Activated BV-2 Microglial Cells Research Articles

Synergy of botanical drug extracts from Dracaena cochinchinensis stemwood and Ardisia elliptica fruit in multifunctional effects on neuroprotection and anti-inflammation.

Combination therapy is one of the promising approaches in developing therapeutics to cure complex diseases, such as Alzheimer's disease (AD). In Thai traditional medicines, the clinical application often comprises multiple botanical drugs as a formulation. The synergistic interactions between botanical drugs in combination therapies are proposed to have several advantages, including increased therapeutic efficacy, and decreased toxicity and/or adverse effects. This study aimed to explore the therapeutic functions of a botanical hybrid preparation (BHP) of two botanical drugs within a traditional multi-herbal formulation. The synergistic actions of BHP of Dracaena cochinchinensis stemwood (DCS) and Ardisia elliptica fruit (AEF) at a specific ratio of 1:9 w/w were illustrated in neuroprotection and anti-inflammation. In cultured PC12 cells, BHP of DCS and AEF showed synergistic functions in inducing neuronal differentiation, characterized by neurofilament expression and neurite outgrowth. In addition, BHP of DCS and AEF exhibited a synergistic effect in inhibiting the aggregation of Aβ, a hallmark of AD pathology. The activated BV2 microglial cells induced by LPS were synergistically suppressed by the BHP of DCS and AEF, as evaluated by the expression of pro-inflammatory markers, including TNF-α, IL-1β, and iNOS, as well as the morphological change of microglial cells. The findings suggested that the effects of BHP of DCS and AEF were greater than individual botanical drugs in a specific ratio of 1:9 w/w to enhance neuroprotective and anti-inflammatory functions.

Ser9p-GSK3β Modulation Contributes to the Protective Effects of Vitamin C in Neuroinflammation.

The prolonged activation of microglia and excessive production of pro-inflammatory cytokines can lead to chronic neuroinflammation, which is an important pathological feature of Parkinson's disease (PD). We have previously reported the protective effect of Vitamin C (Vit C) on a mouse model of PD. However, its effect on microglial functions in neuroinflammation remains to be clarified. Glycogen synthase kinase 3β (GSK3β) is a serine/threonine kinase having a role in driving inflammatory responses, making GSK3β inhibitors a promising target for anti-inflammatory research. In this study, we investigated the possible involvement of GSK3β in Vit C neuroprotective effects by using a well-known 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced animal model of PD and a cellular model of neuroinflammation, represented by Lipopolysaccharide (LPS)-activated BV-2 microglial cells. We demonstrated the ability of Vit C to decrease the expression of different mediators involved in the inflammatory responses, such as TLR4, p-IKBα, and the phosphorylated forms of p38 and AKT. In addition, we demonstrated for the first time that Vit C promotes the GSK3β inhibition by stimulating its phosphorylation at Ser9. This study evidenced that Vit C exerts an anti-inflammatory function in microglia, promoting the upregulation of the M2 phenotype through the activation of the Wnt/β-catenin signaling pathway.

Crude extracts of <i>Momordica cochinchinensis</i> (Lour.) Spreng exerts antioxidant and anti-neuroinflammatory properties in LPS-stimulated BV2 microglia

Purpose: To evaluate the antioxidant and anti-neuroinflammatory properties of the leaves, seeds and pulp of Momordica cochinchinensis Spreng (commonly known as Gac) in vitro and in lipopolysaccharide (LPS)-activated BV2 microglial cells.
 Methods: Lipopolysaccharide was applied to BV2 cells for 24 h in the presence or absence of Gac extracts. Then, levels of reactive oxygen species (ROS) were assayed using CM-H2DCFDA method, while nitric oxide (NO) level was determined with Griess reagent. The levels of TNF-α and IL-6 were determined using ELISA. Three different assays were used for determination of in vitro antioxidant activities of the extracts, namely, 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging; 2,2’-azinobis-3- ethylbenzthiazoline-6-sulfonic acid (ABTS) scavenging, and ferric-reducing antioxidant power (FRAP). Total flavonoids content (TFC) and total phenolic content (TPC) were also determined. 
 Results: There were significantly higher levels of ROS, NO, TNF and IL-6 in LPS-treated BV2 cells than in control cells (p < 0.05). However, exposure of LPS treated BV2 cells to the leaf, seed and pulp extracts of Gac led to marked decreases in levels of ROS and inflammatory mediators, when compared with untreated cells, with the seed extract producing significantly larger decreases in levels of ROS and inflammatory mediators than the leaf and pulp extracts (p < 0.05). The DPPH and ABTS radical scavenging activities and total reducing power activities of the seed extract were superior to those of the leaf and pulp extracts. Moreover, the seed extract showed higher contents of TFC and TPC than the other two extracts.
 Conclusion: Gac extracts exert antioxidant and anti-neuroinflammatory effects on LPS-activated BV2 cells and in an in vitro model. Therefore, Gac extracts, especially seed extract, may be beneficial in the preparation of innovative antioxidative and anti-neuroinflammatory herbal remedies, as well as dietary supplements.

