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ObjectiveTo investigate the mechanism of Baihuan Xiaoyao Decoction （Xiaoyaosan added with Lilii Bulbus and Albiziae Cortex） in alleviating depression-like behaviors of juvenile rats by regulating the polarization of microglia. MethodSixty juvenile SD rats were randomized into normal control， model， fluoxetine， and low-， medium-， and high-dose （5.36， 10.71， 21.42 g·kg-1， respectively） Baihuan Xiaoyao decoction groups. The rat model of juvenile depression was established by chronic unpredictable mild stress （CUMS）. The sucrose preference test （SPT） was carried out to examine the sucrose preference of rats. Forced swimming test （FST） was carried out to measure the immobility time of rats. The open field test （OFT） was conducted to measure the total distance， the central distance， the number of horizontal crossings， and the frequency of rearing. Morris water maze （MWM） was used to measure the escape latency and the number of crossing the platform. The immunofluorescence assay was employed to detect the expression of inducible nitric oxide synthase （iNOS， the polarization marker of M1 microglia） and CD206 （the polarization marker of M2 microglia）. Real-time polymerase chain reaction was employed to determine the mRNA levels of iNOS， CD206， pro-inflammatory cytokines ［tumor necrosis factor （TNF）-α， interleukin （IL）-1β， and IL-6］ and anti-inflammatory cytokines （IL-4 and IL-10） in the hippocampus. Western blotting was employed to determine the protein levels of iNOS and CD206 in the hippocampus. The levels of IL-4 and IL-6 in the hippocampus were detected by enzyme-linked immunosorbent assay. ResultCompared with the normal control group， the model rats showed a reduction in sucrose preference （P<0.05）， an increase in immobility time （P<0.05）， decreased motor and exploratory behaviors （P<0.05）， and weakened learning and spatial memory （P<0.05）. In addition， the model rats showed up-regulated mRNA and protein levels of iNOS and mRNA levels of IL-1β， IL-6， and TNF-α （P<0.05）. Compared with the model group， Baihuan Xiaoyao decoction increased the sucrose preference value （P<0.05）， shortened the immobility time （P<0.01）， increased the motor and exploratory behaviors （P<0.05）， and improved the learning and spatial memory （P<0.01）. Furthermore， the decoction down-regulated the positive expression and protein level of iNOS， lowered the levels of TNF-α， IL-1β， and IL-6 （P<0.01）， promoted the positive expression of CD206， and elevated the levels of IL-4 and IL-10 （P<0.01） in the hippocampus of the high dose group. Moreover， the high-dose Baihuan Xiaoyao decoction group had higher sucrose preference value （P<0.01）， shorter immobility time （P<0.01）， longer central distance （P<0.01）， stronger learning and spatial memory （P<0.01）， higher positive expression and protein level of iNOS （P<0.01）， lower levels of TNF-α， IL-1β， and IL-6 （P<0.05， P<0.01）， lower positive expression and mRNA level of iNOS （P<0.05）， and higher levels of IL-4 and IL-10 （P<0.05， P<0.01） than the fluoxetine group. ConclusionBaihuan Xiaoyao decoction can improve the depression-like behavior of juvenile rats by inhibiting the M1 polarization and promoting the M2 polarization of microglia in the hippocampus.
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As resident immune cells of the retina, retinal microglia constantly monitor the changes of their surroundings and maintain homeostasis through signal transduction with other retinal cells. Retinal microglia play a crucial role not only in the development and physiological processes of the retinal vascular system, but also in pathological neovascularization. In certain retinopathies, activated microglia can stimulate abnormal angiogenesis through neurovascular coupling, leading to irreversible damage. However, the exact mechanisms underlying this process are still unclear. In this review, a brief overview of the relationship between microglia and retinal neovascularization was provided, and the involved cellular and molecular signaling mechanisms were reviewed, aiming to offer new and effective strategies for the prevention and treatment of retinal neovascularization diseases.
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ObjectiveTo study whether Chaihu Longgu Mulitang can inhibit hypothalamic inflammation， mitigate anxiety-like behavior， and alleviate anxiety symptoms by regulating the p38 mitogen-activated protein kinase/nuclear factor-κB （p38 MAPK/NF-κB） signaling pathway in the rat model of generalized anxiety disorder （GAD）. MethodTwelve out of 74 Wistar rats were randomly selected as the blank group， and the remaining rats were subjected to chronic restraint stress for the modeling of GAD. The open field test （OFT） and elevated Porteus maze test （PMT） were conducted 14 days after modeling to detect the anxiety-like behaviors. Sixty successfully modeled rats were selected and randomized into model， low-， medium-， and high-dose （6， 12， and 24 g·kg-1， respectively） Chaihu Longgu Mulitang， and diazepam （1 mg·kg-1） groups （n=12） and administrated with corresponding drugs for 14 consecutive days. OFT and PMT were then carried out to examine the anxiety-like behaviors of the rats. The levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and interleukin-1β （IL-1β） in the hypothalamus and serum of rats were determined by the enzyme-linked immunosorbent assay （ELISA）. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）was conducted to determine the mRNA levels of p38 MAPK， NF-κB p65， nuclear factor κB inhibitor α （IκBα）， and ionized calcium binding adaptor molecule 1 （Iba-1）. The protein levels of p38 MAPK， phosphorylated （p）-p38 MAPK， NF-κB p65， p-NF-κB p65， and IκBα in the hypothalamus of rats were determined by Western blot. The expression of Iba-1 in the hypothalamic microglia was detected by immunofluorescence assay. ResultCompared with the blank group， the model group had decreased body weight， scattered dark yellow fur， increased irritability， and preference to hibernation in the corner. In addition， the modeled rats showed increased edge movement distance and time in OFT （P<0.01） and decreased movement distance and time and the number of entries in the open arm in PMT （P<0.01）. The modeling increased the fluorescence intensity of Iba-1 in paraventricular nucleus of hypothalamus （P<0.01）， elevated the levels of IL-1β， IL-6， and TNF-α in the serum and hypothalamus （P<0.01）， up-regulated the protein and mRNA levels of p38 MAPK， NF-κB p65， p-p38 MAPK， p-NF-κB p65， and Iba-1 （P<0.05， P<0.01）， and down-regulated the protein and mRNA levels of IκBα （P<0.01） in the hypothalamus. Compared with the model group， medium- and high-dose Chaihu Longgu Mulitang and diazepam increased the body weight， improved the fur and behaviors， decreased the edge movement distance and time in OFT （P<0.05， P<0.01）， and increased the movement distance and time in the open arm in PMT （P<0.05， P<0.01）. Furthermore， they decreased the fluorescence intensity of Iba-1 in hypothalamic microglia （P<0.05， P<0.01）， lowered the levels of IL-1β， IL-6， and TNF-α in the serum and hypothalamic tissue （P<0.05， P<0.01）， down-regulated the mRNA and protein levels of p38 MAPK， NF-κB p65， p-p38 MAPK， p-NF-κB p65， and Iba-1 （P<0.05， P<0.01）， and up-regulated the mRNA and protein levels of IκBα （P<0.05， P<0.01） in the hypothalamus. ConclusionChaihu Longgu Mulitang can mitigate anxiety-like behaviors and relieve anxiety in GAD rats by inhibiting the p38 MAPK/NF-κB signaling pathway and reducing the activation of microglia and the levels of pro-inflammatory cytokines in the hypothalamus.
