Anti-Neuroinflammatory Effects of Arecae pericarpium on LPS-Stimulated BV2 Cells.

Arecae pericarpium (AP), the fruit peel of the betel palm, is a traditional Oriental herbal medicine. AP is used to treat various diseases and conditions, such as ascites, edema, and urinary retention, in traditional Korean medicine. Recent studies have demonstrated its anti-obesity and antibacterial effects; however, its anti-neuroinflammatory effects have not yet been reported. Therefore, we investigated the anti-neuroinflammatory effects of AP on lipopolysaccharide (LPS)-stimulated mouse microglia in this study. To determine the anti-neuroinflammatory effects of AP on BV2 microglial cells, we examined the production of nitric oxide (NO) using Griess assay and assessed the mRNA expression levels of inflammatory mediators, such as inducible NO synthase (iNOS) and cyclooxygenase (COX)-2, and pro-inflammatory cytokines, such as interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α, using a real-time reverse transcription-polymerase chain reaction. Furthermore, we determined the levels of mitogen-activated protein kinases and IκBα via Western blotting to understand the regulating mechanisms of AP. AP treatment decreased NO production in LPS-stimulated BV2 cells. Additionally, AP suppressed the expression of iNOS and COX-2 and the production of pro-inflammatory cytokines. AP also inhibited the activation of p38 and nuclear factor-kappa B (NF-κB) in LPS-stimulated BV2 cells. Therefore, AP exerts anti-neuroinflammatory effects via inactivation of the p38 and NF-κB pathways.

The ROS/TXNIP/NLRP3 pathway mediates LPS-induced microglial inflammatory response.

BACKGROUND: Sepsis-associated encephalopathy (SAE) is a diffuse brain dysfunction activated by microglia. The potential pathological changes of SAE are complex, and the cellular pathophysiological characteristics remains unclear. This study aims to explore the ROS/TXNIP/NLRP3 pathway mediated lipopolysaccharide (LPS)-induced inflammatory response in microglia. METHODS: BV-2 cells were pre-incubated with 10 µM N-acetyl-L-cysteine (NAC) for 2 h, which were then reacted with 1 µg/mL LPS for 24 h. Western blot assay examined the protein levels of IBA1, CD68, TXNIP, NLRP3, ASC, and Cleaved Caspase-1 in BV-2 cells. The contents of inflammatory factor were detected by ELISA assay. The co-immunoprecipitation assay examined the interaction between TXNIP and NLRP3. RESULTS: LPS was confirmed to promote the positive expressions of IBA1 and CD68 in BV-2 cells. The further experiments indicated that LPS enhanced ROS production and NLRP3 inflammasome activation in BV-2 cells. Moreover, we also found that NAC partially reversed the facilitation of LPS on the levels of ROS, IL-1ß, IL-18, TXNIP, NLRP3, ASC, and Cleaved Caspase-1 in BV-2 cells. NAC treatment also notably alleviated the interaction between TXNIP and NLRP3 in BV-2 cells. CONCLUSION: ROS inhibition mediated NLRP3 signaling inactivation by decreasing TXNIP expression.

Immune-related transcriptomic and epigenetic reconfiguration in BV2 cells after lipopolysaccharide exposure: an in vitro omics integrative study.

BACKGROUND: Molecular alterations affecting microglia have been consistently associated with the inflammatory response. These cells can have pro- or anti-inflammatory activity, phenotypes thought to be regulated by epigenetic mechanisms. Still, little is known about the details on how epigenetic marks regulate the expression of genes in the context of an inflammatory response. METHODS: Through CUT&RUN, we profiled four genome-wide histone marks (HM) (H3K4me1, H3K4me3, H3K27ac, and H3K27me3) in lipopolysaccharide-exposed cells and compared their distributions to control cells. Transcriptomic profiles were determined through RNA-seq and differentially expressed genes were identified and contrasted with the epigenetic landscapes. Other downstream analyses were also included in this study. RESULTS: Our results illustrate an effectively induced M1 phenotype in microglial cells derived from LPS exposure. We observed differential bound regions associated with the genes classically involved in the inflammatory response in the expected direction according to each histone modification. Consistently, our transcriptomic analysis yielded a conspicuous illustration of the LPS-induced immune activity showing the up-regulation of Nf-κB-induced mRNAs (TNF-α, nfκbiz, nfκbia) and other important genes (Marco, Il-6, etc.). Furthermore, we integrated both omics profiles and identified an important reconfiguration of the genome induced by LPS. The latter was depicted by 8 different chromatin states that changed between conditions and that associated with unique clusters of differentially expressed genes, which not only represented regulatory elements, but also underlined distinct biological functions (inhibition of morphogenesis; changes in metabolism, homeostasis, and cytokine regulation; activation of the inflammatory response). CONCLUSION: This study exhibits important differences in the distribution of histone modifications in treated and control BV2 cells, constituting an epigenetic reconfiguration that leads to the inflammatory response. Also, it highlights the importance of these marks' regulatory role in gene expression and provides possible targets for further studies in the context of inflammation.

