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1.
Mol Neurobiol ; 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32613467

RESUMO

Blood-brain barrier disruption is one of the hallmarks of multiple sclerosis. Mesenchymal stem cells showed great potential for the multiple sclerosis therapy. However, the effect of mesenchymal stem cells on blood-brain barrier in multiple sclerosis remains unclear. Here, we investigated whether mesenchymal stem cells transplantation protected blood-brain barrier integrity and further explored possible underlying mechanisms. Adult female C57BL/6 mice were immunized with myelin oligodendrocyte glycoprotein peptide33-55 (MOG33-55) to induce experimental autoimmune encephalomyelitis (EAE). Mesenchymal stem cells (5 × 105) were transplanted via tail vein at disease onset. In the cell culture, we examined lipopolysaccharide-induced AQP4 upregulation in astrocytes. Results indicated that mesenchymal stem cells therapy improved neurobehavioral outcomes in EAE mice, reduced inflammatory cell infiltration, IgG protein leakage, and demyelination in spinal cord. Mesenchymal stem cells therapy also increased tight junction protein expression. In addition, mesenchymal stem cells downregulated AQP4 and A2B adenosine receptor (A2BAR) expression in EAE mice in spinal cord. We found that MSCs-conditioned medium (MCM) reduced the expression of inflammatory cytokines, AQP4 and A2BAR in lipopolysaccharide-activated astrocytes. BAY-60-6583 (a selective A2BAR agonist) reversed the MCM-induced AQP4 downregulation and increased p38 MAPK phosphorylation. Furthermore, the upregulation effects of A2BAR agonist were eliminated when treated with p38 MAPK inhibitor SB203580. Thus, we concluded that mesenchymal stem cells alleviated blood-brain barrier disruption by downregulating AQP4 in multiple sclerosis, possibly through inhibiting the A2BAR/p38 MAPK signaling pathway. Our work suggests that mesenchymal stem cells exert beneficial effect through maintaining blood-brain barrier integrity in EAE mice.

2.
J Neuroinflammation ; 17(1): 164, 2020 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-32450881

RESUMO

BACKGROUND: Farnesoid X receptor (FXR) is a nuclear receptor that plays a critical role in controlling cell apoptosis in diverse diseases. Previous studies have shown that knocking out FXR improved cardiac function by reducing cardiomyocyte apoptosis in myocardial ischemic mice. However, the role of FXR after cerebral ischemia remains unknown. In this study, we explored the effects and mechanisms of FXR knockout (KO) on the functional recovery of mice post cerebral ischemia-reperfusion. METHODS: Adult male C57BL/6 wild type and FXR KO mice were subjected to 90-min transient middle cerebral artery occlusion (tMCAO). The mice were divided into five groups: sham, wild-type tMCAO, FXR KO tMCAO, wild-type tMCAO treated with calcium agonist Bayk8644, and FXR KO tMCAO treated with Bayk8644. FXR expression was examined using immunohistochemistry and Western blot. Brain infarct and brain atrophy volume were examined at 3 and 14 days after stroke respectively. Neurobehavioral tests were conducted up to 14 days after stroke. The protein levels of apoptotic factors (Bcl-2, Bax, and Cleaved caspase-3) and mRNA levels of pro-inflammatory factors (TNF-α, IL-6, IL-1ß, IL-17, and IL-18) were examined using Western blot and RT-PCR. TUNEL staining and calcium imaging were obtained using confocal and two-photon microscopy. RESULTS: The expression of FXR was upregulated after ischemic stroke, which is located in the nucleus of the neurons. FXR KO was found to reduce infarct volume and promote neurobehavioral recovery following tMCAO compared to the vehicle. The expression of apoptotic and pro-inflammatory factors decreased in FXR KO mice compared to the control. The number of NeuN+/TUNEL+ cells declined in the peri-infarct area of FXR KO mice compared to the vehicle. We further demonstrated that inhibition of FXR reduced calcium overload and addition of ionomycin could reverse this neuroprotective effect in vitro. What is more, in vivo results showed that enhancement of intracellular calcium concentrations could aggravate ischemic injury and reverse the neuroprotective effect of FXR KO in mice. CONCLUSIONS: FXR KO can promote neurobehavioral recovery and attenuate ischemic brain injury, inflammatory release, and neuronal apoptosis via reducing calcium influx, suggesting its role as a therapeutic target for stroke treatments.

