Astaxanthin exerts anti-inflammatory and antioxidant effects in macrophages in NRF2-dependent and independent manners

Although anti-inflammatory effects of astaxanthin (ASTX) have been suggested, the underlying mechanisms have not been fully understood. Particularly, the modulatory action of ASTX in the interplay between nuclear factor E2-related factor 2 (NRF2) and nuclear factor κB (NFκB) to exert its anti-inflammatory effect in macrophages is unknown. The effect of ASTX on mRNA and protein expression of pro-inflammatory and antioxidant genes and/or cellular reactive oxygen species (ROS) accumulation were determined in RAW 264.7 macrophages, bone marrow-derived macrophages (BMDM) from wild-type (WT) and Nrf2-deficient mice, and/or splenocytes and peritoneal macrophages of obese mice fed ASTX. The effect of ASTX on M1 and M2 macrophage polarization was evaluated in BMDM. ASTX significantly decreased LPS-induced mRNA expression of interleukin 6 (Il-6) and Il-1β by inhibiting nuclear translocation of NFκB p65; and attenuated LPS-induced ROS with an increase in NRF2 nuclear translocation, concomitantly decreasing NADPH oxidase 2 expression in RAW 264.7 macrophages. In BMDM of WT and Nrf2-deficient mice, ASTX decreased basal and LPS-induced ROS accumulation. The induction of Il-6 mRNA by LPS was repressed by ASTX in both types of BMDM while Il-1β mRNA was decreased only in WT BMDM. Furthermore, ASTX consumption lowered LPS sensitivity of splenocytes in obese mice. ASTX decreased M1 polarization of BMDM while increasing M2 polarization. ASTX exerts its anti-inflammatory effect by inhibiting nuclear translocation of NFκB p65 and by preventing ROS accumulation in NRF2-dependent and -independent mechanisms. Thus, ASTX is an agent with anti-inflammatory and antioxidant properties that may be used for the prevention of inflammatory conditions.     

Evaluation of analgesic and anti-inflammatory activities and molecular docking analysis of steroidal lactones from Datura stramonium L.

BackgroundDatura stramonium L. is widely used across the world for its therapeutic potential to treat inflammatory disorders. The current work was designed to isolate and identify steroidal lactones from D. stramonium leaves and evaluate their anti-inflammatory and analgesic properties.MethodsSeveral compounds were isolated from D. stramonium leaves and characterized by nuclear magnetic resonance and high-resonance electron spray ionization mass spectrometry techniques. Further, anti-inflammatory properties of these compounds were evaluated by in vitro assays, such as release of NO and pro-inflammatory cytokines by lipopolysaccharide (LPS)-activated J774A.1 macrophages. Using in vivo models, anti-inflammatory and analgesic effects were examined by mouse tail-flick, carrageenan-induced inflammation in rat paw model, vascular permeability in rats, and acetic acid-induced writhing in mice. The docking studies were performed for assessing the binding efficiency of the test compounds with cyclooxygenase-1 (COX-1) and COX-2, lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), inducible nitric oxide synthases (iNOS) and nuclear factor-κB (NF-κB).ResultsThree lactones were isolated and confirmed as daturalactone (D1), 12-deoxywithastramonolide (D23), and daturilin (D27). Further, the isolated compounds showed nitric oxide inhibition and pro-inflammatory cytokines released by LPS-activated J774A.1 macrophages. The in vivo results suggest that D1, D23 and D27 (20 mg/kg) were able to reduce the pain and inflammation in various animal models. The docking analysis showed that these three compounds actively bind with COX-1, COX-2, LOX-1, NF-κB, and iNOS, validating the anti-inflammatory effects of the lactones.ConclusionThese findings demonstrate substantial anti-inflammatory and analgesic properties of D. stramonium-derived lactones and their potential as anti-inflammatory agents to treat chronic inflammatory ailments.     

