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1.
PLoS Pathog ; 16(5): e1008204, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32357162

RESUMO

Zika virus (ZIKV) can infect and cause microcephaly and Zika-associated neurological complications in the developing fetal and adult brains. In terms of pathogenesis, a critical question is how ZIKV overcomes the barriers separating the brain from the circulation and gains access to the central nervous system (CNS). Despite the importance of ZIKV pathogenesis, the route ZIKV utilizes to cross CNS barriers remains unclear. Here we show that in mouse models, ZIKV-infected cells initially appeared in the periventricular regions of the brain, including the choroid plexus and the meninges, prior to infection of the cortex. The appearance of ZIKV in cerebrospinal fluid (CSF) preceded infection of the brain parenchyma. Further the brain infection was significantly attenuated by neutralization of the virus in the CSF, indicating that ZIKV in the CSF at the early stage of infection might be responsible for establishing a lethal infection of the brain. We show that cells infected by ZIKV in the choroid plexus were pericytes. Using in vitro systems, we highlight the possibility that ZIKV crosses the blood-CSF barrier by disrupting the choroid plexus epithelial layer. Taken together, our results suggest that ZIKV might exploit the blood-CSF barrier rather than the blood-brain barrier to invade the CNS.


Assuntos
Plexo Corióideo/patologia , Pericitos/patologia , Infecção por Zika virus/patologia , Animais , Barreira Hematoencefálica/patologia , Encéfalo/patologia , Sistema Nervoso Central/patologia , Chlorocebus aethiops , Plexo Corióideo/metabolismo , Plexo Corióideo/virologia , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microcefalia/complicações , Microcefalia/virologia , Doenças do Sistema Nervoso , Pericitos/metabolismo , Pericitos/virologia , Cultura Primária de Células , Células Vero , Zika virus/fisiologia , Infecção por Zika virus/virologia
2.
Stroke ; 51(5): 1578-1586, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32279622

RESUMO

Background and Purpose- Our recent study demonstrated that release of Prx2 (peroxiredoxin 2) from red blood cells (RBCs) is involved in the inflammatory response and brain injury after intracerebral hemorrhage. The current study investigated the role of extracellular Prx2 in hydrocephalus development after experimental intraventricular hemorrhage. Methods- There were 4 parts in this study. First, Sprague-Dawley rats received an intraventricular injection of lysed RBC or saline and were euthanized at 1 hour for Prx2 measurements. Second, rats received an intraventricular injection of Prx2, deactivated Prx2, or saline. Third, lysed RBC was coinjected with conoidin A, a Prx2 inhibitor, or vehicle. Fourth, rats received Prx2 injection and were treated with minocycline or saline (i.p.). The effects of Prx2 and the inhibitors were examined using magnetic resonance imaging assessing ventriculomegaly, histology assessing ventricular wall damage, and immunohistochemistry to assess inflammation, particularly at the choroid plexus. Results- Intraventricular injection of lysed RBC resulted in increased brain Prx2 and hydrocephalus. Intraventricular injection of Prx2 alone caused hydrocephalus, ventricular wall damage, activation of choroid plexus epiplexus cells (macrophages), and an accumulation of neutrophils. Conoidin A attenuated lysed RBC-induced injury. Systemic minocycline treatment reduced the epiplexus cell activation and hydrocephalus induced by Prx2. Conclusions- Prx2 contributed to the intraventricular hemorrhage-induced hydrocephalus, probably by inducing inflammatory responses in choroid plexus and ventricular wall damage.


Assuntos
Hemorragia Cerebral Intraventricular/metabolismo , Plexo Corióideo/metabolismo , Hidrocefalia/metabolismo , Inflamação/metabolismo , Macrófagos/metabolismo , Peroxirredoxinas/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Hemorragia Cerebral Intraventricular/complicações , Plexo Corióideo/efeitos dos fármacos , Plexo Corióideo/patologia , Modelos Animais de Doenças , Epêndima/efeitos dos fármacos , Epêndima/patologia , Feminino , Hidrocefalia/etiologia , Hylobatidae , Inflamação/patologia , Injeções Intraventriculares , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Masculino , Minociclina/farmacologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/patologia , Peroxirredoxinas/antagonistas & inibidores , Peroxirredoxinas/farmacologia , Quinoxalinas/farmacologia , Ratos , Ratos Sprague-Dawley
3.
J Neurosci ; 40(19): 3849-3861, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32269105

