β-Ionone attenuates LPS-induced pro-inflammatory mediators such as NO,PGE2 and TNF-α in BV2 microglial cells via suppression of the NF-κB and MAPK pathway

β-Ionone, a precursor of carotenoids, possesses a variety of biological properties such as anti-cancerous, anti-mutagenic and anti-microbial activity. Nevertheless, anti-inflammatory effects of β-ionone remain unknown. In this study, we investigated whether ION attenuates the expression of lipopolysaccharide (LPS)-induced pro-inflammatory mediators such as nitric oxide (NO), prostaglandin E₂ (PGE₂) and tumor necrosis factor-α (TNF-α) in BV2 microglia cells. Our data showed that β-ionone significantly inhibits secretion of NO, PGE₂ and TNF-α. β-Ionone also inhibits the expression of inducible NO synthesis (iNOS), cyclooxygenase-2 (COX-2) and TNF-α protein and their mRNA in LPS-stimulated BV2 microglia cells. In addition, β-ionone significantly reduced DNA-binding activity of nuclear factor-κB (NF-κB) through suppression of nuclear translocation of p50 and p65. We showed that NF-κB inhibitor N-acetyl-L-cysteine (NAC) effectively attenuates the expression of LPS-stimulated iNOS, COX-2 and TNF-α. We also found that LPS-induced NF-κB activation is significantly regulated through inhibition of Akt phosphorylation in the presence of β-ionone. Finally, we showed that β-ionone substantially inhibits the phosphorylation of mitogen-activated protein kinases (MAPKs), including ERK1/2, p38 and JNK, which are closely related to regulation of pro-inflammatory mediator secretion. Taken together, these data imply that β-ionone regulates LPS-induced NF-κB-dependent inflammatory pathways through suppression of Akt and MAPK activation.     

Narirutin fraction from citrus peels attenuates LPS-stimulated inflammatory response through inhibition of NF-κB and MAPKs activation

In this study, we examined the regulatory activity of narirutin fraction from citrus peels on the production of inflammatory mediators managing acute or chronic inflammatory diseases in macrophages. Narirutin fraction inhibited the release, by lipopolysaccharide (LPS)-stimulated macrophages, of nitric oxide (NO) and prostaglandin E₂ (PGE₂) through suppressing the expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively. The release, by LPS stimulated macrophages, of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) was also reduced by narirutin fraction in a dose-dependent manner. Furthermore, narirutin fraction inhibited the LPS-mediated activation of nuclear factor-κB (NF-κB) and mitogen-activated protein kinases (MAPKs), which are signaling molecules involved in production of pro-inflammatory factors. As a result of these properties, narirutin fraction has the potential to be used as a functional dietary supplement and effective anti-inflammatory agent.     

Exopolysaccharide of Laetiporus sulphureus var. miniatus downregulates LPS-induced production of NO, PGE2, and TNF-α in BV2 microglia cells via suppression of the NF-κB pathway

Our previous study showed that the exopolysaccharide (EPS) of Laetiporus sulphureus var. miniatus was well characterized and prevented cell damage in streptozotocin-induced apoptosis. However, little is known about the molecular mechanisms underlying its anti-inflammatory effects. Therefore, we attempted in this study to determine whether EPS induces a significant inhibition of pro-inflammatory mediators in lipopolysaccharide (LPS)-stimulated murine BV2 microglia cells. Our results showed that EPS significantly inhibited LPS-induced pro-inflammatory mediators, such as nitric oxide (NO), prostaglandin E₂ (PGE₂), and tumor necrosis factor-α (TNF-α), without any significant cytotoxicity. EPS also downregulated mRNA and protein expression of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and TNF-α in LPS-induced BV2 microglia cells. Our data also revealed that EPS treatment significantly reduced translocation of nuclear factor-κB (NF-κB) subunit p65 and its DNA-binding activity in LPS-stimulated BV2 microglia cells. Furthermore, we confirmed by using proteasome inhibitor N-acetyl-l-cysteine (NAC), that the inhibition of NF-κB activity influenced the expression of pro-inflammatory genes in LPS-induced BV2 microglia cells. As expected, NAC suppressed the expression of iNOS, COX-2, and TNF-α by blocking proteasome-mediated degradation. Taken together, our data indicate that EPS inhibits the expression of pro-inflammatory mediators by suppressing NF-κB activity.     

