Microglia Research Articles

Microglial galectin-3 increases with aging in the mouse hippocampus.

Microglial activation during aging is associated with neuroinflammation and cognitive impairment. Galectin-3 plays a crucial role in microglial activation and phagocytosis. However, the role of galectin-3 in the aged brain is not completely understood. In the present study, we investigated aging-related mechanisms and microglial galectin-3 expression in the mouse hippocampus using female 6-, 12-, and 24-month-old C57BL/6 mice. Western blot analysis revealed neurodegeneration, blood-brain barrier leakage, and increased levels of neuroinflammation-related proteins in 24-month-old mice compared to 6- and 12-month-old mice. Immunohistochemistry revealed an increase in activated microglia in the hippocampus of 24-month-old mice compared to 6- and 12-month-old mice. Furthermore, we found more galectin-3 and triggering receptor expressed on myeloid cells-2-positive microglia in 24-month-old mice compared to 6- and 12-month-old mice. Using primary mouse microglial cells, galectin -3 was also increased by lipopolysaccharide treatment. These findings suggest that galectin-3 may play an important role in microglial activation and neuroinflammation during brain aging.

Dark microglia: a therapeutic target to prevent synaptic loss and neurodegeneration?

Dark microglia is a distinct state of microglia progressively increasing with aging and present in high numbers in neurodegenerative diseases. Dark microglia are characterised by various ultrastructural markers of cellular stress, including a condensed, electron-dense cytoplasm and nucleoplasm, giving them a dark appearance in electron microscopy. Expression of classical homeostatic microglia markers such as like IBA1, P2RY12 and TMEM119 is significantly down-regulated in dark microglia. Dark microglial cells are also hyper-ramified and make extensive interactions, phagocytic and non-phagocytic, with synapses, myelinated axons, amyloid plaques and the vasculature, suggesting a role in brain remodelling in aging and disease. Emergence of dark microglia is linked to the activation of integrated stress response. It is suggested that metabolic interventions such as ketogenic diets and supplements could be effective at targeting and normalizing dark microglia, with positive outcomes on brain resilience and healthy cognitive aging. Keywords: microglia; synaptic loss; neurodegeneration; Alzheimer’s disease; integrated stress response; metabolism; magnetic resonance imaging

Long-Term Exposure to Tire-Derived 6-PPD Quinone Causes Neurotoxicity and Neuroinflammation via Inhibition of HTR2A in C57BL/6 Mice.

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine quinone (6-PPDQ), a novel contaminant derived from tire wear, has raised concerns due to its potential neurotoxicity, yet its long-term effects on mammalian neurological health remain poorly understood. This study investigates the neurotoxic and neuroinflammatory impacts of prolonged 6-PPDQ exposure using male C57BL/6 mice. Behavioral assessments revealed significant cognitive deficits, while biochemical analyses demonstrated increased levels of reactive oxygen species, apoptosis, and blood-brain barrier (BBB) disruption. Elevated pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) and activation of microglial cells were observed, indicating a robust neuroinflammatory response. Network pharmacology and molecular docking identified serotonin receptor HTR2A as a key target through which 6-PPDQ mediates its toxic effects. Activation of HTR2A by the agonist DOI (2,5-dimethoxy-4-iodoamphetamine) mitigated these effects, suggesting a potential therapeutic strategy. These findings provide the first evidence of 6-PPDQ-induced neurotoxicity in mammals, underscoring the need for preventive measures to protect neurological health.

Neurosteroids and neurological disorders.

Neurosteroids play an important role as endogenous neuromodulators that are locally produced in the central nervous system and rapidly change the excitability of neurons and the activation of microglial cells and astrocytes. Here we review the mechanisms of synthesis, metabolism, and actions of neurosteroids in the central nervous system. Neurosteroids are able to play a variety of roles in the central nervous system under physiological conditions by binding to membrane ion channels and receptors such as gamma-aminobutyric acid type A receptors, Nmethyl- D-aspartate receptors, L- and T-type calcium channels, and sigma-1 receptors. In addition, numerous neurological disorders, including persistent neuropathic pain, multiple sclerosis, and seizures, have altered the levels of neurosteroids in the central nervous system. Thus, we review how local synthesis and metabolism of neurosteroids are modulated in the central nervous system and describe the role of neurosteroids under pathological conditions. Furthermore, we discuss whether neurosteroids may play a role as a new therapeutic for the treatment of neurological disorders.

