Engeletin alleviates depressive-like behaviours by modulating microglial polarization via the LCN2/CXCL10 signalling pathway.

Microglial polarization and associated inflammatory activity are the key mediators of depression pathogenesis. The natural Smilax glabra rhizomilax derivative engeletin has been reported to exhibit robust anti-inflammatory activity, but no studies to date have examined the mechanisms through which it can treat depressive symptoms. We showed that treatment for 21 days with engeletin significantly alleviated depressive-like behaviours in chronic stress social defeat stress (CSDS) model mice. T1-weighted imaging (T1WI), T2-weighted imaging (T2WI) imaging revealed no significant differences between groups, but the bilateral prefrontal cortex of CSDS mice exhibited significant increases in apparent diffusion coefficient and T2 values relative to normal control mice, with a corresponding reduction in fractional anisotropy, while engeletin reversed all of these changes. CSDS resulted in higher levels of IL-1ß, IL-6, and TNF-a production, enhanced microglial activation, and greater M1 polarization with a concomitant decrease in M2 polarization in the mPFC, whereas engeletin treatment effectively abrogated these CSDS-related pathological changes. Engeletin was further found to suppress the LCN2/C-X-C motif chemokine ligand 10 (CXCL10) signalling axis such that adeno-associated virus-induced LCN2 overexpression ablated the antidepressant effects of engeletin and reversed its beneficial effects on the M1/M2 polarization of microglia. In conclusion, engeletin can alleviate CSDS-induced depressive-like behaviours by regulating the LCN2/CXCL10 pathway and thereby altering the polarization of microglia. These data suggest that the antidepressant effects of engeletin are correlated with the polarization of microglia, highlighting a potential avenue for future design of antidepressant strategies that specifically target the microglia.

A2 reactive astrocyte-derived exosomes alleviate cerebral ischemia-reperfusion injury by delivering miR-628.

Ischemia and hypoxia activate astrocytes into reactive types A1 and A2, which play roles in damage and protection, respectively. However, the function and mechanism of A1 and A2 astrocyte exosomes are unknown. After astrocyte exosomes were injected into the lateral ventricle, infarct volume, damage to the blood-brain barrier (BBB), apoptosis and the expression of microglia-related proteins were measured. The dual luciferase reporter assay was used to detect the target genes of miR-628, and overexpressing A2-Exos overexpressed and knocked down miR-628 were constructed. qRT-PCR, western blotting and immunofluorescence staining were subsequently performed. A2-Exos obviously reduced the infarct volume, damage to the BBB and apoptosis and promoted M2 microglial polarization. RT-PCR showed that miR-628 was highly expressed in A2-Exos. Dual luciferase reporter assays revealed that NLRP3, S1PR3 and IRF5 are target genes of miR-628. After miR-628 was overexpressed or knocked down, the protective effects of A2-Exos increased or decreased, respectively. A2-Exos reduced pyroptosis and BBB damage and promoted M2 microglial polarization through the inhibition of NLRP3, S1PR3 and IRF5 via the delivery of miR-628. This study explored the mechanism of action of A2-Exos and provided new therapeutic targets and concepts for treating cerebral ischemia.

Polyphyllin I alleviates neuroinflammation after cerebral ischemia-reperfusion injury via facilitating autophagy-mediated M2 microglial polarization.

Microglial activation and polarization play a central role in poststroke inflammation and neuronal damage. Modulating microglial polarization from pro-inflammatory to anti-inflammatory phenotype is a promising therapeutic strategy for the treatment of cerebral ischemia. Polyphyllin I (PPI), a steroidal saponin, shows multiple bioactivities in various diseases, but the potential function of PPI in cerebral ischemia is not elucidated yet. In our study, the influence of PPI on cerebral ischemia-reperfusion injury was evaluated. Mouse middle cerebral artery occlusion (MCAO) model and oxygen-glucose deprivation and reoxygenation (OGD/R) model were constructed to mimic cerebral ischemia-reperfusion injury in vivo and in vitro. TTC staining, TUNEL staining, RT-qPCR, ELISA, flow cytometry, western blot, immunofluorescence, hanging wire test, rotarod test and foot-fault test, open-field test and Morris water maze test were performed in our study. We found that PPI alleviated cerebral ischemia-reperfusion injury and neuroinflammation, and improved functional recovery of mice after MCAO. PPI modulated microglial polarization towards anti-inflammatory M2 phenotype in MCAO mice in vivo and post OGD/R in vitro. Besides, PPI promoted autophagy via suppressing Akt/mTOR signaling in microglia, while inhibition of autophagy abrogated the effect of PPI on M2 microglial polarization after OGD/R. Furthermore, PPI facilitated autophagy-mediated ROS clearance to inhibit NLRP3 inflammasome activation in microglia, and NLRP3 inflammasome reactivation by nigericin abolished the effect of PPI on M2 microglia polarization. In conclusion, PPI alleviated post-stroke neuroinflammation and tissue damage via increasing autophagy-mediated M2 microglial polarization. Our data suggested that PPI had potential for ischemic stroke treatment.