Deciphering the mechanism of cimifugin in mitigating LPS-induced neuroinflammation in BV-2 cells.

PURPOSE: Sepsis often triggers a systemic inflammatory response leading to multi-organ dysfunction, with complex and not fully understood pathogenesis. This study investigates the therapeutic effects of cimifugin on BV-2 cells under sepsis-induced stress conditions. METHODS: We utilized a BV-2 microglial cell model treated with lipopolysaccharide (LPS) to mimic sepsis. Assessments included cellular vitality, inflammatory cytokine quantification (6 interleukin [6IL]-1ß, interleukin 6 [IL-6], and tumor necrosis factor-α [TNF-α]) via enzyme-linked-immunosorbent serologic assay, and analysis of mRNA expression using real-time polymerase chain reaction. Oxidative stress and mitochondrial function were also evaluated to understand the cellular effects of cimifugin. RESULTS: Cimifugin significantly attenuated LPS-induced inflammatory responses, oxidative stress, and mitochondrial dysfunction. It enhanced cell viability and modulated the secretion and gene expression of inflammatory cytokines IL-1ß, IL-6, and TNF-α. Notably, cimifugin activated the deacetylase sirtuin 1-nuclear factor erythroid 2-related factor 2 pathway, contributing to its protective effects against mitochondrial damage. CONCLUSION: Cimifugin demonstrates the potential of being an effective treatment for sepsis--induced neuroinflammation, warranting further investigation.

A new cassane diterpenoid from the seed of Caesalpinia sappan.

In this study, a previously undescribed cassane diterpenoid, named caesalpinin JF (1), along with two known cassane diterpenoids caesanine C (2) and tomocinol B (3), was isolated from 95% EtOH extract of the seeds of Caesalpinia sappan Linn. Additionally, three known compounds including pulcherrin R (4), syringaresinol-4'-O-ß-D-glucopyranoside (5) and kaempferol (6) were also identified. The structures of the isolated compounds were elucidated by comprehensive 1D and 2D NMR spectroscopic analyses. Additionally, electronic circular dichroism (ECD) calculation was used to identify the absolute structure of compound 1. Among the isolated compounds, compound 1 displayed a potent anti-neuroinflammation with an IC50 value of 9.87 ± 1.71 µM.

MiRNA-338-3p Inhibits Neuroinflammation in the Corpus Callosum of LCV-LPS Rats Via STAT1 Signal Pathway.

