Activation Of Microglial Cells Research Articles

Protecting effects of 4-octyl itaconate on neonatal hypoxic-ischemic encephalopathy via Nrf2 pathway in astrocytes

BackgroundNeonatal hypoxic-ischemic encephalopathy (HIE) is one of the most common neurological problems occurring in the perinatal period. However, there still is not a promising approach to reduce long-term neurodevelopmental outcomes of HIE. Recently, itaconate has been found to exhibit anti-oxidative and anti-inflammatory effects. However, the therapeutic efficacy of itaconate in HIE remains inconclusive. Therefore, this study attempts to explore the pathophysiological mechanisms of oxidative stress and inflammatory responses in HIE as well as the potential therapeutic role of a derivative of itaconate, 4-octyl itaconate (4OI).MethodsWe used 7-day-old mice to induce hypoxic-ischemic (HI) model by right common carotid artery ligation followed by 1 h of hypoxia. Behavioral experiments including the Y-maze and novel object recognition test were performed on HI mice at P60 to evaluate long-term neurodevelopmental outcomes. We employed an approach combining non-targeted metabolomics with transcriptomics to screen alterations in metabolic profiles and gene expression in the hippocampal tissue of the mice at 8 h after hypoxia. Immunofluorescence staining and RT-PCR were used to evaluate the pathological changes in brain tissue cells and the expression of mRNA and proteins. 4OI was intraperitoneally injected into HI model mice to assess its anti-inflammatory and antioxidant effects. BV2 and C8D1A cells were cultured in vitro to study the effect of 4OI on the expression and nuclear translocation of Nrf2. We also used Nrf2-siRNA to further validate 4OI-induced Nrf2 pathway in astrocytes.ResultsWe found that in the acute phase of HI, there was an accumulation of pyruvate and lactate in the hippocampal tissue, accompanied by oxidative stress and pro-inflammatory, as well as increased expression of antioxidative stress and anti-inflammatory genes. Treatment of 4OI could inhibit activation and proliferation of microglial cells and astrocytes, reduce neuronal death and relieve cognitive dysfunction in HI mice. Furthermore, 4OI enhanced nuclear factor erythroid-2-related factor (Nfe2l2; Nrf2) expression and nuclear translocation in astrocytes, reduced pro-inflammatory cytokine production, and increased antioxidant enzyme expression.ConclusionOur study demonstrates that 4OI has a potential therapeutic effect on neuronal damage and cognitive deficits in HIE, potentially through the modulation of inflammation and oxidative stress pathways by Nrf2 in astrocytes.

Prenatal treatment with preimplantation factor improves early postnatal neurogenesis and cognitive impairments in a mouse model of Down syndrome

Down syndrome (DS) is a genetic disease characterized by a supernumerary chromosome 21. Intellectual deficiency (ID) is one of the most prominent features of DS. Central nervous system defects lead to learning disabilities, motor and language delays, and memory impairments. At present, a prenatal treatment for the ID in DS is lacking. Subcutaneous administration of synthetic preimplantation factor (sPIF, a peptide with a range of biological functions) in a model of severe brain damage has shown neuroprotective and anti-inflammatory properties by directly targeting neurons and microglia. Here, we evaluated the effect of PIF administration during gestation and until weaning on Dp(16)1Yey mice (a mouse model of DS). Possible effects at the juvenile stage were assessed using behavioral tests and molecular and histological analyses of the brain. To test the influence of perinatal sPIF treatment at the adult stage, hippocampus-dependent memory was evaluated on postnatal day 90. Dp(16)1Yey pups showed significant behavioral impairment, with impaired neurogenesis, microglial cell activation and a low microglial cell count, and the deregulated expression of genes linked to neuroinflammation and cell cycle regulation. Treatment with sPIF restored early postnatal hippocampal neurogenesis, with beneficial effects on astrocytes, microglia, inflammation, and cell cycle markers. Moreover, treatment with sPIF restored the level of DYRK1A, a protein that is involved in cognitive impairments in DS. In line with the beneficial effects on neurogenesis, perinatal treatment with sPIF was associated with an improvement in working memory in adult Dp(16)1Yey mice. Perinatal treatment with sPIF might be an option for mitigating cognitive impairments in people with DS.

