RESUMO
Alzheimer's disease (AD) is the most common cause of dementia, characterized by memory loss, cognitive decline, personality changes, and various neurological symptoms. The role of blood-brain barrier (BBB) injury, extracellular matrix (ECM) abnormalities, and oligodendrocytes (ODCs) dysfunction in AD has gained increasing attention, yet the detailed pathogenesis remains elusive. This study integrates single-cell sequencing of AD patients' cerebrovascular system with a genome-wide association analysis. It aims to elucidate the associations and potential mechanisms behind pericytes injury, ECM disorder, and ODCs dysfunction in AD pathogenesis. Finally, we identified that abnormalities in the pericyte PI3K-AKT-FOXO signaling pathway may be involved in the pathogenic process of AD. This comprehensive approach sheds new light on the complex etiology of AD and opens avenues for advanced research into its pathogenesis and therapeutic strategies.
Assuntos
Doença de Alzheimer , Barreira Hematoencefálica , Estudo de Associação Genômica Ampla , Pericitos , Doença de Alzheimer/patologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/etiologia , Humanos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Pericitos/patologia , Pericitos/metabolismo , Transdução de Sinais , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Matriz Extracelular/metabolismo , Microvasos/patologia , Microvasos/metabolismo , Análise de Célula Única , Feminino , Masculino , Fosfatidilinositol 3-Quinases/metabolismoRESUMO
BACKGROUND: The choroid plexus (ChP) helps maintain the homeostasis of the brain by forming the blood-CSF barrier via tight junctions (TJ) at the choroid plexus epithelial cells, and subsequently preventing neuroinflammation by restricting immune cells infiltration into the central nervous system. However, whether chronic cerebral hypoperfusion causes ChP structural damage and blood-CSF barrier impairment remains understudied. METHODS: The bilateral carotid stenosis (BCAS) model in adult male C57BL/6 J mice was used to induce cerebral hypoperfusion, a model for vascular contributions to cognitive impairment and dementia (VCID). BCAS-mediated changes of the blood-CSF barrier TJ proteins, apical secretory Na+-K+-Cl- cotransporter isoform 1 (NKCC1) protein and regulatory serine-threonine kinases SPAK, and brain infiltration of myeloid-derived immune cells were assessed. RESULTS: BCAS triggered dynamic changes of TJ proteins (claudin 1, claudin 5) accompanied with stimulation of SPAK-NKCC1 complex and NF-κB in the ChP epithelial cells. These changes impacted the integrity of the blood-CSF barrier, as evidenced by ChP infiltration of macrophages/microglia, neutrophils and T cells. Importantly, pharmacological blockade of SPAK with its potent inhibitor ZT1a in BCAS mice attenuated brain immune cell infiltration and improved cognitive neurological function. CONCLUSIONS: BCAS causes chronic ChP blood-CSF damage and immune cell infiltration. Our study sheds light on the SPAK-NKCC1 complex as a therapeutic target in neuroinflammation.
Assuntos
Estenose das Carótidas , Camundongos Endogâmicos C57BL , Doenças Neuroinflamatórias , Animais , Camundongos , Masculino , Doenças Neuroinflamatórias/patologia , Doenças Neuroinflamatórias/metabolismo , Doenças Neuroinflamatórias/etiologia , Estenose das Carótidas/patologia , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/metabolismo , Plexo Corióideo/patologia , Plexo Corióideo/metabolismoRESUMO
Diabetic encephalopathy (DE) is a severe complication that occurs in the central nervous system (CNS) and leads to cognitive impairment. DE involves various pathophysiological processes, and its pathogenesis is still unclear. This review summarised current research on the pathogenesis of diabetic encephalopathy, which involves neuroinflammation, oxidative stress, iron homoeostasis, blood-brain barrier disruption, altered gut microbiota, insulin resistance, etc. Among these pathological mechanisms, neuroinflammation has been focused on. This paper summarises some of the molecular mechanisms involved in neuroinflammation, including the Mammalian Target of Rapamycin (mTOR), Lipocalin-2 (LCN-2), Pyroptosis, Advanced Glycosylation End Products (AGEs), and some common pro-inflammatory factors. In addition, we discuss recent advances in the study of potential therapeutic targets for the treatment of DE against neuroinflammation. The current research on the pathogenesis of DE is progressing slowly, and more research is needed in the future. Further study of neuroinflammation as a mechanism is conducive to the discovery of more effective treatments for DE in the future.