Tumor necrosis factor α-induced protein 8-like-2 controls microglia phenotype via metabolic reprogramming in BV2 microglial cells and responses to neuropathic pain

Microglial are major players in neuroinflammation that have recently emerged as potential therapeutic targets for neuropathic pain. Glucose metabolic programming has been linked to differential activation state and function in microglia. Tumor necrosis factor α-induced protein 8-like-2 (TNFAIP8L2) is an important component in regulating the anti-inflammatory response. However, the role of TNFAIP8L2 in microglia differential state during neuropathic pain and its interplay with glucose metabolic reprogramming in microglia has not yet been determined. Thus, we aimed to investigate the role of TNFAIP8L2 in the status of microglia in vitro and in vivo. BV2 microglial cells were treated with lipopolysaccharides plus interferon-gamma (LPS/IFNγ) or interleukin-4 (IL-4) to induce the two different phenotypes of microglia in vitro. In vivo experiments were conducted by chronic constriction injury of the sciatic nerve (CCI). We investigated whether TNFAIP8L2 regulates glucose metabolic programming in BV2 microglial cells. The data in vitro showed that TNFAIP8L2 lowers glycolysis and increases mitochondrial oxidative phosphorylation (OXPHOS) in inflammatory microglia. Blockade of glycolytic pathway abolished TNFAIP8L2-mediated differential activation of microglia. TNFAIP8L2 suppresses inflammatory microglial activation and promotes restorative microglial activation in BV2 microglial cells and in spinal cord microglia after neuropathic pain. Furthermore, TNFAIP8L2 controls differential activation of microglia and glucose metabolic reprogramming through the MAPK/mTOR/HIF-1α signaling axis. This study reveals that TNFAIP8L2 plays a critical role in neuropathic pain, providing important insights into glucose metabolic reprogramming and microglial phenotypic transition, which indicates that TNFAIP8L2 may be used as a potential drug target for the prevention of neuropathic pain.

MiRNA-192-5p targets Dyrk1a to attenuate cerebral injury in MCAO mice by suppressing neuronal apoptosis and neuroinflammation.

Ischemic stroke (IS) is a leading cause of disability and mortality worldwide. Several studies have demonstrated the involvement of microRNAs (miRNAs) in brain diseases. miRNA-192-5p is a regulatory molecule in neurodegenerative diseases and its expression was found to be significantly downregulated in the whole blood samples of IS patients, but the specific role of miRNA-192-5p in IS not fully understood. Here, we investigated the role of miRNA-192-5p in a murine model of acute cerebral injury after IS. Male C57BL/6J mice received an intracerebroventricular (i.c.v.) injection of agomir-192-5p or antagomir-192-5p 2 h before middle cerebral artery occlusion (MCAO). Infarct volume was assessed by 2,3,5 triphenyltetrazolium chloride (TTC) staining. Brain slices were subjected to Fluoro-Jade B, TUNEL, and immunofluorescence stainings. Contents of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) were measured using enzyme-linked immunosorbent assay (ELISA) kits. In vitro, murine microglial BV-2 cells were subjected to oxygen-glucose deprivation (OGD), and the contents of pro-inflammatory cytokines were measured in cell lysates. miRNA-192-5p was downregulated in the ischemic penumbra of the cerebral cortex. Pretreatment with agomir-192-5p attenuated neurological deficits and reduced cerebral edema and infarct volume in MCAO mice. Agomir-192-5p-treated animals had fewer degenerating and apoptotic neurons in the ischemic penumbra. Additionally, agomir-192-5p significantly suppressed neuroinflammation as evidenced by decreased immunostaining for GFAP and Iba1 and decreased levels of pro-inflammatory cytokines. Antagomir-192-5p pretreatment showed the opposite effect. Furthermore, dual specificity tyrosine phosphorylation regulated kinase 1A (Dyrk1a) was identified as a target gene of miRNA-192-5p, and the elevated Dyrk1a expression in the ischemic penumbra was markedly reduced by agomir-192-5p. Dyrk1a overexpression in BV-2 microglial cells impaired miRNA-192-5p-mediated inhibition of OGD-induced activation of BV-2 microglial cells. Opposite results were obtained using miRNA-192-5p inhibitor and Dyrk1a siRNA. We found that intracerebroventricular administration of miRNA-192-5p before MCAO attenuatedacute cerebral injury by suppressing neuronal apoptosis and neuroinflammation in mice, and these protective effects might be mediated by downregulation of Dyrk1a. This study would help identify novel therapeutic targets for IS.

