Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract(MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of ...Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract(MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of MLE was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide assay. The inflammatory response of BV-2 cells were induced with lipopolysaccharide. The generation of nitric oxide levels was determined by using Griess assay and the level of pro-inflammatory cytokines(IL-1β, IL-6 and TNF-α) was evaluated by ELISA kit. The expression of iNOS, COX-2 as well as IκB-ααwas carried out by immunoblot analysis. Results: MLE reduced the nitric oxide production in concentration-dependent manner, and maintained the viability of BV-2 microglial cells which indicated absence of toxicity. In addition, MLE repressed the activation of nuclear factor kappa B by arresting the deterioration of IκB-α, consequently resulted in suppression of cytokines expression such as COX-2 and iNOS. Conclusions: MLE inhibitory activities are associated with the inhibition of nuclear factor kappa B transcriptional activity in BV2 microglial cells. Thus MLE may offer a substantial treatment for neuroinflammatory diseases.展开更多
Objective To study the therapeutic effects of Shenyuan Gan(参远苷,SYG)on the inflammat-ory response in BV2 microglial cells induced by lipopolysaccharide(LPS).Methods The cytotoxicity of SYG to BV2 microglial cells wa...Objective To study the therapeutic effects of Shenyuan Gan(参远苷,SYG)on the inflammat-ory response in BV2 microglial cells induced by lipopolysaccharide(LPS).Methods The cytotoxicity of SYG to BV2 microglial cells was evaluated using a Cell Counting Kit-8(CCK-8)assay,and the effect of SYG concentrations on LPS-induced BV2 microglial cells was studied.The morphological changes were observed using an optical microscope.The nitric oxide(NO)concentration in cell culture supernatant was determined using Griess re-agent.The expression of cytokines and inflammatory mediators were also measured by an en-zyme-linked immunosorbent assay(ELISA).Western blot analysis was used to determine the levels of inducible NO synthase(iNOS),nuclear factor-kappa B(NF-κB)p65,alpha inhibitor of NF-κB(IκB-α),phosphorylation-IκB-α(p-IκB-α),NOD-like receptor 3(NLRP3),and cas-pase-1 expression.Moreover,the expression of iNOS,NLRP3,and ionized calcium binding adapter molecule 1(Iba1)was also observed using immunofluorescent staining.Results SYG had a low cytotoxic effect on BV2 microglial cells and could significantly decr-ease LPS-induced morphological changes of BV2 microglial cells(P<0.05).ELISA results showed that SYG significantly inhibited the LPS-induced increase in interleukin(IL)-1βand IL-6 in BV2 microglia cells(P<0.05),and Western blot analysis showed that the phosphoryla-tion levels of iNOS,NF-κB p65,and IκB-αas well as NLRP3 and caspase-1 expression were also significantly decreased,and IκB-αexpression was increased after SYG treatment(P<0.05,compared with the LPS-treated group).The immunofluorescence results were consist-ent with the Western blot results,and Iba1 staining indicated that the cell morphology tended to be resting.These results indicate that SYG has a certain inhibitory effect on LPS-induced inflammation in BV2 microglial cells.Conclusion SYG can inhibit LPS-induced release of inflammatory factors in BV2 microglial cells by affecting the phosphorylation levels of NF-κB p65 and IκB-α.SYG is a valuable candid-ate for treating neuroinflammation-related diseases.展开更多