3.
Medicine (Baltimore) ; 99(9): e19247, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32118730

RESUMO

RATIONALE: The capsular warning syndrome (CWS) is a rare and special type of transient ischemic attacks (TIAs) syndrome. The pathophysiology of CWS is very complicate, and intracranial atherosclerotic stenosis (ICAS) is rare cause. Moreover, the effective and standard therapy has not yet been established. PATIENT CONCERNS: A 47-year-old man experienced repeated and exacerbated TIAs of right hemiparesis and dysarthria. Fourteen hours after the first episode of TIAs, he developed more severe right hemiparesis and dysarthria, the National Institute of Health Stroke Scale (NIHSS) score was 12 points, and did not recover in a long time. DIAGNOSIS: The computed tomography (CT) angiography displayed high stenosis in the M1 segment of the left middle cerebral artery. The patient was diagnosed as CWS with ICAS. INTERVENTIONS: Loading dose of clopidogrel and aspirin were started but were ineffective, then we used recombinant tissue plasminogen (r-tPA) for thrombolysis therapy after repeat CT scan that showed small acute infarcts in the right putamen and no bleeding. OUTCOMES: The patient was successfully treated by r-tPA intravenous thrombolysis after loading dose of dual-anti-platelet. He recovered rapidly, and the NIHSS score was 0 point, modified Rankin Scale score was 0 point, and Barthel Index score was 100 points at 3-month follow-up. LESSONS: r-tPA combined with loading dose of dual antiplatelet appears safe and effective in carefully selected CWS patients with ICAS. The collection of similar cases and further randomized controlled trial research would be desirable.


Assuntos
Fibrinolíticos/uso terapêutico , Arteriosclerose Intracraniana/tratamento farmacológico , Ataque Isquêmico Transitório/tratamento farmacológico , Aspirina/administração & dosagem , Aspirina/uso terapêutico , Clopidogrel/administração & dosagem , Clopidogrel/uso terapêutico , Angiografia por Tomografia Computadorizada , Diagnóstico Diferencial , Fibrinolíticos/administração & dosagem , Humanos , Arteriosclerose Intracraniana/complicações , Arteriosclerose Intracraniana/diagnóstico por imagem , Ataque Isquêmico Transitório/complicações , Ataque Isquêmico Transitório/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Artéria Cerebral Média/diagnóstico por imagem , Síndrome , Ativador de Plasminogênio Tecidual/administração & dosagem , Ativador de Plasminogênio Tecidual/uso terapêutico , Tomografia Computadorizada por Raios X
4.
CNS Neurosci Ther ; 26(4): 416-429, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32154670

RESUMO

INTRODUCTION: Clearance of damaged cells and debris is beneficial for the functional recovery after ischemic brain injury. However, the specific phagocytic receptor that mediates microglial phagocytosis after ischemic stroke is unknown. AIM: To investigate whether P2Y6 receptor-mediated microglial phagocytosis is beneficial for the debris clearance and functional recovery after ischemic stroke. RESULTS: The expression of the P2Y6 receptor in microglia increased within 3 days after transient middle cerebral artery occlusion. Inhibition of microglial phagocytosis by the selective inhibitor MRS2578 enlarged the brain atrophy and edema volume after ischemic stroke, subsequently aggravated neurological function as measured by modified neurological severity scores and Grid walking test. MRS2578 treatment had no effect on the expression of IL-1α, IL-1ß, IL-6, IL-10, TNF-α, TGF-ß, and MPO after ischemic stroke. Finally, we found that the expression of myosin light chain kinase decreased after microglial phagocytosis inhibition in the ischemic mouse brain, which suggested that myosin light chain kinase was involved in P2Y6 receptor-mediated phagocytosis. CONCLUSION: Our results indicate that P2Y6 receptor-mediated microglial phagocytosis plays a beneficial role during the acute stage of ischemic stroke, which can be a therapeutic target for ischemic stroke.