CD36 deletion prevents white matter injury by modulating microglia polarization through the Traf5-MAPK signal pathway.

BACKGROUND: White matter injury (WMI) represents a significant etiological factor contributing to neurological impairment subsequent to Traumatic Brain Injury (TBI). CD36 receptors are recognized as pivotal participants in the pathogenesis of neurological disorders, including stroke and spinal cord injury. Furthermore, dynamic fluctuations in the phenotypic polarization of microglial cells have been intimately associated with the regenerative processes within the injured tissue following TBI. Nevertheless, there is a paucity of research addressing the impact of CD36 receptors on WMI and microglial polarization. This investigation aims to elucidate the functional role and mechanistic underpinnings of CD36 in modulating microglial polarization and WMI following TBI. METHODS: TBI models were induced in murine subjects via controlled cortical impact (CCI). The spatiotemporal patterns of CD36 expression were examined through quantitative polymerase chain reaction (qPCR), Western blot analysis, and immunofluorescence staining. The extent of white matter injury was assessed via transmission electron microscopy, Luxol Fast Blue (LFB) staining, and immunofluorescence staining. Transcriptome sequencing was employed to dissect the molecular mechanisms underlying CD36 down-regulation and its influence on white matter damage. Microglial polarization status was ascertained using qPCR, Western blot analysis, and immunofluorescence staining. In vitro, a Transwell co-culture system was employed to investigate the impact of CD36-dependent microglial polarization on oligodendrocytes subjected to oxygen-glucose deprivation (OGD). RESULTS: Western blot and qPCR analyses revealed that CD36 expression reached its zenith at 7 days post-TBI and remained sustained at this level thereafter. Immunofluorescence staining exhibited robust CD36 expression in astrocytes and microglia following TBI. Genetic deletion of CD36 ameliorated TBI-induced white matter injury, as evidenced by a reduced SMI-32/MBP ratio and G-ratio. Transcriptome sequencing unveiled differentially expressed genes enriched in processes linked to microglial activation, regulation of neuroinflammation, and the TNF signaling pathway. Additionally, bioinformatics analysis pinpointed the Traf5-p38 axis as a critical signaling pathway. In vivo and in vitro experiments indicated that inhibition of the CD36-Traf5-MAPK axis curtailed microglial polarization toward the pro-inflammatory phenotype. In a Transwell co-culture system, BV2 cells treated with LPS + IFN-γ exacerbated the damage of post-OGD oligodendrocytes, which could be rectified through CD36 knockdown in BV2 cells. CONCLUSIONS: This study illuminates that the suppression of CD36 mitigates WMI by constraining microglial polarization towards the pro-inflammatory phenotype through the down-regulation of the Traf5-MAPK signaling pathway. Our findings present a potential therapeutic strategy for averting neuroinflammatory responses and ensuing WMI damage resulting from TBI.

The activation impact of lactobacillus-derived extracellular vesicles on lipopolysaccharide-induced microglial cell.