Little is known about whether tamoxifen (TAM) can affect resting state microglia apoptosis and about the cellular mechanism that may account for this. To explore this question, we incubated the microglia cell line BV-...Little is known about whether tamoxifen (TAM) can affect resting state microglia apoptosis and about the cellular mechanism that may account for this. To explore this question, we incubated the microglia cell line BV-2 cells with TAM at different concentrations. Cell viability was assessed by the MTT assay, and flow cytometric analysis was performed to detect the cell apoptosis rate. Furthermore, mitochondrial membrane potential (Δψm) was tested by flow cytometry, and Bax, Bcl-2, Fas, and Fas-L expression was detected by Western blot. The results demonstrated that TAM decreased cell viability and induced apoptosis of BV-2 cells in a concentration- and time-dependent manner. In addition, disruption of Δψm was followed by up-regulated expression of pro-apoptotic Bax, Fas and Fas-L, and down-regulated expression of anti-apoptotic Bcl-2. These results indicate that TAM may induce apoptosis of BV-2 cells through both mitochondria- and death receptor-mediated pathways.展开更多
Prion diseases are infectious and fatal neurodegenerative diseases.The pathogenic agent is an abnormal prion protein aggregate.Microglial activation in the centre nervous system is a characteristic feature of prion di...Prion diseases are infectious and fatal neurodegenerative diseases.The pathogenic agent is an abnormal prion protein aggregate.Microglial activation in the centre nervous system is a characteristic feature of prion disease.In this study,we examined the effect of PrP 106-126 on PrP mRNA gene expression in Mouse microglia cells BV-2 by real-time quantitative PCR.PrP mRNA expression level was found to be significantly increased after 18 h exposure of BV-2 cells to PrP 106-126,with 3-fold increase after 18 h and 4.5-fold increase after 24 h and BV-2 cells proliferating occurred correspondingly.Our results provide the first in vitro evidence of the increase of PrP mRNA levels in microglial cells exposed to PrP 106-126,and indicate that microglial cells might play a critical role in prion pathogenesis.展开更多
Microglial cells are the key innate immune cells in the brain and they are crucial in maintaining brain parenchyma homeostasis.Under physiological conditions,microglial cells assume a ramified morphology with a small ...Microglial cells are the key innate immune cells in the brain and they are crucial in maintaining brain parenchyma homeostasis.Under physiological conditions,microglial cells assume a ramified morphology with a small cell body and an extensive network of fine processes,which secrete neurotrophic factors and patrol the surroundings in search for pathogens and eliminate cellular debris via phagocytosis.Microglial cells express a repertoire of pattern recognition receptors(PRRs)that enable them to detect diverse danger-associated molecular patterns(DAMPs)released from damaged cells or cells under stress,or pathogen-associated molecular patterns generated by pathogens during infection.展开更多
Microglial cells are important resident innate immune components in the central nervous system that are often activated during neuroinflammation.Activated microglia can display one of two phenotypes,M1 or M2,which eac...Microglial cells are important resident innate immune components in the central nervous system that are often activated during neuroinflammation.Activated microglia can display one of two phenotypes,M1 or M2,which each play distinct roles in neuroinflammation.Rutin,a dietary flavonoid,exhibits protective effects against neuroinflammation.However,whether rutin is able to influence the M1/M2 polarization of microglia remains unclear.In this study,in vitro BV-2 cell models of neuroinflammation were established using 100 ng/mL lipopolysaccharide to investigate the effects of 1-hour rutin pretreatment on microglial polarization.The results revealed that rutin pretreatment reduced the expression of the proinflammatory cytokines tumor necrosis factor-α,interleukin-1β,and interleukin-6 and increased the secretion of interleukin-10.Rutin pretreatment also downregulated the expression of the M1 microglial markers CD86 and inducible nitric oxide synthase and upregulated the expression of the M2 microglial markers arginase 1 and CD206.Rutin pretreatment inhibited the expression of Toll-like receptor 4 and myeloid differentiation factor 88 and blocked the phosphorylation of I kappa B kinase and nuclear factor-kappa B.These results showed that rutin pretreatment may promote the phenotypic switch of microglia M1 to M2 by inhibiting the Toll-like receptor 4/nuclear factor-kappa B signaling pathway to alleviate lipopolysaccharide-induced neuroinflammation.展开更多
