Polygonum viviparum L. inhibits the lipopolysaccharide‐induced inflammatory response in RAW264.7 macrophages through haem oxygenase‐1 induction and activation of the Nrf2 pathway

BACKGROUND: Polygonum viviparum L. (PV) is a member of the family Polygonaceae and is widely distributed in high‐elevation areas. It is used as a folk remedy to treat inflammation‐related diseases. This study was focused on the anti‐inflammatory response of PV against lipopolysaccharide (LPS)‐induced inflammation in RAW264.7 macrophages. RESULTS: Treatment with PV did not cause cytotoxicity at 0–50 µg mL^?1 in RAW264.7 macrophages, and the IC₅₀ value was 270 µg mL^?1. PV inhibited LPS‐stimulated nitric oxide (NO), prostaglandin (PG)E₂, interleukin (IL)‐1β and tumour necrosis factor (TNF)‐α release and inducible NO synthase (iNOS) and cyclooxygenase (COX)‐2 protein expression. In addition, PV suppressed the LPS‐induced p65 expression of nuclear factor (NF)‐κB, which is associated with the inhibition of IκB‐α degradation. These results suggest that, among mechanisms of the anti‐inflammatory response, PV inhibits the production of NO and these cytokines by down‐regulating iNOS and COX‐2 gene expression. Furthermore, PV can induce haem oxygenase (HO)‐1 protein expression through nuclear factor E2‐related factor 2 (Nrf2) activation. A specific inhibitor of HO‐1, zinc(II) protoporphyrin IX, inhibited the suppression of iNOS and COX‐2 expression by PV. CONCLUSION: These results suggest that PV possesses anti‐inflammatory actions in macrophages and works through a novel mechanism involving Nrf2 actions and HO‐1. Thus PV could be considered for application as a potential therapeutic approach for inflammation‐associated disorders. © 2012 Society of Chemical Industry     

Anti‐inflammatory properties of high pressure‐assisted extracts of Grifola frondosa in lipopolysaccharide‐activated RAW 264.7 macrophages

The aim of this study was to determine the in vitro anti‐inflammatory properties of the shake extract (SE) and the high pressure‐assisted extract (PE) of the mycelia of Grifola frondosa in a lipopolysaccharide‐stimulated RAW 264.7 macrophage model. The content of total polysaccharides and β‐glucans of PE at 600 MPa (PE‐600) was 41.2 and 6.2 mg g^?1 dry weight, respectively, which were significantly higher than SE extracts. The results showed that treatment with 500 μg mL^?1 of PE by 600 MPa (PE‐600) did not reduce RAW 264.7 cell viability but did significantly inhibit the production of LPS‐induced NO, PGE₂ and intracellular ROS. The PE‐600 inhibited the activation of NF‐kB and then reduced the production of LPS‐induced TNF‐α, IL‐6 and IL‐1β in a dose‐dependent manner. Thus, the PE could be used as an alternative extraction method for improving the extraction efficacy of G. frondosa and serve as an alternative source of anti‐inflammatory agents.     

Enhanced Anti‐Inflammatory Activities by the Combination of Luteolin and Tangeretin

Dietary components in combination may act synergistically and produce enhanced biological activities. Herein, we investigated the anti‐inflammatory effects of 2 flavonoids, that is luteolin (LUT) and tangeretin (TAN) in combination. Lipopolysaccharide (LPS)‐stimulated RAW 264.7 macrophages were treated with noncytotoxic concentrations of LUT, TAN, and their combinations. The results showed that LUT/TAN in combination produced synergistic inhibitory effects on LPS‐stimulated production of nitric oxide (NO). ELISA results demonstrated that LUT/TAN in combination caused stronger suppression on the LPS‐induced overexpression of proinflammatory mediators, such as prostaglandin E₂ (PGE₂), interleukin (IL)‐1β, and IL‐6 than LUT or TAN alone. Immunoblotting and Real‐Time PCR analyses showed that LUT/TAN combination significantly decreased LPS‐induced protein and mRNA expression of inducible nitric oxide synthase and cyclooxygenase‐2. These inhibitory effects of the combination treatment were stronger than those produced by LUT or TAN alone. Overall, our results demonstrated for the first time that combination of LUT and TAN produced synergistic anti‐inflammatory effects in LPS‐stimulated RAW 264.7 macrophages.     