Nobiletin and Eriodictyol Suppress Release of IL-1β, CXCL8, IL-6, and MMP-9 from LPS, SARS-CoV-2 Spike Protein, and Ochratoxin A-Stimulated Human Microglia.

Neuroinflammation is involved in various neurological and neurodegenerative disorders in which the activation of microglia is one of the key factors. In this study, we examined the anti-inflammatory effects of the flavonoids nobiletin (5,6,7,8,3',4'-hexamethoxyflavone) and eriodictyol (3',4',5,7-tetraxydroxyflavanone) on human microglia cell line activation stimulated by either lipopolysaccharide (LPS), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) full-length Spike protein (FL-Spike), or the mycotoxin ochratoxin A (OTA). Human microglia were preincubated with the flavonoids (10, 50, and 100 µM) for 2 h, following which, they were stimulated for 24 h. The inflammatory mediators interleukin-1 beta (IL-1β), chemokine (C-X-C motif) ligand 8 (CXCL8), IL-6, and matrix metalloproteinase-9 (MMP-9) were quantified in the cell culture supernatant by enzyme-linked immunosorbent assay (ELISA). Both nobiletin and eriodictyol significantly inhibited the LPS, FL-Spike, and OTA-stimulated release of IL-1β, CXCL8, IL-6, and MMP-9 at 50 and 100 µM, while, in most cases, nobiletin was also effective at 10 µM, with the most pronounced reductions at 100 µM. These findings suggest that both nobiletin and eriodictyol are potent inhibitors of the pathogen-stimulated microglial release of inflammatory mediators, highlighting their potential for therapeutic application in neuroinflammatory diseases, such as long COVID.

Netrin-1's Protective Effect on A-Burdened BV2 Microglial Cells Enhanced Its Viability of Peptide Drugs for Alzheimers Disease Treatment

The rising prevalence of Alzheimers Disease (AD) has stimulated extensive research into novel therapeutic strategies. Current studies suggest peptide-based drugs are emerging as promising candidates targeting various aspects of AD pathophysiology. Microglia play a crucial role in AD due to their functions as the resident immune cells in the central nervous system, and are a prominent target for the development of peptide drugs. Netrin-1, a brain-derived extracellular factor, demonstrated therapeutic potential in AD by modulating microglial function. Here, we investigated the effects of netrin-1 on amyloid-beta (A)-burden microglia using BV2 microglial cells as a model. Cell counting was performed to assess the cell viability under different concentrations of netrin-1. Immunofluorescence was employed to examine morphological changes associated with microglial activation. To examine the polarization towards M1 or M2 phenotypes, fluorescence detection was performed after amplification. ELISA quantified A concentration to assess the protective effect of netrin-1. Microglial gene expression influenced by A was identified in the Gene Expression Omnibus (GEO) database (GSE227221). Expression levels of netrin-1 receptors were analyzed to identify potential targets. Protein structures from the RCSB database were docked with netrin-1 using PyMOL to identify the amino acids on the binding interface that may stimulate the receptors and mitigate A-induced damage. Specifically, a concentration of 100 ng/mL of netrin-1 optimally maintained cell viability in the presence of A. Netrin-1 could lessen A-induced microglial activation, especially M2 activation phenotype. ELISA data showed that netrin-1 treatment significantly improved the phagocytic abilities of A-burden BV2 cells. Analysis of the GEO database revealed that the expressions of DCC, DSCAM, and UNC5a were significantly elevated following A treatment in microglia. Eight potential polypeptides were identified as the potential targets for netrin-1 binding. In conclusion, netrin-1 mitigated the A-induced damage in microglia through its protective effects. The eight identified peptides of netrin-1 may represent a significant advancement toward developing drugs targeting A-related AD pathology. Netrin-1 based peptide drugs hold novel therapeutic promise for AD

Ghrelin Induces Ferroptosis Resistance and M2 Polarization of Microglia to Alleviate Neuroinflammation and Cognitive Impairment in Alzheimer's Disease.