CD73 Activity is Dispensable for the Polarization of M2 Macrophages

The ectoenzyme CD73 catalyzes the hydrolysis of AMP, and is one of the most important producers of extracellular adenosine. On regulatory T cells, CD73 is necessary for immunosuppressive functions, and on Th17 cells CD73-generated adenosine exerts anti-inflammatory effects. However, the expression and function of CD73 in pro-inflammatory M1 and in immunosuppressive M2 macrophages is largely unknown. Here we show that CD73 expression and enzyme activity were induced in in vitro polarized pro-inflammatory human M(LPS+TNF) monocytes/macrophages, while CD73 was absent from immunosuppressive M(IL-4+M-CSF)-polarized macrophages. Inhibition of CD73 activity with the inhibitor AMPCP did not affect the polarization of human monocytes. In mice, CD73 was present on resident peritoneal macrophages. In striking contrast, elicited peritoneal macrophages remained CD73 negative regardless of their polarization towards either a pro-inflammatory M(LPS) or anti-inflammatory M(IL-4c) direction. Finally, the ability of peritoneal macrophages to polarize to pro- and anti-inflammatory cells was perfectly normal in CD73-deficient mice in vivo. These data indicate that, in contrast to other major leukocyte subpopulations, CD73 activity on macrophages does not play a major role in their polarization and that in mice host CD73 on any cell type is not required in vivo for peritoneal macrophage polarization towards either a pro- or an anti-inflammatory direction.     

Piliation of Lactobacillus rhamnosus GG Promotes Adhesion,Phagocytosis, and Cytokine Modulation in Macrophages

Recently, spaCBA-encoded pili on the cell surface of Lactobacillus rhamnosus GG were identified to be key molecules for binding to human intestinal mucus and Caco-2 intestinal epithelial cells. Here, we investigated the role of the SpaCBA pilus of L. rhamnosus GG in the interaction with macrophages in vitro by comparing the wild type with surface mutants. Our results show that SpaCBA pili play a significant role in the capacity for adhesion to macrophages and also promote bacterial uptake by these phagocytic cells. Interestingly, our data suggest that SpaCBA pili also mediate anti-inflammatory effects by induction of interleukin-10 (IL-10) mRNA and reduction of interleukin-6 (IL-6) mRNA in a murine RAW 264.7 macrophage cell line. These pili appear to mediate these effects indirectly by promoting close contact with the macrophages, facilitating the exertion of anti-inflammatory effects by other surface molecules via yet unknown mechanisms. Blockage of complement receptor 3 (CR3), previously identified to be a receptor for streptococcal pili, significantly decreased the uptake of pilus-expressing strains in RAW 264.7 cells, while the expression of IL-10 and IL-6 mRNA by these macrophages was not affected by this blocking. On the other hand, blockage of Toll-like receptor 2 (TLR2) significantly reduced the expression of IL-6 mRNA irrespective of the presence of pili.     

Docosahexaenoic acid attenuates macrophage-induced inflammation and improves insulin sensitivity in adipocytes-specific differential effects between LC n-3 PUFA

ObjectiveAdipose tissue inflammation with immune cell recruitment plays a key role in obesity-induced insulin resistance (IR). Long-chain (LC) n-3 polyunsaturated fatty acids (PUFA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have anti-inflammatory potential; however, their individual effects on adipose IR are ill defined. We hypothesized that EPA and DHA may differentially affect macrophage-induced IR in adipocytes.MethodsJ774.2 macrophages pretreated with EPA or DHA (50 μM for 5 days) were stimulated with lipopolysaccharide (LPS, 100 ng/ml for 30 min–48 h). Cytokine secretion profiles and activation status of macrophages were assessed by enzyme-linked immunosorbent assay and flow cytometry. Pretreated macrophages were seeded onto transwell inserts and placed over 3T3-L1 adipocytes for 24–72 h; effects on adipocyte–macrophage cytokine cross-talk and insulin-stimulated ³H-glucose transport into adipocytes were monitored.ResultsDHA had more potent anti-inflammatory effects relative to EPA, with marked attenuation of LPS-induced nuclear factor (NF)κB activation and tumor necrosis factor (TNF)α secretion in macrophages. DHA specifically enhanced anti-inflammatory interleukin (IL)-10 secretion and reduced the expression of proinflammatory M1 (F4/80⁺/CD11⁺) macrophages. Co-culture of DHA-enriched macrophages with adipocytes attenuated IL-6 and TNFα secretion while enhancing IL-10 secretion. Conditioned media (CM) from DHA-enriched macrophages attenuated adipocyte NFκB activation. Adipocytes co-cultured with DHA-enriched macrophages maintained insulin sensitivity with enhanced insulin-stimulated ³H-glucose transport, GLUT4 translocation and preservation of insulin-receptor substrate-1 expression compared to co-culture with untreated macrophages. We confirmed that IL-10 expressed by DHA-enriched macrophages attenuates the CM-induced proinflammatory IR phenotype in adipocytes.ConclusionsWe demonstrate an attenuated proinflammatory phenotype of DHA-pretreated macrophages, which when co-cultured with adipocytes partially preserved insulin sensitivity.     