Neuroinflammation is a common characteristic of intracranial infection (ICI), which is associated with the activation of astrocytes and microglia. MiRNAs are involved in the process of neuroinflammation. This study aimed to investigate the potential mechanism by which miR-338-3p negatively modulate the occurrence of neuroinflammation. We here reported that the decreased levels of miR-338-3p were detected using qRT-PCR and the upregulated expression of TNF-α and IL-1ß was measured by ELISA in the cerebrospinal fluid (CSF) in patients with ICI. A negative association between miR-338-3p and TNF-α or IL-1ß was revealed by Pearson correlation analysis. Sprague-Dawley (SD) rats were injected with LPS (50 µg) into left cerebral ventricule (LCV), following which the increased expression of TNF-α and IL-1ß and the reduction of miR-338-3p expression were observed in the corpus callosum (CC). Moreover, the expression of TNF-α and IL-1ß in the astrocytes and microglia in the CC of LCV-LPS rats were saliently inhibited by the overexpression of miR-338-3p. In vitro, cultured astrocytes and BV2 cells transfected with mimic-miR-338-3p produced less TNF-α and IL-1ß after LPS administration. Direct interaction between miR-338-3p and STAT1 mRNA was validated by biological information analysis and dual luciferase assay. Furthermore, STAT1 pathway was found to be implicated in inhibition of neuroinflammation induced by mimic miR-338-3p in the astrocytes and BV2 cells. Taken together, our results suggest that miR-338-3p suppress the generation of proinflammatory mediators in astrocyte and BV2 cells induced by LPS exposure through the STAT1 signal pathway. MiR-338-3p could act as a potential therapeutic strategy to reduce the neuroinflammatory response. Diagram describing the cellular and molecular mechanisms associated with LPS-induced neuroinflammation via the miR-338-3p/STAT1 pathway. LPS binds to TLRs on astrocytes or microglia to activate the STAT1 pathway and upregulate the production of pro-inflammatory cytokines. However, miR-338-3p inhibits the expression of STAT1 and reduces the production of inflammatory mediators.

Adiponectin deficiency accelerates brain aging via mitochondria-associated neuroinflammation.

BACKGROUND: A wide spectrum of changes occurs in the brain with age, from molecular to morphological aspects, and inflammation accompanied by mitochondria dysfunction is one of the significant factors associated with age. Adiponectin (APN), an essential adipokine in glucose and lipid metabolism, is involved in the aging; however, its role in brain aging has not been adequately explored. Here, we aimed to explore the relationship between APN deficiency and brain aging using multiple biochemical and pharmacological methods to probe APN in humans, KO mice, primary microglia, and BV2 cells. RESULTS: We found that declining APN levels in aged human subjects correlated with dysregulated cytokine levels, while APN KO mice exhibited accelerated aging accompanied by learning and memory deficits, anxiety-like behaviors, neuroinflammation, and immunosenescence. APN-deficient mice displayed aggravated mitochondrial dysfunction and HDAC1 upregulation. In BV2 cells, the APN receptor agonist AdipoRon alleviated the mitochondrial deficits and aging markers induced by rotenone or antimycin A. HDAC1 antagonism by Compound 60 (Cpd 60) improved mitochondrial dysfunction and age-related inflammation, as validated in D-galactose-treated APN KO mice. CONCLUSION: These findings indicate that APN is a critical regulator of brain aging by preventing neuroinflammation associated with mitochondrial impairment via HDAC1 signaling.

Oxyberberine an oxoderivative of berberine confers neuroprotective effects in controlled-cortical impact mouse model of traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) is known as a silent epidemic that causes many deaths and disabilities worldwide. We examined the response of oxyberberine (OBB) in lipopolysaccharide-stimulated BV2 microglial cells and a controlled-cortical impact (CCI) mouse model of TBI. METHODS: We synthesized OBB from berberine, and also prepared OBB-nanocrystals (OBB-NC). Male C57BL/6 mice were used for CCI surgery, and post-CCI neurobehavioral deficits were assessed from 1 h after injury through 21 days post-injury (dpi). RESULTS: OBB treatment reduced the lipopolysaccharide-triggered elevated levels of reactive oxygen species, nitric oxide, and nuclear factor kappa B (NF-κB) in BV2 microglial cells, indicating a neuroprotective potential. CCI-operated mice exhibited significant neurological deficits on 1, 3, and 5 dpi in neurological severity scoring and rotarod assay. OBB (25 and 50 mg/kg/day) and OBB-NC (3 mg/kg/day) ameliorated these neurological aberrations. Mice subjected to CCI surgery also displayed anxiogenic- and depression-like behaviours, and cognitive impairments in forced-swimming test and elevated-zero maze, and novel object recognition task, respectively. Administration of OBB reduced these long-term neuropsychiatric complications, and also levels of toll-like receptor 4 (TLR4), high-motility group protein 1 (HMGB1), NF-κB, tumour necrosis factor-alpha and interleukin 6 cytokines in the ipsilateral cortex of mice. CONCLUSION: We suggest that the administration of OBB offers neuroprotective effects via inhibition of HMGB1-mediated TLR4/NFκB pathway.

