• 대한본초학회지
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• 제30권1호
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• pp.77-84
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• 2015

Objectives : Brain inflammation early activates the microglia and activated microglia secrete a variety of pro-inflammatory cytokines. Kaempferol, which is a flavonoid in Cuscutae Semen, shows a wide range of physiological activities, including neurons protection and anti-inflammatory actions through inhibition of pro-inflammatory mediators. The present study examined the modulatory effect of kaempferol on cytokines [tumor necrosis factor- alpha ($TNF-{\alpha}$), interleukin-1beta ($IL-1{\beta}$) and interleukin-6 (IL-6)] and cyclooxygenase-2 (COX-2) mRNA expression and microglia activation in the brain tissue of the mouse. Methods : Kaempferol was administered orally three doses of 10, 20 and 30 mg/kg respectively, once 1 hour before the lippolysaccharide(LPS) (3 mg/kg, i.p.) injection. Brain tissue was removed at 4 hours after LPS injection. Cytokines and COX-2 mRNA expression in the brain tissue was measured by the quantitative real-time polymerase chain reaction (PCR) method. Iba1 expression was calculated by western blotting method. Microglia was observed with immunohistochemistry. Immunohistochemistry stained microglia was analyzed by using ImageJ software. Results : Kaempferol 20 and 30 mg/kg was significantly attenuated the expression of $TNF-{\alpha}$, $IL-1{\beta}$ and IL-6 mRNA. Kaempfrol 10, 20 and 30 mg/kg significantly attenuated COX-2 mRNA expression in the brain tissue. Kaempferol 30 mg/kg significantly suppressed the increase of Iba1 protein expression by LPS. Kaempferol 30 mg/kg significantly decreased the number of microglia in the cerebral cortex and the number and cell size of microglia in the hypothalamic region and the area percentage of ionized calcium binding adaptor molecule 1(Iba1)-expressed microglia in the hippocampus. Conclusions : This results indicate that kaempferol plays an anti-inflammatory role in the brain.

• Biomolecules & Therapeutics
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• 제21권1호
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• pp.21-28
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• 2013

Microglia play a role in maintaining and resolving brain tissue homeostasis. In pathological conditions, microglia release pro-inflammatory cytokines and cytotoxic factors, which aggravate the progression of neurodegenerative diseases. Autophagy pathway might be involved in the production of pro-inflammatory cytokines and cytotoxic factors in microglia, though details of the mechanism remain largely unknown. In the present study, we examined the role of the autophagy pathway in activated BV2 microglia cells. In BV2 cells, rapamycin treatment activated the formation of anti-LC3-labeled autophagosomes, whereas the ATG5 depletion using siRNA-ATG5 prevented the formation of LC3-labeled autophagosomes, indicating that BV2 cells exhibit an active classical autophagy system. When treated with LPS, BV2 cells expressed an increase of anti-LC3-labeled dots. The levels of LC3-labeled dots were not suppressed, instead tended to be enhanced, by the inhibition of the autophagy pathway with siRNA-ATG5 or wortmannin, suggesting that LPS-induced LC3-labeled dots in nature were distinct from the typical autophagosomes. The levels of LPS-induced expression of iNOS and IL6 were suppressed by treatment with rapamycin, and conversely, their expressions were enhanced by siRNA-ATG5 treatment. Moreover, the activation of the autophagy pathway using rapamycin inhibited cell death of LPS-stimulated microglia. These results suggest that although microglia possess a typical autophagy pathway, the glial cells express a non-typical autophagy pathway in response to LPS, and the activation of the autophagy pathway suppresses the expression of iNOS and IL6, and the cell death of LPS-stimulated microglia.