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OBJECTIVE To study the repair effect of ephedrine on lipopolysaccharide （LPS）-induced microglia function injury and its mechanism. METHODS Human microglia cells （HMC3） were used as research objects to investigate the effects of different concentrations of ephedrine （75， 150， 300， 600 μg/mL） on the viability and apoptosis of HMC3 cells. HMC3 cells were divided into control group （without drug intervention）， LPS group （1 μg/mL）， ephedrine group （1 μg/mL LPS+300 μg/mL ephedrine）， BAY11-7082 group [1 μg/mL LPS+5 μmol/L nuclear factor-κB （NF-κB） pathway inhibitor BAY11-7082]， inhibitor group （1 μg/mL LPS+300 μg/mL ephedrine+5 μmol/L BAY11-7082） and activator group （1 μg/mL LPS+300 μg/mL ephedrine+1 μmol/L NF-κB pathway activator Prostratin）. After 24 hours of drug treatment， cell migration， the levels of soluble interleukin-6（sIL-6）， interleukin-10（IL-10）， superoxide dismutase（SOD）and malondialdehyde（MDA）， and the expressions of NF-κB pathway-related proteins were all detected. RESULTS The viability of HMC3 cells could be increased significantly by 300 μg/mL ephedrine， while the apoptotic rate was decreased significantly （P＜0.05）. Compared with the control group， the number of migrating cells was increased significantly in the LPS group； the levels of sIL-6 and MDA， the phosphorylation of NF-κB protein were increased significantly， while the levels of IL-10 and SOD were decreased significantly （P＜0.05）. Compared with the LPS group， the above indexes were reversed significantly in the ephedrine group and BAY11-7082 group （P＜0.05）. Compared with the ephedrine group， the number of migrating cells was decreased significantly in the inhibitor group； the levels of sIL-6 and MDA， the phosphorylation of NF-κB protein were decreased significantly， while the levels of IL-10 and SOD were increased significantly （P＜0.05）. The above indexes were reversed significantly in the activator group （P＜0.05）can repair cell injury by inhibiting LPS induced apoptosis， migration， inflammation and oxidant stress of HMC3 cells， the mechanism of which may be associated with inhibiting the activity of the NF-κB signaling pathway.
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Neuroinflammation is a common pathological feature of neurodegenerative diseases （NDs）. Microglia （MG）， a resident macrophage in the brain with a unique developmental origin， is the core driver of neuroinflammation. It can participate in the occurrence and development of NDs through different polarization states and play a key role in regulating neurogenesis and synapse shaping and maintaining homeostasis. MG can be divided into M1 pro-inflammatory phenotype and M2 anti-inflammatory phenotype according to its function. The inflammatory mediators released by the M1 phenotype can lead to nerve degeneration and myelin sheath damage， while the activation of the M2 phenotype is required to inhibit the inflammatory response and promote tissue repair. With the advantages of multi-pathway， multi-target， and bidirectional regulation， traditional Chinese medicine can regulate the polarization balance of MG and has dual effects on NDs such as Alzheimer's disease， Parkinson's disease， and multiple sclerosis. The active components of traditional Chinese medicine and its compound can inhibit the activation of MG by regulating phosphatidylinositol-3-kinases/protein kinase B（PI3K/Akt）， NOD-like receptor thermal protein domain associated protein 3（NLRP3）， signal transducer and activator of transcription factor1（STAT1）， nuclear transcription factor kappa B（NF-κB）， and other pathways， promote the polarization of M1 phenotype to M2 phenotype， reduce the expression of interleukin（IL）-6， tumor necrosis factor-α（TNF-α）， and other pro-inflammatory factors， and increase the secretion of IL-10， arginase-1（Arg-1）， and other anti-inflammatory factors. It can also reduce β-amyloid deposition and tau protein expression in Alzheimer's disease， alleviate dopaminergic neuronal damage in Parkinson's disease， and relieve demyelination， inflammatory cell infiltration， and related clinical symptoms of multiple sclerosis. The bidirectional regulation of the M1/M2 polarization balance of MG by traditional Chinese medicine is a potential strategy for the treatment of NDs. This paper focused on the targets of the regulation of MG polarization balance by traditional Chinese medicine monomer and its compound in the treatment of NDs， so as to further study and summarize the existing research results and provide ideas and basis for the future treatment of NDs.