6.
Cell Death Dis ; 11(1): 9, 2020 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-31907363

RESUMO

Blood-brain barrier damage is a critical pathological feature of ischemic stroke. Oligodendrocyte precursor cells are involved in maintaining blood-brain barrier integrity during the development. However, whether oligodendrocyte precursor cell could sustain blood-brain barrier permeability during ischemic brain injury is unknown. Here, we investigate whether oligodendrocyte precursor cell transplantation protects blood-brain barrier integrity and promotes ischemic stroke recovery. Adult male ICR mice (n = 68) underwent 90 min transient middle cerebral artery occlusion. After ischemic assault, these mice received stereotactic injection of oligodendrocyte precursor cells (6 × 105). Oligodendrocyte precursor cells transplantation alleviated edema and infarct volume, and promoted neurological recovery after ischemic stroke. Oligodendrocyte precursor cells reduced blood-brain barrier leakage via increasing claudin-5, occludin and ß-catenin expression. Administration of ß-catenin inhibitor blocked the beneficial effects of oligodendrocyte precursor cells. Wnt7a protein treatment increased ß-catenin and claudin-5 expression in endothelial cells after oxygen-glucose deprivation, which was similar to the results of the conditioned medium treatment of oligodendrocyte precursor cells on endothelial cells. We demonstrated that oligodendrocyte precursor cells transplantation protected blood-brain barrier in the acute phase of ischemic stroke via activating Wnt/ß-catenin pathway. Our results indicated that oligodendrocyte precursor cells transplantation was a novel approach to the ischemic stroke therapy.

7.
Theranostics ; 10(1): 74-90, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31903107

RESUMO

Microglial activation participates in white matter injury after cerebral hypoperfusion. However, the underlying mechanism is unclear. Here, we explore whether activated microglia aggravate white matter injury via complement C3-C3aR pathway after chronic cerebral hypoperfusion. Methods: Adult male Sprague-Dawley rats (n = 80) underwent bilateral common carotid artery occlusion for 7, 14, and 28 days. Cerebral vessel density and blood flow were examined by synchrotron radiation angiography and three-dimensional arterial spin labeling. Neurobehavioral assessments, CLARITY imaging, and immunohistochemistry were performed to evaluate activation of microglia and C3-C3aR pathway. Furthermore, C3aR knockout mice were used to establish the causal relationship of C3-C3aR signaling on microglia activation and white matter injury after hypoperfusion. Results: Cerebral vessel density and blood flow were reduced after hypoperfusion (p<0.05). Spatial learning and memory deficits and white matter injury were shown (p<0.05). These impairments were correlated with aberrant microglia activation and an increase in the number of reactive microglia adhering to and phagocytosed myelin in the hypoperfusion group (p<0.05), which were accompanied by the up-regulation of complement C3 and its receptors C3aR (p<0.05). Genetic deletion of C3ar1 significantly inhibited aberrant microglial activation and reversed white matter injury after hypoperfusion (p<0.05). Furthermore, the C3aR antagonist SB290157 decreased the number of microglia adhering to myelin (p<0.05), attenuated white matter injury and cognitive deficits in chronic hypoperfusion rats (p<0.05). Conclusions: Our results demonstrated that aberrant activated microglia aggravate white matter injury via C3-C3aR pathway during chronic hypoperfusion. These findings indicate C3aR plays a critical role in mediating neuroinflammation and white matter injury through aberrant microglia activation, which provides a novel therapeutic target for the small vessel disease and vascular dementia.