Perioperative neurocognitive dysfunction (PND) emerges as a common postoperative complication among elderly patients. Currently, the mechanism of PND remains unclear, but there exists a tendency to believe that inflammation plays a significant role in PND. Alterations in the abundance of intestinal microbiota can increase the permeability of the intestinal mucosal barrier and incite extraintestinal inflammatory responses. Metabolites from these microbiota can be absorbed by the intestinal mucosa into the bloodstream, exerting influence upon the central nervous system (CNS). Lactobacillus (Lac), serving as an intestinal probiotic bacterium, possesses the capacity to modulate emotional behavior and cognitive functions. Extracellular vesicles (EVs) are recognized as novel therapeutic carriers for targeted delivery to regulate physiology and pathogenesis. While the mechanism governing the primary function of Lac-EVs in the CNS remains uncertain. Therefore, we established an in vitro neuroinflammation model to induce PND and then treated the mice with Lac-EVs to observe the effect of these EVs on neuroinflammation, particularly on microglial (MG) polarization. Our research unveils that Lac-EVs reduced inflammation induced by LPS in microglia and the activation of related proteins, including the mRNA expression of M1 labeled protein (iNOS). Moreover, the mRNA expression of M2-labeled protein (Arg1) increased. In addition, flow cytometry revealed that the ratio of M1/M2 microglia also changed significantly. Therefore, Lac-EVs promoted the differentiation of M2 microglia by inducing the preferential expression of specific markers related to M2 macrophages and inflammation. In terms of inflammatory cytokine expression, Lac-EVs decreased the secretion of proinflammatory cytokines (IL-1ß and IL-6) and increased IL-10 production after lipopolysaccharide (LPS) stimulation. Therefore, Lac-EVs induce the activation of M2 microglial cells without inducing cellular harm in vitro, and they demonstrate anti-inflammatory effects against lipopolysaccharide-induced neuroinflammation. This finding suggested that it is an effective anti-inflammatory strategy for alleviating inflammation-driven PNDs.

Neutrophil extracellular traps facilitate sympathetic hyperactivity by polarizing microglia toward M1 phenotype after traumatic brain injury.

Traumatic brain injury (TBI), particularly diffuse axonal injury (DAI), often results in sympathetic hyperactivity, which can exacerbate the prognosis of TBI patients. A key component of this process is the role of neutrophils in causing neuroinflammation after TBI by forming neutrophil extracellular traps (NETs), but the connection between NETs and sympathetic excitation following TBI remains unclear. Utilizing a DAI rat model, the current investigation examined the role of NETs and the HMGB1/JNK/AP1 signaling pathway in this process. The findings revealed that sympathetic excitability intensifies and peaks 3 days post-injury, a pattern mirrored by the activation of microglia, and the escalated NETs and HMGB1 levels. Subsequent in vitro exploration validated that HMGB1 fosters microglial activation via the JNK/AP1 pathway. Moreover, in vivo experimentation revealed that the application of anti-HMGB1 and AP1 inhibitors can mitigate microglial M1 polarization post-DAI, effectively curtailing sympathetic hyperactivity. Therefore, this research elucidates that post-TBI, NETs within the PVN may precipitate sympathetic hyperactivity by stimulating M1 microglial polarization through the HMGB1/JNK/AP1 pathway.

Knockdown of trem2 promotes proinflammatory microglia and inhibits glioma progression via the JAK2/STAT3 and NF-κB pathways.

BACKGROUND: In the tumor immune microenvironment (TIME), triggering receptor expressed on myeloid cells 2 (trem2) is widely considered to be a crucial molecule on tumor-associated macrophages(TAMs). Multiple studies have shown that trem2 may function as an immune checkpoint in various malignant tumors, mediating tumor immune evasion. However, its specific molecular mechanisms, especially in glioma, remain elusive. METHODS: Lentivirus was transfected to establish cells with stable knockdown of trem2. A Transwell system was used for segregated coculture of glioma cells and microglia. Western blotting, quantitative real-time polymerase chain reaction (qRT‒PCR), and immunofluorescence (IF) were used to measure the expression levels of target proteins. The proliferation, invasion, and migration of cells were detected by colony formation, cell counting kit-8 (CCK8), 5-ethynyl-2'-deoxyuridine (EdU) and transwell assays. The cell cycle, apoptosis rate and reactive oxygen species (ROS) level of cells were assessed using flow cytometry assays. The comet assay and tube formation assay were used to detect DNA damage in glioma cells and angiogenesis activity, respectively. Gl261 cell lines and C57BL/6 mice were used to construct the glioma orthotopic transplantation tumor model. RESULTS: Trem2 was highly overexpressed in glioma TAMs. Knocking down trem2 in microglia suppressed the growth and angiogenesis activity of glioma cells in vivo and in vitro. Mechanistically, knockdown of trem2 in microglia promoted proinflammatory microglia and inhibited anti-inflammatory microglia by activating jak2/stat1 and inhibiting the NF-κB p50 signaling pathway. The proinflammatory microglia produced high concentrations of nitric oxide (NO) and high levels of the proinflammatory cytokines TNF-α, IL-6, and IL-1ß, and caused further DNA damage and promoted the apoptosis rate of tumor cells. CONCLUSIONS: Our findings revealed that trem2 in microglia plays a significant role in the TIME of gliomas. Knockdown of trem2 in microglia might help to improve the efficiency of inhibiting glioma growth and delaying tumor progression and provide new ideas for further treatment of glioma.