Microglial polarization and ferroptosis are important pathological features in Alzheimer's disease (AD). Ghrelin, a brain-gut hormone, has potential neuroprotective effects in AD. This study aimed to explore the potential mechanisms by which ghrelin regulates the progression of AD, as well as the crosstalk between microglial polarization and ferroptosis.Mouse BV2 microglial cells and male mice were treated with beta-amyloid (Aβ) (1-42) to simulate the AD environment. Microglia ferroptosis was measured by detecting levels of ferroptosis-related proteins (SLC7A11, GPX4, FTL1, and FTH1), metabolic markers (ROS, MDA, GSH, SOD), and observing mitochondrial morphological changes. Microglial polarization was evaluated by measuring levels of inflammatory markers and surface markers. The impact of ghrelin on Aβ1-42-exposed microglia was assessed by coupling with the ferroptosis activator Erastin. Cognitive impairment in AD mice was evaluated through behavioral tests. Tissue staining was applied to determine neuronal damage.In Aβ1-42-exposed microglia, ghrelin upregulated the protein expression of SLC7A11, GPX4, FTL1 and FTH1, reduced ROS and MDA levels, and elevated GSH and SOD levels through the BMP6/SMAD1 pathway. Ghrelin alleviated mitochondrial structural damage. Additionally, ghrelin reduced levels of pro-inflammatory factors and CD86, while increasing levels of anti-inflammatory factors and CD206. Erastin reversed the effects of ghrelin on ferroptosis and phenotypic polarization in Aβ1-42-exposed microglia. In AD mice, ghrelin ameliorated abnormal behavior, neuroinflammation, and plaque deposition. Ghrelin attenuated iNOS/IBA1-positive expression and enhanced Arg-1/IBA1-positive expression in the hippocampus. Ghrelin induces microglial M2 polarization by inhibiting microglia ferroptosis, thereby alleviating neuroinflammation. Our results indicate that ghrelin may serve as a promising potential agent for treating cognitive impairment in AD.

SMOC2 promotes microglia activity and neuroinflammation in Alzheimer's disease.

Alzheimer's disease (AD), the leading cause of dementia, is characterized by cognitive decline and the accumulation of amyloid-β (Aβ). It affects millions, with numbers expected to double by 2050. SMOC2, implicated in inflammation and fibrosis, may play a role in AD pathogenesis, particularly in microglial cell function, offering a potential therapeutic target. Alzheimer's disease (AD) leads to neurodegeneration, affecting cognition, language, and personality, underscoring the urgency for effective treatments. Our study investigates the role of secreted modular calcium-binding protein 2 (SMOC2) in microglial cells and its impact on AD pathology. We introduced SMOC2 overexpression and interference vectors into microglial cells treated with Aβ. Activity and phagocytosis were assessed using CCK8 and flow cytometry. SMOC2 mRNA levels were quantified by qPCR, and protein levels of SMOC2, TGF-β1, p-NF-κB/NF-κB were analyzed by western blot. Aβ content was determined by ELISA, and immunofluorescence detected TNF-α, IL-1β, CD163, and CD206. Aβ treatment inhibited microglial activity and phagocytosis, but SMOC2 disruption enhanced these functions (p < 0.05). SMOC2 overexpression increased its expression and Aβ levels, while interference reduced them (p < 0.001). SMOC2 overexpression also decreased TGF-β1, CD163, and CD206, and increased p-NF-κB/NF-κB, TNF-α, and IL-1β (p < 0.05). SMOC2 plays a crucial role in microglial cell activity, phagocytosis, and polarization, potentially through the TGF-β1/NF-κB pathway, offering insights into AD pathogenesis.