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Resumen La obesidad es considerada actualmente como un problema de salud pública global y se caracteriza por la hipertrofia e hiperplasia del tejido adiposo debido a la ingesta hipercalórica y la falta de actividad física, disfunción metabólica, inflamación sistémica crónica de bajo grado y gradualmente neuroinflamación hipotalámica. El tejido adiposo actúa como un órgano endocrino secretando adipocinas y citocinas que actúan como reguladores del metabolismo. Sin embargo, la presencia de niveles elevados de ácidos grasos libres y de moléculas inflamatorias derivadas de los adipocitos, pueden alterar la respuesta inmunitaria sistémica, generando inflamación crónica, comprometiendo la integridad de la barrera hematoencefálica y estimulando la respuesta de la glía, especialmente en regiones específicas del hipotálamo, centro de regulación de la homeostasis energética. Las células gliales hipotalámicas son importantes en la transmisión de señales inflamatorias relacionadas con la dieta, pueden modular la actividad neuronal, responder a las señales inmunológicas periféricas e iniciar una respuesta inflamatoria local y gliosis. Esta revisión se enfoca en la descripción general de la disfunción metabólica asociada a la obesidad y su participación en la alteración de la regulación hipotalámica, provocando neuroinflamación y modificaciones en la conducta alimentaria.
Abstract Nowadays, obesity is considered a worldwide rising health problem and is characterized by adipose tissue hypertrophy and hyperplasia due to hypercaloric intake and lack of physical activity, promoting the development of metabolic dysfunction, low-grade systemic chronic inflammation, and gradually hypothalamic neuroinflammation. Adipose tissue acts as an endocrine organ secreting adipokines and cytokines around peripheral organs, functioning as a master metabolism regulator. However, high levels of adipocyte-derived free fatty acids and inflammatory molecules promote impairments in systemic immune response, generate chronic inflammation, disrupt the blood-brain barrier, and stimulate glia, specifically in some hypothalamic regions, the master regulators of energetic homeostasis. Hypothalamic glial cells are essential in diet-related inflammatory signals transmission and can modulate neuronal activity, also respond to peripheral inflammatory signals and begin local inflammatory response and gliosis. This review aims to analyze obesity-related metabolic dysfunction and how it participates in the hypothalamic regulation impairments due to neuroinflammation and impairment in food intake behavior.
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BACKGROUND The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants can infect common mice inducing significant pathological lung lesions and inflammatory responses. This substantially mimics coronavirus disease 19 (COVID-19) infection and pathogenesis in humans. OBJECTIVES To characterise the effects of recombinant SARS-CoV-2 S1 receptor-binding domain (RBD) peptide in murine macrophage and microglial cells' immune activation compared with classical PAMPs in vitro. METHODS Murine RAW 264.7 macrophages and BV2 microglial cells were exposed to increasing concentrations of the RBD peptide (0.01, 0.05, and 0.1 µg/mL), Lipopolysaccharide (LPS) and Poly(I:C) and evaluated after two and 24 h for significant markers of macrophage activation. We determined the effects of RBD peptide on cell viability, cleaved caspase 3 expressions, and nuclear morphometry analysis. FINDINGS In RAW cells, RBD peptide was cytotoxic, but not for BV2 cells. RAW cells presented increased arginase activity and IL-10 production; however, BV2 cells expressed iNOS and IL-6 after RBD peptide exposure. In addition, RAW cells increased cleaved-caspase-3, apoptosis, and mitotic catastrophe after RBD peptide stimulation but not BV2 cells. CONCLUSION RBD peptide exposure has different effects depending on the cell line, exposure time, and concentration. This study brings new evidence about the immunogenic profile of RBD in macrophage and microglial cells, advancing the understanding of SARS-Cov2 immuno- and neuropathology.
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OBJECTIVE Na+/K+-ATPase(NKA)is a large membrane protein expressed universally in all cells.It is indispensable for the maintenance of ionic gradient.We previously reported that the dysfunction of this pump in neurons and astrocytes contributes to stroke and neurodegenerative diseases,respectively.However,its roles in the microglia and stress-related diseases are still unclear.METHODS Two classical models,chronic restraint stress(CRS)model and electronic foot shock(ES)model,were used to study the pathogenesis of anxi-ety in either NKAα1 global knockout(NKAα1 GKO)mice or NKA α1 conditional knockout(NKAα1 CKO)mice.Behavioral tests like open-field test,elevated plus maze,Morris water maze,novel object recognition test and gait imaging test were performed.A variety of molecular bio-logical methods were employed,including RNA sequenc-ing(RNA-seq)analyses,immunofluorescence and elec-trophysiological recordings etc.RESULTS NKAα1 defi-ciency had a broad impact on physical stress-induced anxiety-like behavior,but failed to exacerbate CRS induced memory deficits.Electrophysiology experiment showed that NKAα1 GKO and NKAα1 CKO mice exhibit-ed neuronal hyperexcitability under chronic stress.The underlying mechanisms may involve neuroinflammation,as NKAα1 deficiency exacerbated stress-induced microg-lia activation in vivo.Similarly,inhibition or downregula-tion of NKA α 1 aggravated LPS + ATP-induced inflam-mation in vitro.DR5-12D,a monoclonal antibody against the DR-region of NKAa1,improved stress-induced anxiety-like behavior through amelioration of the neuronal hyper-excitability and neurogenesis deficit in the ventral hippo-campus of mice.CONCLUSION NKA is closely related to neuroinflammation in microglia and DR-region of NKA a1 subunit may serve as a novel target to treat stress-induced anxiety.