RESUMO

Neonatal stroke is as frequent as stroke in the elderly, but many pathophysiological injury aspects are distinct in neonates, including immune signaling. While myeloid cells can traffic into the brain via multiple routes, the choroid plexus (CP) has been identified as a uniquely educated gate for immune cell traffic during health and disease. To understand the mechanisms of myeloid cell trafficking via the CP and their influence on neonatal stroke, we characterized the phenotypes of CP-infiltrating myeloid cells after transient middle cerebral artery occlusion (tMCAO) in neonatal mice of both sexes in relation to blood-brain barrier permeability, injury, microglial activation, and CX3CR1-CCR2 signaling, focusing on the dynamics early after reperfusion. We demonstrate rapid recruitment of multiple myeloid phenotypes in the CP ipsilateral to the injury, including inflammatory CD45+CD11b+Ly6chighCD86+, beneficial CD45+CD11b+Ly6clowCD206+, and CD45+CD11b+Ly6clowLy6ghigh cells, but only minor leukocyte infiltration into acutely ischemic-reperfused cortex and negligible vascular albumin leakage. We report that CX3CR1-CCR2-mediated myeloid cell recruitment contributes to stroke injury. Considering the complexity of inflammatory cascades triggered by stroke and a role for TLR2 in injury, we also used direct TLR2 stimulation as an independent injury model. TLR2 agonist rapidly recruited myeloid cells to the CP, increased leukocytosis in the CSF and blood, but infiltration into the cortex remained low over time. While the magnitude and the phenotypes of myeloid cells diverged between tMCAO and TLR2 stimulation, in both models, disruption of CX3CR1-CCR2 signaling attenuated both monocyte and neutrophil trafficking to the CP and cortex.SIGNIFICANCE STATEMENT Stroke during the neonatal period leads to long-term disabilities. The mechanisms of ischemic injury and inflammatory response differ greatly between the immature and adult brain. We examined leukocyte trafficking via the choroid plexus (CP) following neonatal stroke in relation to blood-brain barrier integrity, injury, microglial activation, and signaling via CX3CR1 and CCR2 receptors, or following direct TLR2 stimulation. Ischemia-reperfusion triggered marked unilateral CX3CR1-CCR2 dependent accumulation of diverse leukocyte subpopulations in the CP without inducing extravascular albumin leakage or major leukocyte infiltration into the brain. Disrupted CX3CR1-CCR2 signaling was neuroprotective in part by attenuating monocyte and neutrophil trafficking. Understanding the migratory patterns of CP-infiltrating myeloid cells with intact and disrupted CX3CR1-CCR2 signaling could identify novel therapeutic targets to protect the neonatal brain.


Assuntos
Quimiotaxia de Leucócito/fisiologia , Plexo Corióideo/metabolismo , Células Mieloides/metabolismo , Acidente Vascular Cerebral/fisiopatologia , Animais , Animais Recém-Nascidos , Receptor 1 de Quimiocina CX3C/metabolismo , Plexo Corióideo/imunologia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células Mieloides/imunologia , Receptores CCR2/metabolismo , Acidente Vascular Cerebral/imunologia , Acidente Vascular Cerebral/metabolismo , Receptor 1 Toll-Like/metabolismo , Receptor 2 Toll-Like/metabolismo
4.
PLoS Pathog ; 16(3): e1008371, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32130281

RESUMO

The human polyomavirus, JCPyV, is the causative agent of progressive multifocal leukoencephalopathy (PML) in immunosuppressed and immunomodulated patients. Initial infection with JCPyV is common and the virus establishes a long-term persistent infection in the urogenital system of 50-70% of the human population worldwide. A major gap in the field is that we do not know how the virus traffics from the periphery to the brain to cause disease. Our recent discovery that human choroid plexus epithelial cells are fully susceptible to virus infection together with reports of JCPyV infection of choroid plexus in vivo has led us to hypothesize that the choroid plexus plays a fundamental role in this process. The choroid plexus is known to relay information between the blood and the brain by the release of extracellular vesicles. This is particularly important because human macroglia (oligodendrocytes and astrocytes), the major targets of virus infection in the central nervous system (CNS), do not express the known attachment receptors for the virus and do not bind virus in human tissue sections. In this report we show that JCPyV infected choroid plexus epithelial cells produce extracellular vesicles that contain JCPyV and readily transmit the infection to human glial cells. Transmission of the virus by extracellular vesicles is independent of the known virus attachment receptors and is not neutralized by antisera directed at the virus. We also show that extracellular vesicles containing virus are taken into target glial cells by both clathrin dependent endocytosis and macropinocytosis. Our data support the hypothesis that the choroid plexus plays a fundamental role in the dissemination of virus to brain parenchyma.


Assuntos
Plexo Corióideo/metabolismo , Células Epiteliais/metabolismo , Vesículas Extracelulares/metabolismo , Vírus JC/metabolismo , Leucoencefalopatia Multifocal Progressiva/metabolismo , Neuroglia/metabolismo , Receptores Virais/metabolismo , Linhagem Celular Transformada , Plexo Corióideo/patologia , Plexo Corióideo/virologia , Células Epiteliais/patologia , Células Epiteliais/virologia , Vesículas Extracelulares/patologia , Vesículas Extracelulares/virologia , Humanos , Leucoencefalopatia Multifocal Progressiva/patologia , Neuroglia/patologia , Neuroglia/virologia
5.
Biochem Biophys Res Commun ; 524(1): 231-235, 2020 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-31983432