Early hyperbaric oxygen therapy through regulating the HIF-1α signaling pathway attenuates Neuroinflammation and behavioral deficits in a mouse model of Sepsis-associated encephalopathy

BackgroundSepsis-associated encephalopathy (SAE) presents a significant clinical challenge, associated with increased mortality and healthcare expenses. Hyperbaric oxygen therapy (HBOT), involving inhaling pure or highly concentrated oxygen under pressures exceeding one atmosphere, has demonstrated neuroprotective effects in various conditions. However, the precise mechanisms underlying its protective actions against sepsis-associated brain injury remain unclear. This study aimed to determine whether HBOT protects against SAE and to elucidate the impact of the hypoxia-inducible factor-1α (HIF-1α) signaling pathway on SAE. MethodsThe experiment consisted of two parts. In the first part, C57BL/6 J male mice were divided into five groups using a random number table method: control group, sham surgery group, sepsis group, HBOT + sepsis group, and HBOT + sham surgery group. In the subsequent part, C57BL/6 J male mice were divided into four groups: sepsis group, HBOT + sepsis group, HIF-1α + HBOT + sepsis group, and HIF-1α + sepsis group. Sepsis was induced via cecal ligation and puncture (CLP). Hyperbaric oxygen therapy was administered at 1 h and 4 h post-CLP. After 24 h, blood and hippocampal tissue were collected for cytokine measurements. HIF-1α, TNF-α, IL-1β, and IL-6 expression were assessed via ELISA and western blotting. Microglial expression was determined by immunofluorescence. Blood-brain barrier permeability was quantified using Evans Blue. Barnes maze and fear conditioning were conducted 14 days post-CLP to evaluate learning and memory. ResultsOur findings reveal that CLP-induced hippocampus-dependent cognitive deficits coincided with elevated HIF-1α and increased TNF-α, IL-1β, and IL-6 levels in both blood and hippocampus. Observable activation of microglial cells in the hippocampus and increased blood-brain barrier (BBB) permeability were also evident. HBOT mitigated HIF-1α, TNF-α, IL-1β, and IL-6 levels, attenuated microglial activation in the hippocampus, and significantly improved learning and memory deficits in CLP-exposed mice. Additionally, these outcomes were corroborated by injecting a lentivirus that overexpressed HIF-1α into the hippocampal region of the mice. ConclusionHIF-1α escalation induced peripheral and central inflammatory factors, promoting microglial activation, BBB impairment, and cognitive dysfunction. However, HBOT ameliorated these effects by reducing HIF-1α levels in Sepsis-Associated Encephalopathy.

Comparative study of virus and lymphocyte distribution with clinical data suggests early high dose immunosuppression as potential key factor for the therapy of patients with BoDV-1 infection

ABSTRACT Borna disease virus 1 (BoDV-1) was just recently shown to cause predominantly fatal encephalitis in humans. Despite its rarity, bornavirus encephalitis (BVE) can be considered a model disease for encephalitic infections caused by neurotropic viruses and understanding its pathomechanism is of utmost relevance. Aim of this study was to compare the extent and distribution pattern of cerebral inflammation with the clinical course of disease, and individual therapeutic procedures. For this, autoptic brain material from seven patients with fatal BVE was included in this study. Tissue was stained immunohistochemically for pan-lymphocytic marker CD45, the nucleoprotein of BoDV-1, as well as glial marker GFAP and microglial marker Iba1. Sections were digitalized and counted for CD45-positive and BoDV-1-positive cells. For GFAP and Iba1, a semiquantitative score was determined. Furthermore, detailed information about the individual clinical course and therapy were retrieved and summarized in a standardized way. Analysis of the distribution of lymphocytes shows interindividual patterns. In contrast, when looking at the BoDV-1-positive glial cells and neurons, a massive viral involvement in the brain stem was noticeable. Three of the seven patients received early high-dose steroids, which led to a significantly lower lymphocytic infiltration of the central nervous tissue and a longer survival compared to the patients who were treated with steroids later in the course of disease. This study highlights the potential importance of early high-dose immunosuppressive therapy in BVE. Our findings hint at a promising treatment option which should be corroborated in future observational or prospective therapy studies. ABBREVIATIONS: BoDV-1: Borna disease virus 1; BVE: bornavirus encephalitis; Cb: cerebellum; CNS: central nervous system; FL: frontal lobe; GFAP: glial fibrillary acid protein; Hc: hippocampus; Iba1: ionized calcium-binding adapter molecule 1; Iba1act: general activation of microglial cells; Iba1nod: formation of microglial nodules; IL: insula; Me: mesencephalon; Mo: medulla oblongata; OL: occipital lobe; pASS: per average of 10 screenshots; patearly: patients treated with early high dose steroid shot; patlate: patients treated with late or none high dose steroid shot; Po: pons; So: stria olfactoria; Str: striatum