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OBJECTIVE To investigate the role of the complement C3/C3aR signaling pathway in the prefrontal cortex and colon neuroglia cell interactions during meth-amphetamine(METH)addiction,to observe the effects of TLR4 inhibitors as well as complement C3 elimination on METH reward and relapse behavior,and to explore the neuroinflammatory mechanisms of complement C3 acti-vation in METH addiction.METHODS ①A 14 d and 28 d rat METH addiction model was established to observe the effects of TLR4 antagonist ibudilast 3 mg·kg-1 and 10 mg·kg-1 on self-administration,reward motivation,relapse,and natural reward behavior in METH-trained 14 d rats and the effects of 0.02 mg·kg-1 complement C3 antago-nist on self-administration behavior in METH-trained 28 d rats.② Differences in the expression of TLR4,NF-κB,GRP94,C3,cathepsin L,CD68,and GFAP in the pre-frontal cortex of each group were examined using West-ern blotting.③ In addition,the expression of ATF6 in the prefrontal cortex of each group and the effects on neuro-nal and microglia/macrophage INOS,CD206 GRP94,and complement C3/C3aR.RESULTS ① Endoplasmic reticulum stress occurred in neurons and microglia after METH exposure depending on GRP94 and unfolded pro-tein responses to the ATF6 pathway.In addition,it acti-vates the TLR4-NF-κB pathway.② Microglia with high complement C3/C3aR expression in the prefrontal cortex were recruited to synaptic pruning and phagocytic responses around neurons with high GRP94,comple-ment C3/C3aR expression and these effects were blocked by complement C3 antagonists.③ In the rec-tum,GRP94 functions as a molecular chaperone for com-plement C3 and cathepsin L.Crosstalk occurs between enteric neurons high in GRP94,complement C3,and macrophages high in C3aR,located in the submucosa,lamina propria,and muscular,respectively,and all of these effects are blocked by complement C3 antago-nists.④ Treatment with the TLR4 antagonist ibudilast inhibits self-administration,reward motivation,and cue-or METH-priming in METH-trained 14 d rats,but fails to affect natural reward behavior.Ibudilast treatment attenu-ates the TLR4-NF-κB inflammatory pathway and comple-ments C3/C3aR pathway in the prefrontal cortex.CON-CLUSION Activation of the complement C3/C3aR signal-ing pathway by TLR4-NF-κB inflammatory signaling in the prefrontal cortex mediates the METH addiction pro-cess,providing an experimental basis for the clinical treatment of METH addiction,and targeting TLR4/NF-κB inflammatory signaling and complement C3/C3aR may be a new way to intervene in METH addiction.
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OBJECTIVE In this study,the effects of live Lactobacillus murinus(L.m)and heat-killed L.muri-nus(H-k L.m)on DA neuronal damage in rats and the underlying mechanisms were investigated.METHODS Male SD rats were randomly divided into five groups:vehicle group,L.m/H-k L.m(1×109 cfu)group,6-OHDH group,6-OHDH + L.m/H-k L.m(1×107 cfu)group,and 6-OHDH + L.m/H-k L.m(1×109 cfu)group.Wild-type and NLRP3 knockout mice were divided into three groups:sham(vehicle),6-OHDH,and 6-OHDH + H-k L.m(1×109 cfu).The model was established after five weeks of pre-administration.Motor ability of experimental mice was assessed by rotarod,mine,and stepping experiments;the expression of dopaminergic neuron markers—tyro-sine hydroxylase(TH),microglial cell markers—ionized calnexin 1(IBA-1),and NOD-like receptor family protein 3(NLRP3)in the substantia nigra was detected by immunohistochemistry and immunofluorescence experi-ments.The expression changes of TH,IBA-1,NLRP3,apoptosis-associated microparticle protein(ASC),cas-pase 1,and inflammatory factors such as interleukin-1β(IL-1β),IL-18,and tumor necrosis factor-α(TNF-α)were detected by immunoblotting experiments.RESULTS H-k L.m ameliorated 6-OHDH-induced motor dysfunctions and loss of substantia nigra DA neurons,while no protec-tion was shown in live L.m treatment.At the same time,H-k L.m reduced the activation of NLRP3 inflammasome in microglia and the secretion of pro-inflammatory factors,thus inhibiting the development of neuroinflammation.Fur-thermore,H-k L.m failed to display its original neuropro-tective properties in NLRP3 inflammasome knockout mice.CONCLUSION H-k L.m conferred neuroprotec-tion against DA neuronal loss via the inhibition of microglial NLRP3 inflammasome activation,these findings provide a promising potential for future applications of L.m,and also beneficial strategy for PD treatment.
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OBJECTIVE Vascular dementia(VaD)is associated with cerebral hypoperfusion,which results in long-term cognitive impairment and memory loss.Neuroin-flammation is an important mechanism of vascular demen-tia.Cornel iridoid glycoside(CIG)is the major active con-stituent isolated from the ripe fruit of Cornus officinalis.Previous studies have shown that CIG enhances neuro-logical function in VaD rats.In the present research,we attempted to clarify the molecular processes underlying the role of CIG on neuroinflammation in VaD.METHODS In vivo,we created a chronic cerebral ischemia rat model by ligation of the bilateral common carotid arteries(2VO).The rats were divided into sham operation,2VO,2VO + CIG(60 and120 mg·kg-1·d-1),and 2VO+ butylphthalide(100 mg·kg-1·d-1)groups and then treated rats with differ-ent concentrations of CIG.In vitro,BV2 microglia cells were induced with bacterial lipopolysaccharide(LPS)and interferon-γ(IFN-γ)to construct the model of microglias with analog neuroinflammation.Histopathology and biel-schowsky silver staining were used to detect myelin integrity and neuronal loss.Immunofluorescence was used to observe changes in microglia.Magnetic Luminex Assay was used to detect changes in inflammatory fac-tors.Western blotting,ELISA or calpain activity assay was used to measure the expression and activity of cal-pain,as well as the expression of NLRP3 inflammasome protein.Furthermore,NLRP3 overexpressing cells were used to further elucidate the potential anti-inflammatory molecular mechanism of CIG.RESULTS ① CIG improved neuronal impairment in the brain of 2VO rats.②CIG increased white matter(WM)integrity in 2VO rats.③ CIG reduced microglia inflammatory response in the cortex and hippocampus of 2VO rats.④ CIG inhibited calpain activity in the cortex and hippocampus of 2VO rats.⑤ CIG exerted anti-inflammatory effects on BV2 cells stimulated by LPS and IFN-γ.⑥ CIG Inhibited the expression and activity of calpain in LPS/IFN-γ-activated BV2 cells.⑦ The main component of CIG had a weak binding force to calpain1.⑧ CIG inhibited the activation of the NLRP3 inflammasome.⑨CIG reduced the activity of calpain induced by NLRP3 overexpression.CONCLU-SION CIG inhibits microglial polarization into a proinflam-matory state by attenuating the assembly of the NLRP3 inflammasome and calpain activation,thus reducing brain inflammation,WM injury,and the loss of neurons.To sum up,the present study suggests that CIG inhibits neuroinflammation.The NLRP3/calpain pathway may be the main pathway by which CIG protects against neuroin-flammation.