Spinal microglial M1 polarization contributes paclitaxel-induced neuropathic pain by triggering cells necroptosis.

Paclitaxel (PTX) is a chemotherapeutic agent that is widely used for the treatment of several types of tumors. However, PTX-induced peripheral neuropathy (PIPN) is an adverse effect generally induced by long-term PTX use that significantly impairs the quality of life. Necroptosis has been implicated in various neurodegenerative disorders. Necroptosis of dorsal root ganglion neurons triggers the pathogenesis of PIPN. Therefore, the present study aims to investigate the role of spinal neuronal necroptosis in PIPN. It also explores the potential role of microglial polarization in necroptosis. We established rat models of PIPN via quartic PTX administration on alternate days (accumulated dose: 8 mg/kg). PTX induced obvious neuronal necroptosis and upregulated the expression of receptor-interacting protein kinase (RIP3) and mixed lineage kinase domain-like protein (MLKL) in the spinal dorsal horn. These effects were inhibited with a necroptosis pathway inhibitor, necrostatin-1 (Nec-1). The effect of microglial polarization on the regulation of spinal necroptosis was elucidated by administering minocycline to inhibit PTX-induced M1 polarization of spinal microglia caused by PTX. We observed a significant inhibitory effect of minocycline on PTX-induced necroptosis in spinal cord cells, based on the downregulation of RIP3 and MLKL expression, and suppression of tumor necrosis factor-α and IL-ß synthesis. Additionally, minocycline improved hyperalgesia symptoms in PIPN rats. Overall, this study suggests that PTX-induced polarization of spinal microglia leads to RIP3/MLKL-regulated necroptosis, resulting in PIPN. These findings suggest a potential target for the prevention and treatment of neuropathic pain.

Mxene-bpV plays a neuroprotective role in cerebral ischemia-reperfusion injury by activating the Akt and promoting the M2 microglial polarization signaling pathways.

Studies have shown that the inhibition of phosphatase and tensin homolog deleted on chromosome 10 (PTEN)was neuroprotective against ischemia/reperfusion(I/R) injury. Bisperoxovanadium (bpV), a derivative of vanadate, is a well-established inhibitor of PTEN. However, its function islimited due to its general inadequacy in penetrating cell membranes. Mxene(Ti3C2Tx) is a novel two-dimensional lamellar nanomaterial with an excellent ability to penetrate the cell membrane. Yet, the effects of this nanomaterial on nervous system diseases have yet to be scrutinized. Here, Mxene(Ti3C2Tx) was used for the first time to carry bpV(HOpic), creating a new nanocomposite Mxene-bpV that was probed in a cerebral I/R injury model. The findings showed that this synthetic Mxene-bpV was adequately stable and can cross the cell membraneeasily. We observed that Mxene-bpV treatment significantly increased the survival rate of oxygen glucose deprivation/reperfusion(OGD/R)--insulted neurons, reduced infarct sizes and promoted the recovery of brain function after mice cerebral I/R injury. Crucially, Mxene-bpV treatment was more therapeutically efficient than bpV(HOpic) treatment alone over the same period. Mechanistically, Mxene-bpV inhibited the enzyme activity of PTEN in vitro and in vivo. It also promoted the expression of phospho-Akt (Ser473) by repressing PTEN and then activated the Akt pathway to boost cell survival. Additionally, in PTEN transgenic mice, Mxene-bpV suppressed I/R-induced inflammatory response by promoting M2 microglial polarization through PTEN inhibition. Collectively, the nanosynthetic Mxene-bpV inhibited PTEN' enzymatic activity by activating Akt pathway and promoting M2 microglial polarization, and finally exerted neuroprotection against cerebral I/R injury.