Macrophage-mediated controlled release of cysteine protease inhibitor from PLGA-PEG/hydroxyapatite microspheres for targeting cathepsin S in Alzheimer’s disease

The cause of Alzheimer’s disease (AD) remains unknown; however, studies have indicated the increased expression of cathepsin S (CTSS) in AD brains. Cathepsin S acts as a β-secretase, prompting the release of amyloid-β peptides and stimulating microglial cell activation, causing microglial migration, and contributing to neuroinflammation and disease progression. In this study, CT001, an inhibitor of the cysteine protease CTSS, was encapsulated in poly(lactic-co-glycolic acid)-polyethylene glycol microspheres combined with hydroxyapatite microspheres (CPPMs/HAMs) to achieve a constant and long-term drug release for AD. The results showed that CPPMs/HAMs had a uniform diameter of approximately 1 μm, enabling macrophage endocytosis and facilitating sustained CT001 release into the bloodstream, ultimately reaching the brain for AD treatment. The developed CPPMs/HAMs demonstrated excellent anti-inflammatory properties and mitigated CTSS expression in an in vitro inflammation microglial model. Intramuscular administration of CPPMs/HAMs improved learning and memory in AD mice during behavioral assessments. Furthermore, CPPMs/HAMs reduced CTSS expression, amyloid-beta accumulation, and deposition in vivo. This study demonstrated the potential of CT001 delivered via CPPMs/HAMs as a promising approach for managing AD in patients.

The combined administration of LNC-encapsulated retinoic acid and calcitriol stimulates oligodendrocyte progenitor cell differentiation in vitro and in vivo after intranasal administration

Intranasal administration is an efficient strategy for bypassing the BBB, favoring drug accumulation in the brain, and improving its efficiency. Lipid nanocapsules (LNC) are suitable nanocarriers for the delivery of lipophilic drugs via this route and can be used to encapsulate lipophilic molecules such as retinoic acid (RA) and calcitriol (Cal). As the hallmarks of multiple sclerosis (MS) are neuroinflammation and oligodendrocyte loss, our hypothesis was that by combining two molecules known for their pro-differentiating properties, encapsulated in LNC, and delivered by intranasal administration, we would stimulate oligodendrocyte progenitor cells (OPC) differentiation into oligodendrocytes and provide a new pro-remyelinating therapy. LNC loaded with RA (LNC-RA) and Cal (LNC-Cal) were stable for at least 8 weeks. The combination of RA and Cal was more efficient than the molecules alone, encapsulated or not, on OPC differentiation in vitro and decreased microglia cell activation in a dose-dependent manner. After the combined intranasal administration of LNC-RA and LNC-Cal in a mouse cuprizone model of demyelination, increased MBP staining was observed in the corpus callosum. In conclusion, intranasal delivery of lipophilic drugs encapsulated in LNC is a promising strategy for myelinating therapies.

Single-cell transcriptome reveals highly complement activated microglia cells in association with pediatric tuberculous meningitis.

Tuberculous meningitis (TBM) is a devastating form of tuberculosis (TB) causing high mortality and disability. TBM arises due to immune dysregulation, but the underlying immune mechanisms are unclear. We performed single-cell RNA sequencing on peripheral blood mononuclear cells (PBMCs) and cerebrospinal fluid (CSF) cells isolated from children (n=6) with TBM using 10 xGenomics platform. We used unsupervised clustering of cells and cluster visualization based on the gene expression profiles, and validated the protein and cytokines by ELISA analysis. We revealed for the first time 33 monocyte populations across the CSF cells and PBMCs of children with TBM. Within these populations, we saw that CD4_C04 cells with Th17 and Th1 phenotypes and Macro_C01 cells with a microglia phenotype, were enriched in the CSF. Lineage tracking analysis of monocyte populations revealed myeloid cell populations, as well as subsets of CD4 and CD8 T-cell populations with distinct effector functions. Importantly, we discovered that complement-activated microglial Macro_C01 cells are associated with a neuroinflammatory response that leads to persistent meningitis. Consistently, we saw an increase in complement protein (C1Q), inflammatory markers (CRP) and inflammatory factor (TNF-α and IL-6) in CSF cells but not blood. Finally, we inferred that Macro_C01 cells recruit CD4_C04 cells through CXCL16/CXCR6. We proposed that the microglial Macro_C01 subset activates complement and interacts with the CD4_C04 cell subset to amplify inflammatory signals, which could potentially contribute to augment inflammatory signals, resulting in hyperinflammation and an immune response elicited by Mtb-infected tissues.