The antidepressive mechanism of Longya Lilium combined with Fluoxetine in mice with depression-like behaviors

Traditional Chinese medicine is one of the most commonly used complementary and alternative medicine therapies for depression. Integrated Chinese-western therapies have been extensively applied in numerous diseases due to their superior efficiency in individual treatment. We used the meta-analysis, network pharmacology, and bioinformatics studies to identify the putative role of Longya Lilium combined with Fluoxetine in depression. Depression-like behaviors were mimicked in mice after exposure to the chronic unpredictable mild stress (CUMS). The underlying potential mechanism of this combination therapy was further explored based on in vitro and in vivo experiments to analyze the expression of COX-2, PGE2, and IL-22, activation of microglial cells, and neuron viability and apoptosis in the hippocampus. The antidepressant effect was noted for the combination of Longya Lilium with Fluoxetine in mice compared to a single treatment. COX-2 was mainly expressed in hippocampal CA1 areas. Longya Lilium combined with Fluoxetine reduced the expression of COX-2 and thus alleviated depression-like behavior and neuroinflammation in mice. A decrease of COX-2 curtailed BV-2 microglial cell activation, inflammation, and neuron apoptosis by blunting the PGE2/IL-22 axis. Therefore, a combination of Longya Lilium with Fluoxetine inactivates the COX-2/PGE2/IL-22 axis, consequently relieving the neuroinflammatory response and the resultant depression.

Iridoids from the aerial parts of Adenosma caeruleum with their nitric oxide inhibitory effects

Three new iridoids, adenocaeroids A−C (1−3), and seven known analogs, umbellatolide B (4), pedicularis-lactone (5), iridolactone (6), artselaenin C (7), eucommiol (8), rehmaglutin D (9), and 3-deoxy-artselaenin C (10) were isolated from the aerial parts of Adenosma caeruleum. Their chemical structures were identified by the comprehensive spectroscopic methods, including 1D and 2D NMR and HRESI mass spectra. The relative configurations of 1−3 were established by analyzing their NOESY spectra as well as by DFT-GIAO NMR calculations followed by ML-J-DP4, DP4+, and DU8 methods. The absolute configurations of 1−3 were determined by comparing experimental and calculated ECD spectra and specific optical rotation values. All of the compounds, except 7 and 10, inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-activated BV2 microglial cells in a dose-dependent manner, with IC50 values ranging from 28.4−61.8 µM. However, none of the compounds showed inhibitory effect on protein-tyrosine phosphatase 1B (PTP1B) activity.

Identification of Anti-Neuroinflammatory Bioactive Compounds in Essential Oils and Aqueous Distillation Residues Obtained from Commercial Varieties of Cannabis sativa L.

Neuroinflammation, which is mainly triggered by microglia, is a key contributor to multiple neurodegenerative diseases. Natural products, and in particular Cannabis sativa L., due to its richness in phytochemical components, represent ideal candidates to counteract neuroinflammation. We previously characterized different C. sativa commercial varieties which showed significantly different chemical profiles. On these bases, the aim of this study was to evaluate essential oils and aqueous distillation residues from the inflorescences of three different hemp varieties for their anti-neuroinflammatory activity in BV-2 microglial cells. Cells were pretreated with aqueous residues or essential oils and then activated with LPS. Unlike essential oils, aqueous residues showed negligible effects in terms of anti-inflammatory activity. Among the essential oils, the one obtained from ‘Gorilla Glue’ was the most effective in inhibiting pro-inflammatory mediators and in upregulating anti-inflammatory ones through the modulation of the p38 MAPK/NF-κB pathway. Moreover, the sesquiterpenes (E)-caryophyllene, α-humulene, and caryophyllene oxide were identified as the main contributors to the essential oils’ anti-inflammatory activity. To our knowledge, the anti-neuroinflammatory activity of α-humulene has not been previously described. In conclusion, our work shows that C. sativa essential oils characterized by high levels of sesquiterpenes can be promising candidates in the prevention/counteraction of neuroinflammation.