Jatrophane polyesters from the leaves of Euphorbia peplus with anti-inflammatory activity

Three new jatrophane polyesters, Euphopepluanones M-O (1–3), along with eight known ones (4-11), were isolated from the leaves of Euphorbia peplus Linn. Their chemical structures were established based on extensive spectroscopic analysis. The absolute configurations of compound 1 were assigned by X-ray crystallographic analysis. In addition, their anti-inflammatory effects on lipopolysaccharide-induced RAW264.7 macrophages were evaluated in vitro. Among them, 2 and 7 exhibited significant anti-inflammatory activity.     

Tubule-mitophagic secretion of SerpinG1 reprograms macrophages to instruct anti-septic acute kidney injury efficacy of high-dose ascorbate mediated by NRF2 transactivation

High-dose ascorbate confers tubular mitophagy responsible for septic acute kidney injury (AKI) amelioration, yet its biological roles in immune regulation remain poorly understood.Methods: The role of tubular mitophagy in macrophage polarization upon high-dose ascorbate treatment was assessed by fluorescence-activated cell sorter analysis (FACS) in vitro and by immunofluorescence in AKI models of LPS-induced endotoxemia (LIE) from Pax8-cre; Atg7^flox/flox mice. The underlying mechanisms were revealed by RNA-sequencing, gene set enrichment analysis (GSEA), luciferase reporter, chromatin immunoprecipitation (ChIP) and adeno-associated viral vector serotype 9 (AAV9) delivery assays.Results: High-dose ascorbate enables conversion of macrophages from a pro-inflammatory M1 subtype to an anti-inflammatory M2 subtype in murine AKI models of LIE, leading to decreased renal IL-1β and IL-18 production, reduced mortality and alleviated tubulotoxicity. Blockade of tubular mitophagy abrogates anti-inflammatory macrophages polarization under the high-dose ascorbate-exposed coculture systems. Similar abrogations are verified in LIE mice with tubular epithelium-specific ablation of Atg7, where the high-dose ascorbate-inducible renal protection and survival improvement are substantially weaker than their control littermates. Mechanistically, high-dose ascorbate stimulates tubular secretion of serpin family G member 1 (SerpinG1) through maintenance of mitophagy, for which nuclear factor-erythroid 2 related factor 2 (NRF2) transactivation is required. SerpinG1 perpetuates anti-inflammatory macrophages to prevent septic AKI, while kidney-specific disruption of SerpinG1 by adeno-associated viral vector serotype 9 (AAV9)-short hairpin RNA (shRNA) delivery thwarts the anti-inflammatory macrophages polarization and anti-septic AKI efficacy of high-dose ascorbate.Conclusion: Our study identifies SerpinG1 as an intermediate of tubular mitophagy-orchestrated myeloid function during septic AKI and reveals a novel rationale for ascorbate-based therapy.     

The antioxidant and anti-inflammatory effects of abalone intestine digest, Haliotis discus hannai in RAW 264.7 macrophages

Abalone is a marine gastropod and an important fishery and food industrial resource that is massively maricultured in Asia, Africa, Australia, and America. However, the health beneficial effects of abalone have rarely been reported. In the present study, we examine the antioxidant and anti-inflammatory effects of abalone Haliotis discus hannai in macrophage cells. The results showed that abalone intestine digest (AID) has antioxidant activities against lipid peroxidation, ROS stress and DNA damage in H₂O₂-treated RAW264.7 macrophages. In the lipopolysaccharide (LPS)-induced macrophages, AID suppresses LPS-induced production of nitric oxide (NO) via inducible nitric oxide synthase (iNOS) expression in a dose-dependent manner. It also significantly reduced the generation of proinflammatory cytokines, such as interleukin (IL-1??), tumor necrosis factor (TNF)-??, and IL-6. Furthermore, AID significantly suppresses the phosphorylation of mitogen-activated protein kinases (MAPKs) such as ERK, JNK, and p38. These results indicated that AID inhibits oxidative damage by ROS and LPS-induced inflammatory response via blocking the MAPK signaling pathway in murine macrophages. The potent antioxidant and anti-inflammatory effects of abalone intestine as byproducts from fishery manufacturing may suggest the possibility of high valuable utilization and application as a nutraceutical and therapeutic substance.     