Pathological high intraocular pressure induces glial cell reactive proliferation contributing to neuroinflammation of the blood-retinal barrier via the NOX2/ET-1 axis-controlled ERK1/2 pathway

BackgroundNADPH oxidase (NOX), a primary source of endothelial reactive oxygen species (ROS), is considered a key event in disrupting the integrity of the blood-retinal barrier. Abnormalities in neurovascular-coupled immune signaling herald the loss of ganglion cells in glaucoma. Persistent microglia-driven inflammation and cellular innate immune system dysregulation often lead to deteriorating retinal degeneration. However, the crosstalk between NOX and the retinal immune environment remains unresolved. Here, we investigate the interaction between oxidative stress and neuroinflammation in glaucoma by genetic defects of NOX2 or its regulation via gp91ds-tat.MethodsEx vivo cultures of retinal explants from wildtype C57BL/6J and Nox2−/− mice were subjected to normal and high hydrostatic pressure (Pressure 60 mmHg) for 24 h. In vivo, high intraocular pressure (H-IOP) was induced in C57BL/6J mice for two weeks. Both Pressure 60 mmHg retinas and H-IOP mice were treated with either gp91ds-tat (a NOX2-specific inhibitor). Proteomic analysis was performed on control, H-IOP, and treatment with gp91ds-tat retinas to identify differentially expressed proteins (DEPs). The study also evaluated various glaucoma phenotypes, including IOP, retinal ganglion cell (RGC) functionality, and optic nerve (ON) degeneration. The superoxide (O2-) levels assay, blood-retinal barrier degradation, gliosis, neuroinflammation, enzyme-linked immunosorbent assay (ELISA), western blotting, and quantitative PCR were performed in this study.ResultsWe found that NOX2-specific deletion or activity inhibition effectively attenuated retinal oxidative stress, immune dysregulation, the internal blood-retinal barrier (iBRB) injury, neurovascular unit (NVU) dysfunction, RGC loss, and ON axonal degeneration following H-IOP. Mechanistically, we unveiled for the first time that NOX2-dependent ROS-driven pro-inflammatory signaling, where NOX2/ROS induces endothelium-derived endothelin-1 (ET-1) overexpression, which activates the ERK1/2 signaling pathway and mediates the shift of microglia activation to a pro-inflammatory M1 phenotype, thereby triggering a neuroinflammatory outburst.ConclusionsCollectively, we demonstrate for the first time that NOX2 deletion or gp91ds-tat inhibition attenuates iBRB injury and NVU dysfunction to rescue glaucomatous RGC loss and ON axon degeneration, which is associated with inhibition of the ET-1/ERK1/2-transduced shift of microglial cell activation toward a pro-inflammatory M1 phenotype, highlighting NOX2 as a potential target for novel neuroprotective therapies in glaucoma management.

Galectin-3 modulates microglial activation and neuroinflammation in early brain injury after subarachnoid hemorrhage

BackgroundAneurysmal subarachnoid hemorrhage (SAH) is a devastating acute cerebrovascular event with high mortality and permanent disability rates. Higher galectin-3 levels on days 1–3 have been shown to predict the development of delayed cerebral infarction or adverse outcomes after SAH. Recent single-cell analysis of microglial transcriptomic diversity in SAH revealed that galectin could influence the development and course of neuroinflammation after SAH. MethodsThis study aimed to investigate the role and mechanism of galectin-3 in SAH and to determine whether galectin-3 inhibition prevents early brain injury by reducing microglia polarization using a mouse model of SAH and oxyhemoglobin-treated activation of mouse BV2 cells in vitro. ResultsWe found that the expression of galectin-3 began to increase 12 h after SAH and continued to increase up to 72 h. Importantly, TD139-inhibited galectin-3 expression reduced the release of inflammatory factors in microglial cells. In the experimental SAH model, TD139 treatment alleviated neuroinflammatory damage after SAH and improved defects in neurological functions. Furthermore, we demonstrated that galectin-3 inhibition affected the activation and M1 polarization of microglial cells after SAH. TD139 treatment inhibited the expression of TLR4, p-NF-κB p65, and NF-κB p65 in microglia activated by oxyhemoglobin as well as eliminated the increased expression and phosphorylation of JAK2 and STAT3. ConclusionThese findings suggest that regulating microglia polarization by galectin-3 after SAH to improve neuroinflammation may be a potential therapeutic target.