Human iPSC-derived microglial cells protect neurons from neurodegeneration in long-term cultured adhesion brain organoids

Brain organoid models have greatly facilitated our understanding of human brain development and disease. However, key brain cell types, such as microglia, are lacking in most brain organoid models. Because microglia have been shown to play important roles in brain development and pathologies, attempts have been made to add microglia to brain organoids through co-culture. However, only short-term microglia-organoid co-cultures can be established, and it remains challenging to have long-lasting survival of microglia in organoids to mimic long-term residency of microglia in the brain. In this study, we developed an adhesion brain organoid (ABO) platform that allows prolonged culture of brain organoids (greater than a year). Moreover, the long-term (LT)-ABO system contains abundant astrocytes and can support prolonged survival and ramification of microglia. Furthermore, we showed that microglia in the LT-ABO could protect neurons from neurodegeneration by increasing synaptic density and reducing p-Tau level and cell death in the LT-ABO. Therefore, the microglia-containing LT-ABO platform generated in this study provides a promising human cellular model for studying neuron-glia and glia-glia interactions in brain development and the pathogenesis of neurodegenerative diseases such as Alzheimer’s disease.

Anti-neuroinflammatory and Neuroprotective Effects of T-006 on Alzheimer's Disease Models by Modulating TLR4-Mediated MyD88/ NF-κB Signaling.

Neuroinflammation derived from the activation of the microglia is considered a vital pathogenic factor of Alzheimer's Disease (AD). T-006, a tetramethylpyrazine derivative, has been found to alleviate cognitive deficits via inhibiting tau expression and phosphorylation in AD transgenic mouse models. Recently, T-006 has been proven to dramatically decrease the levels of total Amyloid β (Aβ) peptide and Glial Fibrillary Acidic Protein (GFAP) and suppress the expression of ionized calcium binding adaptor molecule-1 (Iba-1) in APP/PS1 mice. Therefore, we have further investigated the effects of T-006 on neuroinflammation in AD-like pathology. The anti-inflammatory effects of T-006 and its underlying mechanisms were evaluated in Lipopolysaccharide (LPS)-induced AD rats. The potential protective effects against LPS-activated microglia-mediated neurotoxicity were also measured. T-006 significantly improved the cognitive impairment in LPS-induced AD rats by inhibiting the microglia/astrocyte activation. Further cellular assays found that T-006 significantly reserved the anomalous elevation of inflammatory cytokines in LPS-induced BV2 microglial cells in a concentration-dependent manner, while T-006 treatment alone showed no effects on the normal cultured cells. T-006 also reduced the levels of Toll-like Receptor 4 (TLR4)/Myeloid Differentiation protein-88 (MyD88)/NF-κB signaling-related proteins in BV2 cells exposed to LPS stimulation. TAK242, which selectively inhibits TLR4, slightly lessened the effects of T-006 in LPS-treatment BV2 cells without significance. Importantly, T-006 protected neurons against LPS-induced neuroinflammation by inhibiting the Reactive Oxygen Species (ROS) production and maintaining mitochondrial function. T-006 inhibited TLR4-mediated MyD88/NF-κB signaling pathways to suppress neuroinflammation in the LPS-induced AD rat model.

Carotid bodies mediate glial cell activation and neuroinflammation in the NTS following long-term intermittent hypoxia: role in cardiorespiratory dysfunction.