Caffeine suppresses lipopolysaccharide-stimulated BV2 microglial cells by suppressing Akt-mediated NF-κB activation and ERK phosphorylation

Since the anti-inflammatory effect of caffeine is unclear in microglial cells, we performed whether caffeine attenuates the expression of pro-inflammatory mediators in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. Caffeine substantially suppressed the LPS-induced pro-inflammatory mediators nitric oxide (NO), prostaglandin E₂ (PGE₂) and tumor necrosis factor-α (TNF-α) in BV2 microglial cells. These effects resulted from the inhibition of their regulatory genes inducible NO synthase (iNOS), cycloxygenase-2 (COX-2) and TNF-α. In addition, caffeine significantly decreased LPS-induced DNA-binding activity of nuclear factor-κB (NF-κB) by suppressing the nuclear translocation of p50 and p65 subunits. A specific NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC), attenuated the LPS-induced expression of iNOS, COX-2 and TNF-α genes. In addition, we elucidated that inhibition of Akt phosphorylation plays a crucial role in caffeine-mediated NF-κB regulation in LPS-stimulated BV2 microglial cells. Caffeine also attenuated the LPS-induced phosphorylation of extracellular signal-regulated kinase (ERK) and a specific inhibitor of ERK, PD98059, subsequently downregulated the expression of the pro-inflammatory genes iNOS, COX-2 and TNF-α. Taken together, our data indicate that caffeine suppresses the generation of pro-inflammatory mediators, such as NO, PGE₂ and TNF-α as well as their regulatory genes in LPS-stimulated BV2 microglial cells by inhibiting Akt-dependent NF-κB activation and the ERK signaling pathway.     

Anti-neuroinflammatory effect of aurantiamide acetate from the marine fungus Aspergillus sp. SF-5921: Inhibition of NF-κB and MAPK pathways in lipopolysaccharide-induced mouse BV2 microglial cells

In the course of a search for anti-neuroinflammatory metabolites from marine fungi, aurantiamide acetate (1) was isolated from marine-derived Aspergillus sp. as an anti-neuroinflammatory component. Compound 1 dose-dependently inhibited the production of nitric oxide (NO) and prostaglandin E₂ (PGE₂) in BV2 microglial cells. It also attenuated inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and other pro-inflammatory cytokines, such as interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). In a further study designed to elucidate the mechanism of its anti-neuroinflammatory effect, compound 1 was shown to block the activation of nuclear factor-kappa B (NF-κB) in lipopolysaccharide (LPS)-induced BV2 microglial cells by inhibiting the phosphorylation of the inhibitor kappa B-α (IκB)-α. In addition, compound 1 decreased the phosphorylation levels of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinases (MAPKs). These results suggest that compound 1 has an anti-neuroinflammatory effect on LPS stimulation through its inhibition of the NF-κB, JNK and p38 pathways.     

Theobromine mitigates IL-1β-induced oxidative stress,inflammatory response,and degradation of type II collagen in human chondrocytes

Osteoarthritis is one of the major causes of disability in elderly adults. Chondrocytes are responsible for the formation and remodeling of articular cartilage in joint tissue. The dysfunction of chondrocytes is a significant factor in the development of osteoarthritis. In the current study, we found that theobromine, a constituent of the cacao plant, possesses a preventive effect against interleukin (IL)-1β-induced chondrocyte dysfunction. Theobromine ameliorates IL-1β-induced production of cellular reactive oxygen species (ROS) and inflammatory mediators including cyclooxygenase-2 (COX-2) and prostaglandin E₂ (PGE₂). The presence of theobromine suppresses IL-1β-induced inducible nitro oxide synthase (iNOS) expression and cellular nitro oxide (NO) production. Theobromine also suppresses IL-1β-induced production of the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and monocyte chemoattractant protein-1 (MCP-1), as well as matrix metalloproteinases (MMP)-3 and MMP-13. Additionally, theobromine mitigates IL-1β-induced type II collagen degradation. Mechanistically, we show that theobromine inhibits IL-1β-induced IκBα activation, nuclear factor-κB (NF-κB) protein p65 accumulation, and transfected NF-κB promoter activity, indicating that theobromine suppresses the NF-κB pathway in chondrocytes. Collectively, our study demonstrates that the natural molecule theobromine has a protective effect to counter cytokine-induced chondrocyte dysfunction, implying its beneficial effect in the prevention of osteoarthritis.     

