scholarly journals Notch1 and Galectin-3 Modulate Cortical Reactive Astrocyte Response After Brain Injury

2021 ◽  
Vol 9 ◽  
Author(s):  
Tais Novaki Ribeiro ◽  
Lina Maria Delgado-García ◽  
Marimelia A. Porcionatto
Keyword(s):  
Brain Injury ◽  
Notch Signaling ◽  
Nuclear Translocation ◽  
Reactive Astrocytes ◽  
Reactive Astrocyte ◽  
Notch1 Signaling ◽  
Cortical Astrocytes ◽  
Lesion Core ◽  
Galectin 3

After a brain lesion, highly specialized cortical astrocytes react, supporting the closure or replacement of the damaged tissue, but fail to regulate neural plasticity. Growing evidence indicates that repair response leads astrocytes to reprogram, acquiring a partially restricted regenerative phenotype in vivo and neural stem cells (NSC) hallmarks in vitro. However, the molecular factors involved in astrocyte reactivity, the reparative response, and their relation to adult neurogenesis are poorly understood and remain an area of intense investigation in regenerative medicine. In this context, we addressed the role of Notch1 signaling and the effect of Galectin-3 (Gal3) as underlying molecular candidates involved in cortical astrocyte response to injury. Notch signaling is part of a specific neurogenic microenvironment that maintains NSC and neural progenitors, and Gal3 has a preferential spatial distribution across the cortex and has a central role in the proliferative capacity of reactive astrocytes. We report that in vitro scratch-reactivated cortical astrocytes from C57Bl/6J neonatal mice present nuclear Notch1 intracellular domain (NICD1), indicating Notch1 activation. Colocalization analysis revealed a subpopulation of reactive astrocytes at the lesion border with colocalized NICD1/Jagged1 complexes compared with astrocytes located far from the border. Moreover, we found that Gal3 increased intracellularly, in contrast to its extracellular localization in non-reactive astrocytes, and NICD1/Gal3 pattern distribution shifted from diffuse to vesicular upon astrocyte reactivation. In vitro, Gal3–/– reactive astrocytes showed abolished Notch1 signaling at the lesion core. Notch1 receptor, its ligands (Jagged1 and Delta-like1), and Hes5 target gene were upregulated in C57Bl/6J reactive astrocytes, but not in Gal3–/– reactive astrocytes. Finally, we report that Gal3–/– mice submitted to a traumatic brain injury model in the somatosensory cortex presented a disrupted response characterized by the reduced number of GFAP reactive astrocytes, with smaller cell body perimeter and decreased NICD1 presence at the lesion core. These results suggest that Gal3 might be essential to the proper activation of Notch signaling, facilitating the cleavage of Notch1 and nuclear translocation of NICD1 into the nucleus of reactive cortical astrocytes. Additionally, we hypothesize that reactive astrocyte response could be dependent on Notch1/Jagged1-Hes5 signaling activation following brain injury.

Abstract 386: Notch1 Signaling Negatively Modulates Repolarizing Kv Currents in Cardiomyocytes

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Giulia Borghetti ◽  
Sergio Signore ◽  
Andrea Sorrentino ◽  
Polina Goichberg ◽  
Marcello Rota
Keyword(s):  
Patch Clamp ◽  
Notch Signaling ◽  
Splice Variants ◽  
Postnatal Life ◽  
Wild Type ◽  
Early Postnatal Development ◽  
Electrophysiological Properties ◽  
Notch1 Signaling ◽  
Developing Heart