Hepialiamides A-C: Aminated Fusaric Acid Derivatives and Related Metabolites with Anti-Inflammatory Activity from the Deep-Sea-Derived Fungus Samsoniella hepiali W7.

A systematic investigation combined with a Global Natural Products Social (GNPS) molecular networking approach, was conducted on the metabolites of the deep-sea-derived fungus Samsoniella hepiali W7, leading to the isolation of three new fusaric acid derivatives, hepialiamides A-C (1-3) and one novel hybrid polyketide hepialide (4), together with 18 known miscellaneous compounds (5-22). The structures of the new compounds were elucidated through detailed spectroscopic analysis. as well as TD-DFT-based ECD calculation. All isolates were tested for anti-inflammatory activity in vitro. Under a concentration of 1 µM, compounds 8, 11, 13, 21, and 22 showed potent inhibitory activity against nitric oxide production in lipopolysaccharide (LPS)-activated BV-2 microglia cells, with inhibition rates of 34.2%, 30.7%, 32.9%, 38.6%, and 58.2%, respectively. Of particularly note is compound 22, which exhibited the most remarkable inhibitory activity, with an IC50 value of 426.2 nM.

Gastrodin Regulates PI3K/AKT-Sirt3 Signaling Pathway and Proinflammatory Mediators in Activated Microglia.

Activated microglia and their mediated inflammatory responses play an important role in the pathogenesis of hypoxic-ischemic brain damage (HIBD). Therefore, regulating microglia activation is considered a potential therapeutic strategy. The neuroprotective effects of gastrodin were evaluated in HIBD model mice, and in oxygen glucose deprivation (OGD)-treated and lipopolysaccharide (LPS)activated BV-2 microglia cells. The potential molecular mechanism was investigated using western blotting, immunofluorescence labeling, quantitative realtime reverse transcriptase polymerase chain reaction, and flow cytometry. Herein, we found that PI3K/AKT signaling can regulate Sirt3 in activated microglia, but not reciprocally. And gastrodin exerts anti-inflammatory and antiapoptotic effects through the PI3K/AKT-Sirt3 signaling pathway. In addition, gastrodin could promote FOXO3a phosphorylation, and inhibit ROS production in LPSactivated BV-2 microglia. Moreover, the level P-FOXO3a decreased significantly in Sirt3-siRNA group. However, there was no significant change after gastrodin and siRNA combination treatment. Notably, gastrodin might also affect the production of ROS in activated microglia by regulating the level of P-FOXO3a via Sirt3. Together, this study highlighted the neuroprotective role of PI3K/AKT-Sirt3 axis in HIBD, and the anti-inflammatory, anti-apoptotic, and anti-oxidative stress effects of gastrodin on HIBD.

Microglia-secreted TNF-α affects differentiation efficiency and viability of pluripotent stem cell-derived human dopaminergic precursors.

Disease is a neurodegenerative disorder characterised by the progressive loss of dopaminergic cells of the substantia nigra pars compacta. Even though successful transplantation of dopamine-producing cells into the striatum exhibits favourable effects in animal models and clinical trials; transplanted cell survival is low. Since every transplant elicits an inflammatory response which can affect cell survival and differentiation, we aimed to study in vivo and in vitro the impact of the pro-inflammatory environment on human dopaminergic precursors. We first observed that transplanted human dopaminergic precursors into the striatum of immunosuppressed rats elicited an early and sustained activation of astroglial and microglial cells after 15 days' post-transplant. This long-lasting response was associated with Tumour necrosis factor alpha expression in microglial cells. In vitro, conditioned media from activated BV2 microglial cells increased cell death, decreased Tyrosine hydroxylase-positive cells and induced morphological alterations on human neural stem cells-derived dopaminergic precursors at two differentiation stages: 19 days and 28 days. Those effects were ameliorated by inhibition of Tumour necrosis factor alpha, a cytokine which was previously detected in vivo and in conditioned media from activated BV-2 cells. Our results suggest that a pro-inflammatory environment is sustained after transplantation under immunosuppression, providing a window of opportunity to modify this response to increase transplant survival and differentiation. In addition, our data show that the microglia-derived pro-inflammatory microenvironment has a negative impact on survival and differentiation of dopaminergic precursors. Finally, Tumour necrosis factor alpha plays a key role in these effects, suggesting that this cytokine could be an interesting target to increase the efficacy of human dopaminergic precursors transplantation in Parkinson's Disease.