Characterization,Anti‐Inflammatory and Antiproliferative Activities of Natural and Sulfonated Exo‐Polysaccharides from Streptococcus thermophilus ASCC 1275

Exo‐polysaccharides (EPS) isolated from Streptococcus thermophilus ASCC 1275 were sulfated (31%). High‐performance liquid chromatography identified that EPS was composed of mannose (30.19%), galactose (20.10%), glucose (18.05%), glucosamine (16.04%), galactosamine (9.06%), glucuronic acid (3.55%), and ribose (3.01%). Pro‐/anti‐inflammatory cytokine secretion ratios (IL‐1β/IL‐10, IL‐6/IL‐10, and TNF‐α/IL‐10) of lipopolysaccharide stimulated RAW 264.7 macrophages were significantly decreased by EPS and S.EPS treatments in a dose dependent manner. Furthermore, anti‐inflammatory activities of S.EPS improved 49.3% and 24.0% than those of EPS before or after LPS treatment. The reactive oxygen species were inhibited by EPS and S.EPS by 49.6% and 55.1% at 50 μg/mL, respectively. Inhibition activities of S.EPS on nitric oxide production were 12.9% and 55.4% higher than those of EPS at 10 and 50 μg/mL. Additionally, S.EPS exhibited stronger antiproliferative activity on Caco‐2 and HepG2 cells. Our results indicated that anti‐inflammatory and antiproliferative activities of EPS were significantly (P < 0.01) improved by sulfonation.     

Effects of Catechol O‐Methyl Transferase Inhibition on Anti‐Inflammatory Activity of Luteolin Metabolites

Although luteolin is known to have potent anti‐inflammatory activities, much less information has been provided on such activities of its hepatic metabolites. Luteolin was subjected to hepatic metabolism in HepG2 cells either without or with catechol O‐methyl transferase (COMT) inhibitor. To identify hepatic metabolites of luteolin without (luteolin metabolites, LMs) or with COMT inhibitor (LMs+CI), metabolites were treated by β‐glucuronidase and sulfatase, and found that they were composed of glucuronide and sulfate conjugates of diosmetin in LMs or these conjugates of luteolin in LMs+CI. LMs and LMs+CI were examined for their anti‐inflammatory activities on LPS stimulated Raw 264.7 cells. Expression of iNOS and production of nitric oxide and pro‐inflammatory cytokines such as TNF‐α, IL‐1β, and IL‐6 were suppressed more effectively by the treatment with LMs+CI than LMs. Our data provide a new insight on possible improvement in functional properties of luteolin on target cells by modifying their metabolic pathway in hepatocytes.     

Phenethyl isothiocyanate suppresses nitric oxide production via inhibition of phosphoinositide 3‐kinase/Akt‐induced IFN‐γ secretion in LPS‐activated peritoneal macrophages

Phenethyl isothiocyanate (PEITC), a constituent of many cruciferous vegetables, is well known to have versatile physiological activities, including chemopreventive effects. On the other hand, its anti‐inflammatory effects are poorly reported. Nitric oxide (NO) is associated with a wide variety of inflammatory diseases. In this study, we investigated the effects of PEITC on NO production in LPS‐activated peritoneal macrophages from ICR mice. The signaling pathway of LPS‐induced NO production was examined using neutralizing antibodies [anti‐interferon (IFN)‐γ and anti‐interleukin (IL‐12)] and specific protein kinase inhibitors, as well as others. The activity of PEITC toward NOx production was assessed in mice that received LPS via intraperitoneal administration. The neutralizing antibody of anti‐IFN‐γ, but not anti‐IL‐12, suppressed LPS‐induced NO production by 90%. LY294002, a specific inhibitor of phosphoinositide‐3‐kinase, suppressed Akt and IFN‐γ mRNA expression up‐regulated by LPS, whereas PEITC exhibited a similar inhibition profile. Furthermore, oral administration of PEITC significantly suppressed the serum concentration of NOx in ICR mice. Our results suggest that PEITC suppresses LPS‐induced NO production via inhibition of Akt activation and the resultant decrease in expression of IFN‐γ. This is one of the first reports to demonstrate a marked anti‐inflammatory effect of PEITC following its oral administration.     