Chronic intermittent hypoxia (CIH), the main feature of obstructive sleep apnea, heightened chemosensory discharges of the carotid body (CB), which contributes to potentiate the ventilatory hypoxic response and elicits hypertension. We aimed to determine: 1) whether the persistence of cardiorespiratory alterations found in long-term CIH depend on the inputs from the CB and, 2) in what extension the activation of glial cells and neuroinflammation in the caudal region of the nucleus of the Solitary Tract (NTS) requires functional CB chemosensory activity. To evaluate these hypotheses, we exposed male mice to CIH for 60 days. At 50 days of CIH, CBs were denervated and animals were kept in CIH for 10 additional days. At the end of the experiments, we measured arterial blood pressure, breathing regularity, and hypoxic ventilatory response (HVR) and assessed astrocyte and microglia cell activation. Compared to Sham, CIH induces hypertension (MABP: 83.47±1.39 vs. 95.00±2.18 mmHg), disordered breathing (IS: 7.77±0.49 vs. 12.56±1.66), increased the HVR (1.69±0.17 vs. 4.31±0.87 ΔVE/min), and produced an early transient activation of astrocytes followed by a late and persistent activation of microglia in the NTS. In addition, CIH increased IL-1β, IL-6, and TNF-α levels in the NTS. Bilateral CB denervation after 50 days of CIH results in the restoration of normal glial cell activation in the NTS, lower levels of IL-6 and TNF-α, reductions in arterial blood pressure (83.47±1.38 mmHg) and HVR (1.63±0.43 ΔVE/min). Present results suggest that CB inputs to the NTS during long-term CIH contributes to maintain the cardiorespiratory alterations and the formation of a neuroinflammatory niche at the NTS by modifying glial cell activity.

A Network Pharmacology Study and Experimental Validation to Identify the Potential Mechanism of Heparan Sulfate on Alzheimer's Disease-Related Neuroinflammation.

Heparan sulfate (HS) is a polysaccharide that is found on the surface of cells and has various biological functions in the body. The purpose of this study was to predict the pharmacological effects and molecular mechanisms of HS on Alzheimer's disease (AD) and neuroinflammation (NI) through a network pharmacology analysis and to experimentally verify them. We performed functional enrichment analysis of common genes between HS target genes and AD-NI gene sets and obtained items such as the "Cytokine-Mediated Signaling Pathway", "Positive Regulation Of MAPK Cascade", and "MAPK signaling pathway". To confirm the predicted results, the anti-inflammatory effect of HS was investigated using lipopolysaccharide (LPS)-stimulated BV2 microglia cells. HS inhibited the production of nittic oxide, interleukin (IL)-6, and tumor necrosis factor-α in LPS-stimulated BV2 cells, but not IL-1β. In addition, HS inactivated P38 in the MAPK signaling pathway. These findings suggest the potential for HS to become a new treatment for AD and NI.

Protein kinase R induced by type I interferons is a main regulator of reactive microglia in Zika virus infection.

Microglial cells are the phagocytic cells of the brain that under physiological conditions participate in brain homeostasis and surveillance. Under pathogenic states, microglia undergoes strong morphological and transcriptional changes potentially leading to sustained neuroinflammation, brain damage, and cognitive disorders. Postnatal and adult Zika virus (ZIKV) brain infection is characterized by the induction of reactive microglia associated with brain inflammation, synapse loss and neuropathogenesis. Contrary to neurons, microglial cells are not infected by ZIKV thus raising the question of the mechanism governing ZIKV-induced microglia's reactivity. In this work, we have questioned the role of exogenous, neuronal type I interferons (IFNs-I) in regulating ZIKV-induced microglia's reactivity. Primary cultured microglial cells were either treated with conditioned media from ZIKV-infected mature neurons or co-cultured with ZIKV-infected neurons. Using either an antibody directed against the IFNAR receptor that neutralizes the IFNs-I response or Ifnar-/-microglial cells, we demonstrate that IFNs-I produced by ZIKV-infected neurons are the main regulators of the phagocytic capacity and the pro-inflammatory gene expression profile of reactive, non-infected microglial cells. We identify protein kinase R (PKR), whose expression is activated by IFNs-I, as a major regulator of the phagocytic capacity, pro-inflammatory response, and morphological changes of microglia induced by IFNs-I while up-regulating STAT1 phosphorylation and IRF1 expression. Results obtained herein in vitro with primary cultured cells and in vivo in ZIKV-infected adult immunocompetent mice, unravel a role for IFNs-I and PKR in directly regulating microglia's reactivity that could be at work in other infectious and non-infectious brain pathologies.