The bexarotene derivative OAB-14 ameliorates cognitive decline in APP/PS1 transgenic mice by suppressing microglia-mediated neuroinflammation through the PPAR-γ pathway

Neuroinflammation is believed to be a critical process involved in the pathophysiology of Alzheimer's disease (AD). In this study, we investigated the pharmacological ability of OAB-14, a small molecule compound derived from bexarotene, to reduce neuroinflammation and improve cognitive decline in an AD mouse model (in vivo) and its ability to regulate signaling pathways implicated in neuroinflammation in vitro. It was found that OAB-14 significantly improved the cognitive function of 11-month-old AD mice (APP/PS1 transgenic mice) in a dose-dependent manner. Simultaneously, OAB-14 dramatically inhibited the activation of microglia in the cerebral cortex and hippocampus of AD mice and dose-dependently downregulated the expression of nuclear factor kappa B (NF-κB) and NOD-like receptor protein 3 (NLRP3) in the cerebral cortex. At the cellular level, OAB-14 reversed the downregulation of M2 phenotypic markers, including mannose receptor C-type 1 (MRC1) and arginase 1 (ARG1), in lipopolysaccharide (LPS)- or amyloid-β protein oligomer (oAβ1-42)-activated BV2 microglial cells and partially restored their ability to clear Aβ. However, these effects were suppressed when peroxisome proliferator-activated receptor-γ (PPAR-γ) was specifically inhibited by GW9662, a selective PPAR-γ antagonist. These results suggested that OAB-14 could regulate microglial polarization by regulating PPAR-γ signaling, thereby mitigating neuroinflammation and improving cognitive function in AD mice.

Involvement of Nrf2 Activation and NF-kB Pathway Inhibition in the Antioxidant and Anti-Inflammatory Effects of Hesperetin in Activated BV-2 Microglial Cells.

Alzheimer's disease is a progressive neurodegenerative disorder leading to cognitive decline and memory loss. The incidence of this disease continues to increase due to the limited number of novel therapeutics that prevent or slow down its progression. Flavonoids have been investigated for their potential effects on cellular damage triggered by excessive reactive oxygen species (ROS) and neuroinflammatory conditions. This study investigated the effect of the flavonoid hesperetin on LPS-activated murine BV-2 microglial cells. Results show that hesperetin reduced nitric oxide levels and increased catalase, glutathione, and superoxide dismutase levels, suggesting its potential to reduce neuroinflammation and oxidative stress. Moreover, RT-PCR arrays showed that hesperetin modulated multiple genes that regulate oxidative stress. Hesperetin downregulated the mRNA expression of ERCC6, NOS2, and NCF1 and upregulated HMOX1 and GCLC. RT-PCR results showed that hesperetin-induced Nrf2 mRNA and protein expression in LPS-activated BV-2 microglial cells is involved in the transcription of several antioxidant genes, suggesting that hesperetin's antioxidant effects may be exerted via the Keap1/Nrf2 signaling pathway. Furthermore, the data demonstrated that hesperetin reduced the gene expression of PD-L1, which is upregulated as an individual ages and during chronic inflammatory processes, and inhibited the expression of genes associated with NF-kB signaling activation, which is overactivated during chronic inflammation. It was concluded from this investigation that hesperetin may have therapeutic potential to prevent or slow down the progression of neurodegenerative diseases, such as Alzheimer's disease, by reducing chronic oxidative stress and modulating neuroinflammation.

Acrylamide induces the activation of BV2 microglial cells through TLR2/4-mediated LRRK2-NFATc2 signaling cascade.