Assuntos
Doenças Neuroinflamatórias , Humanos , Doenças Neuroinflamatórias/etiologia , Doenças Neuroinflamatórias/patologia , Animais , Estresse Oxidativo/fisiologia , Complicações do Diabetes/etiologia , Encefalopatias/etiologia , Encefalopatias/patologia , Barreira Hematoencefálica/patologia , Inflamação/patologiaRESUMO
Progression of acute traumatic brain injury (TBI) into chronic neurodegeneration is a major health problem with no protective treatments. Here, we report that acutely elevated mitochondrial fission after TBI in mice triggers chronic neurodegeneration persisting 17 months later, equivalent to many human decades. We show that increased mitochondrial fission after mouse TBI is related to increased brain levels of mitochondrial fission 1 protein (Fis1) and that brain Fis1 is also elevated in human TBI. Pharmacologically preventing Fis1 from binding its mitochondrial partner, dynamin-related protein 1 (Drp1), for 2 weeks after TBI normalizes the balance of mitochondrial fission/fusion and prevents chronically impaired mitochondrial bioenergetics, oxidative damage, microglial activation and lipid droplet formation, blood-brain barrier deterioration, neurodegeneration, and cognitive impairment. Delaying treatment until 8 months after TBI offers no protection. Thus, time-sensitive inhibition of acutely elevated mitochondrial fission may represent a strategy to protect human TBI patients from chronic neurodegeneration.
Assuntos
Lesões Encefálicas Traumáticas , Dinaminas , Mitocôndrias , Dinâmica Mitocondrial , Proteínas Mitocondriais , Lesões Encefálicas Traumáticas/metabolismo , Lesões Encefálicas Traumáticas/patologia , Animais , Dinaminas/metabolismo , Dinaminas/genética , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Humanos , Camundongos , Mitocôndrias/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Estresse Oxidativo , Encéfalo/patologia , Encéfalo/metabolismo , Microglia/metabolismo , Microglia/patologia , Doença Crônica , Modelos Animais de Doenças , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologiaRESUMO
The central nervous system (CNS) represents a site of sanctuary for many metastatic tumors when systemic therapies that control the primary tumor cannot effectively penetrate intracranial lesions. Non-small cell lung cancers (NSCLCs) are the most likely of all neoplasms to metastasize to the brain, with up to 60% of patients developing CNS metastases during the disease process. Targeted therapies such as tyrosine kinase inhibitors (TKIs) have helped reduce lung cancer mortality but vary considerably in their capacity to control CNS metastases. The ability of these therapies to effectively target lesions in the CNS depends on several of their pharmacokinetic properties, including blood-brain barrier permeability, affinity for efflux transporters, and binding affinity for both plasma and brain tissue. Despite the existence of numerous preclinical models with which to characterize these properties, many targeted therapies have not been rigorously tested for CNS penetration during the discovery process, whereas some made it through preclinical testing despite poor brain penetration kinetics. Several TKIs have now been engineered with the characteristics of CNS-penetrant drugs, with clinical trials proving these efforts fruitful. This Review outlines the extent and variability of preclinical evidence for the efficacy of NSCLC-targeted therapies, which have been approved by the US Food and Drug Administration (FDA) or are in development, for treating CNS metastases, and how these data correlate with clinical outcomes.
Assuntos
Neoplasias do Sistema Nervoso Central , Terapia de Alvo Molecular , Humanos , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , Neoplasias do Sistema Nervoso Central/patologia , Neoplasias do Sistema Nervoso Central/secundário , Neoplasias do Sistema Nervoso Central/metabolismo , Animais , Avaliação Pré-Clínica de Medicamentos , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/farmacocinética , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/patologiaRESUMO
Viral encephalitis is characterized by inflammation of the brain parenchyma caused by a variety of viruses, among which the Japanese encephalitis (JE) virus (JEV) is a typical representative arbovirus. Neuronal death, neuroinflammation, and breakdown of the blood brain barrier (BBB) constitute vicious circles of JE progression. Currently, there is no effective therapy to prevent this damage. Growth arrest specific gene 6 (GAS6) is a secreted growth factor that binds to the TYRO3, AXL, and MERTK (TAM) family of receptor tyrosine kinases and has been demonstrated to participate in neuroprotection and suppression of inflammation in many central nervous system (CNS) diseases which has great potential for JE intervention. In this study, we found that GAS6 expression in the brain was decreased and was reversely correlated with viral load and neuronal loss. Mice with GAS6/TAM signalling deficiency showed higher mortality and accelerated neuroinflammation during peripheral JEV infection, accompanied by BBB breakdown. GAS6 directly promoted the expression of tight junction proteins in bEnd.3 cells and strengthened BBB integrity, partly via AXL. Mice administered GAS6 were more resistant to JEV infection due to increased BBB integrity, as well as decreased viral load and neuroinflammation. Thus, targeted GAS6 delivery may represent a strategy for the prevention and treatment of JE especially in patients with impaired BBB.