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OBJECTIVE To investigate the effects of pharmacological inhibition of STING by C-176,a STING selective inhibitor,in experimental model of Parkinson's disease.METHODS The acute and sub-acute mice mod-els of Parkinson's disease(PD)were established by in-traperitoneal injection of 1-methyl-4-(2′-methylphenyl)-1,2,3,6-tetrahydrophine(MPTP).The selective STING inhibitor C-176 was administered by intraperitoneal injec-tion.The potential neuroprotective effects of C-176 were evaluated by behavioral test,tyrosine hydroxylase(TH)immunostaining,Nissl staining,Western blotting,qPCR and immunofluorescence.For in vitro study,the effects of C-176 on LPS/MPP+-induced inflammatory responses in BV2 microglial cells were determined by real time RT-PCR and Western blotting analysis.RESULTS Our study revealed that C-176 significantly inhibited STING signaling activation,ameliorated MPTP-induced dopami-nergic neurotoxicity,motor deficit and associated neuroin-flammation.Furthermore,pharmacological inhibition of STING in BV2 microglia treated with LPS/MPP+ exhibited decreased inflammatory responses.More importantly,C176 also reduced NLRP3 inflammasome activation both in vitro and in vivo.CONCLUSION The results of our study suggest that pharmacologic inhibition of STING protects against neuroinflammation that may act at least in part through suppressing NLRP3 inflammasome acti-vation and thus ameliorated dopaminergic neurodegener-ation.STING signaling may holds great promise for the development of new treatment strategy for PD as an effective therapeutic target.
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Ischemic stroke is an acute and serious cerebrovascular accident.Neurodegenerative disorders are characterized by progressive degeneration of neu-rons in the central nervous system(CNS),resulting in severe disability and death.Myelin is essential for the health and function of neuronal axons.Oligodendrocytes,the myelinating cells of CNS,are vulnerable to ischemia and neurodegenerative disorders.G protein-coupled receptor 17(GPR17)is a dual receptor activated by uracil nucleotides/cysteinyl leukotrienes(CysLTs).Abnormal GPR17 activation contributes to oligodendrocyte dysfunc-tion and axonal damage.Gelosa et al.reported that CysLT1 receptor antagonist montelukast increased the recruitment and proliferation of oligodendrocyte precursor cells(OPCs)at the acute phase after ischemic stroke.Similarly,a study showed that montelukast stimulated neural progenitor proliferation and hippocampal neuro-genesis of aged rats through inhibition of GPR17.These results were supported by several studies on neurode-generative diseases.The authors showed that pharmaco-logical blockade of GPR17 with CysLT1 or CysLT2 recep-tor antagonists(montelukast or HAMI3379)improved oli-godendrocyte function and fiber connectivity,highlighting GPR17 as a potential therapeutic target in oligodendro-cyte protection and remyelination.Recently,growing evi-dence has revealed a significant interaction between mi-croglia and oligodendrocytes in CNS injury and disease.It was reported that M2 microglia promoted,while M1 microglia inhibited oligodendrogenesis,OPCs maturation and remyelination.Microglia-mediated neuroinflamma-tion,considered as an important pathological event,neg-atively affected OPCs fate and function in experimental neurological disorders.This was further corroborated by later studies.It was recently reported that montelukast enhanced OPCs differentiation and maturation by upreg-ulating the number of M2 microglia at chronic phase of brain ischemia.In line with the above results,inhibition of microglial inflammation by montelukast was shown to be responsible for neurite outgrowth.Although the exact mechanisms were not fully clarified,these results indi-cate that montelukast may indirectly promote OPCs dif-ferentiation and remyelination by a microglia-dependent manner.It has been widely accepted that CysLT1,CysLT2 and GPR17 receptors are localized in various cell types and their expression are upregulated after brain damage.Therefore,it is likely that CysLT receptor antagonists confer neuroprotection by targeting different receptors and multiple cell functions.Many studies have reported that CysLT receptor antagonists promote protec-tion of oligodendrocytes by inhibiting GPR17.Moreover,they may improve OPCs differentiation and neuronal sur-vival by regulating CysLTs-mediated microglial activation.Altogether,these data open novel perspectives in the treatment of cerebral ischemia and neurodegenerative diseases.
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Depression has become one of the most important causes of disability worldwide. Although traditional treatment methods such as drug therapy, physical therapy and psychological intervention have positive effects, they still have limitations such as high treatment cost, high drug side effects, high recurrence rate and poor compliance. Exercise therapy, as a treatment recommended by many guidelines, has many advantages such as simple and convenient, few side effects, and strong persistence. Studies have shown that exercise can relieve depression through a variety of mechanisms, but there is still a lack of effective targets for the treatment of depression. Microglia is an innate immune cell in the central nervous system. In recent years, the role of microglia in neuroinflammation in depression and other mental diseases has attracted more and more attention. Thus, microglia may be a key target for exercise to alleviate depression. This article discusses the relationship between exercise and microglia phenotypes as well as depression, and proposes future research directions on microglia-mediated exercise to improve depression, in order to provide new ideas and directions for the clinical treatment of depression.