RESUMO

Major depressive disorder (MDD) has become a potential cause of death and disability among young people worldwide. Numerous studies have indicated that the different cerebrospinal fluid (CSF) proteins may be used as mediums for MDD. Given the emergent interest of CSF proteins in MDD, we validated proteins expression in the choroid plexus (CP), the brain region that produces CSF in the lateral ventricle, the third ventricle, and the fourth ventricle of the central nervous system (CNS). The CSF constantly exchanges molecular substances with the brain tissue, which can dynamically reflect the metabolic microenvironment of the brain. In our previous study, Pepsin A (PGA) and periostin (POSTN) was associated with depressive-like behaviors of depressed macaca fascicularis models in CSF. Moreover, proteins that are expressed in the CP can be secreted into the CSF and may be associated MDD. This study sought to demonstrate the discrepancy of PGA and POSTN in the CP between Lipopolysaccharide (LPS)-induce depressed mice models and wild type (WT) mice. Our findings suggest that PGA and POSTN expression in CP of mice could be a possible candidate pathogenesis involved in MDD, which may contribute to a better understanding and treatment of MDD.


Assuntos
Moléculas de Adesão Celular/metabolismo , Plexo Corióideo/metabolismo , Depressão/metabolismo , Prostaglandinas A/metabolismo , Animais , Comportamento Animal , Peso Corporal , Comportamento Alimentar , Hipocampo/metabolismo , Imobilização , Masculino , Camundongos Endogâmicos C57BL , Sacarose , Fatores de Tempo
6.
J Neuroinflammation ; 16(1): 232, 2019 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-31752904

RESUMO

BACKGROUND: Echovirus 30 (E-30) is one of the most frequently isolated pathogens in aseptic meningitis worldwide. To gain access to the central nervous system (CNS), E-30 and immune cells have to cross one of the two main barriers of the CNS, the epithelial blood-cerebrospinal fluid barrier (BCSFB) or the endothelial blood-brain barrier (BBB). In an in vitro model of the BCSFB, it has been shown that E-30 can infect human immortalized brain choroid plexus papilloma (HIBCPP) cells. METHODS: In this study we investigated the migration of different T cell subpopulations, naive and effector T cells, through HIBCPP cells during E-30 infection. Effects of E-30 infection and the migration process were evaluated via immunofluorescence and flow cytometry analysis, as well as transepithelial resistance and dextran flux measurement. RESULTS: Th1 effector cells and enterovirus-specific effector T cells migrated through HIBCPP cells more efficiently than naive CD4+ T cells following E-30 infection of HIBCPP cells. Among the different naive T cell populations, CD8+ T cells crossed the E-30-infected HIBCPP cell layer in a significantly higher number than CD4+ T cells. A large amount of effector T cells also remained attached to the basolateral side of the HIBCPP cells compared with naive T cells. Analysis of HIBCPP barrier function showed significant alteration after E-30 infection and trans- as well as paracellular migration of T cells independent of the respective subpopulation. Morphologic analysis of migrating T cells revealed that a polarized phenotype was induced by the chemokine CXCL12, but reversed to a round phenotype after E-30 infection. Further characterization of migrating Th1 effector cells revealed a downregulation of surface adhesion proteins such as LFA-1 PSGL-1, CD44, and CD49d. CONCLUSION: Taken together these results suggest that naive CD8+ and Th1 effector cells are highly efficient to migrate through the BCSFB in an inflammatory environment. The T cell phenotype is modified during the migration process through HIBCPP cells.


Assuntos
Movimento Celular/imunologia , Plexo Corióideo/metabolismo , Plexo Corióideo/virologia , Infecções por Echovirus/imunologia , Linfócitos T/imunologia , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/virologia , Humanos , Linfócitos T/metabolismo , Células Tumorais Cultivadas
7.
PLoS One ; 14(9): e0221555, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31479465

RESUMO

The unknown role of the carrier protein transthyretin (TTR) in mechanisms of functional recovery in the postischemic brain prompted us to study its expression following experimental stroke. Male C57/B6 mice (age 9 to 10 weeks) were subjected to permanent focal ischemia induced by photothrombosis (PT) and brain tissues were analyzed for ttr expression and TTR levels at 24 hours, 48 hours, 7 days and 14 days following the insult by RT-PCR, Western blot and immunohistochemistry. Fourteen days after PT, non-specific TTR-like immunoreactive globules were found in the ischemic core and surrounding peri-infarct region by immunohistochemistry that could not be allocated to DAPI positive cells. No TTR immunoreactivity was found when stainings were performed with markers for neurons (Neuronal Nuclei, NeuN), reactive astrocytes (glial fibrillary acidic protein, GFAP) or microglia (cluster of differentiation 68, CD68). In addition, we could not find TTR by immunoblotting in protein extracts obtained from the ischemic territory nor ttr expression by RT-PCR at all time points following PT. In all experiments, ttr expression in the choroid plexus and TTR in the mouse serum served as positive controls and recombinant legumain peptide as negative control. Together, our results indicate that TTR is not synthesized in brain resident cells in the ischemic infarct core and adjacent peri-infarct area. Thus, it seems unlikely that in situ synthesized TTR is involved in mechanisms of tissue reorganization during the first 14 days following PT.