Polystyrene Microplastics Exacerbate Systemic Inflammation in High-Fat Diet-Induced Obesity.

Microplastics (MPs) are recognized as environmental pollutants with potential implications for human health. Considering the rapid increase in obesity rates despite stable caloric intake, there is a growing concern about the link between obesity and exposure to environmental pollutants, including MPs. In this study, we conducted a comprehensive investigation utilizing in silico, in vitro, and in vivo approaches to explore the brain distribution and physiological effects of MPs. Molecular docking simulations were performed to assess the binding affinity of three plastic polymers (ethylene, propylene, and styrene) to immune cells (macrophages, CD4+, and CD8+ lymphocytes). The results revealed that styrene exhibited the highest binding affinity for macrophages. Furthermore, in vitro experiments employing fluorescence-labeled PS-MPs (fPS-MPs) of 1 µm at various concentrations demonstrated a dose-dependent binding of fPS-MPs to BV2 murine microglial cells. Subsequent oral administration of fPS-MPs to high-fat diet-induced obese mice led to the co-existence of fPS-MPs with immune cells in the blood, exacerbating impaired glucose metabolism and insulin resistance and promoting systemic inflammation. Additionally, fPS-MPs were detected throughout the brain, with increased activation of microglia in the hypothalamus. These findings suggest that PS-MPs significantly contribute to the exacerbation of systemic inflammation in high-fat diet-induced obesity by activating peripheral and central inflammatory immune cells.

MiR-145 enriched exosomes derived from bone marrow-derived mesenchymal stem cells protects against cerebral ischemia-reperfusion injury through downregulation of FOXO1.

BACKGROUND: Mesenchymal stem cells-derived exosomes (MSCs-Exo) were able to exert neuroprotective effects in brain injury after ischemic stroke (IS). In addition, exosomes containing microRNAs (miRNAs) can be transported to recipient cells to mediate intercellular communication. It has been shown that the level of miR-145 was significantly downregulated in brain tissues of rats subjected to middle cerebral artery occlusion (MCAO). However, the role of MSCs-derived exosomal miR-145 in IS progression remains largely unknown. METHODS: Microglial BV2 cell exposed to oxygen-glucose deprivation/reperfusion (OGD/R) was applied to mimic cerebral ischemia/reperfusion (I/R) injury conditions in vitro. In addition, a rat model of MCAO was established to induce I/R injury. Meanwhile, exosomes were isolated from miR-145-transfected bone marrow MSCs, and then these isolated exosomes were used to treat OGD/R-stimulated BV-2 cell and rats subject to MCAO/R. RESULTS: In this study, we found that miR-145 could be transferred from MSCs to BV2 cells via exosomes. In addition, exosomal miR-145-derived from MSCs was able to shift microglia polarization toward anti-inflammatory M2 phenotype in OGD/R-stimulated BV2 cells. Moreover, exosomal miR-145 markedly suppressed the apoptosis, cell cycle arrest and oxidative stress in OGD/R-treated BV2 cells. Additionally, exosomal miR-145 notably decreased the expression of FOXO1 in BV2 cell exposed to OGD/R and in brain tissues of MCAO rats. Furthermore, exosomal miR-145 remarkably decreased infarct area in MCAO rats. CONCLUSION: Collectively, exosomal miR-145-derived from MSCs was able to attenuate cerebral I/R injury through downregulation of FOXO1. These studies may serve as a potential approach for treating of cerebral I/R injury.

Anti-inflammatory role of GM1 and other gangliosides on microglia.