8.
Stroke ; 51(2): 619-627, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31822249

RESUMO

Background and Purpose- Blood-brain barrier (BBB) disruption is a critical pathological feature after stroke. MicroRNA-126 (miR-126) maintains BBB integrity by regulating endothelial cell function during development. However, the role of miR-126-3p and -5p in BBB integrity after stroke is unclear. Here, we investigated whether miR-126-3p and -5p overexpression regulates BBB integrity after cerebral ischemia. Methods- A lentivirus carrying genes encoding miR-126-3p or -5p was stereotactically injected into adult male Institute of Cancer Research mouse brains (n=36). Permanent middle cerebral artery occlusion was performed 2 weeks after virus injection. Brain infarct volume, edema volume, and modified neurological severity score were assessed at 1 and 3 days after ischemia. Immunostaining of ZO-1 (zonula occludens-1) and occludin was used to evaluate BBB integrity. IL-1ß (interleukin-1ß), TNF-α (tumor necrosis factor-α), VCAM-1 (vascular cell adhesion molecule-1), and E-selectin expression levels were determined by real-time polymerase chain reaction and Western blot analysis. Results- The expression of miR-126-3p and -5p decreased at 1 and 3 days after ischemia (P<0.05). Injection of lentiviral miR-126-3p or -5p reduced brain infarct volume and edema volume (P<0.05) and attenuated the decrease in ZO-1/occludin protein levels and IgG leakage at 3 days after stroke (P<0.05). Injection of lentiviral miR-126-5p improved behavioral outcomes at 3 days after stroke (P<0.05). miR-126-3p and -5p overexpression downregulated the expression of proinflammatory cytokines IL-1ß and TNF-α and adhesion molecules VCAM-1 and E-selectin, as well as decreased MPO+ (myeloperoxidase positive) cell numbers at 3 days after ischemia (P<0.05). Conclusions- miR-126-3p and -5p overexpression reduced the expression of proinflammatory cytokines and adhesion molecules, and attenuated BBB disruption after ischemic stroke, suggesting that miR-126-3p and -5p are new therapeutic targets in the acute stage of stroke.

9.
J Cereb Blood Flow Metab ; : 271678X19892777, 2019 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-31865842

RESUMO

Endothelial progenitor cell transplantation is a potential therapeutic approach in brain ischemia. However, whether the therapeutic effect of endothelial progenitor cells is via affecting complement activation is unknown. We established a mouse focal ischemia model (n = 111) and transplanted endothelial progenitor cells into the peri-infarct region immediately after brain ischemia. Neurological outcomes and brain infarct/atrophy volume were examined after ischemia. Expression of C3, C3aR and pro-inflammatory factors were further examined to explore the role of endothelial progenitor cells in ischemic brain. We found that endothelial progenitor cells improved neurological outcomes and reduced brain infarct/atrophy volume after 1 to 14 days of ischemia compared to the control (p < 0.05). C3 and C3aR expression in the brain was up-regulated at 1 day up to 14 days (p < 0.05). Endothelial progenitor cells reduced astrocyte-derived C3 (p < 0.05) and C3aR expression (p < 0.05) after ischemia. Endothelial progenitor cells also reduced inflammatory response after ischemia (p < 0.05). Endothelial progenitor cell transplantation reduced astrocyte-derived C3 expression in the brain after ischemic stroke, together with decreased C3aR and inflammatory response contributing to neurological function recovery. Our results indicate that modulating complement C3/C3aR pathway is a novel therapeutic target for the ischemic stroke.

10.
Brain Res ; 1724: 146440, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31513789

RESUMO

BACKGROUND: Previous studies have estimated that the risk of recurrent stroke was nearly 20% shortly after a transient ischemic attack (TIA) or minor stroke. A missing or hypoplastic (<0.5 mm) anterior communicating artery can have deleterious effects on the brain. Our study aimed to investigate the effect of anterior communicating artery flow on neurovascular injury and neurobehavioral outcomes in mice with recurrent stroke and to identify its underlying mechanisms. METHODS: A recurrent stroke model was established by an initial cortical infarction followed by a corticostriatal infarction 3 days later. The vascular structure was visualized using synchrotron radiation angiography & magnetic resonance angiography in vivo and transparent endovascular perfusion imaging in vitro. Microvessel perfusion was assessed via fluorescein isothiocyanate perfusion. The infarct volume was measured by magnetic resonance imaging. RESULTS: The finding that anterior communicating artery flow facilitates pial artery patency in the ipsilateral hemisphere in mice with recurrent stroke suggests that compensatory collateral patency contributes to increased regional cerebral blood flow, enhanced microcirculatory perfusion, improved neurological function and reduced infarct volume. CONCLUSIONS: The results of this study demonstrate that anterior communicating artery flow alleviates recurrent stroke-induced neurovascular injury and improves neurobehavioral outcomes by promoting the establishment of collateral circulation.