The unanticipated contribution of Zap70 in retinal degeneration: Implications for microglial inflammatory activation.

Inflammation is a major mechanism of photoreceptor cell death in the retina during macular degeneration leading to the blindness. In this study, we investigated the role of the kinase molecule Zap70, which is an inflammatory regulator of the systemic immune system, to elucidate the control mechanism of inflammation in the retina. We observed activated microglial cells migrated and populated the retinal layer following blue LED-induced photoreceptor degeneration and activated microglial cells in the LED-injured retina expressed Zap70, unlike the inactive microglial cells in the normal retina. Visual function was considerably decreased in blue-LED light-exposed mice, and animals with Zap70 mutations were adversely affected. Furthermore, extensive photoreceptor cell death was observed in the SKG mice, bearing a Zap70 mutation that induces autoimmune disease. In the blue-LED light-exposed groups, SKG retinas had significantly higher levels of inflammatory cytokines than those in wild-type mice. Furthermore, regulating Zap70 activity has a significant influence on microglial inflammatory state. We discovered that active microglial cells expressing Zap70 could modify vascular endothelial growth factor A (Vegfa) signaling in primary retinal pigment epithelial (RPE) cells. Our novel study revealed that the production of Zap70 by retinal microglial cells is responsible for inflammatory signals that promote apoptosis in photoreceptor cells. Furthermore, Zap70-positive microglial cells were capable of regulating Vegfa signaling in RPE cells, which matches the hallmark of macular degeneration. Overall, we discovered Zap70's inflammatory activity in the retina, which is necessary for upregulating multiple inflammatory cytokines and cell death. Zap70 represents a novel therapeutic target for treating retinal degeneration.

Single-nucleus transcriptome profiling provides insights into the pathophysiology of adhesive arachnoiditis.

Adhesive arachnoiditis (AA) is a rare form of chronic degenerative pathology associated with persistent inflammation in the arachnoid matter of the spinal cord. Despite the existing knowledge, the detailed pathological mechanisms underlying AA are not fully understood. This study aimed to elucidate through comprehensive single nuclei RNA sequencing (snRNA-seq) to delineate the transcriptomic landscape of AA. From six arachnoid membrane samples, a total of 52,886 cells met the quality control standards for analysis. The main cell populations identified with specific gene markers were as follows: fibroblasts, glial cells, microglial cells, endothelial cells, mural cells, plasma cells, and T cells. Downstream analysis of fibroblasts, glial cells, and microglial cells was performed. Notably, fibroblast subsets 1 and 3 demonstrated a strong association with AA. Among them, subcluster 3 demonstrated elevated expression of genes COL1A1, COL3A1, and FN1, indicative of enhanced Wnt/β-catenin and extracellular matrix (ECM) synthesis pathways. Subcluster 3 was predicted to progressively transform into subcluster 1. In subcluster 1, there was a significant upregulation of genes such as BMP and ALPL, signaling enhanced activation of calcification-related pathways. This was highly relevant to end-stage arachnoid ossification formation. After being activated, microglial cells transformed into inflammatory disease-associated microglial cells and continued to express high levels of chemokines CCL2, CCL4, IL-1β, and other inflammatory factors NAMPT, INPP5D and NLRP3. This might be the main reason why AA recurrence is frequently observed in patients. These insights enhance our understanding of the pathological progression of AA and may contribute to the identification of novel therapeutic targets.

Regional brain iron correlates with transcriptional and cellular signatures in Alzheimer's disease.