Protective effect of dried safflower petal aqueous extract and its main constituent,carthamus yellow,against lipopolysaccharide‐induced inflammation in RAW264.7 macrophages

BACKGROUND: Safflower, whose botanic name is Carthamus tinctorius L., is a member of the family Compositae or Asteraceae. Carthamus yellow (CY) is the main constituent of safflower and is composed of safflomin A and safflomin B. Dried safflower petals are used in folk medicine and have been shown to invigorate blood circulation, break up blood stasis, and promote menstruation. In addition, dried safflower petals contain yellow dyes that are used to color food and cosmetics. In this study, we investigated the effects of dried safflower petals aqueous extracts (SFA) and CY on lipopolysaccharide (LPS)‐induced inflammation using RAW264.7 macrophages. RESULTS: Our data showed that treatment with SFA (1–1000 µg mL^?1) and CY (1–2000 µg mL^?1) does not cause cytotoxicity in cells. SFA and CY inhibited LPS‐stimulated nitric oxide (NO), prostaglandin E₂ (PGE₂), and interleukin 1β (IL‐1β) release, through attenuation of inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2) protein expression. Further, SFA and CY suppressed the LPS‐induced phosphorylation of nuclear factor‐κB, which was associated with the inhibition of IκB‐α degradation. CONCLUSION: These results suggest that SFA and CY provide an anti‐inflammatory response through inhibiting the production of NO and PGE₂ by the downregulation of iNOS and COX‐2 gene expression. Thus safflower petals have the potential to provide a therapeutic approach to inflammation‐associated disorders. Copyright © 2010 Society of Chemical Industry     

Bovine casein peptides co‐stimulate naive macrophages with lipopolysaccharide for proinflammatory cytokine production and nitric oxide release

Three bovine casein peptides, ie LLY, PGPIPN and TTMPLW, were used to investigate their effects on cytokine (TNF‐α and IL‐6) production and nitric oxide (NO) release by murine bone marrow macrophages (BMMs). The results showed that these peptides alone were incapable of stimulating cytokine production or NO release in naive or IFN‐γ‐primed BMMs. However, when BMMs were co‐incubated with the peptides at a concentration of 1.0 µM and lipopolysaccharide (LPS; 100 ng ml⁻¹), an augmentative effect on TNF‐α, IL‐6 and NO production was observed. Of the three peptides, TTMPLW had the greatest augmentative effect on NO production by LPS‐stimulated BMMs and induced the highest amount of TNF‐α production at a concentration of 1.0 µM . All the peptides at a concentration of 1.0 µM stimulated IL‐6 production by BMMs. TNF‐α was neutralised by anti‐TNF‐α monoclonal antibody and the release of NO was reduced by about 33.3% (p < 0.01). These results demonstrate that bovine casein peptides can co‐stimulate naive macrophages with LPS for proinflammatory cytokine production and NO release and may play a role in host defence against pathogens. © 2000 Society of Chemical Industry     

Mung Bean Protein Hydrolysate Modulates the Immune Response Through NF‐κB Pathway in Lipopolysaccharide‐Stimulated RAW 264.7 Macrophages

The objective of this study was to evaluate the immunomodulatory activity of mung bean protein hydrolysate (MBPH) in lipopolysaccharide (LPS)‐induced RAW 264.7 cells and discuss the possible immune regulatory mechanism. MBPH was prepared by alcalase, trypsin, neutrase, and flavourzyme. The 3‐h alcalase‐hydrolyzed hydrolysate with a molecular weight less than 1,450 Da was selected for the immunological tests. Results showed that MBPH possessed strong suppressing activity to proinflammatory mediators in a dose‐dependent manner. Compared to the LPS alone group, MBPH (200 µg/mL) significantly reduced nitric oxide (NO), inducible nitric oxide synthase, interleukin (IL)‐6, and IL‐1β secretion levels by 52.6%, 53.2%, 48.4%, and 49.7%, respectively, in LPS‐induced macrophages. It also enhanced IL‐10 secretion from 789 to 3,678 pg/mL. MBPH blocked nuclear factor‐kappa B (NF‐κB) translocation in LPS‐induced macrophages through the prevention of IκBα phosphorylation, and this process further prevented p65 translocation into the nucleus. A possible mechanism of MBPH is that it regulated the expression of inflammatory factors via the NF‐κB pathway, thus inhibiting inflammatory reactions. The results suggested that MBPH is of application potential in the development of immunomodulatory functional food to ameliorate immunosuppression.     