11.
Theranostics ; 9(17): 4923-4934, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31410191

RESUMO

Rationale: Brain collaterals contribute to improving ischemic stroke outcomes. However, dynamic and timely investigations of collateral blood flow and collateral restoration in whole brains of living animals have rarely been reported. Methods: Using multiple modalities of imaging, including synchrotron radiation angiography, laser speckle imaging, and micro-CT imaging, we dynamically explored collateral circulation throughout the whole brain in the rodent middle cerebral artery occlusion model. Results: We demonstrated that compared to control animals, 4 neocollaterals gradually formed between the intra- and extra-arteries in the skull base of model animals after occlusion (p<0.05). Two main collaterals were critical to the supply of blood from the posterior to the middle cerebral artery territory in the deep brain (p<0.05). Abundant small vessel and capillary anastomoses were detected on the surface of the cortex between the posterior and middle cerebral artery and between the anterior and middle cerebral artery (p<0.05). Collateral perfusion occurred immediately (≈15 min) and was maintained for up to 14 days after occlusion. Further study revealed that administration of rapamycin at 15 min after MCAO dilated the existing collateral vessels and promoted collateral perfusion. Principal conclusions: Our results provide evidence of collateral functional perfusion in the skull base, deep brain, and surface of the cortex. Rapamycin was capable of enlarging the diameter of collaterals, potentially extending the time window for ischemic stroke therapy.

12.
Stroke Vasc Neurol ; 4(2): 61-62, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31338210
13.
Chin Med J (Engl) ; 132(14): 1700-1705, 2019 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-31283648

RESUMO

BACKGROUND: More than ten genome-wide association studies have identified the significant association between the gap junction delta-2 (GJD2) gene and myopia. However, no functional studies have been performed to confirm that this gene is correlated with myopia. This study aimed to observe how this gene changed in mRNA and protein level in the form-deprivation myopia (FDM) animal model. METHODS: Four-week-old guinea pigs were randomly divided into two groups: control and FDM groups (n = 12 for each group). The right eyes of the FDM group were covered with opaque hemispherical plastic lenses for 3 weeks. For all the animals, refractive status, axial length (AL), and corneal radius of curvature were measured at baseline and 3 weeks later by streak retinoscope, A-scan ultrasonography, and keratometer, respectively. Retinal GJD2 mRNA expression and connexin 36 (Cx36) levels in FDM and control groups were measured by quantitative real-time PCR and Western blot analyses, respectively. Those results were compared using independent t test, Mann-Whitney U test, or paired t test. A significance level of P < 0.05 was used. RESULTS: Three weeks later, the FDM group (form-deprived eyes) showed about a myopic shift of approximately -6.75 (-7.94 to -6.31) D, while the control group remained hyperopic with only a shift of -0.50 (-0.75 to 0.25) D (Z = -3.38, P < 0.01). The AL increased by 0.74 (0.61-0.76) and 0.10 (0.05-0.21) mm in FDM and control groups, respectively (Z = -3.37, P < 0.01). The relative mRNA expression of GJD2 in the FDM group decreased 31.58% more than the control group (t = 11.44, P < 0.01). The relative protein expression of CX36 on the retina was lowered by 37.72% in form-deprivation eyes as compared to the controls (t = 17.74, P < 0.01). CONCLUSION: Both the mRNA expression of GJD2 and Cx36 protein amount were significantly decreased in the retina of FDM guinea pigs. This indicates that Cx36 is involved in FDM development, providing compensating evidence for the results obtained from genome-wide association studies.