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Objective:To investigate the effects of human umbilical cord mesenchymal stem cells exosomes (hucMSC-Exo) on depression-like behavior and polarization dynamic transition of hippocampal microglia in chronic unpredictable mild stress(CUMS) mice.Methods:The hucMSC was isolated, cultured, and the 5th generation hucMSC-Exo was extracted by ultracentrifugation and identified.Biological markers of CD9 and CD63 of hucMSC-Exo were detected by Western blot.A total of 12 SPF grade male C57BL/6J mice were randomly selected to establish depression model by CUMS method at the same time, and relevant behavioral experiments, including opening field test (OFT) and forced swimming test (FST) were used to detect changes of depression-like behavior in CUMS mice. After modeling, 12 mice were randomly divided into two groups, with 6 mice in each group. One group of mice was the exosome treatment group (CUMS+ Exo group) after stereotactic brain injection of hucMSC-Exo in the hippocampus, and the other group was the CUMS group. The expression of inflammatory signals in the hippocampus of mice in both groups were detected by micro PET/CT scanning technique. The proportion of M2-type microglia and M1-type microglia in the hippocampus of mice was detected by tissue immunofluorescence. β3-tubulin immunofluorescence staining was used to detect the length changes of neuronal axons in the depressed cell model constructed with corticosterone (CORT). EdU staining was used to detect neuronal proliferation. TUNEL staining was used to observe the apoptosis of neurons in the hippocampus of mice.Statistical analysis was conducted by GraphPad 8.0 software, and t-test was used for inter group comparison. Results:Under the electron microscope, hucMSC-Exo showed typical " double-layer cup-like" small vesicle-like changes and the particle diameter was about 100 nm.Western blot confirmed the expression of lconic proteins of CD9 and CD63. In the micro PET/CT scans, the uptake of [18F]DPA-714 in the CUMS+ Exo group was lower than that of the CUMS group, and the difference was statistically significant ((0.91±0.02)g/mL, (0.81±0.05)g/mL, t=4.54, P=0.001 1). In the opening field test, the percentage of central path length ((3.40±0.44)%, (5.17±0.90)%, t=4.33, P=0.001 5) and the time spent on the central path ((7.04±0.60)s, (10.22±1.41)s, t=6.02, P=0.000 1) in CUMS+ Exo group were higher than those in CUMS group, while the total distances of the two groups were not statistically significant ( t=0.48, P>0.05). In the forced swimming test, the immobility time in the CUMS+ Exo group was less than that in the CUMS group ((152.33±7.28) s, (133.50±4.32) s, t=5.45, P=0.000 3). In tissue immunofluorescence experiments, compared with the CUMS group, the proportion of M2-type microglia of hippocampus in the CUMS+ Exo group((0.33±0.04), (0.59±0.12), t=5.11, P=0.000 5)increased and the proportion of M1-type microglia ((0.56±0.06), (0.41±0.03), t=5.15, P=0.000 4) decreased in the CUMS+ Exo group. β3-tubulin-labeled immunofluorescence results showed an increase of neuronal axon length in the CORT+ Exo group compared with that in the CORT group((3.99±0.99) μm, (6.76±1.11) μm, t=6.10, P=0.000 1). The results of cell proliferation test showed an increase of proliferation rate in the CORT+ Exo group compared with that in the CORT group((0.74±0.07), (0.64±0.03), t=3.32, P=0.001 8). In the TUNEL staining experiment, the apoptosis rate of neurons in the hippocampal region of mice in the CUMS+ Exo group was lower than that of CUMS group ((0.24±0.04), (0.39±0.04), t=6.11, P=0.000 1). Conclusion:hucMSC-Exo can promote the conversion of M1 polarized microglia to M2 type microglia to alleviate depression-like behavior and reduce neuronal apoptosis in CUMS mice.
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Restricted and repetitive behaviors is the characteristic clinical manifestation of many neuropsychiatric diseases, which seriously affects the work, study and daily communication of patients, and brings huge mental and economic burden to the family and society.Its causes are complex and its manifestations are diverse.Current studies have shown that microglia is closely related to the occurrence of restricted and repetitive behaviors, and the in-depth study of microglia has become a research hotspot to explore the mechanism of restricted and repetitive behaviors.In recent years, studies have found that restricted and repetitive behaviors of various neuropsychiatric diseases (such as frontotemporal dementia, obsessive-compulsive disorder, autism spectrum disorder) are related to microglia.However, reliable evidence for the exact mechanism of microglia involved in restricted and repetitive behaviors remains lacking.This article reviews the recent research progress of microglia involved in restricted and repetitive behaviors.It is expected to provide a new theoretical basis and therapeutic targeting cell for the development and treatment of neuropsychiatric diseases related to restricted and repetitive behaviors in the future.