目的:探讨XMU-MP-1(Xiamen University-inhibitor of mammalian sterile 20-like kinase protein 1)对氧糖剥夺(OGD)损伤后小胶质细胞M1/M2极化平衡的调节作用。方法采用OGD法诱导BV2细胞损伤。实验分为6组:对照组、模型组、MST1/2 siRN...目的:探讨XMU-MP-1(Xiamen University-inhibitor of mammalian sterile 20-like kinase protein 1)对氧糖剥夺(OGD)损伤后小胶质细胞M1/M2极化平衡的调节作用。方法采用OGD法诱导BV2细胞损伤。实验分为6组:对照组、模型组、MST1/2 siRNA组和低、中、高剂量实验组(分别给予1.25、5.0和20.0μg·mL^(-1) XMU-MP-1)。采用MTT法测细胞活力,ELISA测细胞上清液中TNF-α、IL-6和IL-1β表达,qRT-PCR测M1和M2标志物的mRNA表达,流式细胞术测CD206表达,蛋白印迹法测MST1、LATS1和YAP蛋白表达。结果与模型组相比,XMU-MP-1抑制BV2细胞增殖,显著降低TNF-α、IL-6和IL-1β的表达水平,下调MCP-1、IL-6、TNF-α和i NOS mRNA的表达,上调CD206、IL-10、TGF-β、IL-10和YM1 mRNA表达,降低MST1和LAST 1蛋白表达,上调YAP和CD206表达。结论XMU-MP-1通过调控MST1/2的磷酸化,调节OGD损伤后的BV2细胞M1/M2极化平衡,为神经炎症靶点药物研发提供理论基础。展开更多
P2X4 and P2X7 receptors play an important role in neuropathic pain after spinal cord injury. Regulation of P2X4 and P2X7 receptors can obviously reduce pain hypersensitivity after injury. To investigate the role of ne...P2X4 and P2X7 receptors play an important role in neuropathic pain after spinal cord injury. Regulation of P2X4 and P2X7 receptors can obviously reduce pain hypersensitivity after injury. To investigate the role of neural stem cell transplantation on P2X receptor-mediated neuropathic pain and explore related mechanisms, a rat model of spinal cord injury was prepared using the free-falling heavy body method with spinal cord segment 10 as the center. Neural stem cells were injected into the injured spinal cord segment using a micro-syringe. Expression levels of P2X4 and P2X7 receptors, neurofilament protein, and glial fibrillary acidic protein were determined by immunohistochemistry and western blot assay. In addition, sensory function was quantitatively assessed by current perception threshold. The Basso-Beattie-Bresnahan locomotor rating scale was used to assess neuropathological pain. The results showed that 4 weeks after neural stem cell transplantation, expression of neurofilament protein in the injured segment was markedly increased, while expression of glial fibrillary acidic protein and P2X4 and P2X7 receptors was decreased. At this time point, motor and sensory functions of rats were obviously improved, and neuropathic pain was alleviated. These findings demonstrated that neural stem cell transplantation reduced overexpression of P2X4 and P2X7 receptors, activated locomotor and sensory function reconstruction, and played an important role in neuropathic pain regulation after spinal cord injury. Therefore, neural stem cell transplantation is one potential option for relieving neuropathic pain mediated by P2X receptors.展开更多
Inflammation plays an important role in the pathological process of ischemic stroke,and systemic inflammation affects patient prognosis.As resident immune cells in the brain,microglia are significantly involved in imm...Inflammation plays an important role in the pathological process of ischemic stroke,and systemic inflammation affects patient prognosis.As resident immune cells in the brain,microglia are significantly involved in immune defense and tissue repair under various pathological conditions,including cerebral ischemia.Although the differentiation of M1 and M2 microglia is certainly oversimplified,changing the activation state of microglia appears to be an intriguing therapeutic strategy for cerebral ischemia.Recent evidence indicates that both mesenchymal stem cells(MSCs)and MSC-derived extracellular vesicles(EVs)regulate inflammation and modify tissue repair under preclinical stroke conditions.However,the precise mechanisms of these signaling pathways,especially in the context of the mutual interaction between MSCs or MSC-derived EVs and resident microglia,have not been sufficiently unveiled.Hence,this review summarizes the state-ofthe-art knowledge on MSC-and MSC-EV-mediated regulation of microglial activity under ischemic stroke conditions with respect to various signaling pathways,including cytokines,neurotrophic factors,transcription factors,and microRNAs.展开更多