Assuntos
Conexinas/metabolismo , Miopia/metabolismo , RNA Mensageiro/metabolismo , Animais , Western Blotting , Conexinas/genética , Córnea/metabolismo , Modelos Animais de Doenças , Junções Comunicantes/metabolismo , Estudo de Associação Genômica Ampla , Cobaias , Miopia/genética , RNA Mensageiro/genética
14.
Theranostics ; 9(10): 2910-2923, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31244932

RESUMO

Rationale: Microglia play a critical role in modulating cell death and neurobehavioral recovery in response to brain injury either by direct cell-cell interaction or indirect secretion of trophic factors. Exosomes secreted from cells are well documented to deliver bioactive molecules to recipient cells to modulate cell function. Here, we aimed to identify whether M2 microglia exert neuroprotection after ischemic attack through an exosome-mediated cell-cell interaction. Methods: M2 microglia-derived exosomes were intravenously injected into the mouse brain immediately after middle cerebral artery occlusion. Infarct volume, neurological score, and neuronal apoptosis were examined 3 days after ischemic attack. Exosome RNA and target protein expression levels in neurons and brain tissue were determined for the mechanistic study. Results: Our results showed that the M2 microglia-derived exosomes were taken up by neurons in vitro and in vivo. M2 microglia-derived exosome treatment attenuated neuronal apoptosis after oxygen-glucose deprivation (p<0.05). In vivo results showed that M2 microglia-derived exosome treatment significantly reduced infarct volume and attenuated behavioral deficits 3 days after transient brain ischemia (p<0.05), whereas injection of miR-124 knockdown (miR-124k/d) M2 microglia-derived exosomes partly reversed the neuroprotective effect. Our mechanistic study further demonstrated that ubiquitin-specific protease 14 (USP14) was the direct downstream target of miR-124. Injection of miR-124k/d M2 exosomes plus the USP14 inhibitor, IU1, achieved comparable neuroprotective effect as injection of M2 exosomes alone. Conclusions: We demonstrated that M2 microglia-derived exosomes attenuated ischemic brain injury and promoted neuronal survival via exosomal miR-124 and its downstream target USP14. M2 microglia-derived exosomes represent a promising avenue for treating ischemic stroke.

15.
CNS Neurosci Ther ; 25(9): 1030-1041, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31218845

RESUMO

INTRODUCTION: L-glutamine is an antioxidant that plays a role in a variety of biochemical processes. Given that oxidative stress is a key component of stroke pathology, the potential of L-glutamine in the treatment of ischemic stroke is worth exploring. AIMS: In this study, we investigated the effect and mechanisms of action of L-glutamine after cerebral ischemic injury. RESULTS: L-glutamine reduced brain infarct volume and promoted neurobehavioral recovery in mice. L-glutamine administration increased the expression of heat-shock protein 70 (HSP70) in astrocytes and endothelial cells. Such effects were abolished by the coadministration of Apoptozole, an inhibitor of the ATPase activity of HSP70. L-glutamine also reduced oxidative stress and neuronal apoptosis, and increased the level of superoxide dismutase, glutathione, and brain-derived neurotrophic factor. Cotreatment with Apoptozole abolished these effects. Cell culture study further revealed that the conditioned medium from astrocytes cultured with L-glutamine reduced the apoptosis of neurons after oxygen-glucose deprivation. CONCLUSION: L-glutamine attenuated ischemic brain injury and promoted functional recovery via HSP70, suggesting its potential in ischemic stroke therapy.