BACKGROUND: Gangliosides are glycosphingolipids highly enriched in the brain, with important roles in cell signaling, cell-to-cell communication, and immunomodulation. Genetic defects in the ganglioside biosynthetic pathway result in severe neurodegenerative diseases, while a partial decrease in the levels of specific gangliosides was reported in Parkinson's disease and Huntington's disease. In models of both diseases and other conditions, administration of GM1-one of the most abundant gangliosides in the brain-provides neuroprotection. Most studies have focused on the direct neuroprotective effects of gangliosides on neurons, but their role in other brain cells, in particular microglia, is not known. In this study we investigated the effects of exogenous ganglioside administration and modulation of endogenous ganglioside levels on the response of microglia to inflammatory stimuli, which often contributes to initiation or exacerbation of neurodegeneration. METHODS: In vitro studies were performed using BV2 cells, mouse, rat, and human primary microglia cultures. Modulation of microglial ganglioside levels was achieved by administration of exogenous gangliosides, or by treatment with GENZ-123346 and L-t-PDMP, an inhibitor and an activator of glycolipid biosynthesis, respectively. Response of microglia to inflammatory stimuli (LPS, IL-1ß, phagocytosis of latex beads) was measured by analysis of gene expression and/or secretion of pro-inflammatory cytokines. The effects of GM1 administration on microglia activation were also assessed in vivo in C57Bl/6 mice, following intraperitoneal injection of LPS. RESULTS: GM1 decreased inflammatory microglia responses in vitro and in vivo, even when administered after microglia activation. These anti-inflammatory effects depended on the presence of the sialic acid residue in the GM1 glycan headgroup and the presence of a lipid tail. Other gangliosides shared similar anti-inflammatory effects in in vitro models, including GD3, GD1a, GD1b, and GT1b. Conversely, GM3 and GQ1b displayed pro-inflammatory activity. The anti-inflammatory effects of GM1 and other gangliosides were partially reproduced by increasing endogenous ganglioside levels with L-t-PDMP, whereas inhibition of glycolipid biosynthesis exacerbated microglial activation in response to LPS stimulation. CONCLUSIONS: Our data suggest that gangliosides are important modulators of microglia inflammatory responses and reveal that administration of GM1 and other complex gangliosides exerts anti-inflammatory effects on microglia that could be exploited therapeutically.

Zileuton, a 5-Lipoxygenase Inhibitor, Attenuates Haemolysate-Induced BV-2 Cell Activation by Suppressing the MyD88/NF-κB Pathway.

M1 microglia induce neuroinflammation-related neuronal death in animal models of spontaneous subarachnoid haemorrhage. Zileuton is a 5-lipoxygenase inhibitor that reduces the levels of downstream pro-inflammatory cytokines. This study aimed to investigate whether zileuton inhibits microglial activation and describe its underlying mechanisms. BV-2 cells were exposed to 1 mg/mL haemolysate for 30 min, followed by treatment with different concentrations (5, 10, 15, or 20 µM) of zileuton for 24 h. The cells were then assessed for viability, polarisation, and protein expression levels. Haemolysate increases the viability of BV-2 cells and induces M1 polarisation. Subsequent exposure to high concentrations of zileuton decreased the viability of BV-2 cells, shifted the polarisation to the M2 phenotype, suppressed the expression of 5-lipoxygenase, decreased tumour necrosis factor α levels, and increased interleukin-10 levels. Furthermore, high concentrations of zileuton suppressed the expression of myeloid differentiation primary response protein 88 and reduced the phosphorylated-nuclear factor-kappa B (NF-kB)/NF-kB ratio. Therefore, phenotype reversal from M1 to M2 is a possible mechanism by which zileuton attenuates haemolysate-induced neuroinflammation after spontaneous subarachnoid haemorrhage.

Identification of Potential Anti-Neuroinflammatory Inhibitors from Antarctic Fungal Strain Aspergillus sp. SF-7402 via Regulating the NF-κB Signaling Pathway in Microglia.