狐臭柴茎挥发油体外抗炎活性及基于NF-κB和MAPKs信号通路作用机制的研究

目的:探究狐臭柴茎挥发油的体外抗炎活性及其作用机制。方法:通过水蒸气蒸馏法制备狐臭柴茎挥发油,采用GC-MS对其化学成分进行分析。通过Griess法和ELISA法测定挥发油对LPS诱导RAW264.7细胞上清液中一氧化氮(NO)、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)水平的影响;qRT-PCR检测iNOS、COX-2、TNF-α、IL-6、IL-1β mRNA的表达;Western blot测定挥发油对iNOS、COX-2、NF-κB和MAPKs信号通路蛋白的影响。结果:从挥发油中共鉴定出74种化学成分,其中主要的化学成分是茅术醇(13.50%)。挥发油显著抑制NO、TNF-α和IL-6的分泌(P＜0.01),同时降低了促炎因子(TNF-α、IL-6和IL-1β)和促炎酶(iNOS和COX-2)的mRNA表达水平(P＜0.01)。Western blot研究表明挥发油能下调NF-κB信号通路细胞核p65蛋白表达、p65和IκBα磷酸化(P＜0.05或P＜0.01)。此外,还降低了MAPKs信号通路p38、JNK和ERK蛋白的磷酸化(P＜0.01)。结论:狐臭柴茎挥发油对LPS诱导RAW264.7细胞炎症模型具有较好的抗炎效果,其内在的分子机制与下调NF-κB和MAPKs信号通路有关。     

A supercritical CO₂ extract from seabuckthorn leaves inhibits pro-inflammatory mediators via inhibition of mitogen activated protein kinase p38 and transcription factor nuclear factor-κB

In the present study, we have demonstrated the anti-inflammatory properties of supercritical CO₂ extract of seabuckthorn leaves (SCE) on mouse alveolar macrophage cell line (MH-S), human peripheral blood mononuclear cells (hPBMCs) in-vitro and in-vivo. Treatment of MH-S cells with SCE (0.5–100 μg/ml) significantly inhibited lipopolysaccharide (LPS)-stimulated nitric oxide (NO) production. It also inhibited the release of LPS-induced pro-inflammatory cytokines IL-6 and TNF-α, which was further confirmed by suppression of LPS induced TNF-α in hPBMCs by ELISPOT assay. In addition, western blot analysis demonstrated that SCE decreased LPS-induced inducible nitric-oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression in MH-S cells. Furthermore, SCE treatment also reduced nuclear factor-κB (NF-κB) translocation in nucleus induced by LPS in MH-S cells. To elucidate the molecular mechanism for inhibition of pro-inflammatory mediators by SCE (100 μg/ml), we further studied the effect of SCE on LPS-induced p38 mitogen-activated protein kinase (MAPK). It was observed that the phosphorylation of p38 MAPK in LPS-stimulated MH-S cells was significantly inhibited by SCE, which was further proven by suppression of LPS induced CD40 expression. The in-vivo model of AIA mice also showed a significant reduction in the inflammation of paw edema. These data collectively suggest that SCE suppressed the LPS-induced production of NO, IL-6, and TNF-α and expression of CD40, iNOS and COX-2 proteins by inhibiting NF-κB activation and phosphorylation of p38 MAPK. Hence, the SCE has potent anti-inflammatory activity and might be useful in chronic inflammatory diseases.     

Britanin suppresses LPS-induced nitric oxide,PGE2 and cytokine production via NF-κB and MAPK inactivation in RAW 264.7 cells

Little is known about the biological properties of britanin, which is isolated from the flowers of Inula japonica (Inulae Flos). Based on our previous studies that Inulae Flos had anti-inflammation and anti-asthmatic activities, we tried to find the bioactive compounds from it. In this study, the anti-inflammatory effects of britanin on the inflammatory mediators as well as on nuclear factor (NF)-кB and mitogen-activated protein (MAP) kinase activation were evaluated in RAW 264.7 cells. Britanin inhibited the production of nitric oxide (NO) and prostaglandin E2 (PGE₂) along with the expression of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. In addition, britanin reduced the release of pro-inflammatory cytokines, such as TNF-α, IL-1β, and IL-6. Furthermore, the phosphorylations of MAP kinases (p38 and JNK) in LPS-stimulated RAW 264.7 cells were suppressed by britanin. Moreover, britanin inhibited the NF-κB activation induced by LPS, which was associated with the abrogation of IκBα degradation and subsequent decreases in nuclear p65 levels. This study suggests that the anti-inflammatory activities of britanin might be attributed to the inhibition of iNOS and COX-2 and cytokine expression at least in part, through the attenuation of the phosphorylations of MAP kinases and NF-κB activation via IκBα degradation in macrophages. We conclude that britanin may have potential for the treatment of inflammatory diseases through the down-regulation of MAP kinases and NF-κB mediated activation of macrophages.     