Assuntos
Encefalite Japonesa , Peptídeos e Proteínas de Sinalização Intercelular , Doenças Neuroinflamatórias , Animais , Camundongos , Receptor Tirosina Quinase Axl , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Modelos Animais de Doenças , Encefalite Japonesa/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Doenças Neuroinflamatórias/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Receptores Proteína Tirosina Quinases/genéticaRESUMO
Systemic inflammation has been implicated in the development and progression of neurodegenerative conditions such as cognitive impairment and dementia. Recent clinical studies indicate an association between sepsis, endothelial dysfunction, and cognitive decline. However, the investigations of the role and therapeutic potential of the cerebral microvasculature in sepsis-induced cognitive dysfunction have been limited by the lack of standardized experimental models for evaluating the alterations in the cerebral microvasculature and cognition induced by the systemic inflammatory response. Herein, we validated a mouse model of endotoxemia that recapitulates key pathophysiology related to sepsis-induced cognitive dysfunction, including the induction of an acute systemic hyperinflammatory response, blood-brain barrier (BBB) leakage, neurovascular inflammation, and memory impairment after recovery from the systemic inflammation. In the acute phase, we identified novel molecular (e.g., upregulation of plasmalemma vesicle-associated protein, PLVAP, a driver of endothelial permeability, and the procoagulant plasminogen activator inhibitor-1, PAI-1) and functional perturbations (i.e., albumin and small-molecule BBB leakage) in the cerebral microvasculature along with neuroinflammation. Remarkably, small-molecule BBB permeability, elevated levels of PAI-1, intra-/perivascular fibrin/fibrinogen deposition, and microglial activation persisted 1 month after recovery from sepsis. We also highlight molecular neuronal alterations of potential clinical relevance following systemic inflammation including changes in neurofilament phosphorylation and decreases in postsynaptic density protein 95 and brain-derived neurotrophic factor, suggesting diffuse axonal injury, synapse degeneration, and impaired neurotrophism. Our study serves as a standardized mouse model to support future mechanistic studies of sepsis-associated cognitive dysfunction and to identify novel endothelial therapeutic targets for this devastating condition.
Assuntos
Barreira Hematoencefálica , Disfunção Cognitiva , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Microvasos , Sepse , Animais , Disfunção Cognitiva/etiologia , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/fisiopatologia , Microvasos/metabolismo , Microvasos/patologia , Camundongos , Masculino , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Sepse/complicações , Sepse/fisiopatologia , Encéfalo/metabolismoRESUMO
Alcohol consumption leads to neuroinflammation and bloodâbrain barrier (BBB) damage, resulting in neurological impairment. We previously demonstrated that ethanol-induced disruption of barrier function in human brain endothelial cells was associated with mitochondrial injury, increased ATP and extracellular vesicle (EV) release, and purinergic receptor P2 × 7R activation. Therefore, we aimed to evaluate the effect of P2 × 7R blockade on peripheral and neuro-inflammation in ethanol-exposed mice. In a chronic intermittent ethanol (CIE)-exposed mouse model, P2 × 7R was inhibited by two different methods: Brilliant Blue G (BBG) or gene knockout. We assessed blood ethanol concentration (BEC), brain microvessel gene expression by using RT2 PCR array, plasma P2 × 7R and P-gp, serum ATP, EV-ATP, number of EVs, and EV mtDNA copy numbers. An RT2 PCR array of brain microvessels revealed significant upregulation of proinflammatory genes involved in apoptosis, vasodilation, and platelet activation in CIE-exposed wild-type animals, which were decreased 15-50-fold in BBG-treated-CIE-exposed animals. Plasma P-gp levels and serum P2 × 7R shedding were significantly increased in CIE-exposed animals. Pharmacological or genetic suppression of P2 × 7R decreased receptor shedding to levels equivalent to those in control group. The increase in EV number and EV-ATP content in the CIE-exposed mice was significantly reduced by P2 × 7R inhibition. CIE mice showed augmented EV-mtDNA copy numbers which were reduced in EVs after P2 × 7R inhibition or receptor knockout. These observations suggested that P2 × 7R signaling plays a critical role in ethanol-induced brain injury. Increased extracellular ATP, EV-ATP, EV numbers, and EV-mtDNA copy numbers highlight a new mechanism of brain injury during alcohol exposure via P2 × 7R and biomarkers of such damage. In this study, for the first time, we report the in vivo involvement of P2 × 7R signaling in CIE-induced brain injury.