M1 polarization induction by lead and amyloid peptides in microglial cells: Implications for neurodegeneration process.

Neurodegeneration in conditions like Alzheimer's and Parkinson's disease is influenced by genetic and environmental factors. This study explores the potential neurodegenerative effects of lead (Pb) toxicity and amyloid beta peptides (Aßp 1-40 and Aßp 25-35) by promoting M1 polarization in microglial cells. To this end, we investigated and observed that IC50 concentrations of Pb (22.8 µM) and Aßp 25-35(29.6 µM). Our results demonstrated significant Pb uptake (31.13% at 25 µM Pb) and increased intracellular ROS levels (77.1%) upon treatment with Pb in combination of both Aßp 1-40 and Aßp 25-35. Protein carbonylation significantly increased (73.12 nmol/mL) upon treatment with Pb in combination of both Aßp 1-40 and Aßp 25-35, indicating oxidative damage and compromised cellular defenses against oxidative stress along with elevated DNA oxidative damage (164.9 pg/mL of 8-OH-dG) upon treatment with Pb in combination with both Aßp 1-40 and Aßp 25-35. Microglial polarization showed elevated M1 markers (inducible nitric oxide synthase and cyclooxygenase 2) and reduced M2 markers (arginase-1 and cluster of differentiation 206), suggesting Pb's role in inducing neurodegenerative microglial polarization. These findings provide insights into the complex molecular events contributing to Pb-induced neurotoxicity and neurodegenerative diseases.

Electroacupuncture attenuates middle cerebral artery occlusion-induced learning and memory impairment by regulating microglial polarization in hippocampus.

BACKGROUND: As a traditional medical therapy, electroacupuncture (EA) has been demonstrated to have beneficial effects on ischemic stroke-induced cognitive impairment. However, the underlying mechanism is largely unclear. METHODS: Adult rats received occlusion of the middle cerebral artery and reperfusion (MCAO/R) to establish the ischemic stroke model. Morris water maze test was performed following EA stimulation at the GV20, PC6, and KI1 acupoints in rats to test the learning and memory ability. Western blot, immunofluorescent staining, and enzyme-linked immunosorbent assay were conducted to assess the cellular and molecular mechanisms. RESULTS: EA stimulation attenuated neurological deficits. In the Morris water maze test, EA treatment ameliorated the MCAO/R-induced learning and memory impairment. Moreover, we observed that MCAO/R induced microglial activation and polarization in the ischemic hippocampus, whereas, EA treatment dampened microglial activation and inhibited M1 microglial polarization but enhanced M2 microglial polarization. EA treatment inhibited the increased expression of proinflammatory cytokines and enhanced the increased expression of anti-inflammatory cytokines. Finally, we found that EA treatment dampened microglial p38 mitogen-activated protein kinase (MAPK) phosphorylation. CONCLUSION: Collectively, our data suggested that EA treatment ameliorated cognitive impairment induced by MCAO/R and the underlying mechanism may be p38-mediated microglia polarization and neuroinflammation.

Circadian modulation of microglial physiological processes and immune responses.

Microglia is considered the central nervous system (CNS) resident macrophages that establish an innate immune response against pathogens and toxins. However, the recent studies have shown that microglial gene and protein expression follows a circadian pattern; several immune activation markers and clock genes are expressed rhythmically without the need for an immune stimulus. Furthermore, microglia responds to an immune challenge with different magnitudes depending on the time of the day. This review examines the circadian control of microglia function and the possible physiological implications. For example, we discuss that synaptic prune is performed in the cortex at a certain moment of the day. We also consider the implications of daily microglial function for maintaining biological rhythms like general activity, body temperature, and food intake. We conclude that the developmental stage, brain region, and pathological state are not the only factors to consider for the evaluation of microglial functions; instead, emerging evidence indicates that circadian time as an essential aspect for a better understanding of the role of microglia in CNS physiology.

METTL14 regulates microglia/macrophage polarization and NLRP3 inflammasome activation after ischemic stroke by the KAT3B-STING axis.