Acrylamide (ACR), a potential neurotoxin, is generated from the Maillard reaction between reducing sugars and free amino acids during food processing. Our work focuses on clarifying the role of the leucine-rich repeat kinase 2 (LRRK2) and nuclear factor of activated T cells, cytoplasmic 2 (NFATc2) in the polarization of BV2 cells to the M1 proinflammatory type induced by ACR. Specifically, ACR promoted the phosphorylation of LRRK2 and NFATc2 in BV2 microglia. Furthermore, selectively phosphorylated LRRK2 by ACR induced nuclear translocation of NFATc2 to trigger a neuroinflammatory cascade. Knock-down of LRRK2 by silencing significantly diminished ACR-induced microglial neurotoxic effect with the decline of IL-1β, IL-6, and iNOS levels and the decrease of NFATc2 expression in BV2 cells. After pretreated with Toll-Like Receptor 2 (TLR2) and TLR4 inhibitors separately, both the activation of LRRK2 and the release of pro-inflammatory factors were inhibited in BV2 cells. Gallic acid (GA) is ubiquitous in most parts of the medicinal plant. GA alleviated the increased CD11b expression, IL-6 and iNOS levels induced by ACR in BV2 microglia. In conclusion, this study shows that ACR leads to the cascade activation of LRRK2-NFATc2 mediated by TLR2 and TLR4 to induce microglial toxicity.

Chemical constituents from leaves of Jatropha curcas

ObjectiveTo investigate the chemical constituents from the leaves of Jatropha curcas and evaluate their inhibition on lipopolysaccharide (LPS)-activated BV-2 microglia cells. MethodsThe n-BuOH extract of the leaves of J. curcas was isolated by macroporous adsorption resin, silica gel, ODS, column chromatography and semi-preparative HPLC. The structures of the compounds were identified by MS, NMR, ECD, and other spectroscopic methods. In addition, anti-neuroinflammatory effects of isolated compounds were evaluated by measuring the production of nitric oxide (NO) in over-activated BV-2 cells. ResultsSeventeen compounds, including (7R,8S)-crataegifin A-4-O-β-D-glucopyranoside (1), (8R,8′R)-arctigenin (2), arctigenin-4′-O-β-D-glucopyranoside (3), (-)-syringaresinol (4), syringaresinol-4′-O-β-D-glucopyranoside (5), (-)-pinoresinol (6), pinoresinol-4′-O-β-D-glucopyranoside (7), buddlenol D (8), (2R,3R)-dihydroquercetin (9), (2S,3S)-epicatechin (10), (2R,3S)-catechin (11), isovitexin (12), naringenin-7-O-β-D-glucopyranoside (13), chamaejasmin (14), neochamaejasmin B (15), isoneochamaejasmin A (16), and tomentin-5-O-β-D-glucopyranoside (17) were isolated and identified. Compounds 2, 4 and 8 significantly inhibited the release of NO in BV-2 microglia activated by LPS, with IC50 values of 18.34, 29.33 and 26.30 μmol/L, respectively. ConclusionCompound 1 is a novel compound, and compounds 2, 3, 8, 14–17 are isolated from Jatropha genus for the first time. In addition, the lignans significantly inhibited NO release and the inhibitory activity was decreased after glycosylation.

Сyclooxygenase-2 Inhibitor Parecoxib Reduces LPS-Induced Activation of BV2 Microglia Cells.

We studied the inhibitory effect of cyclooxygenase-2 inhibitor parecoxib on LPS-induced activation of BV2 microglia cells. The optimal dose of parecoxib (80 μmol/liter) was evaluated by the Cell Counting Kit-8. The cells were divided into the following groups: control (intact cells without treatment); LPS (treatment with 1 μg/ml LPS for 6 h), and experimental (pretreatment with 80 μmol/liter parecoxib for 24 h followed by incubation with 1 μg/ml LPS for 6 h). Cell morphology and proliferation and the expression of NLRP3, caspase-1, pro-caspase-1, and IL-1β were assessed. LPS induced significant morphological changes and decreased proliferation of primary BV2 cells in comparison with the control. These changes were prevented by parecoxib pretreatment. LPS significantly increased NLRP3 inflammatory vesicle activation and expression of NLRP3, caspase-1, pro-caspase-1, and IL-1β in comparison with the control group; pretreatment with parecoxib prevented all these changes. Our results suggest that pretreatment with parecoxib inhibited LPS-induced activation of BV2 microglial cells and probably inhibited NLRP3 inflammasome activation.

Isoschaftoside Inhibits Lipopolysaccharide-Induced Inflammation in Microglia through Regulation of HIF-1α-Mediated Metabolic Reprogramming.