Water extract of processed Hydrangea macrophylla (Thunb.) Ser. leaf attenuates the expression of pro-inflammatory mediators by suppressing Akt-mediated NF-κB activation

Although Hydrangea macrophylla is native to Northeast Asia and widely cultivated in many parts of the world, no studies on its anti-inflammatory effects have been reported. In this study, we evaluated the anti-inflammatory effect of a water extract of processed H. macrophylla leaf (WH) in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage cells. WH inhibited the expression of LPS-stimulated pro-inflammatory mediators such as nitric oxide (NO), prostaglandin E₂ (PGE₂), and tumor necrosis factor-α (TNF-α), as well as their regulatory genes inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and TNF-α without any accompanying cytotoxicity. Moreover, WH significantly suppressed the LPS-induced DNA-binding activity of nuclear factor-κB (NF-κB), as well as the nuclear translocation of the NF-κB subunits, p65 and p50 by suppressing of IκBα phosphorylation and degradation. WH also increased Akt dephosphorylation, leading to the suppression of the DNA-binding activity of NF-κB in LPS-stimulated RAW264.7 macrophage cells. Our results indicate that WH downregulates the expression of pro-inflammatory mediators such as NO, PGE₂, and TNF-α by suppressing the Akt-mediated NF-κB activity in LPS-stimulated RAW264.7 macrophage cells.     

Effects of volatile oils of Angelica sinensis on an acute inflammation rat model

Context Despite several pharmacological studies of volatile oils of Angelica sinensis (Oliv.) Diels (Umbelliferae) (VOAS), its anti-inflammatory mechanism remains unknown.

Objective The study investigates the effects of VOAS on the lipopolysaccharide (LPS)-induced acute inflammation rat model and analyzes its possible anti-inflammatory mechanisms.

Materials and methods Fourty rats were randomly divided into the control, model, VOAS and dexamethasone (Dex) groups. The VOAS and Dex groups were given VOAS (0.176?mL/kg) and Dex (40?μg/kg), respectively. Rats in all groups except the control group were intraperitoneally injected with LPS (100?μg/kg), their exterior behaviour and liver pathological changes were observed, and the level of white blood cell (WBC), the number of neutrophils (NE)%, glutamic oxalacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), alkaline phosphatase (ALP), tumour necrosis factor (TNF-α), interleukin (IL)-1β, IL-6, IL-10, histamine (HIS), 5-hydroxytryptamine (5-HT), nitric oxide (NO), prostaglandin E₂ (PGE₂), inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) were detected.

Results Compared with the model group, VOAS and Dex significantly accelerated the recovery of the exterior behaviour, the liver pathological changes of rats, and increased the level of IL-10, but decreased the level of WBC, NE%, GOT, GPT, ALP, TNF-α, IL-1β, IL-6, HIS, 5-HT, NO, PGE₂, iNOS and COX-2 (p?<?0.05).

Conclusion VOAS exhibits anti-inflammatory and liver protection effects by inhibiting the secretion of the pro-inflammatory cytokines (TNF-α, IL-1β and IL-6), the inflammatory mediators (HIS, 5-HT, PGE₂ and NO), the inflammation-related enzymes (iNOS and COX-2), as well as promoting the production of the anti-inflammatory cytokines IL-10.     

Pro-inflammatory effects of commercial alpha-lactalbumin on RAW 264.7 macrophages is due to endotoxin contamination

This study investigated the effects of alpha-lactalbumin (α-LA) on cellular signaling molecules associated with inflammatory responses in RAW 264.7 macrophages. The results indicated that commercial α-LA could increase prostaglandin E₂ (PGE₂) and the expression of COX-2 via increased phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2), p38 mitogen-activated protein kinase (MAPK) and jun N-terminal kinase (JNK), and increase nitric oxide (NO) and the expression of iNOS via the activation of ERK1/2 and JNK. Furthermore, commercial α-LA could increase nuclear translocation of p65 nuclear factor-kappa B (p65 NF-κB) through stimulation on inhibitor kappa B-alpha (IκB-α) degradation. Since endotoxin also has these effects, we assayed the content of endotoxin in the commercial α-LA. We found to our surprise that endotoxin was there and that α-LA-induced NO and PGE₂ production could be suppressed by polymyxin B, a specific inhibitor of endotoxin. Thus, the pro-inflammatory effects of commercial α-LA might be caused by endotoxin contamination through activation and expression of iNOS and COX-2 which were upregulated by MAPKs or nuclear translocation of p65 NF-κB in RAW 264.7 cells. It is therefore crucial to assess the possibility of endotoxin contamination within any biological product being studied for immune augmenting activities before a meaning result can be obtained.     