Peripheral Macrophage-derived Exosomes promote repair after Spinal Cord Injury by inducing Local Anti-inflammatory type Microglial Polarization via Increasing Autophagy

Treatment for spinal cord injury (SCI) remains a challenge worldwide, and inflammation is a major cause of secondary injury after SCI. Peripheral macrophages (PMs) have been verified as a key factor that exert anti-inflammatory effects after SCI, but the mechanism is unidentified. As local macrophages, microglia also exert significant effects after SCI, especially polarization. Exosomes show source cell-like biological functions to target cells and have been the subject of much research in recent years. Thus, we hypothesized the PM-derived exosomes (PM-Exos) play an important role in signal transmission with local microglia and can be used therapeutic agents for SCI in a series of in vivo and in vitro studies. For the in vivo experiment, three groups of Sprague-Dawley (SD) rats subjected to spinal cord contusion injury were injected with 200 µg/ml PM-Exos, 20 µg/ml PM-Exos or PBS via the tail vein. Recovery of the rats and of spinal cord function were observed. In vitro, we investigated the potential anti-inflammatory mechanism of PM-Exos and evaluated microglial autophagy, anti-inflammatory type microglia polarization and the upstream signaling pathway. The results showed that spinal cord function and recovery were better in the PM-Exo groups than the control group. In the in vitro study, microglial autophagy levels and the expression of anti-inflammatory type microglia were higher in the experimental groups than the control group. Moreover, the expression of proteins related to the PI3K/AKT/mTOR autophagic signaling pathway was suppressed in the PM-Exo groups. PM-Exos have a beneficial effect in SCI, and activation of microglial autophagy via inhibition of the PI3K/AKT/mTOR signaling pathway, enhancing the polarization of anti-inflammatory type microglia, that may play a major role in the anti-inflammatory process.     

In vivo and in vitro anti-inflammatory activity of Cryptostegia grandiflora Roxb. ex R. Br. leaves

Background

Despite Cryptostegia grandiflora Roxb. ex R. Br. (Apocynaceae) leaves are widely used in folk Caribbean Colombian medicine for their anti-inflammatory effects, there are no studies that support this traditional use. Therefore, this work aimed to evaluate the effect of the total extract and primary fractions obtained from Cryptostegia grandiflora leaves, using in vivo and in vitro models of inflammation, and further get new insights on the mechanisms involved in this activity.

Results

Ethanolic extract of Cryptostegia grandiflora leaves, and its corresponding ether and dichloromethane fractions, significantly reduced inflammation and myeloperoxidase activity (MPO) in ear tissue of mice treated with 12-O-tetradecanoyl-phorbol-13-acetate (TPA). Histological analysis revealed a reduction of edema and leukocyte infiltration. Complementarily, we demonstrated that extract and fractions reduced nitric oxide (NO•) and prostaglandin E2 (PGE2) production in LPS-stimulated RAW 264.7 macrophages, as well as scavenging activity on DPPH and ABTS radicals.

Conclusions

Our results demonstrated for the first time the anti-inflammatory activity of Cryptostegia grandiflora leaves, supporting its traditional use. This activity was related to inhibition of MPO activity, and PGE2 and NO• production. These mechanisms and its antioxidant activity could contribute, at least in part, to the anti-inflammatory effect showed by this plant.     

Targeting of a Fixed Bacterial Immunogen to Fc Receptors Reverses the Anti-Inflammatory Properties of the Gram-Negative Bacterium,Francisella tularensis,during the Early Stages of Infection

Production of pro-inflammatory cytokines by innate immune cells at the early stages of bacterial infection is important for host protection against the pathogen. Many intracellular bacteria, including Francisella tularensis, the agent of tularemia, utilize the anti-inflammatory cytokine IL-10, to evade the host immune response. It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity. The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date. In the current study, we hypothesized that F. tularensis polarizes antigen presenting cells during the early stages of infection towards an anti-inflammatory status characterized by increased synthesis of IL-10 and decreased production of IL-12p70 and TNF-α in an IFN-ɣ-dependent fashion. In addition, F. tularensis drives an alternative activation of alveolar macrophages within the first 48 hours post-infection, thus allowing the bacterium to avoid protective immunity. Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10. More specifically, we observed that targeting of iFt to FcγRs enhances the classical activation of macrophages not only within the respiratory mucosa, but also systemically, at the early stages of infection. These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.     

Berry anthocyanins suppress the expression and secretion of proinflammatory mediators in macrophages by inhibiting nuclear translocation of NF-κB independent of NRF2-mediated mechanism