Anti-inflammatory activities of aqueous extract of Mesona procumbens in experimental mice

BACKGROUND: Mesona procumbens is consumed as a herbal drink and jelly‐type dessert in Taiwan. The aim of this study was to determine the mechanism of anti‐inflammatory activities of the aqueous extract of M. procumbens (AMP) using the λ‐carrageenin (Carr)‐induced mouse paw oedema model. The fingerprint chromatogram of AMP was obtained by high‐performance liquid chromatography (HPLC) analysis. To investigate the anti‐inflammatory mechanism of AMP, the activities of catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx) and the level of malondialdehyde (MDA) in paw oedema were monitored. Serum nitric oxide (NO), tumour necrosis factor‐α (TNF‐α) and interleukin‐1β (IL‐1β) were also evaluated. RESULTS: The fingerprint chromatogram from HPLC indicated that AMP contained protocatechuic acid, chlorogenic acid, vanillic acid and caffeic acid. In the anti‐inflammatory test, AMP decreased paw oedema after Carr administration and increased the CAT, SOD and GPx activities and decreased the MDA level in paw oedema at 5 h after Carr injection. AMP also affected the serum NO, TNF‐α and IL‐1β levels at 5 h after Carr injection. Western blotting revealed that AMP decreased the expression of Carr‐induced inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2). CONCLUSION: Mesona procumbens has the potential to provide a therapeutic approach to inflammation‐associated disorders. Copyright © 2011 Society of Chemical Industry     

Anti‐inflammatory activity of rosemary extracts obtained by supercritical carbon dioxide enriched in carnosic acid and carnosol

The in vitro anti‐inflammatory activity of supercritical rosemary (Rosmarinus officinalis L.) extracts (rosemary A and B) is been reported in this study. To achieve that, THP‐1 macrophages were activated using lipopolysaccharide or human ox‐LDL and secretion and gene expression of TNF‐α, IL‐1β, IL‐6 and IL‐10 were evaluated, as well as COX‐2 gene expression. Results indicated that both rosemary extracts (A & B) exhibit high anti‐inflammatory activity although at a higher extent in case of rosemary B extract (5 μg mL^?1), representing a higher quantity of carnosic acid and carnosol than rosemary A. When comparing the activity of the extract to the standard itself, the anti‐inflammatory activity of standards of carnosic acid and carnosol was not as intense as that obtained with rosemary B. These data indicated that although carnosic acid content in the extracts is considered as the main anti‐inflammatory compound, a synergistic interaction with other compounds may play a significant role in enhancing its activity. Results provided the grounds for possible increase in the application of supercritical rosemary extracts in food formulations for mitigation or prevention of inflammatory diseases.     

Phase II enzyme induction and anti‐inflammatory effects of crude extracts and secondary fractions obtained from bran from five black glutinous rice cultivars

Methanolic extracts of bran from black glutinous rice cultivars were screened for potential health benefits using cellular bioassays for quinone reductase (QR) induction in murine hepatoma cells and inhibition of NO production in lipopolysaccharide (LPS)‐activated RAW 264.7 macrophages. Fractionation of the crude extract by semi‐preparative HPLC afforded respective fractions enriched in phenolic acids, anthocyanins and proanthocyanidins. The relative potency of QR induction was phenolic acids > proanthocyanidins > anthocyanins, and the activity of the crude extract was conserved among the aggregate of these fractions. In contrast, all three fractions were more potent than the crude extract in terms of anti‐inflammatory effect. This suggests synergism effects among anti‐inflammatory agents through partial purification. Inhibition of macrophage NO production by binary mixtures of gallic acid or caffeic acid with the anthocyanin‐rich fraction indicated potential synergism. These results warrant further efforts to identify the active agents and substantiate synergistic interactions through isobologram analysis.     