Assuntos
Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Encéfalo/metabolismo , Pré-Albumina/genética , Pré-Albumina/metabolismo , Animais , Astrócitos/metabolismo , Encéfalo/patologia , Isquemia Encefálica/patologia , Plexo Corióideo/metabolismo , Plexo Corióideo/patologia , Modelos Animais de Doenças , Expressão Gênica , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/metabolismo , Neurônios/metabolismo , Pré-Albumina/líquido cefalorraquidiano , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
8.
ACS Appl Mater Interfaces ; 11(43): 39633-39647, 2019 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-31532618

RESUMO

Effective and timely delivery of therapeutic agents from the systemic circulation to the central nervous system (CNS) is often precluded by the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB). A new pathway of folate uptake mediated by folate receptor alpha (FRα, molecular weight of 28.29 kg mol-1) occurring in various epithelial cells of the CNS (e.g., choroid plexus) was described. Aiming to investigate this mechanism for the delivery of nanomedicines to the CNS, in this work, we initially produced nanoparticles (NPs) made of a highly hydrophobic poly(ethylene glycol)-b-poly(ε-caprolactone) (PEG-b-PCL) block copolymer functionalized with an amine moiety in the edge of the PEG block by a simple nanoprecipitation method. Hydrophilic PEG blocks migrated to the NP surface during formation, exposing primary amine groups that were used to conjugate the targeting ligand, FRα. The size of the NPs was in the 58-98 nm range and standard deviation (S.D., a measure of the size population peak width) of 26-41 nm, as measured by dynamic light scattering (DLS). The FRα conjugation yield ranged between 50% and 75% (determined indirectly by the bicinchoninic acid protein assay). Pristine and FRα-modified NPs showed good compatibility with primary human choroid plexus epithelial cells (HCPEpiCs). The uptake of FRα-conjugated NPs by HCPEpiCs was qualitatively evaluated in vitro using inverted optical fluorescence and confocal microscopy. FRα-modified NPs were internalized by HCPEpiCs to a greater extent than the unmodified counterparts. Then, their permeability was characterized in standard and inverted HCPEpiC monolayers. In both cases, NPs surface modified with the FRα and complexed to folic acid (FA) showed significantly higher apparent permeability coefficient (Papp) values than the pristine ones. Finally, the biodistribution of unmodified and FRα-FA-modified NPs following intravenous (i.v.) administration was compared in ICR mice. Results indicated that conjugation of the FRα-FA complex to the NP surface promotes higher accumulation in the brain, highlighting the promise of FRα-FA-modified NPs to serve as a platform for the targeting of active molecules to the CNS from the systemic circulation.


Assuntos
Barreira Hematoencefálica/metabolismo , Plexo Corióideo/metabolismo , Sistemas de Liberação de Medicamentos , Células Epiteliais/metabolismo , Receptor 1 de Folato , Nanopartículas/química , Animais , Barreira Hematoencefálica/patologia , Linhagem Celular , Plexo Corióideo/patologia , Células Epiteliais/patologia , Receptor 1 de Folato/química , Receptor 1 de Folato/farmacocinética , Receptor 1 de Folato/farmacologia , Humanos , Camundongos , Camundongos Endogâmicos ICR
9.
Am J Physiol Cell Physiol ; 317(5): C881-C893, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31411921

RESUMO

The choroid plexus (CP), composed of capillaries surrounded by a barrier epithelium, is the main producer of cerebrospinal fluid (CSF). The CP epithelium regulates the transport of ions and water between the blood and the ventricles, contributing to CSF production and composition. Several studies suggest a connection between the cation channel transient receptor potential vanilloid-4 (TRPV4) and transepithelial ion movement. TRPV4 is a nonselective, calcium-permeable cation channel present in CP epithelia reported to be activated by cytokines and inflammatory mediators. Utilizing the PCP-R (porcine choroid plexus-Riems) cell line, we investigated the effects of various cytokines and inflammatory mediators on TRPV4-mediated activity. Select proinflammatory cytokines (TNF-α, IL-1ß, TGF-ß1) had inhibitory effects on TRPV4-stimulated transepithelial ion flux and permeability changes, whereas anti-inflammatory cytokines (IL-10, IL-4, and IL-6) had none. Quantitative mRNA analysis showed that these cytokines had no effect on TRPV4 transcription levels. Inhibition of the transcription factor NF-κB, involved in the production and regulation of several inflammatory cytokines, inhibited TRPV4-mediated activity, suggesting a link between TRPV4 and cytokine production. Contrary to published studies, the proinflammatory mediator arachidonic acid (AA) had inhibitory rather than stimulatory effects on TRPV4-mediated responses. However, inhibition of AA metabolism also caused inhibitory effects on TRPV4, suggesting a complex interaction of AA and its metabolites in the regulation of TRPV4 activity. Together these data imply that TRPV4 activity is involved in the inflammatory response; it is negatively affected by proinflammatory mediators. Furthermore, arachidonic acid metabolites, but not arachidonic acid itself, are positive regulators of TRPV4.