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Objective:To investigate the effects of artesunate ( ART ) on neuronal apoptosis, inflammatory response after stroke in rats and microglia polarization.Methods:(1)Animal experiment: twenty-seven male SD rats of SPF grade were divided into sham operation group, model group and ART treatment group according to the random number table method, with 9 rats in each group.Rats in the model group and ART treatment group were used to establish a stroke model by middle cerebral artery occlusion (MCAO). And rats in the ART treatment group were intraperitoneally injected with ART (25 mg/kg) once a day for three days before modeling, while the rats in sham operation group and the model group were injected with the same amount of solvent.And 24 h after the modeling, TTC staining was used to evaluate the volume of cerebral infarction, Western blot was used to detect the expression of Bcl2 in the infarct area, penumbra and hippocampus, TUNEL method was used to detect neuronal apoptosis, and tissue immunofluorescence was used to observe the expression of tumor necrosis factor-α(TNF-α) in the penumbra region of cerebral cortex.(2)Cell experiments: microglia BV2 were cultured and divided into control group, oxygen-glucose deprivation/reoxygenation group, oxygen-glucose deprivation/reoxygenation + 0.05 μmol/L ART group, oxygen-glucose deprivation/reoxygenation + 0.1 μmol/L ART group and oxygen-glucose deprivation/reoxygenation + 0.5 μmol/L ART group.The levels of inflammatory factors interleukin-6(IL-6), interleukin-1β(IL-1β) and TNF-α were detected by qRT-PCR, the expressions of M2 type microglia marker protein CD206 and ARG1 were detected by Western blot, the BV2 cell medium after treatment in each of the above groups was collected as conditioned medium to culture HT22 hippocampal neuron cells and cell activity was measured by CCK8 method.GraphPad Prism 7 software was used for data analysis.One-way ANOVA was used for comparison of differences among multiple groups, and LSD was used for further two-by-two comparisons.Results:(1)Animal experiment results: TTC staining results showed that the percentage of cerebral infarction volume in the ART treatment group was smaller than that in the model group ((23.09±8.51)%, (39.63±5.71)%, t=33.93, P<0.01). The results of TUNEL staining showed that the number of apoptotic cells in the model group and ART treatment group was higher than that in the sham operation group ((638.90±177.82)cells/mm 2, (72.75±13.21) cells/mm 2, (16.16±2.73) cells/mm 2, both P<0.05), and the number of apoptotic cells in the ART treatment group was lower than that in the model group ( P<0.05). Western blot results showed that the levels of Bcl2 protein in penumbra and infarct area of the model group were both lower than those in sham group(both P<0.05). The levels of Bcl2 protein in penumbra, the hippocampus and infarcted area of the ART treatment group were significantly lower than those of the model group(all P<0.05). The results of tissue immunofluorescence showed that the fluorescence intensities of TNF-α in the model group and ART treatment group were higher than those in the sham group (all P<0.05), while the fluorescence intensity of TNF-α in the ART treatment group was lower than that in the model group ( P<0.05). (2)Cell experiment: qRT-PCR results showed that compared with the control group, the mRNA levels of IL-6, IL-1β and TNF-α (all P<0.05) in oxygen-glucose deprivation/reoxygenation group were significantly higher than those of the control group.And the mRNA levels of IL-1β, IL-6 and TNF-α in oxygen-glucose deprivation/reoxygenation + 0.05 μmol/L ART group, oxygen-glucose deprivation/reoxygenation + 0.1 μmol/L ART group and oxygen-glucose deprivation/reoxygenation + 0.5 μmol/L ART group were significantly lower than those of the oxygen-glucose deprivation/reoxygenation group (all P<0.05). Western blot results showed that compared with the control group, the expression of CD206 ((0.85±0.04), (1.07±0.07), P<0.05) was significantly down-regulated in the oxygen-glucose deprivation/reoxygenation group.The CD206 and ARG in oxygen-glucose deprivation/reoxygenation + 0.1 μmol/L ART group((1.22±0.06), (1.35±0.08)) and oxygen-glucose deprivation/reoxygenation + 0.5 μmol/L ART group((1.24±0.14), (1.14±0.07)) were significantly higer than those of oxygen-glucose deprivation/reoxygenation group((0.85±0.04), (0.85±0.05))(all P<0.05). The results of CCK8 showed that compared with the control group, the cell viability in the oxygen-glucose deprivation/reoxygenation group was significantly decreased( P<0.05). The cell viability of the oxygen-glucose deprivation/reoxygenation + 0.05 μmol/L ART group, the oxygen-glucose deprivation/reoxygenation + 0.1 μmol/L ART group, the oxygen-glucose deprivation/reoxygenation + 0.5 μmol/L ART group were all higher than those of oxygen-glucose deprivation/reoxygenation group(all P<0.05). Conclusion:ART reduces neuronal apoptosis after stroke, decreases the neuroinflammatory response after stroke, and promotes oxygen-glucose deprivation/reoxygenation-activated microglia BV2 polarization to the M2 type.
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【Objective】 To investigate the protective effect and mechanism of platelet-rich plasma (PRP) on lipopolysaccharide (LPS) -induced inflammatory response in BV2 cells. 【Methods】 BV2 microglia were divided into normal control group, 10%PRP control group, LPS group (LPS induction), 3%PRP+ LPS group (LPS induction, 3%PRP pretreatment), 5%PRP+ LPS group (LPS induction, 5%PRP pretreatment), 10%PRP+ LPS group (LPS induction, 10%PRP pretreatment), and the proliferation of BV2 cells was measured by CCK-8. The mitochondrial membrane potential of BV2 cells was measured by confocal microscopy, ROS was measured by fluorescence method, and NO was measured by Griess method. The protein expressions of IL-6, TNF-α, BACH1, GPX4, NRF2 and HO-1 were detected by Western blot. In addition, BV2 microglia were treated with HO-1 inhibitor and divided into normal control group, LPS group, ZnPP+ LPS group, 10%PRP+ LPS group, ZnPP+ LPS+ 10%PRP group, and the protein expressions of HO-1, IL-6 and TNF-α were detected by Western blot. 【Results】 Compared with normal control group, PRP promoted the proliferation of BV2 cells (P<0.01). The mitochondrial membrane potential decreased, ROS production increased, the levels of NO, IL-6, TNF-α and BACH1 increased (P<0.01). However, the expression levels of GPX4, NRF2 and HO-1 decreased (P<0.01) in LPS group. Compared with LPS group, the proliferation activity and mitochondrial membrane potential of BV2 cells in 3%PRP+ LPS, 5%PRP+ LPS and 10%PRP+ LPS groups significantly increased. The levels of ROS, NO, IL-6, TNF-α and BACH1 significantly decreased (P<0.01). The expressions of GPX4, NRF2 and HO-1 in different concentrations of PRP (3%, 5% and 10%) increased (P<0.01). Moreover, the expression of IL-6 and TNF-α in ZnPP+ LPS group was significantly higher than that in LPS group after HO-1 inhibitor treatment. Compared with 10%PRP+ LPS+ ZnPP group, HO-1 inhibitor could reverse the effect of PRP on the expression of IL-6 and TNF-α in LPS-induced BV2 cells (P<0.01). 【Conclusion】 PRP inhibits the inflammatory response of BV2 microglia induced by LPS by activating the NRF2/HO-1 signaling pathway.