Assuntos
Barreira Hematoencefálica , Etanol , Doenças Neuroinflamatórias , Receptores Purinérgicos P2X7 , Transdução de Sinais , Animais , Masculino , Camundongos , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Depressores do Sistema Nervoso Central/toxicidade , Depressores do Sistema Nervoso Central/farmacologia , Etanol/toxicidade , Camundongos Endogâmicos C57BL , Camundongos Knockout , Doenças Neuroinflamatórias/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Blood-brain barrier (BBB) injury and dysfunction following infection with the human immunodeficiency virus (HIV) enables viral entry into the brain, infection of resident brain cells, neuronal injury and subsequent neurodegeneration leading to HIV-associated neurocognitive disorders (HAND). Although combination antiretroviral therapy has significantly reduced the incidence and prevalence of acquired immunodeficiency syndrome and increased the life expectancy of people living with HIV, the prevalence of HAND remains high. With aging of people living with HIV associated with increased comorbidities, the prevalence of HIV-related central nervous system (CNS) complications is expected to remain high. Considering the principal role of the brain endothelium in HIV infection of the CNS and HAND, the purpose of this manuscript is to review the current literature on the pathobiology of the brain endothelium structural and functional dysregulation in HIV infection, including in the presence of HIV-1 and viral proteins (gp120, Tat, Nef, and Vpr). We summarize evidence from human and animal studies, in vitro studies, and associated mechanisms. We further summarize evidence of synergy or lack thereof between commonly abused substances (cocaine, methamphetamine, alcohol, tobacco, opioids, and cannabinoids) and HIV- or viral protein-induced BBB injury and dysfunction.
Assuntos
Barreira Hematoencefálica , Encéfalo , Infecções por HIV , Transtornos Relacionados ao Uso de Substâncias , Humanos , Infecções por HIV/patologia , Infecções por HIV/complicações , Transtornos Relacionados ao Uso de Substâncias/patologia , Transtornos Relacionados ao Uso de Substâncias/complicações , Transtornos Relacionados ao Uso de Substâncias/metabolismo , Encéfalo/patologia , Encéfalo/metabolismo , Encéfalo/virologia , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/metabolismo , Animais , Endotélio/patologia , Endotélio/metabolismo , HIV-1RESUMO
The neurovascular unit (NVU) is a complex multicellular structure that helps maintain cerebral homeostasis and blood-brain barrier (BBB) integrity. While extensive evidence links NVU alterations to cerebrovascular diseases and neurodegeneration, the underlying molecular mechanisms remain unclear. Here, we use zebrafish embryos carrying a mutation in Scavenger Receptor B2, a highly conserved endolysosomal protein expressed predominantly in Radial Glia Cells (RGCs), to investigate the interplay among different NVU components. Through live imaging and genetic manipulations, we demonstrate that compromised acidification of the endolysosomal compartment in mutant RGCs leads to impaired Notch3 signaling, thereby inducing excessive neurogenesis and reduced glial differentiation. We further demonstrate that alterations to the neuron/glia balance result in impaired VEGF and Wnt signaling, leading to severe vascular defects, hemorrhages, and a leaky BBB. Altogether, our findings provide insights into NVU formation and function and offer avenues for investigating diseases involving white matter defects and vascular abnormalities.
Assuntos
Barreira Hematoencefálica , Lisossomos , Neurogênese , Proteínas de Peixe-Zebra , Peixe-Zebra , Animais , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Lisossomos/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Células Ependimogliais/metabolismo , Células Ependimogliais/patologia , Endossomos/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Fator A de Crescimento do Endotélio Vascular/genética , Receptores Notch/metabolismo , Receptores Notch/genética , Neuroglia/metabolismo , Neuroglia/patologia , Diferenciação Celular , Células-Tronco/metabolismo , Via de Sinalização Wnt , Mutação , Neovascularização Fisiológica , Animais Geneticamente Modificados , Encéfalo/metabolismo , Encéfalo/patologia , Encéfalo/irrigação sanguínea , Transdução de Sinais , AngiogêneseRESUMO
CNS diseases associated with compromised blood supply and/or vascular integrity are one of the leading causes of mortality and disability in adults worldwide and are also among 10 most common causes of death in children. Angiogenesis is an essential element of regeneration processes upon nervous tissue damage and can play a crucial role in neuroprotection. Here we review the features of cerebral vascular regeneration after ischemic stroke, including the complex interactions between endothelial cells and other brain cell types (neural stem cells, astrocytes, microglia, and oligodendrocytes). The mechanisms of reciprocal influence of angiogenesis and neurogenesis, the role of astrocytes in the formation of the blood-brain barrier, and roles of microglia and oligodendrocytes in vascular regeneration are discussed. Understanding the mechanisms of angiogenesis regulation in CNS is of critical importance for the development of new treatments of neurovascular pathologies.