N6-methyladenosine (m6A) plays a crucial role in ischemic stroke, whereas the role of methyltransferase-like 14 (METTL14) in ischemic stroke remains unknown. A model of middle cerebral artery occlusion (MCAO) in rats and oxygen-glucose deprivation/reperfusion (OGD/R) model in HAPI cells were used to simulate ischemic stroke in vivo and in vitro. We found that METTL14 level was upregulated in microglia/macrophage after MCAO and OGD/R. METTL14 enhanced the expression of KAT3B by promoting the m6A modification of KAT3B mRNA. STING has been identified as a target for KAT3B and KAT3B increased STING expression by enhancing H3K27ac in the STING promoter. METTL14 promoted M1 polarization and NLRP3 inflammasome/pyroptosis axis by the KAT3B-STING signaling after OGD/R. METTL14 depletion relieved brain injury by inhibiting M1-like microglia/macrophage polarization and NLRP3 inflammasome/pyroptosis axis in MCAO rats. These findings indicate that METTL14 depletion relieves MCAO-induced brain injury, probably via switching microglia/macrophage from M1 towards M2 and restraining NLRP3 inflammasome/pyroptosis axis in microglia/macrophage.

Nanoparticulate MgH2 ameliorates anxiety/depression-like behaviors in a mouse model of multiple sclerosis by regulating microglial polarization and oxidative stress.

Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system (CNS). Anxiety and depression are the most common psychiatric comorbidities of MS, which seriously affect patients' quality of life, treatment compliance, and prognosis. However, current treatments for anxiety and depression in MS show low therapeutic efficacy and significant side effects. In the present study, we explored the therapeutic effects of a novel low-toxic anti-inflammatory drug, nanoparticulate magnesium hydride (MgH2), on mood disorders of MS. We observed that anxiety/depression-like behaviors in experimental autoimmune encephalomyelitis (EAE) mice were alleviated by MgH2 treatment. In addition, disease severity and inflammatory demyelination were also diminished. Furthermore, we confirmed the suppressive effect of MgH2 on depression in the acute restraint stress model. Mechanistically, MgH2 may play a therapeutic role by promoting microglial M2 polarization, inhibiting microglial M1 polarization, and reducing oxidative stress and mitochondrial damage. Therefore, nanoparticulate MgH2 may be a promising therapeutic drug for psychiatric comorbidities of MS.

Multifunctional Supramolecular Vesicles as Zn2+ -Triggered Microglial Modulator Alleviates Alzheimer's Disease.

Zn2+ -induced ß-amyloid protein (Aß) aggregation and microglia activation are the predominant contributors in Alzheimer's disease (AD). Regulating intracephalic excessive Zn2+ is a promising therapeutic strategy for AD treatment. However, only inhibition of Zn2+ is hardly to repair continuous damages caused by activated microglia. Herein, an intelligent resveratrol-loaded supramolecular vesicles (RES-loaded vesicles) with zinc ion chelation function and responsive release capability are constructed to alleviate Aß fibrillation, oxidative stress, and microglial dysfunction. The resveratrol encapsulation efficiency and drug loading efficiency are calculated to be 49.67% and 7.87%, respectively. In vitro studies demonstrate that the RES-loaded vesicles can modulate Zn2+ -dependent Aß aggregation. More importantly, the cargoes will be released in zinc environment and further reprograms microglia from proinflammatory M1 phenotype toward anti-inflammatory M2 phenotype, which prevents spontaneous neuroinflammation and alleviates cytotoxicity of cultured cells from 29% to 12%. With the stereotactic or intranasal administration, RES-loaded vesicles can overcome the blood brain barrier, alleviate neuronal apoptosis, neuroinflammation, and ultimately ameliorate cognitive impairment in two AD mouse models. This work provides a new sight for taking advantage of Zn2+ to treat CNS disorders.

Muscone promotes functional recovery by facilitating microglia polarization into M2 phenotype through PPAR-γ pathway after ischemic stroke.