Microglia Mitigate Neuronal Activation in a Zebrafish Model of Dravet Syndrome.

It has been known for a long time that epileptic seizures provoke brain neuroinflammation involving the activation of microglial cells. However, the role of these cells in this disease context and the consequences of their inflammatory activation on subsequent neuron network activity remain poorly understood so far. To fill this gap of knowledge and gain a better understanding of the role of microglia in the pathophysiology of epilepsy, we used an established zebrafish Dravet syndrome epilepsy model based on Scn1Lab sodium channel loss-of-function, combined with live microglia and neuronal Ca2+ imaging, local field potential (LFP) recording, and genetic microglia ablation. Data showed that microglial cells in scn1Lab-deficient larvae experiencing epileptiform seizures displayed morphological and biochemical changes characteristic of M1-like pro-inflammatory activation; i.e., reduced branching, amoeboid-like morphology, and marked increase in the number of microglia expressing pro-inflammatory cytokine Il1β. More importantly, LFP recording, Ca2+ imaging, and swimming behavior analysis showed that microglia-depleted scn1Lab-KD larvae displayed an increase in epileptiform seizure-like neuron activation when compared to that seen in scn1Lab-KD individuals with microglia. These findings strongly suggest that despite microglia activation and the synthesis of pro-inflammatory cytokines, these cells provide neuroprotective activities to epileptic neuronal networks, making these cells a promising therapeutic target in epilepsy.

IL-1β, the first piece to the puzzle of sepsis-related cognitive impairment?

Sepsis is a leading cause of death resulting from an uncontrolled inflammatory response to an infectious agent. Multiple organ injuries, including brain injuries, are common in sepsis. The underlying mechanism of sepsis-associated encephalopathy (SAE), which is associated with neuroinflammation, is not yet fully understood. Recent studies suggest that the release of interleukin-1β (IL-1β) following activation of microglial cells plays a crucial role in the development of long-lasting neuroinflammation after the initial sepsis episode. This review provides a comprehensive analysis of the recent literature on the molecular signaling pathways involved in microglial cell activation and interleukin-1β release. It also explores the physiological and pathophysiological role of IL-1β in cognitive function, with a particular focus on its contribution to long-lasting neuroinflammation after sepsis. The findings from this review may assist healthcare providers in developing novel interventions against SAE.

Emodin relieves morphine-stimulated BV2 microglial activation and inflammation through the TLR4/NF-κB/NLRP3 pathway.

The objective of this study is to disclose the role of emodin, a natural anthraquinone derivative that has been proposed to suppress microglial activation and inflammation, in morphine tolerance. Here, cell counting kit-8 method assayed the viability of BV2 microglial cells treated by ascending concentrations of emodin. In emodin-pretreated BV2 microglial cells challenged with morphine with or without transfection of toll-like receptor 4 (TLR4) overexpression plasmids, transwell assay measured cell migration. Immunofluorescence staining and western blot detected the expression of microglial markers. Inflammatory levels were subjected to ELISA and western blot. BODIPY 581/591 C11 assay estimated lipid reactive oxygen species activity. Iron assay kit examined total iron content. Western blot tested the expression of ferroptosis- and TLR4/nuclear factor-kappaB (NF-κB)/NOD-like receptor 3 (NLRP3) pathway-associated proteins. Molecular docking predicted the binding affinity of emodin to TLR4. Emodin was noted to obstruct the migration, activation, inflammatory response, and ferroptosis of BV2 microglial cells induced by morphine. In addition, emodin had a high binding affinity with TLR4 and inactivated TLR4/NF-κB/NLRP3 pathway in morphine-challenged BV2 microglial cells. Upregulation of TLR4 partially countervailed the protective role of emodin against morphine-elicited BV2 microglial cell migration, activation, inflammation, and ferroptosis. Accordingly, emodin might target TLR4 and act as an inactivator of TLR4/NF-κB/NLRP3 pathway, thus inhibiting BV2 microglial activation and inflammation to mitigate morphine tolerance.