Assuntos
Astrócitos , Barreira Hematoencefálica , AVC Isquêmico , Neovascularização Fisiológica , Células-Tronco Neurais , Neurogênese , Humanos , AVC Isquêmico/fisiopatologia , AVC Isquêmico/metabolismo , AVC Isquêmico/patologia , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/fisiopatologia , Neovascularização Fisiológica/fisiologia , Neurogênese/fisiologia , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Astrócitos/fisiologia , Células-Tronco Neurais/metabolismo , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Oligodendroglia/fisiologia , Microglia/patologia , Microglia/metabolismo , Microglia/fisiologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Neuroglia/metabolismo , Neuroglia/patologia , Isquemia Encefálica/fisiopatologia , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Sistema Nervoso Central/irrigação sanguínea , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/patologia , Encéfalo/irrigação sanguínea , Encéfalo/patologia , Encéfalo/metabolismo , Encéfalo/fisiopatologia , AngiogêneseRESUMO
Subarachnoid hemorrhage (SAH) significantly compromises the blood-brain barrier (BBB) and impairs patient recovery. This study elucidates the critical role of astrocytic Neogenin-1 (NEO1) in BBB integrity post-SAH and examines the regulatory effects of hepcidin on endothelial cell (EC) function amid NEO1-mediated disruptions in iron homeostasis. Proteomic analyses of cerebrospinal fluid (CSF) from SAH patients revealed a substantial decrease in NEO1 expression, identifying it as a key factor in BBB integrity. 111 CSF proteins were significantly reduced in early SAH stages (days 1-3), with NEO1 among the most significantly altered. This dysregulation was linked to poorer patient outcomes, as indicated by a negative correlation between NEO1 levels and Modified Rankin Scale scores six months post-SAH (R = -0.4743, P < 0.0001). Experimental models further highlighted the importance of NEO1: SAH model and NEO1GFAP-Cre mice exhibited exacerbated EC dysfunction and increased BBB permeability, evidenced by significant Evans Blue retention and dextran leakage in the parietal cortex, effects that were mitigated by hepcidin administration. Our findings highlight the complex interplay between astrocytic signaling and endothelial function in SAH pathophysiology. The loss of astrocytic NEO1 led to increased EC proliferation and altered BBB structure, as confirmed by transmission electron microscopy and immunostaining for PECAM-1, indicating heightened blood vessel density in the affected cortex. Hepcidin treatment effectively reversed the EC dysfunction and BBB disruption in both NEO1-cKO mice and the SAH model, highlighting its potential as a therapeutic agent to enhance recovery and improve prognosis following SAH.
Assuntos
Astrócitos , Barreira Hematoencefálica , Hepcidinas , Hemorragia Subaracnóidea , Hemorragia Subaracnóidea/metabolismo , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/efeitos dos fármacos , Animais , Hepcidinas/metabolismo , Hepcidinas/genética , Astrócitos/metabolismo , Humanos , Camundongos , Masculino , Camundongos Endogâmicos C57BL , Células Endoteliais/metabolismo , Modelos Animais de Doenças , Feminino , Pessoa de Meia-Idade , Proteínas de Membrana/metabolismoRESUMO
PURPOSE: Glioblastoma (GBM) is the most prevalent, malignant, primary brain tumor in adults, characterized by limited treatment options, frequent relapse, and short survival after diagnosis. Until now, none of the existing therapy and treatment approaches have proven to be an effective cure. The availability of predictive human blood-tumor barrier (BTB) test systems that can mimic in-vivo pathophysiology of GBM would be of great interest in preclinical research. Here, we present the establishment of a new BTB in-vitro test system combining GBM spheroids and BBB models derived from human induced pluripotent stem cells (hiPSCs). METHODS: We co-cultured hiPSC-derived brain capillary endothelial-like cells (iBCECs) with GBM spheroids derived from U87-MG and U373-MG cell lines in a cell culture insert-based format. Spheroids were monitored over 168 hours (h) of culture, characterized for GBM-specific marker expression and treated with standard chemotherapeutics to distinguish inhibitory effects between 2D mono-culture and 3D spheroids. GBM-induced changes on iBCECs barrier integrity were verified via measurement of transendothelial electrical resistance (TEER), immunocytochemical staining of tight junction (TJ) proteins claudin-5 and occludin as well as the glucose transporter-1 (Glut-1). GBM-induced secretion of vascular endothelial growth factor (VEGF) was additionally quantified. RESULTS: Our hypothesis was validated by reduced expression of TJ proteins, occludin and claudin-5 together with significant barrier breakdown in iBCECs after only 24 h of co-culture, demonstrated by reduction in TEER from 1313 ± 265 Ω*cm2 to 712 ± 299 Ω*cm2 (iBCECs + U87-MG) and 762 ± 316 Ω*cm2 (iBCECs + U373-MG). Furthermore, 3D spheroids show more resistance to standard GBM chemotherapeutics in-vitro compared to 2D cultures. CONCLUSIONS: We demonstrate the establishment of a simplified, robust in-vitro BTB test system, with potential application in preclinical therapeutic screening and in studying GBM-induced pathological changes at the BBB.