Microglia play a significant role in immune defense and tissue repair in the central nervous system (CNS). Microglial activation and the resulting neuroinflammation play a key role in the pathogenesis of neurodegenerative disorders. Recently, inflammation reduction strategies in neurodegenerative diseases have attracted increasing attention. Herein, we discovered and evaluated the anti-neuroinflammatory potential of compounds from the Antarctic fungi strain Aspergillus sp. SF-7402 in lipopolysaccharide (LPS)-stimulated BV2 cells. Four metabolites were isolated from the fungi through chemical investigations, namely, 5-methoxysterigmatocystin (1), sterigmatocystin (2), aversin (3), and 6,8-O-dimethylversicolorin A (4). Their chemical structures were elucidated by extensive spectroscopic analysis and HR-ESI-MS, as well as by comparison with those reported in literature. Anti-neuroinflammatory effects of the isolated metabolites were evaluated by measuring the production of nitric oxide (NO), tumor necrosis factor (TNF)-α, and interleukin (IL)-6 in LPS-activated microglia at non-cytotoxic concentrations. Sterigmatocystins (1 and 2) displayed significant effects on NO production and mild effects on TNF-α and IL-6 expression inhibition. The molecular mechanisms underlying this activity were investigated using Western blot analysis. Sterigmatocystin treatment inhibited NO production via downregulation of inducible nitric oxide synthase (iNOS) expression in LPS-stimulated BV2 cells. Additionally, sterigmatocystins reduced nuclear translocation of NF-κB. These results suggest that sterigmatocystins present in the fungal strain Aspergillus sp. are promising candidates for the treatment of neuroinflammatory diseases.

Anti-Neuroinflammatory Components from Clausena lenis Drake.

Clausena lenis Drake (C. lenis) is a folk medicinal herb to treat influenza, colds, bronchitis, and malaria. The 95% and 50% ethanol extract of C. lenis showed significant nitric oxide (NO) inhibition activity in BV-2 microglial cells stimulated by lipopolysaccharide (LPS). Bio-guided isolation of the active extract afforded five new compounds, including a chlorine-containing furoquinoline racemate, (±)-claulenine A (1), an amide alkaloid, claulenine B (2), a prenylated coumarin, claulenin A (3), a furocoumarin glucoside, clauleside A (4), and a multi-prenylated p-hydroxybenzaldehyde, claulenin B (5), along with 33 known ones. Their structures were determined via spectroscopic methods, and the absolute configurations of new compounds were assigned via the electronic circular dichroism (ECD) calculations and single-crystal X-ray diffraction analysis. Compounds 2, 23, 27, 28, 33, and 34 showed potent anti-neuroinflammatory effects on LPS-induced NO production in BV-2 microglial cells, with IC50 values in the range of 17.6-40.9 µM. The possible mechanism was deduced to interact with iNOS through molecular docking.

Platinum nanoparticles Protect Against Lipopolysaccharide-Induced Inflammation in Microglial BV-2 Cells via Decreased Oxidative Damage and Increased Phagocytosis.

Neuroinflammation and oxidative stress cooperate to compromise the function of the central nervous system (CNS). Colloidal platinum nanoparticles (Pt NPs) are ideal candidates for reducing the deleterious effects of neuroinflammation since they act as free radical scavengers. Here we evaluated the effects of Pt NPs on several markers of lipopolysaccharide (LPS)-induced inflammation in cultured BV-2 microglial cells. BV-2 cells were treated with increased dilutions (1-100 ppm) of Colloidal Pt and/or LPS (1-10 µg/mL) at different exposure times. Three different protocols of exposure were used combining Pt NPs and LPS: (a) conditioning-protective effect (pre-post-treat), (b) therapeutic effect (co-treat) and (c) conditioning-therapeutic effect (pre-co-treat). After exposure to LPS for 24 h, cells were used for assessment of cell viability, reactive oxygen species (ROS) generation, lactate dehydrogenase (LDH) activity, apoptosis and caspase-3 levels, cell proliferation, mitochondrial membrane potential, inducible nitric oxide (iNOS) activity, pro-inflammatory cytokine (IL-1ß, TNF-α and IL-6) levels, and phagocytic activity. Low concentrations (below or equal to 10 ppm) of Colloidal Pt prevented or ameliorated the LPS-induced increase in ROS formation, loss of mitochondrial membrane potential, induction of apoptosis, increase in LDH release, increase in pro-inflammatory cytokines and iNOS, inhibition of phagocytosis linked to microglial persistence in the M1 phase phenotype, loss of cell adhesion, differentiation and/or proliferation, as well as loss of cell viability. These protective effects were evident when cells were preconditioned with Pt NPs prior to LPS treatment. Collectively, the findings demonstrate that at low concentrations, Pt NPs can regulate the function and phenotype of BV-2 cells, activating protective mechanisms to maintain the microglial homeostasis and reduce inflammatory events triggered by the inflammatory insults induced by LPS. These preventive/protective effects on the LPS pro-inflammatory model are linked to the antioxidant properties and phagocytic activity of these NPs.