Notch1 is a critical signaling pathway during embryonic and early postnatal development and activity of this transduction system gradually declines after birth. Conversely, outward K + Kv currents are progressively increased in myocytes during postnatal life leading to shortening of the action potential (AP) duration and acquisition of the mature electrical phenotype. Thus, we tested the possibility that Notch1 signaling modulates the electrophysiological properties of developing cells by interfering with Kv current densities. For this purpose, molecular and physiological studies were conducted in vitro using mouse neonatal myocytes (NMs). These assays were complemented with tests in adult cells obtained from a conditional mouse model of Notch1 intracellular domain (NICD) overexpression. NMs were treated with a γ-secretase inhibitor (DAPT) to prevent the cleavage of Notch1 receptor and subsequent translocation of the active NICD to the nucleus. By RT-PCR, Notch1 targets Hes1 and Hey1 were reduced with DAPT-treatment, confirming the effective perturbation of Notch1 signaling. Importantly, inhibition of Notch signaling led to upregulation of the three splice variants of the Kv channel-interacting proteins 2 (KChIP2) gene, which controls the appearance of outward Kv currents in myocytes. By patch-clamp, Kv currents were identified in only 15% of NMs (n=20) cultured for 1-2 days in in control condition, whereas this fraction increased to 50% (n=18) in cells treated with DAPT. Thus, Notch inhibition promotes the expression of KChIP2 and appearance of Kv currents. To clarify these in vitro findings, the consequences of ectopic activation of Notch1 signaling on the electrical behavior of adult myocytes were established. By patch-clamp, NICD-overexpressing myocytes presented a >2-fold longer duration of the early repolarization phase of the AP, with respect to cells from wild type hearts. This alteration was coupled with a >50% reduction of Kv currents I to and I Kslow1 in NICD-overexpressing cells, with respect to wild-type myocytes. Thus, Notch signaling represses KChIP2 and Kv currents in cardiomyocytes representing an important modulator of the electrical phenotype of the developing heart.

scholarly journals Activation of Notch1 signaling in podocytes by glucose-derived AGEs contributes to proteinuria

2020 ◽  
Vol 8 (1) ◽  
pp. e001203
Author(s):  
Rajkishor Nishad ◽  
Prajakta Meshram ◽  
Ashish Kumar Singh ◽  
G Bhanuprakash Reddy ◽  
Anil Kumar Pasupulati
Keyword(s):  
Notch Signaling ◽  
Epithelial To Mesenchymal Transition ◽  
Mesenchymal Transition ◽  
Notch1 Signaling ◽  
Reverse Transcription Pcr ◽  
Filtration Barrier ◽  
Foot Process ◽  
Design And Methods

IntroductionAdvanced glycation end-products (AGEs) are implicated in the pathogenesis of diabetic nephropathy (DN). Previous studies have shown that AGEs contribute to glomerulosclerosis and proteinuria. Podocytes, terminally differentiated epithelial cells of the glomerulus and the critical component of the glomerular filtration barrier, express the receptor for AGEs (RAGE). Podocytes are susceptible to severe injury during DN. In this study, we investigated the mechanism by which AGEs contribute to podocyte injury.Research design and methodsGlucose-derived AGEs were prepared in vitro. Reactivation of Notch signaling was examined in AGE-treated human podocytes (in vitro) and glomeruli from AGE-injected mice (in vivo) by quantitative reverse transcription-PCR, western blot analysis, ELISA and immunohistochemical staining. Further, the effects of AGEs on epithelial to mesenchymal transition (EMT) of podocytes and expression of fibrotic markers were evaluated.ResultsUsing human podocytes and a mouse model, we demonstrated that AGEs activate Notch1 signaling in podocytes and provoke EMT. Inhibition of RAGE and Notch1 by FPS-ZM1 (N-Benzyl-4-chloro-N-cyclohexylbenzamide) and DAPT (N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenyl glycine t-butylester), respectively, abrogates AGE-induced Notch activation and EMT. Inhibition of RAGE and Notch1 prevents AGE-induced glomerular fibrosis, thickening of the glomerular basement membrane, foot process effacement, and proteinuria. Furthermore, kidney biopsy sections from people with DN revealed the accumulation of AGEs in the glomerulus with elevated RAGE expression and activated Notch signaling.ConclusionThe data suggest that AGEs activate Notch signaling in the glomerular podocytes. Pharmacological inhibition of Notch signaling by DAPT ameliorates AGE-induced podocytopathy and fibrosis. Our observations suggest that AGE-induced Notch reactivation in mature podocytes could be a novel mechanism in glomerular disease and thus could represent a novel therapeutic target.

scholarly journals Safflower Yellow B Protects Brain against Cerebral Ischemia Reperfusion Injury through AMPK/NF-kB Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Shibin Du ◽  
Youliang Deng ◽  
Hongjie Yuan ◽  
Yanyan Sun
Keyword(s):  
Brain Injury ◽  
Cerebral Ischemia ◽  
Pc12 Cells ◽  
Inflammatory Cytokines ◽  
Nuclear Translocation ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Dependent Manner ◽  
Secondary Damage