Microglial apoptosis is associated with neuroinflammation and no effective strategies are currently available to protect microglia against inflammation-induced apoptosis. Mouse microglial BV-2 cells(5 × 10^6) wer...Microglial apoptosis is associated with neuroinflammation and no effective strategies are currently available to protect microglia against inflammation-induced apoptosis. Mouse microglial BV-2 cells(5 × 10^6) were incubated with 10 μg/mL lipopolysaccharides for 12 hours to mimic an inflammatory environment. Then the cells were co-cultured with mitochonic acid 5(MA-5) for another 12 hours. MA-5 improved the survival of lipopolysaccharide-exposed cells. MA-5 decreased the activity of caspase-3, which is associated with apoptosis. MA-5 reduced the number of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells, and increased adenosine triphosphate levels in cells. MA-5 decreased the open state of the mitochondrial permeability transition pore and reduced calcium overload and diffusion of second mitochondria-derived activator of caspase(Smac). MA-5 decreased the expression of apoptosis-related proteins(mitochondrial Smac, cytoplasmic Smac, pro-caspase-3, cleaved-caspase-3, and caspase-9), and increased the levels of anti-apoptotic proteins(Bcl2 and X-linked inhibitor of apoptosis protein), mitochondria-related proteins(mitochondrial fusion protein 2, mitochondrial microtubule-associated proteins 1 A/1 B light chain 3 B II), and autophagy-related proteins(Beclin1, p62 and autophagy related 5). However, MA-5 did not promote mitochondrial homeostasis or decrease microglial apoptosis when Mitofusin 2 expression was silenced. This shows that MA-5 increased Mitofusin 2-related mitophagy, reversed cellular energy production and maintained energy metabolism in BV-2 cells in response to lipopolysaccharide-induced inflammation. These findings indicate that MA-5 may promote the survival of microglial cells via Mitofusin 2-related mitophagy in response to lipopolysaccharide-induced inflammation.展开更多
目的探讨激活哺乳动物雷帕霉素靶蛋白复合物2(mTORC2)/Akt信号通路对6-羟基多巴胺(6-OHDA)模型小鼠多巴胺能神经元和行为学的影响及可能的机制。方法将36只体重20~25 g 3月龄Nestin-CreERTM::ROSA26-LacZ雄性C57BL/6J小鼠分为NS+玉米油...目的探讨激活哺乳动物雷帕霉素靶蛋白复合物2(mTORC2)/Akt信号通路对6-羟基多巴胺(6-OHDA)模型小鼠多巴胺能神经元和行为学的影响及可能的机制。方法将36只体重20~25 g 3月龄Nestin-CreERTM::ROSA26-LacZ雄性C57BL/6J小鼠分为NS+玉米油组、6-OHDA+玉米油组、6-OHDA+PP242组、6-OHDA+A-443654组,并在小鼠右侧纹状体注射6-OHDA制备帕金森病(PD)小鼠模型以及每日腹腔注射mTORC2/Akt信号通路激动剂A-443654或抑制剂PP242。通过ELISA测定血清肿瘤坏死因子α(TNF-α)和白细胞介素1β(IL-1β)的水平;免疫组织化学和免疫荧光染色考察黑质(SN)-纹状体小胶质细胞、脑室周围神经前体细胞(NPCs)和多巴胺能神经元数目,Western blotting检测中脑水管mTORC2/Akt信号通路各相关蛋白Rictor,p-Akt和DNA损伤反应调节1(REDD1)的表达并通过免疫共沉淀验证它们之间的相互作用,最后观察各组小鼠行为学的变化。结果6-OHDA模型小鼠伴随小胶质细胞的激活和炎症因子的增加,黑质多巴胺能阳性神经元数目明显下降,小鼠的运动功能发生障碍,但NPCs数目较对照组小鼠明显增加,mTORC2/Akt信号通路相关蛋白表达也明显上调,在激动剂A-443654处理后随着相关蛋白的表达进一步上调,上述各指标均有明显改善,而抑制剂PP242处理组则呈现与激动剂A-443654完全相反的情况。结论A-443654通过上调mTORC2/Akt信号通路关键蛋白促进NPCs的增殖,增加新生多巴胺能神经元的数目并减少小胶质细胞的激活和炎症反应最终导致PD模型小鼠黑质-纹状体多巴胺能神经元和小鼠行为学的改善。展开更多
The treatment of microglial BV-2 cells with sodium arsenate（As（V）：0.1-400 μmol/L — 48 hr）induces a dose-dependent response.The neurotoxic effects of high concentrations of As（V）（100,200 and 400 μmol/L） are...The treatment of microglial BV-2 cells with sodium arsenate（As（V）：0.1-400 μmol/L — 48 hr）induces a dose-dependent response.The neurotoxic effects of high concentrations of As（V）（100,200 and 400 μmol/L） are characterized by increased levels of mitochondrial complexesⅠ,Ⅱ,and Ⅳ followed by increased superoxide anion generation.Moreover,As（V） triggers an apoptotic mode of cell death,demonstrated by an apoptotic SubG1 peak,associated with an alteration of plasma membrane integrity.There is also a decrease in transmembrane mitochondrial potential and mitochondrial adenosine triphosphate ATP.It is therefore tempting to speculate that As（V） triggers mitochondrial dysfunction,which may lead to defective oxidative phosphorylation subsequently causing mitochondrial oxidative damage,which in turn induces an apoptotic mode of cell death.展开更多