The link between overload brain iron and transcriptional/cellular signatures in Alzheimer's disease (AD) remains inconclusive. Iron deposition in 41 cortical and subcortical regions of 30 AD patients and 26 healthy controls (HCs) was measured using quantitative susceptibility mapping (QSM). The expression of 15,633 genes was estimated in the same regions using transcriptomic data from the Allen Human Brain Atlas (AHBA). Partial least square (PLS) regression was used to identify the association between the healthy brain gene transcription and aberrant regional QSM signal in AD. The biological processes and cell types associated with the linked genes were evaluated. Gene ontological analyses showed that the first PLS component (PLS1) genes were enriched for biological processes relating to the "protein phosphorylation" and "metal ion transport". Additionally, these genes were expressed in microglia (MG) and glutamatergic neurons (GLUs). Our findings provide mechanistic insights from transcriptional and cellular signatures into regional iron accumulation measured by QSM in AD. Spatial patterns of iron deposition changes in AD correlate with cortical spatial expression genes in healthy subjects. The identified gene transcription profile underlies aberrant iron accumulation in AD was enriched for biological processes relating to "protein phosphorylation" and "metal ion transport". The related genes were predominantly expressed in MG and GLUs.

Multimodal and serial MRI monitors brain peri-hematomal injury and repair mechanisms after experimental intracerebral hemorrhage

The peri-hematomal area (PHA) emerges as a key but puzzling interface where edematous and neuroinflammatory events co-occur after intracerebral hemorrhage (ICH), while being considered either as deleterious or protective. We aimed at unraveling the pathogeny and natural history of PHA over time after experimental ICH. Male and female rats were longitudinally followed up to day 7 using multimodal brain MRI. MRI measures were compared to neuropathological and behavioural results. While the peak of PHA volume at day 3 was predictive for spontaneous locomotor deficit without sex-effect, its drop at day 7 fitted with locomotor recovery and hematoma resorption. The PHA highest water density was observed at onset despite microvascular hypoperfusion, taken over by blood-brain barrier (BBB) leakage at day 3. Water density dropped at day 7, when vascular integrity was normalized, and the highest number of reactive astrocytes, microglial cells, and siderophages found. This study shows that the PHA with edematous component is hematoma-driven at onset and BBB-driven at day 3, but this excess neuroinflammation enabled PHA volume reduction and significant hematoma resorption as soon as day 7. Therapeutic interventions should consider this pathogeny, and be monitored by multimodal MRI in preclinical ICH models.

Acrylamide Exposure Promotes the Progression of Depression-like Behavior in Mice with CUMS via GSDMD-Mediated Pyroptosis

AIMWe investigated the mechanism by which the environmental toxin acrylamide (AM) promotes depression. MethodsA depression mouse model was constructed using the chronic unpredictable mild stress method, AM was administered orally to simulate the exposure state. Depressive-like behavioral changes were assessed by open field test, elevated plus maze test, swimming test, and sucrose preference test. Enzyme-linked immunosorbent assay (ELISA) was used to detect tissue inflammatory factor levels, hematoxylin and eosin (H&E) and Nissl staining to detect neuronal damage, immunohistochemical staining to detect IBA-1 expression, and Western-blotting to detect protein levels. GSDMD knockout (KO) mice and the GSDMD inhibitor LDC7559 were used to inhibit GSDMD. In vitro, primary microglia were used, and AM intervention was applied to detect the levels of cellular inflammatory factors, and fluorescence staining was used to detect GSDMD-NT, and propidium iodide (PI) was used to detect the level of pyroptosis. ResultsAM can exacerbate CUMS-like depression in mice, increase the levels of inflammatory factors in brain tissue, and worsen neuronal damage, with upregulation of IBA-1 expression, and can increase the expression of NLRP3, GSDMD, and GSDMD-NT. When GSDMD-KO or LDC7559 intervention was applied, it could antagonize the effects of AM and improve CUMS-like depression. In microglial cell experiments, AM could promote pyroptosis in microglial cells, increase the expression of inflammatory factors, and when GSDMD-KO was applied, it could inhibit the effects of AM. ConclusionAM can promote the progression of depression in CUMS-like mice via GSDMD-mediated pyroptosis, while also increasing tissue inflammatory levels. GSDMD is an important target for the neurotoxicity of AM.

Inhibition of Kv1.1 channels ameliorates Cu(II)-induced microglial activation and cognitive impairment in mice.