P2X7 receptor: A receptor closely linked with sepsis-associated encephalopathy.

Sepsis is defined as a dysregulated host response to infection resulting in life-threatening organ dysfunction. Sepsis-associated encephalopathy (SAE) is the main manifestation of sepsis. Inflammation, peroxidation stress injury, and apoptosis are the main factors involved in the pathogenesis of SAE. A growing body of evidence has proved that P2X7 receptor (P2X7R), a cationic channel receptor that is widely distributed in the body, plays a major role in the occurrence and development of inflammatory injury. Therefore, this review mainly describes the activation of P2X7R in sepsis, which leads to the recruitment of inflammatory cells to the cerebral vasculature, the destruction of the blood-brain barrier, the activation of microglial cells in the brain, the apoptosis of brain cells, and other damage processes. This review also illustrates the potential therapeutic value of P2X7R inhibition in SAE.

Dose-Related Side Effects of Intravitreal Injections of Humanized Anti-Vascular Endothelial Growth Factor in Rats: Glial Cell Reactivity and Retinal Ganglion Cell Loss.

In a previous study, we documented that the Intravitreal injections (IVIs) of bevacizumab in rats caused a retinal inflammatory response. We now study whether the IVI of other humanized anti-VEGF: ranibizumab and aflibercept also cause an inflammatory reaction in the rat retina and if it depends on the dose administered. Finally, we study whether this reaction affects retinal ganglion cell (RGC) survival. Albino Sprague-Dawley rats received a single IVI of 5 µL of PBS or ranibizumab or aflibercept at the concentration used in clinical practice (10 µg/µL or 40 µg/µL) or at a lower concentration (0.38 µg/µL and 1.5 µg/µL) calculated to obtain within the rat eye the same concentration as in the human eye in clinical practice. Others received a single 5 µL IVI of a polyclonal goat anti-rat VEGF (0.015 µg/µL) or of vehicle (PBS). Animals were processed 7days or 1month later. Retinal whole mounts were immunolabeled for the detection of microglial, macroglial, RGCs, and intrinsically photosensitive RGCs (ipRGCs). Fluorescence and confocal microscopy were used to examine retinal changes, and RGCs and ipRGCs were quantified automatically or semiautomatically, respectively. All the injected substances including the PBS induced detectable side effects, namely, retinal microglial cell activation and retinal astrocyte hypertrophy. However, there was a greater microglial and macroglial response when the higher concentrations of ranibizumab and aflibercept were injected than when PBS, the antibody anti-rat VEGF and the lower concentrations of ranibizumab or aflibercept were injected. The higher concentration of ranibizumab and aflibercept resulted also in significant RGC death, but did not cause appreciable ipRGC death. The IVI of all the substances had some retinal inflammatory effects. The IVI of humanized anti-VEGF to rats at high doses cause important side effects: severe inflammation and RGC death, but not ipRGC death.

Proteomics on human cerebral cavernous malformations reveals novel biomarkers in neurovascular dysfunction for the disease pathology

BackgroundCerebral cavernous malformation (CCM) is a disease associated with an elevated risk of focal neurological deficits, seizures, and hemorrhagic stroke. The disease has an inflammatory profile and improved knowledge of CCM pathology mechanisms and exploration of candidate biomarkers will enable new non-invasive treatments. MethodsWe analyzed protein signatures in human CCM tissue samples by using a highly specific and sensitive multiplexing technique, proximity extension assay. FindingsData analysis revealed CCM specific proteins involved in endothelial dysfunction/inflammation/activation, leukocyte infiltration/chemotaxis, hemostasis, extracellular matrix dysfunction, astrocyte and microglial cell activation. Biomarker expression profiles matched bleeding status, especially with higher levels of inflammatory markers and activated astrocytes in ruptured than non-ruptured samples, some of these biomarkers are secreted into blood or urine. Furthermore, analysis was also done in a spatially resolving manner by separating the lesion area from the surrounding brain tissue. Our spatial studies revealed that although appearing histologically normal, the CCM border areas were pathological when compared to control brain tissues. Moreover, the functional relevance of CD93, ICAM-1 and MMP9, markers related to endothelial cell activation and extracellular matrix was validated by a murine pre-clinical CCM model. InterpretationHere we present a novel strategy for proteomics analysis on human CCMs, offering a possibility for high-throughput protein screening acquiring data on the local environment in the brain. Our data presented here describe CCM relevant brain proteins and specifically those which are secreted can serve the need of circulating CCM biomarkers to predict cavernoma's risk of bleeding.