• 한국식생활문화학회지
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• 제38권3호
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• pp.176-183
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• 2023

In this study, we investigated the effect of the extracts of Cyrtomium fortunei J.Sm. (CFJ) on lipopolysaccharide (LPS) induced inflammation in mouse BV-2 microglial cells. Nitric oxide (NO) production and cell viability were measured using the Griess reagent and the (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) (MTT) assay. Inflammatory cytokines were detected by quantitative polymerase chain reaction (qPCR) in BV-2 microglial cells with and without CFJ extracts. Subsequently, mitogen-activated protein kinases (MAPKs) and antioxidant markers were assessed by western blot analysis. It was found that the CFJ extract significantly decreased the production of pro-inflammatory cytokines (interleukin [IL]-6, tumor necrosis factor [TNF]-α, and IL-1β) and NO in BV-2 microglial cells that were stimulated with LPS. In addition, the expression levels of the phosphorylation of the MAPK family (p38, c-Jun N-terminal kinases [JNK], and extracellular-signal regulated kinase [ERK]) were reduced by CFJ. Also, treatment with CFJ significantly increased the activities of superoxide dismutase type 1(SOD1) and Catalase in BV-2 microglial cells. Our results indicate that CFJ has a potent suppressive effect on the pro-inflammatory responses of activated BV-2 microglia. Therefore, CFJ has the potential to be an effective treatment for neurodegenerative diseases, as it can inhibit the production of inflammatory mediators in activated BV-2 microglial cells.

• 대한본초학회지
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• 제23권3호
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• pp.93-102
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• 2008

Objectives : Forsythiae Fructus (Forsythia koreana Nakai) has been used anti-inflammatory, diuretics, antidote, and antibacterials in traditional herbal medicine. The present study is focused on the inhibitory effect of Forsythiae Fructus ethanol extract (FF-E) on the production of inflammatory mediators such as NO, iNOS and proinflammatory cytokines ($TNF-{\alpha}$, $IL-1{\beta}$ and IL-6) in LPS-stimulated BV-2 cells, a mouse microglial cell line, and investigated the scavenging activity of FF-E. Methods : BV-2 cells were pre-incubated with FF-E for 30 min and then stimulated with LPS (1 ${\mu}g/m{\ell}$) at indicated times. Cell toxicity of GCF was determined by MTT assay. The levels of NO, PGE2 and cytokines were measured by Griess assay and ELISA. The mRNA and protein expressions of iNOS and cytokines were determined by RT-PCR and Western blotting. Free radical scavenging activity of GCF was determined by DPPH assay in tube test. Results : FF-E significantly inhibited the excessive production of NO, $PGE_2$, $TNF-{\alpha}$, and $IL-1{\beta}$ in LPS-stimulated BV-2 cells. In addition, FF-E attenuated the mRNA and protein expressions of iNOS, and proinflammatory cytokines. FF-E also significantly scavenged the DPPH free radicals in a dose-dependent manner. Conclusions : These results indicate that FF-E exhibits anti-inflammatory property by suppressing the transcription of inflammatory mediator genes, suggesting the anti-inflammatory property of FF-E may make it useful as a therapeutic agent for the treatment of human neurodegenerative diseases.

• 대한본초학회지
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• 제28권2호
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• pp.93-101
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• 2013

Objects : Baicalein is a major bioactive flavonoid component of Scutellaria baicalensis Georgi that shows a wide range of biological activities, including neuroprotections and anti-inflammatory actions. Hence it is a potential therapeutic material for the treatment of neuroinflammation. In this study, we investigated the modulatory effect of baicalein on neuroinflammation. Method : Pro-inflammatory cytokines (TNF-${\alpha}$, IL-$1{\beta}$ and IL-6 mRNA), COX-2 mRNA expression and microglial activation in the brain tissue is induced by systemic lipopolysaccharide (LPS) treatment in C57BL/6 mice. Baicalein was treated orally with 10, 20, and 30 mg/kg 1 hour prior to the LPS (3 mg/kg, i.p.) injection. TNF-${\alpha}$, IL-$1{\beta}$, IL-6 and COX-2 mRNA expression in the brain tissue was measured by the quantitative real-time polymerase chain reaction(PCR) method. Iba1 expression in the brain was measured by western blotting method. Microglia was observed with immunohistochemistry. Results : Baicalein 30 mg/kg significantly attenuated the expression of TNF-${\alpha}$, IL-$1{\beta}$, IL-6 and COX-2 mRNA in the brain tissue. Baicalein 20 mg/kg significantly attenuated the expression of IL-6 mRNA in the brain tissue. Baicalein 30 mg/kg significantly attenuated the expression of Iba1 protein expression in the brain tissue. Baicalein 30 mg/kg significantly decreased the number and cell size of microglia in the cerebral cortex and hypothalamic region and the area percentage of Iba1-expressed microglia in the hippocampus. Conclusion : These results demonstrated that baicalein attenuates LPS induced neuroinflammation in the mice via reduction of pro-inflammatory cytokines (TNF-${\alpha}$, IL-$1{\beta}$, IL-6), COX-2 mRNA expression and microglial activation.