Inflammation had showed its important role in the pathogenesis of cerebral ischemia and secondary damage. Safflower yellow B (SYB) had neuroprotective effects against oxidative stress-induced brain injuries, but the mechanisms were still largely unknown to us. In this study, we tried to investigate the anti-inflammation effects of SYB and the possible roles of AMPK/NF-κB signaling pathway on these protective effects. In vivo, brain ischemia/reperfusion (I/R) was induced by transient middle cerebral artery occlusion for 2 h and reperfusion for 20 h. Neurofunctional evaluation, infarction area, and brain water contents were measured. Brain injury markers and inflammatory cytokines levels were measured by ELISA kits. In vitro, cell viability, apoptosis, and LDH leakage were measured after I/R in PC12 cells. The expression and phosphorylation levels of AMPK, NF-κB p65, and P-IκB-α in cytoplasm and nuclear were measured by Western blotting. SiRNA experiment was performed to certify the role of AMPK. The results showed SYB reduced infarct size, improved neurological outcomes, and inhibited brain injury after I/R. In vitro test, SYB treatment alleviated PC12 cells injury and apoptosis and inhibited the inflammatory cytokines (IL-1, IL-6, TNF-α, and COX-2) in a dose-dependent manner. SYB treatment induced AMPK phosphorylation and inhibited NF-κB p65 nuclear translocation both in brain and in PC12 cells. Further studies also showed that the inhibition of NF-κB activity of SYB was through AMPK. In conclusion, SYB protected brain I/R injury through reducing expression of inflammatory cytokines and this effect might be partly due to the inhibition of NF-κB mediated by AMPK.

scholarly journals Notch1–STAT3–ETBR signaling axis controls reactive astrocyte proliferation after brain injury

2015 ◽  
Vol 112 (28) ◽  
pp. 8726-8731 ◽  
Author(s):  
Matthew D. LeComte ◽  
Issei S. Shimada ◽  
Casey Sherwin ◽  
Jeffrey L. Spees
Keyword(s):  
Brain Injury ◽  
Signaling Network ◽  
Reactive Astrocytes ◽  
Conditional Knockout ◽  
Reactive Astrocyte ◽  
Astrocyte Proliferation ◽  
Signaling Axis ◽  
Radial Glial ◽  
Transcription 3 ◽  
Notch1 Receptor

Defining the signaling network that controls reactive astrogliosis may provide novel treatment targets for patients with diverse CNS injuries and pathologies. We report that the radial glial cell antigen RC2 identifies the majority of proliferating glial fibrillary acidic protein-positive (GFAP+) reactive astrocytes after stroke. These cells highly expressed endothelin receptor type B (ETBR) and Jagged1, a Notch1 receptor ligand. To study signaling in adult reactive astrocytes, we developed a model based on reactive astrocyte-derived neural stem cells isolated from GFAP-CreER-Notch1 conditional knockout (cKO) mice. By loss- and gain-of-function studies and promoter activity assays, we found that Jagged1/Notch1 signaling increased ETBR expression indirectly by raising the level of phosphorylated signal transducer and activator of transcription 3 (STAT3), a previously unidentified EDNRB transcriptional activator. Similar to inducible transgenic GFAP-CreER-Notch1-cKO mice, GFAP-CreER-ETBR-cKO mice exhibited a defect in reactive astrocyte proliferation after cerebral ischemia. Our results indicate that the Notch1–STAT3–ETBR axis connects a signaling network that promotes reactive astrocyte proliferation after brain injury.

scholarly journals Aberrant Activation of Notch1 Signaling in Glomerular Endothelium Induces Albuminuria

2021 ◽  
Author(s):  
Liqun Li ◽  
Qiang Liu ◽  
Tongyao Shang ◽  
Wei Song ◽  
Dongmei Xu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transcription Factors ◽  
Notch Signaling ◽  
Molecular Mechanisms ◽  
Notch Pathway ◽  
Endothelial Glycocalyx ◽  
Notch1 Signaling ◽  
Filtration Barrier ◽  
Glomerular Endothelium