Lipopolysaccharide induces BV2 microglial cell migration via a decrease in SET8 expression.

Excessively activated microglia exhibit increased migration, resulting in tissue damage and chronic inflammation. Src was confirmed to play an important role in regulation of cell motility following lipopolysaccharide (LPS) treatment. SET8 plays an important part in multiple cellular signal pathways. In this study, we speculated that SET8 is involved in LPS-induced microglial migration via regulation of Src expression. Our study showed that LPS promoted cell migration via augmentation of Src expression in BV2 cells. Moreover, LPS treatment decreased SET8 expression and upregulated the expression of the transcription factor ETS proto-oncogene 1 (ETS1). Overexpression of both SET8 and small interfering ETS1 reversed LPS-induced Src expression and cell migration. The effects of short hairpin SET8 (shSET8) and ETS1 overexpression are the same as the effects of LPS treatment. Decrease of Src expression reversed the shSET8-induced and ETS1 overexpression-induced migration of BV2 cells. Furthermore, SET8 was observed to associate with ETS1. Chromatin immunoprecipitation assay indicated H4K20me1, a downstream target of SET8, in addition to ETS1, was enriched at the Src promoter region. Furthermore, shSET8 increased Src promoter activity and also increased the positive effect of ETS1 overexpression on Src promoter activity. This study shows that SET8 associates with ETS1 to regulate Src expression, which is involved in LPS-induced BV2 cell migration.

Identification of potential circRNA-miRNA-mRNA regulatory networks in response to graphene quantum dots in microglia by microarray analysis.

Along with the increasing application of graphene quantum dots (GQDs) in the fields of biomedicine and neuroscience, it is important to assess the probably adverse effects of GQDs in the central nervous system (CNS) but their underlying toxic mechanisms is still unclear. In this study, we evaluate the molecular mechanisms associated with circular RNAs (circRNAs) of nitrogen-doped GQDs (N-GQDs) and amino-functionalized GQDs (A-GQDs) damaging the cell viability and cellular structure in microglia by an integrative analysis of RNA microarray. The differentially expressed circRNA (DEcircRNAs)-miRNA- differentially expressed mRNA (DEmRNAs) regulatory networks were conducted in BV2 microglial cells treated with 25 µg/mL N-GQDs, 100 µg/mL N-GQDs and 100 µg/mL A-GQDs. Based on that, the protein-coding genes in each ceRNA network were collected to do bio-functional analysis to evaluate signaling pathways that were indirectly mediated by circRNAs. Some pathways that could play indispensable roles in the neurotoxicity of N-GQDs or both two kinds of GQDs were found. Low-dosed N-GQDs exposure mainly induced inflammatory action in microglia, while high-dosed N-GQDs and A-GQDs exposure both affect olfactory transduction and GABAergic synapse. Meanwhile, several classical signaling pathways, including mTOR, ErbB and MAPK, could make diverse contributions to the neurotoxicity of both two kinds of GQDs. These circRNAs could be toxic biomarkers or protective targets in neurotoxicity of GQDs. More importantly, they emphasized the necessity of comprehensive analysis of latent molecular mechanisms through epigenetics approaches in biosafety assessment of graphene-based nanomaterials.

MKP1 reduces neuroinflammation via inhibiting endoplasmic reticulum stress and mitochondrial dysfunction.