Microglia-mediated neuroinflammation plays a critical role in neuronal damage in neurodegenerative disorders such as Alzheimer's disease. Evidence shows that voltage-gated potassium (Kv) channels regulate microglial activation. We previously reported that copper dyshomeostasis causes neuronal injury via activating microglia. This study was designed to explore the role of Kv1.1 channels in copper-evoked microglial neuroinflammation. BV-2 microglial cells were treated with Cu(II). DiBAC4(3) was used to measure membrane potential. Microglial activation and neuronal loss were detected by enzyme-linked immunosorbent assay, Western blotting, and immunostaining. Learning and memory function was assessed with Morris water maze task. Cu(II) caused a hyperpolarized membrane potential in microglial cells, an effect abolished by functional Kv1.1 blockade. Blockade of Kv1.1 and knock-down of Kv1.1 with small interfering RNA repressed Cu(II)-induced microglial production of pro-inflammatory mediators. Also, Kv1.1 inhibition attenuated activation of PI3K/Akt-ERK1/2 signaling pathway and production of mitochondrial reactive oxidative species as well as nuclear factor-κB activation in Cu(II)-stimulated microglia. Moreover, the Cu(II)-caused, microglia-mediated neurotoxicity (indicated by reduced neuronal survival and increased dendritic loss) was attenuated by Kv1.1 knock-down. In an in vivo mouse model, hippocampal injection of Cu(II) caused elevated Kv1.1 mRNA (but not other Kv1 channels) expression and enhanced microglial Kv1.1 immunoreactivity in the hippocampus. Furthermore, blockade of Kv1.1 attenuated Cu(II)-induced microglial activation and neuronal dendritic loss in the hippocampus and learning and memory dysfunction. These findings suggest that inhibition of Kv1.1 ameliorates Cu(II)-induced microglial activation and cognitive impairment. Thus, it might represent a potential molecular target for anti-inflammatory therapy of neurodegenerative disorders.

Danhong injection modulates microglial polarization and neuroinflammation via the JUNB/NF-κB pathway in ischemic stroke.

Ischemic stroke (IS) is a leading cause of death and disability in China. Danhong Injection (DHI) is a traditional Chinese medicine preparation made from Salvia miltiorrhiza var. miltiorrhiza and Carthamus tinctoriusL. It is used for treating stroke in China with proven safety and efficacy. Microglia M1/M2 polarization is a key factor in IS inflammatory response. However, the key transcription factors that regulate microglia polarization are unknown. It is also not clear how DHI exerts its mechanism in the treatment of IS. This research aimed to investigate the effect of DHI on microglial polarization and neuroinflammation associated with IS and to elucidate the underlying mechanisms, with an emphasis on the JUNB/NF-κB signaling pathway. An oxygen-glucose deprivation (OGD) damage cell model and a permanent middle cerebral artery occlusion (pMCAO) model in C57BL/6 mice were employed. Neurological deficits, cerebral infarct volume, and microglial activation were assessed. Non-targeted metabolomics analysis with UHPLC-QE-MS and molecular biology methods, including RT-qPCR and Western blot, were applied to investigate the mechanisms. In vivo, DHI decreased inflammation, reduced brain damage, and enhanced neurological function. DHI also ameliorated microglial activation and OGD-induced apoptosis in vitro. Metabolomics analysis identified significant metabolic changes, particularly in amino acid metabolism. Additionally, DHI treatment decreased the upregulated mRNA levels of ASS1 and ASL after OGD, indicating an influence on the arginine biosynthesis pathway, which is crucial for microglial function. DHI modulated the M1 to M2 phenotypes of microglial polarization and regulated microglial polarization through the JUNB/NF-κB signaling pathway. This was confirmed by JUNB silencing experiments. DHI exhibits neuroprotective effects via suppressing ASS1 through the JUNB/NF-κB pathway, promoting the M2 state of microglia, and lowering the expression of inflammatory cytokines. This research unveils the potential therapeutic target of JUNB for IS treatment and sheds light on the novel intervention mechanism of DHI in microglial cells.