Rationale: Glomerular capillaries are lined with a highly specialized fenestrated endothelium and contribute to the glomerular filtration barrier (GFB). The Notch signaling pathway is involved in regulation of GFB, but its role in glomerular endothelium has not been investigated due to the embryonic lethality of animal models with genetic modification of Notch pathway components in the endothelium. Objective: To determine the effects of aberrant activation of the Notch signaling in glomerular endothelium and the underlying molecular mechanisms. Methods and Results: We established the ZEG-Notch1 intracellular domain (NICD1)/Tie2-tTA/Tet-O-Cre transgenic mouse model to constitutively activate Notch1 signaling in endothelial cells of adult mice. The triple transgenic mice developed severe albuminuria with significantly decreased VE-cadherin expression in the glomerular endothelium. In vitro studies showed that either NICD1 lentiviral infection or treatment with Notch ligand DLL4 markedly reduced VE-cadherin expression and increased monolayer permeability of human renal glomerular endothelial cells (HRGECs). In addition, Notch1 activation or gene knockdown of VE-cadherin reduced the glomerular endothelial glycocalyx. Further investigation demonstrated that activated Notch1 suppression of VE-cadherin was through the transcription factors SNAI1 and ERG, which bind to the -373 E-box and the -134/-118 ETS element of the VE-cadherin promoter, respectively. Conclusions: Our results reveal novel regulatory mechanisms whereby endothelial Notch1 signaling dictates the level of VE-cadherin through the transcription factors SNAI1 and ERG, leading to dysfunction of GFB and induction of albuminuria.

scholarly journals Galectin-3 modulates microglia inflammation in vitro but not neonatal brain injury in vivo under inflammatory conditions

2021 ◽  
Author(s):  
Karin Sävman ◽  
Wei Wang ◽  
Ali Hoseinpoor Rafati ◽  
Pernilla Svedin ◽  
Syam Nair ◽  
...  
Keyword(s):  
Brain Injury ◽  
Neonatal Brain ◽  
Neonatal Brain Injury ◽  
Inflammatory Conditions ◽  
Galectin 3

scholarly journals Chemical inhibition of pathological reactive astrocytes promotes neural protection

2021 ◽  
Author(s):  
Benjamin L. L. Clayton ◽  
James D. Kristell ◽  
Kevin C. Allan ◽  
Molly Karl ◽  
Eric Garrison ◽  
...  
Keyword(s):  
Neurodegenerative Diseases ◽  
Therapeutic Potential ◽  
Cell Loss ◽  
Reactive Astrocytes ◽  
Reactive Astrocyte ◽  
Chemical Inhibitors ◽  
Functional Features ◽  
Screening Platform

Disease, injury, and aging induce reactive astrocyte states with pathological functions. In neurodegenerative diseases, inflammatory reactive astrocytes are abundant and contribute to progressive cell loss. Modulating the state or function of these reactive astrocytes thereby represents an attractive therapeutic goal. Leveraging a cellular phenotypic screening platform, we show that chemical inhibitors of HDAC3 effectively block pathological astrocyte reactivity. Inhibition of HDAC3 reduces molecular and functional features of reactive astrocytes in vitro including inflammatory gene expression, cytokine secretion, and antigen presentation. Transcriptional and chromatin mapping studies show that HDAC3 inhibition mediates a switch between pro-inflammatory and anti-inflammatory states, which disarms the pathological functions of reactive astrocytes. Systemic administration of a blood-brain barrier penetrant chemical inhibitor of HDAC3, RGFP966, blocks reactive astrocyte formation and promotes axonal protection in vivo. Collectively, these results establish a platform for discovering chemical modulators of reactive astrocyte states, inform the mechanisms controlling astrocyte reactivity, and demonstrate the therapeutic potential of modulating astrocyte reactivity for neurodegenerative diseases.

scholarly journals Role of Jagged1-mediated Notch Signaling Activation in the Differentiation and Stratification of the Human Limbal Epithelium

Cells ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 1945
Author(s):  
Sheyla González ◽  
Maximilian Halabi ◽  
David Ju ◽  
Matthew Tsai ◽  
Sophie X. Deng
Keyword(s):  
Notch Signaling ◽  
Progenitor Cell ◽  
Basal Layer ◽  
Notch Signaling Pathway ◽  
Basal Cells ◽  
Proliferation Markers ◽  
Limbal Epithelium ◽  
Asymmetric Divisions ◽  
Signaling Activation