Single-cell RNA sequencing reveals the immune features and viral tropism in the central nervous system of mice infected with Japanese encephalitis virus

Japanese encephalitis virus (JEV) is a neurotropic pathogen that causes lethal encephalitis. The high susceptibility and massive proliferation of JEV in neurons lead to extensive neuronal damage and inflammation within the central nervous system. Despite extensive research on JEV pathogenesis, the effect of JEV on the cellular composition and viral tropism towards distinct neuronal subtypes in the brain is still not well comprehended. To address these issues, we performed single-cell RNA sequencing (scRNA-seq) on cells isolated from the JEV-highly infected regions of mouse brain. We obtained 88,000 single cells and identified 34 clusters representing 10 major cell types. The scRNA-seq results revealed an increasing amount of activated microglia cells and infiltrating immune cells, including monocytes & macrophages, T cells, and natural killer cells, which were associated with the severity of symptoms. Additionally, we observed enhanced communication between individual cells and significant ligand-receptor pairs related to tight junctions, chemokines and antigen-presenting molecules upon JEV infection, suggesting an upregulation of endothelial permeability, inflammation and antiviral response. Moreover, we identified that Baiap2-positive neurons were highly susceptible to JEV. Our findings provide valuable clues for understanding the mechanism of JEV induced neuro-damage and inflammation as well as developing therapies for Japanese encephalitis.

Imidazo[1,2‐α] azine's esters increase selectivity and anti‐inflammatory activity in microglial cells

AbstractChronic neuroinflammation is crucial in many neurodegenerative diseases, including AD. Neuroinflammation activates microglia, the CNS's central effector cells, and immune cells. Toxins and neuroinflammatory reactions can kill or damage neurons, activating microglial cells. Inflammatory mediators from activated microglia can kill neurons like cytokines, chemokines, reactive oxygen species, and reactive nitrogen species. Imidazo[1,2‐α]azines with different acid groups in position two of the heterocycle system were synthesized and tested as anti‐inflammatory and cell immunomodulators. We measured the ability of human neuroblastoma (SHSY‐5Y) and microglial (HMC3) cell lines to decrease PGE2 production under pro‐inflammatory conditions. All derivatives reduced PGE2 production. Inhibitory profiles indicate that compounds 8a and 8b are more effective and potent than 9a and 9b, indicating a steric effect of carboxyphenyl on imidazo[1,2‐α]azines, reducing their potency. However, imidazo[1,2α]pyrimidines (8b and 9b) with nitrogen at position 8 were less effective than those observed by other groups (8a, 9a, and 10a). imidazo[1,2‐α]pyrimidines are more polar due to nitrogen at position 8. The synthetically derivative imidazo[1,2‐α]pyridines decreased PGE2 formation in vitro on neural cells, microglia (HMC3), and neuroblastoma (SH‐SY5Y) cells, proving that they are more effective than imidazo[1,2‐α]pyrimidines. Two product esters selectively reduced HMC3 microglial cell PGE2 production. This finding may help develop neurodegenerative disease anti‐inflammatories.

Decreased hippocampal microglial cell activation by methanolic extract from the leaves of Mallotus oppositifolius (Geiseler) Müll. Arg contributes to its antidepressant-like effect