MAP kinase phosphatase 1 (MKP1) has been identified as an antiapoptotic protein via sustaining mitochondrial function. However, the role of MKP1 in neuroinflammation has not been fully understood. The aim of this study is to figure out the influence of MKP1 in lipopolysaccharide (LPS)-treated microglia BV-2 cells and investigate whether MKP1 reduces BV-2 cell death via modulating endoplasmic reticulum (ER) stress and mitochondrial dysfunction. The results of this study demonstrated that MKP1 was rapidly downregulated after exposure to LPS. However, the transfection of MKP1 adenovirus could reverse cell viability and attenuate LPS-mediated BV-2 cell apoptosis. Mechanistically, MKP1 overexpression alleviated ER stress and corrected LPS-induced calcium overloading. Besides, MKP1 adenovirus transfection also reversed mitochondrial bioenergetics, maintained mitochondrial membrane potential, and blocked mitochondria-initiated apoptosis signals. Furthermore, we found that MKP1 overexpression is associated with inactivation of mitogen-activated protein kinase-c-Jun N-terminal kinase (MAPK-JNK) pathway. Interestingly, the activation of MAPK-JNK pathway could abolish the protective effects of MKP1 on BV-2 cells survival and mitochondrial function in the presence of LPS. Altogether, our results identified MKP1 as a primary defender of neuroinflammation via modulating ER stress and mitochondrial function in a manner dependent on MAPK-JNK pathway. These findings may open a new window for the treatment of neuroinflammation in the clinical setting.

The glycolytic shift was involved in CdTe/ZnS quantum dots inducing microglial activation mediated through the mTOR signaling pathway.

The excellent optical property and relatively low toxicity of CdTe/ZnS core/shell quantum dots (QDs) make them an advanced fluorescent probe in the application of biomedicines, particularly in neuroscience. Thus, it is important to evaluate the biosafety of CdTe/ZnS QDs on the central nervous system (CNS). Our previous studies have suggested that the high possibility of CdTe/ZnS QDs being transported into the brain across the blood-brain barrier resulted in microglial activation and a shift of glycometabolism, but their underlying mechanism remains unclear. In this study, when mice were injected intravenously with CdTe/ZnS QDs through tail veins, the microglial activation, polarized into both M1 phenotype and M2 phenotype, and the neuronal impairment were observed in the hippocampus. Meanwhile, the increased pro- and anti-inflammatory cytokines released from BV2 microglial cells treated with CdTe/ZnS QDs also indicated that QD exposure was capable of inducing microglial activation in vitro. We further demonstrated that the glycolytic shift from oxidative phosphorylation switching into aerobic glycolysis was required in the microglial activation into M1 phenotype induced by CdTe/ZnS QD treatment, which was mediated through the mTOR signaling pathway. The findings, taken together, provide a mechanistic insight regarding the CdTe/ZnS QDs inducing microglial activation and the role of the glycolytic shift in it.

Luteolin protects microglia against rotenone-induced toxicity in a hormetic manner through targeting oxidative stress response, genes associated with Parkinson's disease and inflammatory pathways.

Rotenone, an environmental toxin, triggers Parkinson's disease (PD)-like pathology through microglia-mediated neuronal death. The effects and molecular mechanisms of flavonoid luteolin against rotenone-induced toxicity was assessed in microglial BV2 cells. Cells were pretreated with luteolin (1-50 µM) for 12 h and then was co-treated with 20 µM of rotenone for an additional 12 h in the presence of luteolin. The viability (MTT), IL-1ß and TNF-α levels and lactate dehydrogenase (LDH) release (ELISA), and Park2, Lrrk2, Pink1, Nrf2 and Trx1 mRNA levels (qRT-PCR) were measured. In rotenone exposed microglia, luteolin increased viability significantly at lower concentrations (1-5 µM) compared to higher concentrations (25-50 µM). Rotenone increased LDH release and IL-1ß levels in a dose-dependent manner (1-20 µM). Luteolin inhibited rotenone-induced LDH release, however the activity decreased in concentration-dependent manner Neither rotenone nor luteolin altered TNF-α levels, but luteolin reduced IL-1ß levels in a concentration dependent manner in rotenone exposed cells. The mRNA levels of Nrf2 and Trx1, which are the master regulators of redox state, were increased by rotenone, as well as by luteolin, which exhibited an inverse relationship between its concentration and effect (1-20 µM). Park2 mRNA levels increased by luteolin, but decreased by rotenone. Pink1 mRNA levels was not altered by rotenone or luteolin. Lrrk2 mRNA levels reduced by luteolin, while it was increased by rotenone. Results suggest that luteolin have favorable effects on regulation of oxidative stress response, genes associated with PD and inflammatory pathways, hence protects microglia against rotenone toxicity in a hormetic manner.