16.
Front Cell Neurosci ; 13: 223, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31164807

RESUMO

Endothelial progenitor cells (EPCs) are multipotential stem cells considered to have immense clinical value for revascularization. However, the clinical application of EPCs has been hampered by their clinical potency in ischemic anoxic environments. This study aimed to explore the effect of microRNA-210 (miR-210) on EPCs under oxygen-glucose deprivation (OGD) conditions. We generated a model of EPCs cultured under OGD conditions to simulate ischemia and explore the expression of miR-210 in vitro. With longer exposure to hypoxia, we found that miR-210-3p expression was highly upregulated in OGD groups compared to that in controls from 4 to 24 h, but not miR-210-5p. We then transfected a miR-210-3p mimic and inhibitor into EPCs, and after 24 h, we exposed them to OGD conditions for 4 h to simulate ischemia. We detected miR-210 by real-time polymerase chain reaction (RT-PCR) and tested the proliferation, migration, and tube formation of normal EPCs and OGD-treated EPCs by CCK-8, transwell chamber, and Matrigel assays, respectively. The direct targets of miR-210-3p were predicted using miRWalk. Compared to that in normal EPCs, higher miR-210-3p expression was found in OGD-treated EPCs (p < 0.05). Moreover, upregulation of miR-210-3p was found to promote proliferation, migration, and tube formation in EPCs under normal and OGD conditions (p < 0.05), whereas down-regulation inhibited these abilities in OGD-treated EPCs (p < 0.05). Repulsive guidance molecule A (RGMA), a negative regulator of angiogenesis, was predicted to be a target of miR-210-3p. Accordingly, upregulation of miR-210-3p was found to inhibit its expression at the protein level in OGD-treated EPCs, whereas downregulation of miR-210-3p inhibited its expression (p < 0.05). A dual-luciferase reporter system confirmed that RGMA is a direct target of miR-210-3p. MicroRNA-210-3p overexpression enhances the angiogenic properties of OGD-treated EPCs by inhibiting RGMA.

17.
J Psychiatr Res ; 115: 129-141, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31129437

RESUMO

We aimed to explore the circulating microRNAs biomarkers in the acute stage following cerebral ischemia to earlier warn late-onset post-stroke depression (PSD). A total of 251 consecutive patients with acute ischemic stroke were recruited. They were divided into three groups depending on whether PSD had occurred at 2 weeks or 3 months since stroke: early-onset PSD, late-onset PSD, and non-depressed group. Microarray assay was conducted to identify the different expression profiles of plasma miRNAs. Comprehensive bioinformatics analysis for their integrating putative target genes was performed. The key miRNA was validated in a larger cohort and its function was further studied in ischemic mice brain. We screened three differentially expressed miRNAs in the late-onset PSD individuals, miR-140-5p and miR-221-3p were significantly upregulated while miR-1246 was downregulated. The bioinformatics analysis demonstrated that their predicted target genes were mainly enriched in axon development and Ras signaling pathway. Logistic regression analysis revealed that miR-140-5p was an independent risk factor for late-onset PSD (P = 0.017, OR = 2.313, 95%CI 1.158 to 4.617). The miR-140-5p expression on admission was significantly positively correlated with HDRS scores assessed at 3 months after stroke (P = 0.0007). The predictive value of miR-140-5p for late-onset PSD is 83.3% sensitivity and 72.6% specificity (AUC = 0.8127, P < 0.0001). AAV-mediated overexpression of miR-140-5p decreased the protein level of IL1rap, IL1rapl1, VEGF, and MEGF10 in the ischemic mouse hippocampus and inhibited neurogenesis and capillary density. MiR-140-5p might be involved in the pathogenesis of late-onset PSD and used as a novel early warning biomarker.

18.
Neurosci Lett ; 705: 251-258, 2019 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-30928480

RESUMO

BACKGROUND AND PURPOSE: Studies demonstrated that oxidative damage decreased intracellular ATP level in astrocytes. However, the pathway mediated ATP level decrease is obscure. Our previous study found intracellular ATP could be released via lysosome exocytosis in astrocytes. Here, we explored whether lysosome exocytosis was involved in ATP release during oxidative stress induced by H2O2 in astrocytes. METHODS: Astrocytes were isolated from the cortex of neonatal rats. Intracellular lysosomes and calcium signals were stained in astrocytes before and after H2O2 stimulation. ATP molecules location and ATP level were detected by immunostaining and bioluminescence method, respectively. Extracellular ß-Hexosaminidase and LDH were examined by colorimetric method. RESULTS: We found that ATP located in lysosome of astrocytes. H2O2 stimulation resulted in the decrease of lysosomes staining and the increase of extracellular ATP, compared to the control (p < 0.05). At the same time, intracellular Fluo4 signals and ß-Hexosaminidase level were also increased (p < 0.05). Extracellular LDH level did not show an increase, suggesting that there is no cell membrane damage after H2O2 stimulation. Glycyl-phenylalanine 2-naphthylamide blocked lysosome exocytosis and inhibited ATP release in astrocytes after H2O2-treatment (p < 0.05). CONCLUSION: Our results indicated that H2O2 induced ATP release from intracellular to extracellular via lysosome exocytosis. The increase of intracellular Ca2+ was necessary for lysosome release under oxidative stress induced by H2O2.