羊栖菜多酚通过核转录因子-κB/丝裂原活化蛋白激酶通路缓解脂多糖诱导的RAW264.7细胞炎症反应

目的：研究羊栖菜多酚对脂多糖（lipopolysaccharide,LPS）诱导小鼠单核巨噬细胞白血病细胞RAW264.7细胞炎症反应的影响。方法：噻唑蓝法测定细胞活力;Griess法测定一氧化氮（NO）水平;实时荧光定量聚合酶链式反应测定白细胞介素（interleukin,IL）-6、IL-1β、肿瘤坏死因子（tumor necrosis factor,TNF）-α、环氧合酶2（cyclooxygenase-2,COX-2）和诱导型一氧化氮合酶（inducible nitric oxide synthase,iNOS）的基因相对表达量;流式细胞术测定细胞吞噬能力;蛋白免疫印迹法测定信号通路丝裂原活化蛋白激酶（mitogen-activated protein kinases,MAPKs）和核转录因子（nuclear factor,NF）-κB信号通路关键蛋白表达水平。结果：羊栖菜多酚对RAW264.7细胞的安全质量浓度范围为0～160 μg/mL。与LPS组相比,羊栖菜多酚剂量依赖性降低巨噬细胞吞噬能力并抑制NO的生成。同时,羊栖菜多酚下调促炎细胞因子（IL-1β、IL-6、TNF-α）和炎症诱导酶（iNOS、COX-2）的mRNA水平,且作用效果与给药剂量及LPS刺激时间相关。这些炎性介质的表达与羊栖菜多酚抑制p38 MAPK和NF-κB p65的激活有关。结论：羊栖菜多酚可通过减弱p38 MAPK和NF-κB p65信号通路的激活水平,抑制下游炎症介质的转录表达,从而缓解LPS诱导的巨噬细胞炎症反应。     

Anti‐inflammatory and anticancer activities of water‐soluble peptide extracts of buffalo and cow milk Cheddar cheeses

This article aimed to assess the anti‐inflammatory and anticancer potential of water‐soluble peptide (WSP) extracts from buffalo and cow milk Cheddar cheeses. Anti‐inflammatory activity was evaluated on the basis of nitric oxide (NO) production in lipopolysaccharide‐stimulated macrophage (RAW‐264.7) cells. A cell viability assay, cell cycle arrest and apoptosis were performed to explore anticancer activity in a colon cancer model (HT‐29). The WSP extracts of both Cheddar cheeses effectively inhibited NO production in activated macrophages. Maximum growth inhibition was observed in the HT‐29 cells at concentrations of 400 and 500 μg/mL. A significant increase in cell population at G₀/G₁ phase of the cell cycle was observed. Moreover, the WSP extracts also induced extensive apoptosis in colon cancer cells.     

The preventive role of breadfruit against inflammation‐associated epithelial carcinogenesis in mice

Artocarpus communis has been identified as a rich source of flavonoids and has been gaining attention for its potential chemopreventive abilities. In this study, methanol extracts from the fruit of A. communis (MEFA) and leaf of A. communis (MELA) were prepared, and their effects on inflammation‐associated skin tumorigenesis were assessed using mouse models, including 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) induced cutaneous inflammation as well as 7,12‐dimethylbenz[α]anthracene (DMBA) initiated and TPA‐promoted skin tumorigenesis. According to the results, both MEFA and MELA decreased the intensity of leukocyte infiltration in mouse dorsal skin and cutaneous edema induced by TPA, which appeared to be mediated by inhibition of proinflammatory genes (inducible nitric oxide synthase, cyclooxygenase‐2 (COX‐2), tumor necrosis factor‐α (TNF‐α), IL‐1β, and IL‐6) and proinflammatory mediators (TNF‐α, IL‐1β, and Prostaglandin E₂). In addition, topical application with MEFA or MELA effectively attenuated tumor incidence, multiplicity, volume, malignancy as well as angiogenesis of TPA‐stimulated skin tumor promotion in DMBA‐initiated mice. Notably, immunohistochemical stain showed that MEFA and MELA attenuated COX‐2 expression of both skin and tumor tissues in different animal tests, which may be closely related to the suppression of nuclear factor kappa B/activator protein signaling networks. These findings first demonstrate that flavonoid‐rich A. communis may exert potent anti‐inflammatory activity through modulation of COX‐2 in TPA‐activated skin and tumor tissues.