基金supported by the Basic Science Research Program through the National Research Foundation of Korea(NRF) funded by the Ministry of Science and ICT(NRF-2017R1C1B2010276 and 2017R1A2A2A07001035)
文摘Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract(MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of MLE was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide assay. The inflammatory response of BV-2 cells were induced with lipopolysaccharide. The generation of nitric oxide levels was determined by using Griess assay and the level of pro-inflammatory cytokines(IL-1β, IL-6 and TNF-α) was evaluated by ELISA kit. The expression of iNOS, COX-2 as well as IκB-ααwas carried out by immunoblot analysis. Results: MLE reduced the nitric oxide production in concentration-dependent manner, and maintained the viability of BV-2 microglial cells which indicated absence of toxicity. In addition, MLE repressed the activation of nuclear factor kappa B by arresting the deterioration of IκB-α, consequently resulted in suppression of cytokines expression such as COX-2 and iNOS. Conclusions: MLE inhibitory activities are associated with the inhibition of nuclear factor kappa B transcriptional activity in BV2 microglial cells. Thus MLE may offer a substantial treatment for neuroinflammatory diseases.
基金The Space Medical Experiment Project of the China Manned Space Program(HYZHXM05003)National Natural Science Foundation of China(82171493)+2 种基金Natural Science Foundation of Hunan province(2021JJ30504)Scientific and Technological Innovation Project of the China Academy of Chinese Medical Sciences(CI2021A04905)Scientific Research Fund of Hunan Provincial Education of the Hunan University of Traditional Chinese Medicine First-class Discipline Project of Chinese Medicine(19B422)。
文摘Objective To study the therapeutic effects of Shenyuan Gan(参远苷,SYG)on the inflammat-ory response in BV2 microglial cells induced by lipopolysaccharide(LPS).Methods The cytotoxicity of SYG to BV2 microglial cells was evaluated using a Cell Counting Kit-8(CCK-8)assay,and the effect of SYG concentrations on LPS-induced BV2 microglial cells was studied.The morphological changes were observed using an optical microscope.The nitric oxide(NO)concentration in cell culture supernatant was determined using Griess re-agent.The expression of cytokines and inflammatory mediators were also measured by an en-zyme-linked immunosorbent assay(ELISA).Western blot analysis was used to determine the levels of inducible NO synthase(iNOS),nuclear factor-kappa B(NF-κB)p65,alpha inhibitor of NF-κB(IκB-α),phosphorylation-IκB-α(p-IκB-α),NOD-like receptor 3(NLRP3),and cas-pase-1 expression.Moreover,the expression of iNOS,NLRP3,and ionized calcium binding adapter molecule 1(Iba1)was also observed using immunofluorescent staining.Results SYG had a low cytotoxic effect on BV2 microglial cells and could significantly decr-ease LPS-induced morphological changes of BV2 microglial cells(P<0.05).ELISA results showed that SYG significantly inhibited the LPS-induced increase in interleukin(IL)-1βand IL-6 in BV2 microglia cells(P<0.05),and Western blot analysis showed that the phosphoryla-tion levels of iNOS,NF-κB p65,and IκB-αas well as NLRP3 and caspase-1 expression were also significantly decreased,and IκB-αexpression was increased after SYG treatment(P<0.05,compared with the LPS-treated group).The immunofluorescence results were consist-ent with the Western blot results,and Iba1 staining indicated that the cell morphology tended to be resting.These results indicate that SYG has a certain inhibitory effect on LPS-induced inflammation in BV2 microglial cells.Conclusion SYG can inhibit LPS-induced release of inflammatory factors in BV2 microglial cells by affecting the phosphorylation levels of NF-κB p65 and IκB-α.SYG is a valuable candid-ate for treating neuroinflammation-related diseases.