• Archives of Pharmacal Research
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• 제27권9호
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• pp.954-960
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• 2004

Expression of the NF-$textsc{k}$B-dependent genes responsible for inflammation, such as TNF-$\alpha$, IL-1$\beta$, and nitric oxide synthase (NOS), contributes to chronic inflammation which is a major cause of neurodegenerative diseases (i.e. Alzheimer＇s disease). Although NF-$textsc{k}$B plays a biphasic role in different cells like neurons and microglia, controlling the activation of NF-$textsc{k}$B is important for its negative feedback in either activation or inactivation. In this study, we found that ursodeoxycholic acid (UDCA) inhibited I$textsc{k}$B$\alpha$ degradation to block expression of the NF-$textsc{k}$B-dependent genes in microglia when activated by $\beta$-amyloid peptide (A$\beta$). We also showed that when microglia is activated by $A\beta$42, the expression of A20 is suppressed. These findings place A20 in the category of ' protective ' genes, protecting cells from pro-inflammatory reper-toires induced in response to inflammatory stimuli in activated microglia via NF-$textsc{k}$B activation. In light of the gene and proteins for NF-$textsc{k}$B-dependent gene and inactivator for NF-$textsc{k}$B (I$textsc{k}$B$\alpha$), the observations now reported suggest that UDCA plays a role in supporting the attenuation of the production of pro-inflammatory cytokines and NO via inactivation of NF-$textsc{k}$B. Moreover, an NF-$textsc{k}$B inhibitor such as A20 can collaborate and at least enhance the anti-inflammatory effect in microglia, thus giving a potent benefit for the treatment of neurodegenerative diseases such as AD.uch as AD.

• BMB Reports
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• 제42권6호
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• pp.380-386
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• 2009

In neurodegenerative diseases, such as Alzheimer' and Parkinson', microglial cell activation is thought to contribute to CNS injury by producing neurotoxic compounds. Prothrombin and kringle-2 increase levels of NO and the mRNA expression of iNOS, IL-1$\beta$, and TNF-$\alpha$ in microglial cells. In contrast, the human prothrombin kringle-2 derived peptide NSA9 inhibits NO release and the production of pro-inflammatory cytokines such as IL-1$\beta$, TNF-$\alpha$, and IL-6 in LPS-activated EOC2 microglia. In this study, we investigated the anti-inflammatory effects of NSA9 in human prothrombin- and kringle-2-stimulated EOC2 microglia. Treatment with 20-100 ${\mu}M$ of NSA9 attenuated both prothrombin- and kringle-2-induced microglial activation. NO production induced by MAPKs and NF-$\kappa$B was similarly reduced by inhibitors of ERK (PD98059), p38 (SB203580), NF-$\kappa$B (N-acetylcysteine), and NSA9. These results suggest that NSA9 acts independently as an inhibitor of microglial activation and that its effects in EOC2 microglia are not influenced by the presence of kringle-2.