Assuntos
Plexo Corióideo/metabolismo , Citocinas/metabolismo , Células Epiteliais/metabolismo , Mediadores da Inflamação/metabolismo , Canais de Cátion TRPV/fisiologia , Animais , Linhagem Celular , Plexo Corióideo/citologia , Plexo Corióideo/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Leucina/análogos & derivados , Leucina/farmacologia , Sulfonamidas/farmacologia , Suínos , Canais de Cátion TRPV/agonistas
10.
J Physiol Pharmacol ; 70(2)2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31356186

RESUMO

In ewes, the turnover rate of cerebrospinal fluid (CSF), mainly produced by choroid plexus (ChP), is photoperiodically modulated and is higher during short days (SDs) than long days (LDs). We demonstrated that, melatonin from continuous slow-release implants increases the expression of aquaporins, water channel-forming proteins engaged in transcellular water transport (across the plasma membrane of the cells), in the ovine ChP. This study evaluated the effect of slow-release melatonin implants on the expression of claudin-2 (CLDN2), a pore-forming protein that allows the paracellular passage (between the cells) of select inorganic cations and water, in the ovine ChP. The studies were conducted on ovariectomized, estradiol-implanted ewes during seasonal anestrus (May/June). The ewes were implanted with slow-release-melatonin implants (n = 6, Melovine 18 mg) or sham-implanted (n = 6). Blood samples were collected for melatonin and prolactin measurements. The ewes were sacrificed 40 days after the melatonin/sham implantation, and the ChPs from the brain ventricles were collected for real-time PCR and Western blot analyses. Plasma melatonin concentration reached the median value of 120.4 pg/ml (range: min/max = 29.6/447.0) or was below the detection limit 40 days after the melatonin/sham implantation, respectively. The area under the curve of the plasma prolactin concentration was significantly (P < 0.05) higher in sham-implanted ewes than in melatonin-implanted ewes. CLDN2 expression in the ChP was significantly (P < 0.05) higher in melatonin-implanted ewes than in sham-implanted ewes at both the mRNA and protein levels. This is the first evidence for the photoperiodic regulation of CLDN2 expression in the ovine ChP, since it has been shown that slow-release melatonin implants during LDs, mimicking SDs, increased the expression of CLDN2. This may partially explain the higher turnover rate of CSF observed in ewes during SDs.


Assuntos
Plexo Corióideo/efeitos dos fármacos , Claudina-2/metabolismo , Melatonina/administração & dosagem , Regulação para Cima/efeitos dos fármacos , Animais , Plexo Corióideo/metabolismo , Estradiol/sangue , Feminino , Melatonina/sangue , Prolactina/sangue , Próteses e Implantes , Estações do Ano , Ovinos
11.
PLoS One ; 14(6): e0218041, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31173612

RESUMO

There is strong evidence that neuronal hyper-excitability underlies migraine, and may or may not be preceded by cortical spreading depression. However, the mechanisms for cortical spreading depression and/or migraine are not established. Previous studies reported that cerebrospinal fluid (CSF) [Na+] is higher during migraine, and that higher extracellular [Na+] leads to hyper-excitability. We raise the hypothesis that altered choroid plexus Na+, K+-ATPase activity can cause both migraine phenomena: inhibition raises CSF [K+] and initiates cortical spreading depression, while activation raises CSF [Na+] and causes migraine. In this study, we examined levels of specific Na+, K+-ATPase inhibitors, endogenous ouabain-like compounds (EOLC), in CSF from migraineurs and controls. CSF EOLC levels were significantly lower during ictal migraine (0.4 nM +/- 0.09) than from either controls (1.8 nM +/- 0.4) or interictal migraineurs (3.1 nM +/- 1.9). Blood plasma EOLC levels were higher in migraineurs than controls, but did not differ between ictal and interictal states. In a Sprague-Dawley rat model of nitroglycerin-triggered central sensitization, we changed the concentrations of EOLC and CSF sodium, and measured aversive mechanical threshold (von Frey hairs), trigeminal nucleus caudalis activation (cFos), and CSF [Na+] (ultra-high field 23Na MRI). Animals were sensitized by three independent treatments: intraperitoneal nitroglycerin, immunodepleting EOLC from cerebral ventricles, or cerebroventricular infusion of higher CSF [Na+]. Conversely, nitroglycerin-triggered sensitization was prevented by either vascular or cerebroventricular delivery of the specific Na+, K+-ATPase inhibitor, ouabain. These results affirm our hypothesis that higher CSF [Na+] is linked to human migraine and to a rodent migraine model, and demonstrate that EOLC regulates them both. Our data suggest that altered choroid plexus Na+, K+-ATPase activity is a common source of these changes, and may be the initiating mechanism in migraine.