基金supported by a grant from the National Natu-ral Science Foundation of China(No.30900449)
文摘Little is known about whether tamoxifen (TAM) can affect resting state microglia apoptosis and about the cellular mechanism that may account for this. To explore this question, we incubated the microglia cell line BV-2 cells with TAM at different concentrations. Cell viability was assessed by the MTT assay, and flow cytometric analysis was performed to detect the cell apoptosis rate. Furthermore, mitochondrial membrane potential (Δψm) was tested by flow cytometry, and Bax, Bcl-2, Fas, and Fas-L expression was detected by Western blot. The results demonstrated that TAM decreased cell viability and induced apoptosis of BV-2 cells in a concentration- and time-dependent manner. In addition, disruption of Δψm was followed by up-regulated expression of pro-apoptotic Bax, Fas and Fas-L, and down-regulated expression of anti-apoptotic Bcl-2. These results indicate that TAM may induce apoptosis of BV-2 cells through both mitochondria- and death receptor-mediated pathways.
基金National Natural Science Foundations ofChina (30871854)National Science and Technology Supporting Program of China (2006BAD06A13)
文摘Prion diseases are infectious and fatal neurodegenerative diseases.The pathogenic agent is an abnormal prion protein aggregate.Microglial activation in the centre nervous system is a characteristic feature of prion disease.In this study,we examined the effect of PrP 106-126 on PrP mRNA gene expression in Mouse microglia cells BV-2 by real-time quantitative PCR.PrP mRNA expression level was found to be significantly increased after 18 h exposure of BV-2 cells to PrP 106-126,with 3-fold increase after 18 h and 4.5-fold increase after 24 h and BV-2 cells proliferating occurred correspondingly.Our results provide the first in vitro evidence of the increase of PrP mRNA levels in microglial cells exposed to PrP 106-126,and indicate that microglial cells might play a critical role in prion pathogenesis.
基金supported in part by grants from the Disciplinary Group of Psychology and Neuroscience Xinxiang Medical University(2016PN-KFKT-06)a visiting professorship from University of Tours(to LHJ)
文摘Microglial cells are the key innate immune cells in the brain and they are crucial in maintaining brain parenchyma homeostasis.Under physiological conditions,microglial cells assume a ramified morphology with a small cell body and an extensive network of fine processes,which secrete neurotrophic factors and patrol the surroundings in search for pathogens and eliminate cellular debris via phagocytosis.Microglial cells express a repertoire of pattern recognition receptors(PRRs)that enable them to detect diverse danger-associated molecular patterns(DAMPs)released from damaged cells or cells under stress,or pathogen-associated molecular patterns generated by pathogens during infection.
基金This study was supported by the Natural Science and Technology Foundation of Zunyi City,China,No.201915(to GPL)Doctor Startup Foundation of Zunyi Medical University,Nos.[2017]5733-045(to GPL),[2017]5733-044(to YYH)Natural Science and Technology Foundation of Guizhou Province,China,No.[2020]1Y292(to YYH).
文摘Microglial cells are important resident innate immune components in the central nervous system that are often activated during neuroinflammation.Activated microglia can display one of two phenotypes,M1 or M2,which each play distinct roles in neuroinflammation.Rutin,a dietary flavonoid,exhibits protective effects against neuroinflammation.However,whether rutin is able to influence the M1/M2 polarization of microglia remains unclear.In this study,in vitro BV-2 cell models of neuroinflammation were established using 100 ng/mL lipopolysaccharide to investigate the effects of 1-hour rutin pretreatment on microglial polarization.The results revealed that rutin pretreatment reduced the expression of the proinflammatory cytokines tumor necrosis factor-α,interleukin-1β,and interleukin-6 and increased the secretion of interleukin-10.Rutin pretreatment also downregulated the expression of the M1 microglial markers CD86 and inducible nitric oxide synthase and upregulated the expression of the M2 microglial markers arginase 1 and CD206.Rutin pretreatment inhibited the expression of Toll-like receptor 4 and myeloid differentiation factor 88 and blocked the phosphorylation of I kappa B kinase and nuclear factor-kappa B.These results showed that rutin pretreatment may promote the phenotypic switch of microglia M1 to M2 by inhibiting the Toll-like receptor 4/nuclear factor-kappa B signaling pathway to alleviate lipopolysaccharide-induced neuroinflammation.