The Notch signaling pathway plays a key role in proliferation and differentiation. We investigated the effect of Jagged 1 (Jag1)-mediated Notch signaling activation in the human limbal stem/progenitor cell (LSC) population and the stratification of the limbal epithelium in vitro. After Notch signaling activation, there was a reduction in the amount of the stem/progenitor cell population, epithelial stratification, and expression of proliferation markers. There was also an increase of the corneal epithelial differentiation. In the presence of Jag1, asymmetric divisions were decreased, and the expression pattern of the polarity protein Par3, normally present at the apical-lateral membrane of basal cells, was dispersed in the cells. We propose a mechanism in which Notch activation by Jag1 decreases p63 expression at the basal layer, which in turn reduces stratification by decreasing the number of asymmetric divisions and increases differentiation.

scholarly journals TRAF3 mediates neuronal apoptosis in early brain injury following subarachnoid hemorrhage via targeting TAK1-dependent MAPKs and NF-κB pathways

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yan Zhou ◽  
Tao Tao ◽  
Guangjie Liu ◽  
Xuan Gao ◽  
Yongyue Gao ◽  
...  
Keyword(s):  
Subarachnoid Hemorrhage ◽  
Brain Injury ◽  
Therapeutic Target ◽  
Cortical Neurons ◽  
Neuronal Apoptosis ◽  
Early Brain Injury ◽  
Neurological Deficits ◽  
Human Brain Tissue

AbstractNeuronal apoptosis has an important role in early brain injury (EBI) following subarachnoid hemorrhage (SAH). TRAF3 was reported as a promising therapeutic target for stroke management, which covered several neuronal apoptosis signaling cascades. Hence, the present study is aimed to determine whether downregulation of TRAF3 could be neuroprotective in SAH-induced EBI. An in vivo SAH model in mice was established by endovascular perforation. Meanwhile, primary cultured cortical neurons of mice treated with oxygen hemoglobin were applied to mimic SAH in vitro. Our results demonstrated that TRAF3 protein expression increased and expressed in neurons both in vivo and in vitro SAH models. TRAF3 siRNA reversed neuronal loss and improved neurological deficits in SAH mice, and reduced cell death in SAH primary neurons. Mechanistically, we found that TRAF3 directly binds to TAK1 and potentiates phosphorylation and activation of TAK1, which further enhances the activation of NF-κB and MAPKs pathways to induce neuronal apoptosis. Importantly, TRAF3 expression was elevated following SAH in human brain tissue and was mainly expressed in neurons. Taken together, our study demonstrates that TRAF3 is an upstream regulator of MAPKs and NF-κB pathways in SAH-induced EBI via its interaction with and activation of TAK1. Furthermore, the TRAF3 may serve as a novel therapeutic target in SAH-induced EBI.

scholarly journals Macropinocytosis requires Gal-3 in a subset of patient-derived glioblastoma stem cells

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Laetitia Seguin ◽  
Soline Odouard ◽  
Francesca Corlazzoli ◽  
Sarah Al Haddad ◽  
Laurine Moindrot ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Survival ◽  
Small Gtpases ◽  
Molecular Signature ◽  
Carbohydrate Binding ◽  
Pancreatic Cancers ◽  
Galectin 3 ◽  
Pharmacologic Inhibition

AbstractRecently, we involved the carbohydrate-binding protein Galectin-3 (Gal-3) as a druggable target for KRAS-mutant-addicted lung and pancreatic cancers. Here, using glioblastoma patient-derived stem cells (GSCs), we identify and characterize a subset of Gal-3high glioblastoma (GBM) tumors mainly within the mesenchymal subtype that are addicted to Gal-3-mediated macropinocytosis. Using both genetic and pharmacologic inhibition of Gal-3, we showed a significant decrease of GSC macropinocytosis activity, cell survival and invasion, in vitro and in vivo. Mechanistically, we demonstrate that Gal-3 binds to RAB10, a member of the RAS superfamily of small GTPases, and β1 integrin, which are both required for macropinocytosis activity and cell survival. Finally, by defining a Gal-3/macropinocytosis molecular signature, we could predict sensitivity to this dependency pathway and provide proof-of-principle for innovative therapeutic strategies to exploit this Achilles’ heel for a significant and unique subset of GBM patients.

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