Assuntos
Líquido Cefalorraquidiano/metabolismo , Íons/metabolismo , Transtornos de Enxaqueca/etiologia , Transtornos de Enxaqueca/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Sódio/metabolismo , Adolescente , Adulto , Idoso , Animais , Plexo Corióideo/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ouabaína/metabolismo , Ratos , Ratos Sprague-Dawley , Adulto Jovem
12.
eNeuro ; 6(2)2019.
Artigo em Inglês | MEDLINE | ID: mdl-31064838

RESUMO

Proliferation and migration during adult neurogenesis are regulated by a microenvironment of signaling molecules originating from local vasculature, from CSF produced by the choroid plexus, and from local supporting cells including astrocytes. Here, we focus on the function of OTX2 homeoprotein transcription factor in the mouse adult ventricular-subventricular zone (V-SVZ), which generates olfactory bulb neurons. We find that OTX2 secreted by choroid plexus is transferred to the supporting cells of the V-SVZ and rostral migratory stream. Deletion of Otx2 in choroid plexus affects neuroblast migration and reduces the number of olfactory bulb newborn neurons. Adult neurogenesis was also decreased by expressing secreted single-chain antibodies to sequester OTX2 in the CSF, demonstrating the importance of non-cell-autonomous OTX2. We show that OTX2 activity modifies extracellular matrix components and signaling molecules produced by supporting astrocytes. Thus, we reveal a multilevel and non-cell-autonomous role of a homeoprotein and reinforce the choroid plexus and astrocytes as key niche compartments affecting adult neurogenesis.


Assuntos
Astrócitos/metabolismo , Líquido Cefalorraquidiano/metabolismo , Plexo Corióideo/metabolismo , Matriz Extracelular/metabolismo , Ventrículos Laterais , Neurogênese/fisiologia , Bulbo Olfatório , Fatores de Transcrição Otx/fisiologia , Transdução de Sinais/fisiologia , Animais , Movimento Celular/fisiologia , Feminino , Ventrículos Laterais/citologia , Ventrículos Laterais/metabolismo , Masculino , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Bulbo Olfatório/citologia , Bulbo Olfatório/metabolismo , Fatores de Transcrição Otx/deficiência , Fatores de Transcrição Otx/metabolismo
13.
eNeuro ; 6(2)2019.
Artigo em Inglês | MEDLINE | ID: mdl-31119189

RESUMO

Neuronal cholinergic circuits have been implicated in cognitive function and neurological disease, but the role of cholinergic signaling in other cellular populations within the brain has not been as fully defined. Here, we show that cholinergic signaling mechanisms are involved in mediating the function of the choroid plexus, the brain structure responsible for generating CSF and releasing various factors into the brain. The choroid plexus was found to express markers of endogenous cholinergic signaling, including multiple nicotinic acetylcholine receptor (nAChR) subtypes in a region-specific manner, and application of nicotine was found to induce cellular activation, as evidenced by calcium influx in primary tissue. During intravenous nicotine self-administration in male rats, nicotine increased expression of transthyretin, a protein selectively produced and released by the choroid plexus, and microRNA-204 (mir-204), a transcript found in high levels in the choroid plexus and CSF. Finally, human choroid plexus tissue from both sexes was found to exhibit similar nAChR, transthyretin and mir-204 expression profiles, supporting the translational relevance of the findings. Together, these studies demonstrate functionally active cholinergic signaling mechanisms in the choroid plexus, the resulting effects on transthyretin and mir-204 expression, and reveal the direct mechanism by which nicotine modulates function of this tissue.


Assuntos
Plexo Corióideo , MicroRNAs , Nicotina/farmacologia , Agonistas Nicotínicos/farmacologia , Pré-Albumina , Receptores Nicotínicos , Transdução de Sinais/efeitos dos fármacos , Animais , Plexo Corióideo/efeitos dos fármacos , Plexo Corióideo/metabolismo , Feminino , Humanos , Masculino , MicroRNAs/efeitos dos fármacos , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Pré-Albumina/efeitos dos fármacos , Pré-Albumina/metabolismo , Ratos , Ratos Wistar , Receptores Nicotínicos/efeitos dos fármacos , Receptores Nicotínicos/metabolismo
14.
Nat Commun ; 10(1): 1498, 2019 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-30940800

RESUMO

WNTs are lipid-modified proteins that control multiple functions in development and disease via short- and long-range signaling. However, it is unclear how these hydrophobic molecules spread over long distances in the mammalian brain. Here we show that WNT5A is produced by the choroid plexus (ChP) of the developing hindbrain, but not the telencephalon, in both mouse and human. Since the ChP produces and secretes the cerebrospinal fluid (CSF), we examine the presence of WNT5A in the CSF and find that it is associated with lipoprotein particles rather than exosomes. Moreover, since the CSF flows along the apical surface of hindbrain progenitors not expressing Wnt5a, we examined whether deletion of Wnt5a in the ChP controls their function and find that cerebellar morphogenesis is impaired. Our study thus identifies the CSF as a route and lipoprotein particles as a vehicle for long-range transport of biologically active WNT in the central nervous system.