Assuntos
Trifosfato de Adenosina/metabolismo , Astrócitos/metabolismo , Exocitose/fisiologia , Peróxido de Hidrogênio/farmacologia , Lisossomos/fisiologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Cálcio/metabolismo , Exocitose/efeitos dos fármacos , L-Lactato Desidrogenase/metabolismo , Lisossomos/efeitos dos fármacos , Cultura Primária de Células , Ratos , beta-N-Acetil-Hexosaminidases/metabolismo
19.
Aging Dis ; 10(2): 429-462, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31011487

RESUMO

The complement system is an essential part of innate immunity, typically conferring protection via eliminating pathogens and accumulating debris. However, the defensive function of the complement system can exacerbate immune, inflammatory, and degenerative responses in various pathological conditions. Cumulative evidence indicates that the complement system plays a critical role in the pathogenesis of ischemic brain injury, as the depletion of certain complement components or the inhibition of complement activation could reduce ischemic brain injury. Although multiple candidates modulating or inhibiting complement activation show massive potential for the treatment of ischemic stroke, the clinical availability of complement inhibitors remains limited. The complement system is also involved in neural plasticity and neurogenesis during cerebral ischemia. Thus, unexpected side effects could be induced if the systemic complement system is inhibited. In this review, we highlighted the recent concepts and discoveries of the roles of different kinds of complement components, such as C3a, C5a, and their receptors, in both normal brain physiology and the pathophysiology of brain ischemia. In addition, we comprehensively reviewed the current development of complement-targeted therapy for ischemic stroke and discussed the challenges of bringing these therapies into the clinic. The design of future experiments was also discussed to better characterize the role of complement in both tissue injury and recovery after cerebral ischemia. More studies are needed to elucidate the molecular and cellular mechanisms of how complement components exert their functions in different stages of ischemic stroke to optimize the intervention of targeting the complement system.

20.
Mol Ther Nucleic Acids ; 16: 15-25, 2019 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-30825669

RESUMO

Studies demonstrate that microRNA-126 plays a critical role in promoting angiogenesis. However, its effects on angiogenesis following ischemic stroke are unclear. Here, we explored the effect of microRNA-126-3p and microRNA-126-5p on angiogenesis and neurogenesis after brain ischemia. We demonstrated that both microRNA (miRNA)-126-3p and microRNA-126-5p increased the proliferation, migration, and tube formation of human umbilical vein endothelial cells (HUVECs) compared with the scrambled miRNA control (p < 0.05). Transferring microRNA-126 into a mouse middle cerebral artery occlusion model via lentivirus, we found that microRNA-126 overexpression increased the number of CD31+/BrdU+ (5-bromo-2'-deoxyuridine-positive) proliferating endothelial cells and DCX+/BrdU+ neuroblasts in the ischemic mouse brain, improved neurobehavioral outcomes (p < 0.05), and reduced brain atrophy volume (p < 0.05) compared with control mice. Western blot results showed that AKT and ERK signaling pathways were activated in the lentiviral-microRNA-126-treated group (p < 0.05). Both PCR and western blot results demonstrated that tyrosine-protein phosphatase non-receptor type 9 (PTPN9) was decreased in the lentiviral-microRNA-126-treated group (p < 0.05). Dual-luciferase gene reporter assay also showed that PTPN9 was the direct target of microRNA-126-3p and microRNA-126-5p in the ischemic brain. We demonstrated that microRNA-126-3p and microRNA-126-5p promoted angiogenesis and neurogenesis in ischemic mouse brain, and further improved neurobehavioral outcomes. Our mechanistic study further showed that microRNA-126 mediated angiogenesis through directly inhibiting its target PTPN9 and activating AKT and ERK signaling pathways.

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