Exploring regimens to facilitate microglia transformation from M1 to M2 phenotype is a feasible strategy to suppress neuroinflammation, therefore reinforcing functional recovery after ischemic stroke. Muscone easily crosses the blood brain barrier (BBB) and distributes throughout the brain. Here, the results illustrated the administration of 8 mg/kg muscone promoted functional recovery through reducing the infarct volume by 2,3,5-triphenyltetrazolium chloride (TTC) staining after ischemic stroke in mice. Then, the expression of pro-inflammatory factors, such as tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß) and interleukin-6 (IL-6), was significantly decreased, whereas the level of anti-inflammatory agents including C-X-C Motif Chemokine Ligand 1 (CXCL1), transforming growth factor-ß (TGF-ß) and interleukin-10 (IL-10) was obviously elevated in penumbra with the treatment of 8 mg/kg muscone using real-time fluorescence quantitative polymerase chain reaction (RT-qPCR), western blot and enzyme-linked immunosorbent assay (ELISA) tests. Subsequently, the results showed the application of muscone upregulated the expression of peroxisome proliferator-activated receptor-γ (PPAR-γ) to facilitate microglia transformation into M2 phenotype using RT-qPCR, western blot and immunofluorescence analysis. Collectively, the present study provides evidence for our hypothesis that muscone intensifies microglia transformation into M2 phenotype via activating PPAR-γ signaling pathway in penumbra after ischemic stroke. These findings demonstrate muscone is a promising candidate for the treatment of ischemic stroke.

Possible Implications of Obesity-Primed Microglia that Could Contribute to Stroke-Associated Damage.

Microglia, the resident macrophages of the central nervous system, are essential players during physiological and pathological processes. Although they participate in synaptic pruning and maintenance of neuronal circuits, microglia are mainly studied by their activity modulating inflammatory environment and adapting their phenotype and mechanisms to insults detected in the brain parenchyma. Changes in microglial phenotypes are reflected in their morphology, membrane markers, and secreted substances, stimulating neighbor glia and leading their responses to control stimuli. Understanding how microglia react in various microenvironments, such as chronic inflammation, made it possible to establish therapeutic windows and identify synergic interactions with acute damage events like stroke. Obesity is a low-grade chronic inflammatory state that gradually affects the central nervous system, promoting neuroinflammation development. Obese patients have the worst prognosis when they suffer a cerebral infarction due to basal neuroinflammation, then obesity-induced neuroinflammation could promote the priming of microglial cells and favor its neurotoxic response, potentially worsening patients' prognosis. This review discusses the main microglia findings in the obesity context during the course and resolution of cerebral infarction, involving the temporality of the phenotype changes and balance of pro- and anti-inflammatory responses, which is lost in the swollen brain of an obese subject. Obesity enhances proinflammatory responses during a stroke. Obesity-induced systemic inflammation promotes microglial M1 polarization and priming, which enhances stroke-associated damage, increasing M1 and decreasing M2 responses.

Anti-inflammatory Action of BT75, a Novel RARα Agonist, in Cultured Microglia and in an Experimental Mouse Model of Alzheimer's Disease.

BT75, a boron-containing retinoid, is a novel retinoic acid receptor (RAR)α agonist synthesized by our group. Previous studies indicated that activation of retinoic acid (RA) signaling may attenuate progression of Alzheimer's disease (AD). Presently, we aimed to examine the anti-inflammatory effect of BT75 and explore the possible mechanism using cultured cells and an AD mouse model. Pretreatment with BT75 (1-25 µM) suppressed the release of nitric oxide (NO) and IL-1ß in the culture medium of mouse microglial SIM-A9 cells activated by LPS. BMS195614, an RARα antagonist, partially blocked the inhibition of NO production by BT75. Moreover, BT75 attenuated phospho-Akt and phospho-NF-κB p65 expression augmented by LPS. In addition, BT75 elevated arginase 1, IL-10, and CD206, and inhibited inducible nitric oxide synthase (iNOS) and IL-6 formation in LPS-treated SIM-A9 cells, suggesting the promotion of M1-M2 microglial phenotypic polarization. C57BL/6 mice were injected intracerebroventricularly (icv) with streptozotocin (STZ) (3 mg/kg) to provide an AD-like mouse model. BT75 (5 mg/kg) or the vehicle was intraperitoneally (ip) injected to icv-STZ mice once a day for 3 weeks. Immunohistochemical analyses indicated that GFAP-positive cells and rod or amoeboid-like Iba1-positive cells, which increased in the hippocampal fimbria of icv-STZ mice, were reduced by BT75 treatment. Western blot results showed that BT75 decreased levels of neuronal nitric oxide synthase (nNOS), GFAP, and phosphorylated Tau, and increased levels of synaptophysin in the hippocampus of icv-STZ mice. BT75 may attenuate neuroinflammation by affecting the Akt/NF-κB pathway and microglial M1-M2 polarization in LPS-stimulated SIM-A9 cells. BT75 also reduced AD-like pathology including glial activation in the icv-STZ mice. Thus, BT75 may be a promising anti-inflammatory and neuroprotective agent worthy of further AD studies.