Assuntos
Lipoproteínas/líquido cefalorraquidiano , Rombencéfalo/embriologia , Proteína Wnt-5a/metabolismo , Animais , Transporte Biológico , Plexo Corióideo/metabolismo , Feminino , Humanos , Masculino , Camundongos Endogâmicos ICR , Morfogênese , Rombencéfalo/metabolismo , Transdução de Sinais , Proteína Wnt-5a/genética
15.
Brain Behav Immun ; 79: 216-227, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30822467

RESUMO

Perinatal infection and inflammation are major risk factors for injury in the developing brain, however, underlying mechanisms are not fully understood. Leukocyte migration to the cerebrospinal fluid (CSF) and brain is a hallmark of many pathologies of the central nervous system including those in neonates. We previously reported that systemic activation of Toll-like receptor (TLR) 2, a major receptor for gram-positive bacteria, by agonist Pam3CSK4 (P3C) resulted in dramatic neutrophil and monocyte infiltration to the CSF and periventricular brain of neonatal mice, an effect that was absent by the TLR4 agonist, LPS. Here we first report that choroid plexus is a route of TLR2-mediated leukocyte infiltration to the CSF by performing flow cytometry and transmission electron microscopy (TEM) of the choroid plexus. Next, we exploited the striking discrepancy between P3C and LPS effects on cell migration to determine the pathways regulating leukocyte trafficking through the choroid plexus. We performed RNA sequencing on the choroid plexus after administration of P3C and LPS to postnatal day 8 mice. A cluster gene analysis revealed a TLR2-specific signature of chemotaxis represented by 80-fold increased expression of the gene Ccl3 and 1000-fold increased expression of the gene Cxcl2. Ingenuity pathway analysis (IPA) revealed TLR2-specific molecular signaling related to cytoskeleton organization (e.g. actin signaling) as well as inositol phospholipids biosynthesis and degradation. This included upregulation of genes such as Rac2 and Micall2. In support of IPA results, ultrastructural analysis by TEM revealed clefting and perforations in the basement membrane of the choroid plexus epithelial cells in P3C-treated mice. In summary, we show that the choroid plexus is a route of TLR2-mediated transmigration of neutrophils and monocytes to the developing brain, and reveal previously unrecognized mechanisms that includes a specific chemotaxis profile as well as pathways regulating cytoskeleton and basement membrane remodeling.


Assuntos
Plexo Corióideo/metabolismo , Plexo Corióideo/ultraestrutura , Receptor 2 Toll-Like/genética , Animais , Animais Recém-Nascidos , Encéfalo/metabolismo , Movimento Celular , Sistema Nervoso Central/metabolismo , Quimiotaxia/genética , Quimiotaxia/fisiologia , Plexo Corióideo/fisiologia , Citoesqueleto/genética , Citoesqueleto/fisiologia , Citometria de Fluxo/métodos , Inflamação/metabolismo , Leucócitos/metabolismo , Leucócitos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Transmissão/métodos , Monócitos/metabolismo , Neutrófilos/metabolismo , Receptor 2 Toll-Like/metabolismo , Transcriptoma
16.
Sci Rep ; 9(1): 203, 2019 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-30659216

RESUMO

The tight junction protein claudin-3 has been identified as a transcriptional target of the Wnt/ß-catenin signaling pathway regulating blood-brain barrier (BBB) maturation. In neurological disorders loss of claudin-3 immunostaining is observed at the compromised BBB and blood-cerebrospinal fluid barrier (BCSFB). Although these observations support a central role of claudin-3 in regulating brain barriers' tight junction integrity, expression of claudin-3 at the brain barriers has remained a matter of debate. This prompted us to establish claudin-3-/- C57BL/6J mice to study the role of claudin-3 in brain barrier integrity in health and neuroinflammation. Bulk and single cell RNA sequencing and direct comparative qRT-PCR analysis of brain microvascular samples from WT and claudin-3-/- mice show beyond doubt that brain endothelial cells do not express claudin-3 mRNA. Detection of claudin-3 protein at the BBB in vivo and in vitro is rather due to junctional reactivity of anti-claudin-3 antibodies to an unknown antigen still detected in claudin-3-/- brain endothelium. We confirm expression and junctional localization of claudin-3 at the BCSFB of the choroid plexus. Our study clarifies that claudin-3 is not expressed at the BBB and shows that absence of claudin-3 does not impair brain barrier function during health and neuroinflammation in C57BL/6J mice.


Assuntos
Barreira Hematoencefálica/metabolismo , Claudina-3/metabolismo , Junções Íntimas/metabolismo , Animais , Transporte Biológico , Encéfalo/metabolismo , Plexo Corióideo/metabolismo , Claudina-3/genética , Células Endoteliais/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Junções Íntimas/genética , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/genética , Via de Sinalização Wnt/fisiologia
17.
Parkinsonism Relat Disord ; 61: 88-93, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30503748