Background Natural remedies with neuroprotective effect are useful in neuroinflammation-associated depression. Although Mallotus oppositifolius extract (MOE) has previously demonstrated antidepressant and anti-inflammatory properties, its neuroprotective effect remains unknown. Thus, the study evaluated the effect of MOE on lipopolysaccharide (LPS)-induced neuroinflammation-associated depression in mice. Methods Antidepressant-like effect of MOE (10 – 100 mg/kg), fluoxetine (20 mg/kg) and minocycline (50 mg/kg) was established in naïve Institute of Cancer Research (ICR) mice using the forced swim (FST), tail suspension (TST) and open-space swim (OSST) tests. In a separate experiment, FST and TST were used to assess the effect of an 11-day pre-treatment with MOE (10 – 100 mg/kg) or minocycline (50 mg/kg) on LPS (1 mg/kg) neuroinflammation at 6 and 24 hours post LPS. Following these tests, mice were sacrificed and their hippocampi isolated to evaluate their resting and activated microglial cells using Golgi-Cox staining technique. Open-field test was used to assess locomotor activity. Results MOE, fluoxetine and minocycline significantly reduced immobility in FST, TST and OSST compared to vehicle (p < 0.05), confirming their antidepressant-like effect. Interestingly, MOE’s antidepressant-like effect was faster than fluoxetine and minocycline. Conversely, LPS treatment increased immobility behavior at 6 and 24 hours, suggestive of neuroinflammation-induced depression. Compared to vehicle group, pre-treatment with MOE and minocycline ameliorated LPS-induced hippocampal microglial activation and reversed increased immobility behavior without affecting locomotor activity (p < 0.05). Resting microglial cell count was significantly increased by MOE pre-treatment in the OSST-challenged mice compared to vehicle group (p < 0.01). Similarly, MOE pre-treatment reversed LPS-induced reduction in resting microglial count, and restored resting microglial count to normal levels compared to LPS naive vehicle group. Conclusions Collectively, the results suggest that MOE exerts neuroprotective effect against LPS-induced neuroinflammation by decreasing the activation of microglia and increasing resting microglial count. This contributes to its antidepressant-like effect.

The effects of urolithin A on poly I:C-induced microglial activation.

Neuroinflammation can be triggered by various stimuli, including viral infections. Viruses can directly invade the brain and infect neuronal cells or indirectly trigger a "cytokine storm" in the periphery that eventually leads to microglial activation in the brain. While this initial activation of microglial cells is important for viral clearance, chronic activation leads to excessive inflammation and oxidative stress, which can be neurotoxic. Remarkebly, recent studies have shown that certain viruses such as influenza A virus, coronavirus, herpes virus and Epstein-Barr virus may be involved in the development of neurodegenerative diseases such as Parkinson's disease, Alzheimer's disease, and multiple sclerosis. Therefore, it is important to find therapeutic strategies against chronic neuroinflammation triggered by viral infections. Here, we investigated the effects of urolithin A (UA) on microglial activation in vitro induced by a viral mimetic, poly I:C, in a triple co-culture system of neurons, astrocytes and microglial cells. Immunocytochemistry was used to perform a comprehensive single-cell analysis of the morphological changes of microglia as an indicator of their reactive state. Treatment with UA significantly prevented the poly I:C-induced reactive state of microglia, which was characterized by increased expression of the microglial activation markers CD68 and IBA-1. UA restored the poly I:C-induced morphology by restoring microglial ramification. In addition, UA was able to reduce the release of the pro-inflammatory mediators CCL2, TNF-α, and IL-1β and showed a trend toward attenuation of cellular ROS production in poly I:C-treated cultures. Overall, this study suggests that UA as a component of a healthy diet may help prevent virus-induced neuroinflammation and may have therapeutic potential for future studies to prevent or treat neurodegenerative diseases by targeting the associated neuroinflammatory processes.

Research of brain targeting nano-metformin on cerebral ischemia-reperfusion injury based on microglial cell activation

The polarization of microglia plays a crucial role in cerebral ischemia-reperfusion injury (CI/RI) and has garnered significant attention from researchers. M1-polarized microglia activation drives harmful inflammatory responses in neuronal cells. Although metformin can inhibit inflammatory reactions and improve CI/RI, it has significant toxic side effects. In this study, we designed a novel nanodrug delivery system, apoferritin nanocages encapsulating metformin (APO@Metformin), to address these issues. On the one hand, apoferritin nanocages are naturally available nanoparticles and widely present in biological systems, which possesses good biocompatibility and safety and reduces the toxic side effects of metformin. On the other hand, the small particle size of apoferritin nanocages enable them to have specific blood-brain barrier penetration to achieve brain targeting delivery. Animal experiments demonstrated that APO@Metformin improved CI/RI and reduced neuronal damage caused by inflammatory reactions. Furthermore, in vitro cell experiments showed that APO@Metformin not only could closely interact with transferrin receptors 1 (TfR1) expression in microglia to achieve superior brain targeting, but also inhibit the activity of M1 polarization and NLRP3 in microglia, thereby suppressing inflammatory responses in CI/RI and protecting neuronal cells.