A combination of umbilical cord mesenchymal stem cells and monosialotetrahexosy 1 ganglioside alleviates neuroinflammation in traumatic brain injury.

Neuro-inflammation and activated microglia play important roles in neuron damage in the traumatic brain injury (TBI). In this study, we determined the effect of neural network reconstruction after human umbilical cord mesenchymal stem cells (UMSCs) combined with monosialotetrahexosy 1 ganglioside (GM1) transplantation and the effect on the neuro-inflammation and polarization of microglia in a rat model of TBI, which was established in male rats using a fluid percussion brain injury device. Rats survived until day 7 after TBI were randomly treated with normal control (NC), saline (NS), GM1, UMSCs, and GM1 plus UMSCs. Modified neurological severity score (mNSS) was assessed on days 7 and 14, and the brain tissue of the injured region was collected. Immunofluorescence, RT-PCR, and western blot analysis found that inhibitory neuro-inflammatory cytokines TGF-ß and CD163 protein expression levels in injured brain tissues were significantly increased in rats treated with GM1 + UMSCs, GM1, or UMSCs and were up-regulated compared to saline-treated rats. Neuro-inflammatory cytokines IL-6, COX-2 and iNOS protein expressions were down-regulated compared to rats treated with saline. The protein expression levels of NE, NF-200, MAP-2 and ß-tubulin III were increased in the injured brain tissues from rats treated with GM1 + UMSCs, or GM1 and UMSCs alone compared to those in the rats treated with NS. The protein expression levels in rats treated with GM1 plus UMSCs were most significant on day 7 following UMSC transplantation. The rats treated with GM1 plus UMSCs had the lowest mNSS compared with that in the other groups. These data suggest that UMSCs and GM1 promote neural network reconstruction and reduce the neuro-inflammation and neurodegeneration through coordinating injury local immune inflammatory microenvironment to promote the recovery of neurological functions in the TBI.

PPAR-γ promotes the polarization of rat retinal microglia to M2 phenotype by regulating the expression of CD200-CD200R1 under hypoxia.

BACKGROUND: Recent reports suggest that peroxisome proliferator-activated receptor-γ (PPAR-γ) could promote microglial M2 polarization to inhibit inflammation. However, the specific molecular mechanisms that trigger PPAR-γ's anti-inflammatory ability in microglia are yet to be expounded. Thus, in this study, we aimed to explore the molecular mechanisms behind the anti-inflammatory effects of PPAR-γ in hypoxia-stimulated rat retinal microglial cells. METHODS AND RESULTS: We used shRNA expressing lentivirus to knock down PPAR-γ and CD200 genes, and we assessed hypoxia-induced polarization markers release - M1 (iNOS, IL-1ß, IL-6, and TNF-α) and M2 (Arg-1, YM1, IL-4, and IL-10) by RT-PCR. We also monitored PPAR-γ-related signals (PPAR-γ, PPAR-γ in cytoplasm or nucleus, CD200, and CD200Rs) by Western blot and RT-PCR. Our results showed that hypoxia enhanced PPAR-γ and CD200 expressions in microglial cells. Moreover, PPAR-γ agonist 15d-PGJ2 elevated CD200 and CD200R1 expressions, whereas sh-PPAR-γ had the opposite effect. Following hypoxia, expressions of M1 markers increased significantly, while those of M2 markers decreased, and the above effects were attenuated by 15d-PGJ2. Conversely, knocking down PPAR-γ or CD200 inhibited the polarization of microglial cells to M2 phenotype. CONCLUSION: Our findings demonstrated that PPAR-γ performed an anti-inflammatory function in hypoxia-stimulated microglial cells by promoting their polarization to M2 phenotype via the CD200-CD200R1 pathway.