文摘目的:探讨XMU-MP-1(Xiamen University-inhibitor of mammalian sterile 20-like kinase protein 1)对氧糖剥夺(OGD)损伤后小胶质细胞M1/M2极化平衡的调节作用。方法采用OGD法诱导BV2细胞损伤。实验分为6组:对照组、模型组、MST1/2 siRNA组和低、中、高剂量实验组(分别给予1.25、5.0和20.0μg·mL^(-1) XMU-MP-1)。采用MTT法测细胞活力,ELISA测细胞上清液中TNF-α、IL-6和IL-1β表达,qRT-PCR测M1和M2标志物的mRNA表达,流式细胞术测CD206表达,蛋白印迹法测MST1、LATS1和YAP蛋白表达。结果与模型组相比,XMU-MP-1抑制BV2细胞增殖,显著降低TNF-α、IL-6和IL-1β的表达水平,下调MCP-1、IL-6、TNF-α和i NOS mRNA的表达,上调CD206、IL-10、TGF-β、IL-10和YM1 mRNA表达,降低MST1和LAST 1蛋白表达,上调YAP和CD206表达。结论XMU-MP-1通过调控MST1/2的磷酸化,调节OGD损伤后的BV2细胞M1/M2极化平衡,为神经炎症靶点药物研发提供理论基础。
基金financially supported by the Natural Science Foundation of Shandong Province of China,No.ZR2014HM046(to ZCZ),ZR2015HL113(to XJD),and ZR2014HL101(to XYW)the Science and Technology Development Project of Taian City of China,No.2015NS2183(to XJD)
文摘P2X4 and P2X7 receptors play an important role in neuropathic pain after spinal cord injury. Regulation of P2X4 and P2X7 receptors can obviously reduce pain hypersensitivity after injury. To investigate the role of neural stem cell transplantation on P2X receptor-mediated neuropathic pain and explore related mechanisms, a rat model of spinal cord injury was prepared using the free-falling heavy body method with spinal cord segment 10 as the center. Neural stem cells were injected into the injured spinal cord segment using a micro-syringe. Expression levels of P2X4 and P2X7 receptors, neurofilament protein, and glial fibrillary acidic protein were determined by immunohistochemistry and western blot assay. In addition, sensory function was quantitatively assessed by current perception threshold. The Basso-Beattie-Bresnahan locomotor rating scale was used to assess neuropathological pain. The results showed that 4 weeks after neural stem cell transplantation, expression of neurofilament protein in the injured segment was markedly increased, while expression of glial fibrillary acidic protein and P2X4 and P2X7 receptors was decreased. At this time point, motor and sensory functions of rats were obviously improved, and neuropathic pain was alleviated. These findings demonstrated that neural stem cell transplantation reduced overexpression of P2X4 and P2X7 receptors, activated locomotor and sensory function reconstruction, and played an important role in neuropathic pain regulation after spinal cord injury. Therefore, neural stem cell transplantation is one potential option for relieving neuropathic pain mediated by P2X receptors.
文摘Inflammation plays an important role in the pathological process of ischemic stroke,and systemic inflammation affects patient prognosis.As resident immune cells in the brain,microglia are significantly involved in immune defense and tissue repair under various pathological conditions,including cerebral ischemia.Although the differentiation of M1 and M2 microglia is certainly oversimplified,changing the activation state of microglia appears to be an intriguing therapeutic strategy for cerebral ischemia.Recent evidence indicates that both mesenchymal stem cells(MSCs)and MSC-derived extracellular vesicles(EVs)regulate inflammation and modify tissue repair under preclinical stroke conditions.However,the precise mechanisms of these signaling pathways,especially in the context of the mutual interaction between MSCs or MSC-derived EVs and resident microglia,have not been sufficiently unveiled.Hence,this review summarizes the state-ofthe-art knowledge on MSC-and MSC-EV-mediated regulation of microglial activity under ischemic stroke conditions with respect to various signaling pathways,including cytokines,neurotrophic factors,transcription factors,and microRNAs.