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Objective To investigate the effect of exposure to lead oxide nanoparticles (PbO NPs) on the polarization of microglia in mouse hippocampus. Methods i) Specific pathogen-free male C57 mice were randomly divided into control group, low-, medium- and high-dose groups, with 10 mice in each group. Mice in these three dose groups were intraperitoneally injected with PbO NPs suspension at doses of 5, 10 and 20 mg/kg per day, respectively, and mice in the control group were intraperitoneally injected with the same volume of 0.9% sodium chloride solution, five days per week for four weeks. ii) BV-2 cells were treated with PbO NPs at doses of 0.0, 2.5, 5.0 and 10.0 mg/L for 24 hours. iii) BV-2 cells were randomly divided into control group, PbO NPs group and triggering receptor expressed on myeloid cells 2 (TREM2) high expression + PbO NPs group. The cells in the control group received no treatment. The cells in PbO NPs group were exposed to 10.0 mg/L PbO NPs suspension for 24 hours. Cells in TREM2 high expression + PbO NPs group were transfected with Trem2 high expression plasmid, and then exposed to 10.0 mg/L PbO NPs suspension for 24 hours. iv) The mRNA expression of M1 markers ［nitric oxide synthase (iNos), cyclooxygenase 2 (Cox2), chemokine receptor 7 (Ccr7)］, M2 markers ［arginin-1 (Arg-1), transforming growth factor-β (Tgf-β), chemokine receptor 2 (Ccr2)］ and Trem2 of microglia was detected by real-time fluorescent quantitative polymerase chain reaction. The protein expression of iNOS, ARG-1 and TREM2 was detected by Western blotting. Results i) During the experiment, there was no significant difference in body weight of mice among these four groups (P>0.05). The relative expression of Cox2 and Ccr7 mRNA in the hippocampus of the mice increased in the low-dose group and the iNos, Cox2 and Ccr7 mRNA increased in the medium- and high-dose groups, compared with the control group (all P<0.05). The relative mRNA expression of Tgf-β in the hippocampus of the mice of low-dose group and Arg-1, Tgf-β and Ccr2 in the medium- and high-dose groups was decreased compared with the control group (all P<0.05). The mRNA relative expression of iNos, Cox2 and Ccr7 was increased (all P<0.05), while the mRNA relative expression of Arg-1, Tgf-β and Ccr2 was decreased (all P<0.05) in the hippocampus of the mice of high-dose group compared with the low-dose group. The relative expression of Trem2 mRNA and TREM2 protein in the hippocampus of mice of the medium- and high-dose groups was lower than those in the control group (all P<0.05). The relative expression of Trem2 mRNA and TREM2 protein in the hippocampus of mice of the high dose group was lower than those in the low- and the medium-dose groups (all P<0.05). With the increase of PbO NPs exposure dose, the relative expression of iNOS protein in hippocampus tissues of mice increased (P<0.01), and the relative expression of ARG-1 protein decreased (P<0.01). ii) With the increase of PbO NPs exposure dose, the relative expression of iNOS protein increased (P<0.01), and the relative expression of ARG-1 protein decreased (P<0.01) in BV-2 cells. The relative expression of iNOS protein in BV-2 cells of PbO NPs group and TREM2 high expression + PbO NPs group was increased (all P<0.05), and the relative expression of ARG-1 protein decreased (all P<0.05) compared with the control group. The relative expression of iNOS protein decreased (P<0.05), and the relative expression of ARG-1 protein increased (P<0.05) in BV-2 cells of TREM2 high expression + PbO NPs group compared with the PbO NPs group. Conclusion Exposure to PbO NPs could increase the M1 polarization and decrease the M2 polarization of microglia, with a dose-effect relationship. The M1 polarization of microglia decreased and M2 polarization increased after overexpression of Trem2 gene. The regulation of microglia polarization by TREM2 may be involved in the neurotoxic effects of PbO NPs.
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@#The aim of the present study was to investigate the effects of exosomes derived from bone marrow mesenchymal stem cells (BMSCs) on the polarization of M1/M2 microglia/macrophages in rats with acute cerebral ischemia.Ultrahigh-speed centrifugation was employed to isolate and identify exosomes; a middle cerebral artery occlusion (MCAO) model was prepared in rats using the intraluminal filament technique; Longa scoring and corner tests were used to evaluate the neurological function of rats; 2, 3, 5-triphenyltetrazole chloride (TTC) staining was used to assess the infarct volume in rat brains; immunofluorescence double-labeling of CD16/32/Iba1 and CD206/Iba1 was performed to detect M1/M2 phenotypes of microglia/macrophages; RT-qPCR was employed to measure the mRNA expression of CD86, inducible nitric oxide synthase (iNOS), tumor necrosis factor-alpha (TNF-α), arginase-1 (Arg-1), interleukin-10 (IL-10), and transforming growth factor beta (TGF-β) in the ischemic penumbra of rat brains.The experimental results showed that BMSC-Exos reduced the number of CD16/32+/Iba1+ positive cells in the ischemic penumbra (P < 0.01) while increasing the number of CD206+/Iba1+positive cells (P < 0.01), and decreased the mRNA expression of iNOS, CD86, and TNF-α, while increasing the mRNA expression of Arg-1, TGF-β, and IL-10 (P < 0.05 or P < 0.01).This research suggests that BMSC-Exos can regulate M1/M2 polarization of microglia/macrophages in rats with acute cerebral ischemia, alleviate neuroinflammation, and improve ischemic brain injury.