Assuntos
Barreira Hematoencefálica , Neoplasias Encefálicas , Técnicas de Cocultura , Glioblastoma , Esferoides Celulares , Glioblastoma/patologia , Glioblastoma/metabolismo , Humanos , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/metabolismo , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/metabolismo , Esferoides Celulares/patologia , Células-Tronco Pluripotentes Induzidas , Células Endoteliais/patologia , Linhagem Celular TumoralRESUMO
Multiple sclerosis (MS) is a chronic inflammatory neurodegenerative disease leading to progressive demyelination and neuronal loss, with extensive neurological symptoms. As one of the most widespread neurodegenerative disorders, with an age onset of about 30 years, it turns out to be a socio-health and economic issue, thus necessitating therapeutic interventions currently unavailable. Loss of integrity in the blood-brain barrier (BBB) is one of the distinct MS hallmarks. Brain homeostasis is ensured by an endothelial cell-based monolayer at the interface between the central nervous system (CNS) and systemic bloodstream, acting as a selective barrier. MS results in enhanced barrier permeability, mainly due to the breakdown of tight (TJs) and adherens junctions (AJs) between endothelial cells. Specifically, proinflammatory mediator release causes failure in cytoplasmic exposure of junctions, resulting in compromised BBB integrity that enables blood cells to cross the barrier, establishing iron deposition and neuronal impairment. Cells with a compromised cytoskeletal protein network, fiber reorganization, and discontinuous junction structure can occur, resulting in BBB dysfunction. Recent investigations on spatial transcriptomics have proven circularRNAs (circRNAs) to be powerful multi-functional molecules able to epigenetically regulate transcription and structurally support proteins. In the present review, we provide an overview of the recent role ascribed to circRNAs in maintaining BBB integrity/permeability via cytoskeletal stability. Increased knowledge of the mechanisms responsible for impairment and circRNA's role in driving BBB damage and dysfunction might be helpful for the recognition of novel therapeutic targets to overcome BBB damage and unrestrained neurodegeneration.
Assuntos
Barreira Hematoencefálica , Epigênese Genética , Esclerose Múltipla , RNA Circular , Humanos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Esclerose Múltipla/patologia , Esclerose Múltipla/genética , Esclerose Múltipla/metabolismo , RNA Circular/genética , RNA Circular/metabolismo , AnimaisRESUMO
Vascular dementia (VaD) is a complex neurodegenerative condition, with cerebral small vessel dysfunctions as the central role in its pathogenesis. Given the lack of suitable animal models to study the disease pathogenesis, we developed a mouse model to closely emulate the clinical scenarios of recurrent transient ischemic attacks (TIAs) leading to VaD using vasoconstricting peptide Endothelin-1(ET-1). We observed that administration of ET-1 led to blood-brain barrier (BBB) disruption and detrimental changes in its components, such as endothelial cells and pericytes, along with neuronal loss and synaptic dysfunction, resulting in irreversible memory loss. Further, in our pursuit of understanding potential interventions, we co-administered pleiotrophin (PTN) alongside ET-1 injections. PTN exhibited remarkable efficacy in preserving vital components of the BBB, including endothelial cells and pericytes, thereby restoring BBB integrity, preventing neuronal loss, and enhancing memory function. Our findings give a valuable framework for understanding the detrimental effects of multiple TIAs on brain health and provide a useful animal model to explore VaD's underlying mechanisms further and pave the way for promising therapies.
Assuntos
Proteínas de Transporte , Citocinas , Endotelina-1 , Camundongos Endogâmicos C57BL , Animais , Camundongos , Proteínas de Transporte/metabolismo , Endotelina-1/toxicidade , Citocinas/metabolismo , Masculino , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/patologia , Disfunção Cognitiva/etiologia , Disfunção Cognitiva/tratamento farmacológico , Disfunção Cognitiva/patologia , Modelos Animais de Doenças , Demência Vascular/patologia , Demência Vascular/tratamento farmacológicoRESUMO
A1 polarization of astrocytes mediated prolonged inflammation contributing to brain injury in ischemic stroke. We have previously shown that AD16 protects against neonatal hypoxic-ischemic brain damage in vivo and oxygen-glucose deprivation in vitro. More recently, AD16 has demonstrated safety, tolerability, and favorable pharmacokinetics in a randomized controlled phase I trial. In this study, we utilized a rat model of transient middle cerebral artery occlusion (tMCAO) to explore whether the anti-inflammatory compound AD16 protects against ischemic brain injury by regulating A1 polarization and its underlying mechanisms. Our results showed that AD16 treatment significantly reduced the brain infarcted volume and improved neurological function in tMCAO rats. GO analysis results show that differential genes among the Sham, tMCAO and AD16 treatment groups are involved in the regulation of cytokine and inflammatory response. KEGG enrichment pathways analysis mainly enriched in cytokine-cytokine receptor interaction, viral protein interaction with cytokine-cytokine receptor, TNF, chemokine, NF-κB and IL-17 signaling pathway. Furthermore, AD16 treatment decreased the permeability of the blood-brain barrier and suppressed neuroinflammation. AD16 treatment also significantly reduced the polarization of A1 and inhibited NF-κB and JAK2/STAT3 signaling pathways. This study demonstrates that AD16 protects against brain injury in ischemic stroke by reducing A1 polarization to suppress neuroinflammation through downregulating NF-κB and JAK2/STAT3 signaling. Our findings uncover a potential molecular mechanism for AD16 and suggest that AD16 holds promising therapeutic potential against cerebral ischemia.