RESUMO

INTRODUCTION: Regenerative therapies in Parkinson's disease aim to slow neurodegeneration and re-establish damaged neuronal circuitry. Neurotrophins are potent endogenous regulators of neuronal survival, development and regeneration. They represent an attractive regenerative treatment option in Parkinson's disease. Porcine choroid plexus produces a number of neurotrophins, and can be safely delivered to the striatum in an encapsulated formulation (termed NTCELL®) to protect them from immune attack. NTCELL® has shown regenerative potential in animal models of stroke, Huntington's disease and Parkinson's disease. Following promising results from an initial open label safety study of intra-striatal delivery of NTCELL® in human subjects, we sought to specifically investigate the safety and efficacy of NTCELL® for the treatment of Parkinson's disease. METHODS: 18 patients aged 56-65 years with idiopathic Parkinson's disease of at least 5 years duration were randomised to receive either sham surgery (general anaesthesia and partial thickness burr holes) or intra-striatal delivery of NTCELL® (the 3 groups in the treatment arm receiving incremental NTCELL® doses). RESULTS: At 26 weeks, we found no significant difference in total UPDRS scores ('on' and 'off'), UPDRS motor scores ('on' and 'off'), PDQ-39, UDysRS, timed walk or modified Hoehn and Yahr stage between patients implanted with NTCELL® and patients undergoing sham procedure. There were no serious adverse events or xenogeneic viral transmission during the study. CONCLUSION: The study did not meet its primary efficacy end-point of a change in UPDRS at 26 weeks post-intervention compared with baseline. Stereotactic NTCELL® implantation was safe and well tolerated.


Assuntos
Encapsulamento de Células , Transplante de Células/métodos , Plexo Corióideo/citologia , Neostriado , Doença de Parkinson/terapia , Idoso , Alginatos , Animais , Plexo Corióideo/metabolismo , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Crescimento Neural/metabolismo , Suínos , Transplante Heterólogo , Resultado do Tratamento
19.
Neurobiol Dis ; 121: 95-105, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30261283

RESUMO

The involvement of the 18kDa translocator protein (TSPO), a marker of neuroinflammation, in Alzheimer's disease (AD) remains controversial. In the present report, we used [125I]-CLINDE, a SPECT TSPO radiotracer never before used in AD, and we investigated the relationship between TSPO and amyloid plaque density (using [125I]-DRM106) in a triple transgenic mouse model of AD (3xTgAD, APPSWE, PS1M146V and TauP301L). Our results show that TSPO increases appear before those of amyloid deposits. Moreover, the different parts of the hippocampus are differentially affected. Indeed, for both TSPO and amyloid, the subiculum is affected earlier and the ventral hippocampus later than the dorsal hippocampus. In the subiculum and the dorsal hippocampus of 3xTgAD mice, a positive correlation between TSPO and of amyloid deposit levels is observed. This data supports the hypothesis that TSPO could be used as a predictive marker of amyloid pathology. In addition, our immunohistochemical data shows a segregation of TSPO in the hippocampus and immunofluorescence imaging revealed a mainly microglial origin of the TSPO expression. Thus, imaging TSPO with CLINDE may be a good alternative to PET radiotracers.


Assuntos
Doença de Alzheimer/metabolismo , Hipocampo/metabolismo , Placa Amiloide/metabolismo , Receptores de GABA/metabolismo , Doença de Alzheimer/complicações , Doença de Alzheimer/diagnóstico por imagem , Animais , Plexo Corióideo/metabolismo , Modelos Animais de Doenças , Encefalite/complicações , Encefalite/metabolismo , Feminino , Hipocampo/diagnóstico por imagem , Humanos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Tomografia Computadorizada de Emissão de Fóton Único/métodos
20.
Mult Scler ; 25(4): 523-531, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-29421990

RESUMO

BACKGROUND: CD59, a broadly expressed glycosylphosphatidylinositol-anchored protein, is the principal cell inhibitor of complement membrane attack on cells. In the demyelinating disorders, multiple sclerosis (MS) and neuromyelitis optica spectrum disorder (NMOSD), elevated complement protein levels, including soluble CD59 (sCD59), were reported in cerebrospinal fluid (CSF). OBJECTIVES: We compared sCD59 levels in CSF and matched plasma in controls and patients with MS, NMOSD and clinically isolated syndrome (CIS) and investigated the source of CSF sCD59 and whether it was microparticle associated. METHODS: sCD59 was quantified using enzyme-linked immunosorbent assay (ELISA; Hycult; HK374-02). Patient and control CSF was subjected to western blotting to characterise anti-CD59-reactive materials. CD59 was localised by immunostaining and in situ hybridisation. RESULTS: CSF sCD59 levels were double those in plasma (CSF, 30.2 ng/mL; plasma, 16.3 ng/mL). Plasma but not CSF sCD59 levels differentiated MS from NMOSD, MS from CIS and NMOSD/CIS from controls. Elimination of microparticles confirmed that CSF sCD59 was not membrane anchored. CONCLUSION: CSF levels of sCD59 are not a biomarker of demyelinating diseases. High levels of sCD59 in CSF relative to plasma suggest an intrathecal source; CD59 expression in brain parenchyma was low, but expression was strong on choroid plexus (CP) epithelium, immediately adjacent the CSF, suggesting that this is the likely source.


Assuntos
Antígenos CD59/líquido cefalorraquidiano , Plexo Corióideo/metabolismo , Doenças Desmielinizantes/líquido cefalorraquidiano , Esclerose Múltipla/líquido cefalorraquidiano , Neuromielite Óptica/líquido cefalorraquidiano , Adulto , Antígenos CD59/sangue , Doenças Desmielinizantes/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Esclerose Múltipla/sangue , Neuromielite Óptica/sangue
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