion for 60 minutes followed by reperfusion for 24 hours. We found that cerebral ischaemia/reperfusion increased the expression levels of autophagy and pyroptosis-related proteins. Treatment with Spautin-1 reduced the infarct size and water content and restored some neurological functions. In vitro experiments were performed using oxygen-glucose deprivation/reoxygenation to model PC12 cells. The results showed that PC12 cells showed a significant decrease in cell viability and a significant increase in ROS and autophagy levels. Spautin-1 treatment reduced autophagy and ROS accumulation and attenuated NLRP3 inflammasome-dependent pyroptosis. However, these beneficial effects were greatly blocked by USP13 overexpression, which significantly counteracted the inhibition of autophagy and NLRP3 inflammasome-dependent ferroptosis by Spautin-1. Together, these results suggest that Spautin-1 may ameliorate cerebral ischaemia-reperfusion injury via the autophagy/pyroptosis pathway. Thus, inhibition of autophagy may be considered as a promising therapeutic approach for cerebral ischaemia-reperfusion injury.

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Autofagia , Isquemia Encefálica/prevención & control , Modelos Animales de Enfermedad , Infarto de la Arteria Cerebral Media/complicaciones , Fármacos Neuroprotectores/farmacología , Piroptosis , Daño por Reperfusión/prevención & control , Animales , Isquemia Encefálica/etiología , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Masculino , Células PC12 , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/etiología , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología

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This study aims to investigate the protective effect of roflumilast, a phosphodiesterase (PDE)-4 enzyme inhibitor, and demonstrate its possible role in the development prevention of cerebral ischemia/reperfusion injury (CI/RI) after stroke induced by carotid artery ligation in juvenile rats. The rats were randomly divided into five groups: healthy group without any treatment, healthy group administered with 1 mg/kg roflumilast, CI group not administered with roflumilast, CI group administered with 0.5 mg/kg roflumilast, and CI group administered with 1 mg/kg roflumilast. In the CI groups, reperfusion was achieved 2h after ischemia induction; in the roflumilast groups, this drug was intraperitoneally administered immediately after reperfusion and at the 12th hour. At the end of 24h, the rats were sacrificed and their brain tissues removed for examination. The mRNA expressions obtained with real-time PCR of IL-1ß, TNF-α, and NLRP3 significantly increased in the CI/RI-induced groups compared with the control group, and this increase was significantly lower in the groups administered with roflumilast compared with the CI/RI-induced groups. Moreover, ELISA revealed that both IL-1 ß and IL-6 brain levels were significantly higher in the CI/RI-induced groups than in the controls. This increase was significantly lower in the groups administered with roflumilast compared with the CI/RI-induced groups. Histopathological studies revealed that the values closest to those of the healthy group were obtained from the roflumilast groups. Nissl staining revealed that the Nissl bodies manifested normal density in the healthy and roflumilast-administered healthy groups, but were rare in the CI/RI-induced groups. Roflumilast treatment increased these decreased Nissl bodies with increasing doses. Observations indicated that the Nissl body density was close to the value in the healthy group in the CI/RI-induced group administered with 1 mg/kg roflumilast. Overall, roflumilast reduced cellular damage caused by CI/RI in juvenile rats, and this effect may be mediated by NLRP3.

Asunto(s)

Aminopiridinas , Benzamidas , Fármacos Neuroprotectores , Animales , Encéfalo , Ciclopropanos , Masculino , Ratas , Daño por Reperfusión

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Cerebral ischaemia/reperfusion (CI/R) injury is a major challenge due to the lack of effective neuroprotective drugs. Hederagenin (HE) is the aglycone part of saponins extracted from Hedera helix Linné that has exhibited anti-apoptotic and anti-inflammatory effects; however, the role of HE in CI/R has not been elucidated. In this study, mice were intraperitoneally (i.p.) injected with HE (26.5, 53, or 106 µmol/kg body weight) for 3 days after middle cerebral artery occlusion (MCAO). Neural function and brain infarct volume were evaluated. HE treatment attenuated CI/R-induced apoptosis and inflammatory cytokine expression within the infarcted areas. HE treatment also decreased the activation of the MLK3 signalling pathway, which potentiates CI/R damage via the MAPK and NFκB pathways. Due to HE's safety profile, it has potential to be used for the clinical treatment of ischaemic stroke.