Assuntos
Astrócitos , Doenças Neuroinflamatórias , Animais , Masculino , Ratos , Anti-Inflamatórios/farmacologia , Astrócitos/metabolismo , Astrócitos/efeitos dos fármacos , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Isquemia Encefálica/tratamento farmacológico , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Polaridade Celular/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Infarto da Artéria Cerebral Média/tratamento farmacológico , Infarto da Artéria Cerebral Média/patologia , Infarto da Artéria Cerebral Média/metabolismo , Doenças Neuroinflamatórias/tratamento farmacológico , Doenças Neuroinflamatórias/metabolismo , Fármacos Neuroprotetores/farmacologia , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
Cerebral small vessel disease (CSVD) is a group of pathologies that affect the cerebral blood vessels. CSVD accounts for 25% of strokes and contributes to 45% of dementia. However, the pathogenesis of CSVD remains unclear, involving a variety of complex mechanisms. CSVD may result from dysfunction in the glymphatic system (GS). The GS contains aquaporin-4 (AQP-4), which is in the perivascular space, at the endfeet of the astrocyte. The GS contributes to the removal of waste products from the central nervous system, occupying perivascular spaces and regulating the exchange and movement of cerebrospinal fluid and interstitial fluid. The GS involves astrocytes and aquaporin channels, which are components of the blood-brain barrier, and problems with them may constitute the pathogenesis of CSVD. Vascular risk factors, including diabetes, dilate the perivascular space, disrupting the glymphatic system and the active regulation of AQP-4. CSVD exacerbation due to disorders of the GS is associated with multiple vasculopathies. Dysfunction of the glymphatic system and AQP-4 interferes with the functioning of the blood-brain barrier, which exacerbates CSVD. In a long-term follow-up of CSVD patients with microbleeds, lacunar infarcts, and white matter hyperintensity, several vascular risk factors, including hypertension, increased the risk of ischemic stroke. Dysfunction of the GS may be the cause of CSVD; however, the underlying treatment needs to be studied further.
Assuntos
Aquaporina 4 , Barreira Hematoencefálica , Doenças de Pequenos Vasos Cerebrais , Sistema Glinfático , Doenças de Pequenos Vasos Cerebrais/metabolismo , Doenças de Pequenos Vasos Cerebrais/patologia , Doenças de Pequenos Vasos Cerebrais/etiologia , Humanos , Sistema Glinfático/metabolismo , Sistema Glinfático/patologia , Aquaporina 4/metabolismo , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Fatores de RiscoRESUMO
BACKGROUND: Cerebral ischemia is characterized by its rapid onset and high rates of recurrence, morbidity, and mortality, with blood-brain barrier (BBB) permeability playing a vital role in brain injury. Therefore, it is important to understand the molecular mechanism which regulates the BBB during cerebral ischemia. MATERIALS AND METHODS: An in vitro model of oxygen-glucose deprivation (OGD) and an in vivo model of cerebral ischemia/reperfusion (I/R) were constructed. PD-1 overexpression vectors and vectors containing si-RNA were transfected and injected into in vitro and in vivo models. Western blotting, real-time quantitative PCR (qPCR), immunofluorescence (IF) analysis, and immunohistochemical staining were employed to evaluate the expression levels of programmed cell death-1 (PD-1), microglia M1 and M2 biomarkers, and tight junction proteins. Flow cytometry and ELISA were used to measure the levels of pro-inflammatory cytokines. The BBB permeability of brain tissues was evaluated by Evans blue dye (EBD) extravasation and transendothelial electrical resistance (TEER). Brain water content was measured to assess the extent of inflammatory exudation. The infarct volume and neurological severity score (NSS) were used to assess the severity of brain injury. Brain cell apoptosis was assessed by the TUNEL assay and hematoxylin-eosin (H&E) staining. RESULTS: PD-1 helped to convert the microglia M1 phenotype to the M2 phenotype and to reduce BBB permeability both in vitro and in vivo. Overexpression of PD-1 promoted a shift of the M1 phenotype to the M2 phenotype and reduced BBB permeability via the ERK and p38 MAPK signaling pathways. PD-1 reduced inflammatory exudation, BBB permeability, cell apoptosis, and brain injury in vivo. CONCLUSION: Our present study verified that PD-1 exerts an anti-inflammatory effect by converting the microglia M1 phenotype to the M2 phenotype, reducing BBB permeability, and thereby relieves brain injury caused by cerebral ischemia. PD-1 is potential therapeutic target for brain injury caused by cerebral ischemia.