基金supported by the Natural Science Foundation of Hunan Province of China,No.2017JJ3273(to ZJX)。
文摘Microglial apoptosis is associated with neuroinflammation and no effective strategies are currently available to protect microglia against inflammation-induced apoptosis. Mouse microglial BV-2 cells(5 × 10^6) were incubated with 10 μg/mL lipopolysaccharides for 12 hours to mimic an inflammatory environment. Then the cells were co-cultured with mitochonic acid 5(MA-5) for another 12 hours. MA-5 improved the survival of lipopolysaccharide-exposed cells. MA-5 decreased the activity of caspase-3, which is associated with apoptosis. MA-5 reduced the number of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells, and increased adenosine triphosphate levels in cells. MA-5 decreased the open state of the mitochondrial permeability transition pore and reduced calcium overload and diffusion of second mitochondria-derived activator of caspase(Smac). MA-5 decreased the expression of apoptosis-related proteins(mitochondrial Smac, cytoplasmic Smac, pro-caspase-3, cleaved-caspase-3, and caspase-9), and increased the levels of anti-apoptotic proteins(Bcl2 and X-linked inhibitor of apoptosis protein), mitochondria-related proteins(mitochondrial fusion protein 2, mitochondrial microtubule-associated proteins 1 A/1 B light chain 3 B II), and autophagy-related proteins(Beclin1, p62 and autophagy related 5). However, MA-5 did not promote mitochondrial homeostasis or decrease microglial apoptosis when Mitofusin 2 expression was silenced. This shows that MA-5 increased Mitofusin 2-related mitophagy, reversed cellular energy production and maintained energy metabolism in BV-2 cells in response to lipopolysaccharide-induced inflammation. These findings indicate that MA-5 may promote the survival of microglial cells via Mitofusin 2-related mitophagy in response to lipopolysaccharide-induced inflammation.
文摘目的探讨激活哺乳动物雷帕霉素靶蛋白复合物2(mTORC2)/Akt信号通路对6-羟基多巴胺(6-OHDA)模型小鼠多巴胺能神经元和行为学的影响及可能的机制。方法将36只体重20~25 g 3月龄Nestin-CreERTM::ROSA26-LacZ雄性C57BL/6J小鼠分为NS+玉米油组、6-OHDA+玉米油组、6-OHDA+PP242组、6-OHDA+A-443654组,并在小鼠右侧纹状体注射6-OHDA制备帕金森病(PD)小鼠模型以及每日腹腔注射mTORC2/Akt信号通路激动剂A-443654或抑制剂PP242。通过ELISA测定血清肿瘤坏死因子α(TNF-α)和白细胞介素1β(IL-1β)的水平;免疫组织化学和免疫荧光染色考察黑质(SN)-纹状体小胶质细胞、脑室周围神经前体细胞(NPCs)和多巴胺能神经元数目,Western blotting检测中脑水管mTORC2/Akt信号通路各相关蛋白Rictor,p-Akt和DNA损伤反应调节1(REDD1)的表达并通过免疫共沉淀验证它们之间的相互作用,最后观察各组小鼠行为学的变化。结果6-OHDA模型小鼠伴随小胶质细胞的激活和炎症因子的增加,黑质多巴胺能阳性神经元数目明显下降,小鼠的运动功能发生障碍,但NPCs数目较对照组小鼠明显增加,mTORC2/Akt信号通路相关蛋白表达也明显上调,在激动剂A-443654处理后随着相关蛋白的表达进一步上调,上述各指标均有明显改善,而抑制剂PP242处理组则呈现与激动剂A-443654完全相反的情况。结论A-443654通过上调mTORC2/Akt信号通路关键蛋白促进NPCs的增殖,增加新生多巴胺能神经元的数目并减少小胶质细胞的激活和炎症反应最终导致PD模型小鼠黑质-纹状体多巴胺能神经元和小鼠行为学的改善。
基金supported by grants from the University of Bourgogne(Dijon,France)the University of Monastir(Monastir,Tunisia)
文摘The treatment of microglial BV-2 cells with sodium arsenate（As（V）：0.1-400 μmol/L — 48 hr）induces a dose-dependent response.The neurotoxic effects of high concentrations of As（V）（100,200 and 400 μmol/L） are characterized by increased levels of mitochondrial complexesⅠ,Ⅱ,and Ⅳ followed by increased superoxide anion generation.Moreover,As（V） triggers an apoptotic mode of cell death,demonstrated by an apoptotic SubG1 peak,associated with an alteration of plasma membrane integrity.There is also a decrease in transmembrane mitochondrial potential and mitochondrial adenosine triphosphate ATP.It is therefore tempting to speculate that As（V） triggers mitochondrial dysfunction,which may lead to defective oxidative phosphorylation subsequently causing mitochondrial oxidative damage,which in turn induces an apoptotic mode of cell death.