• 한방재활의학과학회지
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• 제31권4호
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• pp.1-11
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• 2021

Objectives Hwanggeumjakyak-tang (HJT) has traditionally been used to treat gastrointestinal inflammatory diseases; however, its protective effects against neuronal inflammation are still undiscovered. Methods We investigated the anti-neuroinflammatory effects of HJT water extract on lipopolysaccharide (LPS)-stimulated BV2 mouse microglia cells. BV2 cells were treated with LPS (1 ㎍/mL) 1 hour prior to the addition of HJT. We measured cell viability using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and nitrite production using the Griess assay. We performed a reverse transcription-polymerase chain reaction assay to measure messenger RNA expression of inflammatory cytokines including interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α. Western blot analysis was performed to determine protein expression of mitogen-activated protein kinases (MAPKs) and inhibitor of nuclear factor kappa B (NF-κB)α. Results HJT inhibited excessive nitrite release in LPS-stimulated BV2 cells and also significantly inhibited inflammatory cytokines such as IL-1β, IL-6, and TNF-α in LPS-stimulated BV2 cells. Moreover, HJT significantly suppressed LPS-induced MAPK and NF-κB activation and inhibited the elevation of IL-1β, IL-6, and TNF-α in the brain of LPS-injected mice. Conclusions Our study highlights the anti-neuroinflammatory effects of HJT via MAPK and NF-κB deactivation.

• BMB Reports
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• 제54권11호
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• pp.557-562
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• 2021

Microglial activation is closely associated with neuroinflammatory pathologies. The nucleotide-binding and oligomerization domain-like receptor containing a pyrin domain 3 (NLRP3) inflammasomes are highly organized intracellular sensors of neuronal alarm signaling. NLRP3 inflammasomes activate nuclear factor kappa-B (NF-κB) and reactive oxygen species (ROS), which induce inflammatory responses. Moreover, NLRP3 dysfunction is a common feature of chronic inflammatory diseases. The present study investigated the effect of a novel thiazol derivative, N-cyclooctyl-5-methylthiazol-2-amine hydrobromide (KHG26700), on inflammatory responses in lipopolysaccharide (LPS)-treated BV-2 microglial cells. KHG26700 significantly attenuated the expression of several pro-inflammatory cytokines, including tumor necrosis factor-α, interleukin-1β, and interleukin-6, in these cells, as well as the LPS-induced increases in NLRP3, NF-κB, and phospho-IkBα levels. KHG26700 also suppressed the LPS-induced increases in protein levels of autophagy protein 5 (ATG5), microtubule-associated protein 1 light chain 3 (LC3), and beclin-1, as well as downregulating the LPS-enhanced levels of ROS, lipid peroxidation, and nitric oxide. These results suggest that the anti-inflammatory effects of KHG26700 may be due, at least in part, to the regulation of the NLRP3-mediated signaling pathway during microglial activation.

• Parasites, Hosts and Diseases
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• 제49권3호
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• pp.285-290
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• 2011

Naegleria fowleri, a ubiquitous free-living ameba, causes fatal primary amebic meningoencephalitis in humans. N. fowleri trophozoites are known to induce cytopathic changes upon contact with microglial cells, including necrotic and apoptotic cell death and pro-inflammatory cytokine release. In this study, we treated rat microglial cells with amebic lysate to probe contact-independent mechanisms for cytotoxicity, determining through a combination of light microscopy and scanning and transmission electron microscopy whether N. fowleri lysate could effect on both necrosis and apoptosis on microglia in a time- as well as dose-dependent fashion. A $^{51}Cr$ release assay demonstrated pronounced lysate induction of cytotoxicity (71.5%) toward microglial cells by 24 hr after its addition to cultures. In an assay of pro-inflammatory cytokine release, microglial cells treated with N. fowleri lysate produced TNF-${\alpha}$, IL-6, and IL-$1{\beta}$, though generation of the former 2 cytokines was reduced with time, and that of the last increased throughout the experimental period. In summary, N. fowleri lysate exerted strong cytopathic effects on microglial cells, and elicited pro-inflammatory cytokine release as a primary immune response.