Assuntos
Barreira Hematoencefálica , Isquemia Encefálica , Sistema de Sinalização das MAP Quinases , Microglia , Receptor de Morte Celular Programada 1 , Traumatismo por Reperfusão , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Animais , Microglia/metabolismo , Traumatismo por Reperfusão/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Isquemia Encefálica/metabolismo , Masculino , Camundongos , Sistema de Sinalização das MAP Quinases/fisiologia , Apoptose/fisiologia , Camundongos Endogâmicos C57BL , Polaridade Celular/fisiologiaRESUMO
Traumatic brain injury impairs brain function through various mechanisms. Recent studies have shown that alterations in pericytes in various diseases affect neurovascular function, but the effects of TBI on hippocampal pericytes remain unclear. Here, we investigated the effects of RAGE activation on pericytes after TBI using male C57BL/6 J mice. Hippocampal samples were collected at different time points within 7 days after TBI, the expression of PDGFR-ß, NG2 and the HMGB1-S100B/RAGE signaling pathway was assessed by Western blotting, and the integrity of the hippocampal BBB at different time points was measured by immunofluorescence. RAGE-associated BBB damage in hippocampal pericytes occurred early after cortical impact. By culturing primary mouse brain microvascular pericytes, we determined the different effects of HMGB1-S100B on pericyte RAGE. To investigate whether RAGE blockade could protect neurological function after TBI, we reproduced the process of CCI by administering FPS-ZM1 to RAGE-/- mice. TEM images and BBB damage-related assays showed that inhibition of RAGE resulted in a significant improvement in the number of hippocampal vascular basement membranes and tight junctions and a reduction in perivascular oedema compared with those in the untreated group. In contrast, mouse behavioural testing and doublecortin staining indicated that targeting the HMGB1-S100B/RAGE axis after CCI could protect neurological function by reducing pericyte-associated BBB damage. In conclusion, the present study provides experimental evidence for the strong correlation between the pericyte HMGB1-S100B/RAGE axis and NVU damage in the hippocampus at the early stage of TBI and further demonstrates that pericyte RAGE serves as an important target for the protection of neurological function after TBI.
Assuntos
Barreira Hematoencefálica , Lesões Encefálicas Traumáticas , Hipocampo , Camundongos Endogâmicos C57BL , Pericitos , Receptor para Produtos Finais de Glicação Avançada , Animais , Pericitos/metabolismo , Pericitos/patologia , Lesões Encefálicas Traumáticas/patologia , Lesões Encefálicas Traumáticas/metabolismo , Camundongos , Masculino , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Barreira Hematoencefálica/patologia , Barreira Hematoencefálica/metabolismo , Camundongos Knockout , Proteína HMGB1/metabolismo , Subunidade beta da Proteína Ligante de Cálcio S100/metabolismo , BenzamidasRESUMO
Brain metastases account for more than 50 % of intracranial central nervous system tumors. The blood-brain barrier (BBB) is mainly composed of endothelial cells, which exhibit low endocytosis and high efflux pumps. Although they are connected by continuous tight junctions and serve as a protective insulation, the BBB does not prevent the development of brain metastases from non-small cell lung cancer (NSCLC). Improving understanding on the mechanisms underlying the development of brain metastasis and the differential molecular characteristics relative to the primary tumor are therefore key in the treatment of brain metastases. This study evaluated the differential expression of miR-522-3p in NSCLC and brain metastases using the Gene Expression Omnibus database. NSCLC brain metastasis model was constructed to screen for cell lines that demonstrated high potential for brain metastasis; We also observed differential expression of miRNA-522-3p in the paraffin-embedded specimens of non-small cell lung cancer and brain metastases from our hospital. The molecular biological functions of miRNA-522-3p were verified using 5-ethynyl-2'-deoxyuridine (EdU) proliferation assay and Transwell invasion assays. RNA-seq was employed to identify downstream target proteins, and the dual-luciferase reporter assay confirmed Tensin 1 (TNS1), a protein that links the actin cytoskeleton to the extracellular matrix, as the downstream regulatory target protein. In vitro blood-brain barrier models and co-culture models were constructed to further identify the role of miRNA-522-3p and TNS1; the expression of BBB-related proteins (ZO-1 and OLCN) was also identified. In vivo experiments were performed to verify the effects of miRNA-522-3p on the time and incidence of NSCLC brain metastasis. The results showed significantly high expression in GSE51666; consistent results were obtained in brain metastasis cells and paraffin samples. RNA-seq combined with miRNA target protein prediction demonstrated TNS1 to be directly downstream of miR-522-3p and to be associated with cell proliferation and invasion. By regulating ZO-1 and OCLN expression, mi-522-3p/TNS1 may increase tumor cell penetration through the BBB while decreasing its permeability. In vivo, miR-522-3p was further demonstrated to significantly promote the formation of brain metastases. miR-522-3p/TNS1 can affect BBB permeability and encourage the growth of brain metastases by modifying the BBB TJ proteins. This axis offers new therapeutic targets for the prevention of brain metastasis.