Isoschaftoside is a C-glycosyl flavonoid extracted from the root exudates of Desmodium uncinatum and Abrus cantoniensis. Previous studies suggested that C-glycosyl flavonoid has neuroprotective effects with the property of reducing oxidative stress and inflammatory markers. Microglia are key cellular mediators of neuroinflammation in the central nervous system. The aim of this study was to investigate the effect of isoschaftoside on lipopolysaccharide-induced activation of BV-2 microglial cells. The BV-2 cells were exposed to 10 ng/ml lipopolysaccharide and isoschaftoside (0-1000 μM). Isoschaftoside effectively inhibited lipopolysaccharide-induced nitric oxide production and proinflammatory cytokines including iNOS, TNF-α, IL-1β, and COX2 expression. Isoschaftoside also significantly reduced lipopolysaccharide-induced HIF-1α, HK2, and PFKFB3 protein expression. Induction of HIF-1α accumulation by CoCl2 was inhibited by isoschaftoside, while the HIF-1α specific inhibitor Kc7f2 mitigated the metabolic reprogramming and anti-inflammatory effect of isoschaftoside. Furthermore, isoschaftoside attenuated lipopolysaccharide-induced phosphorylation of ERK1/2 and mTOR. These results suggest that isoschaftoside can suppress inflammatory responses in lipopolysaccharide-activated microglia, and the mechanism was partly due to inhibition of the HIF-1α-mediated metabolic reprogramming pathway.

Inhibiting EGFR/HER-2 ameliorates neuroinflammatory responses and the early stage of tau pathology through DYRK1A.

The FDA-approved EGFR/HER2 inhibitor varlitinib inhibits tumor growth and is used in cancer treatment. However, the neuroinflammatory response associated with EGFR/HER2 and its underlying mechanism have not been elucidated. This study evaluates the impact of varlitinib on LPS- and tau-mediated neuroinflammatory responses for the first time. In BV2 microglial cells, varlitinib reduced LPS-stimulated il-1β and/or inos mRNA levels and downstream AKT/FAK/NF-kB signaling. Importantly, varlitinib significantly diminished LPS-mediated microglial nlrp3 inflammasome activation in BV2 microglial cells. In primary astrocytes, varlitinib downregulated LPS-evoked astroglial il-1β mRNA levels, AKT signaling, and nlrp3 inflammasome activation. In LPS-treated wild-type mice, varlitinib significantly reduced LPS-stimulated glial activation and IL-1β/NLRP3 inflammasome formation. Moreover, varlitinib significantly reduced micro- and astroglial activation and tau hyperphosphorylation in 3-month-old tau-overexpressing PS19 mice by downregulating tau kinase DYRK1A levels. However, in 6-month-old tau-overexpressing PS19 mice, varlitinib only significantly diminished astroglial activation and tau phosphorylation at Thr212/Ser214. Taken together, our findings suggest that varlitinib has therapeutic potential for LPS- and tau-induced neuroinflammatory responses and the early stages of tau pathology.

Exosomes Derived from Human Amniotic Fluid Mesenchymal Stem Cells Preserve Microglia and Neuron Cells from Aβ.

Background: Neuroinflammation is involved in neuronal cell death that occurs in neurodegenerative diseases such as Alzheimer’s disease (AD). Microglia play important roles in regulating the brain amyloid beta (Aβ) levels, so immunomodulatory properties exerted by mesenchymal stem cells may be exploited to treat this pathology. The evidence suggests that the mechanism of action of human amniotic fluid stem cells (hAFSCs) is through their secretome, which includes exosomes (exo). Methods: We examined the effect of exosomes derived from human amniotic fluid stem cells (hAFSCs-exo) on activated BV-2 microglia cells by lipopolysaccharide (LPS) as a neuroinflammation model. To investigate the exo effect on the interplay between AD neurons and microglia, SH-SY5Y neuroblastoma cells treated with Aβ were exposed to a conditioned medium (CM) obtained from activated BV-2 or co-culture systems. Results: We found that the upregulation of the markers of pro-inflammatory microglia was prevented when exposed to hAFSC-exo whereas the markers of the anti-inflammatory macrophage phenotype were not affected. Interestingly, the hAFSC-exo pretreatment significantly inhibited the oxidative stress rise and apoptosis occurring in the neurons in presence of both microglia and Aβ. Conclusion: We demonstrated that hAFSC-exo mitigated an inflammatory injury caused by microglia and significantly recovered the neurotoxicity, suggesting that hAFSC-exo may be a potential therapeutic agent for inflammation-related neurological conditions, including AD.

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.