龙胆苦苷对慢性前列腺炎的作用及对NF-κB和MAPKs信号通路活化的研究

目的探讨龙胆苦苷对慢性前列腺炎大鼠的作用及相关分子机制。方法采用完全弗氏佐剂诱导自身免疫性慢性前列腺炎大鼠模型,评估龙胆苦苷对大鼠前列腺器官指数、组织病理形态,以及大鼠前列腺组织中炎症因子IL-6、IL-1β、TNF-α和炎症介质COX-2、PGE2的表达水平及iNOS酶活性的影响。采用Western blot法检测大鼠前列腺组织中NF-κB和MAPKs通路相关分子蛋白表达水平。结果龙胆苦苷对慢性前列腺炎有保护作用,其可以降低前列腺器官指数,缓解前列腺组织病理损伤,此外,龙胆苦苷下调大鼠前列腺组织中炎症因子IL-6、IL-1β、TNF-α和炎症介质COX-2、PGE2的表达水平及iNOS酶活性,下调p-p65、p-ERK/ERK、p-JNK/JNK、p-p38/p38的表达水平。结论龙胆苦苷对慢性前列腺炎具有明显的保护作用,且这种作用可能与抑制NF-κB和MAPKs信号通路活化有关。     

Ginsenoside-Rd exhibits anti-inflammatory activities through elevation of antioxidant enzyme activities and inhibition of JNK and ERK activation in vivo

Our previous study has reported that ginsenoside-Rd significantly inhibited the production of pro-inflammatory cytokines and mediators in carrageenan (Carr)-induced rat paw edema, which might be due to its blocking of the nuclear factor-κB (NF-κB) signaling pathway. The aim of the present study was to clarify the more detailed mechanisms of anti-inflammatory activity of ginsenoside-Rd in Carr-induced rat paw edema model. Rats were pretreated with dexamethasone or ginsenoside-Rd 1 h before the Carr injection. Six hours after Carr injection, the malondialdehyde (MDA) level and myeloperoxidase (MPO), superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) activities in inflamed paw tissues were determined. The levels of nitric oxide (NO) and prostaglandin E₂ (PGE₂) in serum were measured. The expressions of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and NF-κB were detected by western blot. In addition, the extent of phosphorylation of extracellular signal-regulated protein kinase (ERK), p38 and c-Jun NH₂-terminal kinase (JNK) was analyzed by western blot. The results showed that ginsenoside-Rd significantly attenuated MPO activity and MDA level, increased the activities of SOD, GPx and CAT, lowered the levels of NO and PGE₂, down-regulated the expressions of iNOS, COX-2 and NF-κB, and suppressed the phosphorylation of ERK and JNK. Taken together, the possible mechanisms of anti-inflammatory activity of ginsenoside-Rd were: it could reduce the inflammatory cell infiltration into inflammatory sites, inhibit the tissue lipid peroxidation, increase the antioxidant enzyme activities, and suppress the proinflammatory enzyme expressions through the downregulation of NF-κB activation via suppression of ERK and JNK phosphorylation.     

Anti-inflammatory effects of Cynanchum taiwanianum rhizome aqueous extract in IL-1β-induced NRK-52E cells

Context: Cynanchum taiwanianum T. Yamaza (Asclepiadaceae) is a medicinal herb used in folk medicine for the treatment of several inflammation-related diseases such as hepatitis and dermatitis in Taiwan.

Objective: In the present study, we investigated the anti-inflammatory effect of C. taiwanianum T. Yamaza rhizome aqueous extract (CTAE).

Materials and methods: The present study investigated the anti-inflammatory effect of CTAE using IL-1β-induced NRK-52E cells. Production of NO and PGE₂ by ELISA, the mRNA and protein expression of iNOS and COX-2, phosphorylation of IκBα, and activation of NF-κB by RT-PCR and western blotting were determined.

Results: The CTAE significantly (P?<?0.05) inhibited NO and PGE₂ production (decreased by 46.1% and 51%, respectively), and also significantly (P?<?0.05) attenuated protein and mRNA expression of iNOS and COX-2 (decreased by 90% and 55% for iNOS and by 72% and 74%% for COX-2, respectively) in IL-1β-induced NRK-52E cells, in a dose-dependent manner, without obvious cytotoxic effects. Furthermore, the CTAE suppressed the NF-κB nuclear translocation, in terms of inhibition of IκBα phosphorylation.

Discussion and conclusion: Our results provided evidence for its folkloric uses and suggest that the anti-inflammatory activities of CTAE may result from the inhibition of inflammatory mediators, such as NO and PGE₂, and an upstream suppression of a NF-κB-dependent mechanism, might be involved.