The objectives of this study were to compare the anti-inflammatory effects of anthocyanins from blueberry (BBA), blackberry (BKA), and blackcurrant (BCA) and to determine the relationship between their antioxidant capacity and anti-inflammatory effect in macrophages. Major anthocyanins in BBA, BKA and BCA were malvidin-3-glucoside (16%), cyanidin-3-glucoside (98%) and delphinidin-3-rutinoside (44%), respectively. BKA showed higher total antioxidant capacity than BBA and BCA. RAW 264.7 macrophages were incubated with 0–20 μg/ml of BBA, BKA and BCA, and subsequently activated by lipopolysaccharide (LPS) to measure proinflammatory cytokine production. Interleukin 1β (IL-1β) messenger RNA (mRNA) levels were significantly decreased by all berry anthocyanins at 10 μg/ml or higher. Tumor necrosis factor α (TNFα) mRNA levels and secretion were also significantly decreased in LPS-treated macrophages. The levels of the repression were comparable for all berry anthocyanins. LPS-induced nuclear factor κB (NF-κB) p65 translocation to the nucleus was markedly attenuated by all of the berry anthocyanins. In bone marrow-derived macrophages (BMMs) from nuclear factor E2-related factor 2 wild-type (Nrf2^+/+) mice, BBA, BKA and BCA significantly decreased cellular reactive oxygen species (ROS) levels with a concomitant decrease in IL-1β mRNA levels upon LPS stimulation. However, in the BMM from Nrf2^−/− mice, the anthocyanin fractions were able to significantly decrease IL-1β mRNA despite the fact that ROS levels were not significantly affected. In conclusion, BBA, BKA and BCA exert their anti-inflammatory effects in macrophages, at least in part, by inhibiting nuclear translocation of NF-κB independent of the NRF2-mediated pathways.     

A comparative study on root and bark extracts of Eleutherococcus senticosus and their effects on human macrophages

BackgroundEleutherococcus senticosus or Siberian ginseng is a medicinal plant containing adaptogenic substances believed to regulate immune responses. Both, the root and stem bark are commonly used in traditional medicines.PurposeThe purpose of the present study is to chemically characterize E. senticosus root and bark extracts and to compare their effects on functions of human primary macrophages.Study design and methodsHPLC-DAD-MS analysis was used to characterize chemical constituents of alcoholic extracts from E. senticosus root and bark. The data obtained and available databases were combined for network pharmacology analysis. Involvement of predicted pathways was further functionally confirmed by using monocyte-derived human macrophages and endotoxin-free E. senticosus root and bark extracts.ResultsChemical analysis showed that the root extract contained more syringin, caffeic acid, and isofraxidin than the bark extract. At variance, bark extract contained more sesamin and oleanolic acid. Coniferyl aldehyde and afzelin were below the limit of quantification in both extracts. Network pharmacology analysis indicated that constituents of E. senticosus might affect the immune cell phenotype and signaling pathways involved in cell metabolism and cytoskeleton regulation. Indeed, both extracts promoted actin polymerization, migration, and phagocytosis of E. coli by macrophages pointing to macrophage polarization towards the M2 phenotype. In addition, treatment with E. senticosus root and bark extracts decreased phosphorylation of Akt on Ser473 and significantly reduced expression of the hemoglobin scavenger receptor CD163 by macrophages. Neither extract affected expression of CD11b, CD80, or CD64 by macrophages. In addition, macrophages treated with the bark extract, but not with the root extract, exhibited activated p38 MAPK and NF-κB and released increased, but still moderate, amounts of proinflammatory TNF-α and IL-6, anti-inflammatory IL-10, and chemotactic CCL1, which all together point to a M2b-like macrophage polarization. Differently, the root extract increased the IL-4-induced expression of anti-inflammatory CD200R. These changes in monocytes are in agreement with an increased M2a macrophage polarization.ConclusionThe ability of E. senticosus root and bark extracts to promote polarization of human macrophages towards anti-inflammatory M2a and M2b phenotypes, respectively, might underlay the immunoregulatory activities and point to potential wound healing promoting effects of this medicinal plant.     

Amlexanox exerts anti-inflammatory actions by targeting phosphodiesterase 4B in lipopolysaccharide-activated macrophages

Amlexanox, an anti-inflammatory agent, is widely used for treating aphthous ulcers. Recently, amlexanox has received considerable attention because of its efficacy in mitigating metabolic inflammation via directly suppressing IKKε/TBK1. However, because the knockdown of IKKε/TBK1 has no anti-inflammatory effect on lipopolysaccharide (LPS)-primed RAW264.7 cells, the mechanism of amlexanox against classical inflammation is independent of IKKε/TBK1. In this study, we aim to examine the effects of amlexanox on LPS-treated macrophages and in a mouse model of endotoxemia. We found that amlexanox significantly inhibited the production of pro-inflammatory mediators, both in vitro and in vivo, while increased interleukin-10 level in LPS-activated macrophages. Mechanistically, amlexanox down-regulated nuclear factor κB and extracellular signal-regulated kinase/activator protein-1 signaling by elevating intracellular 3′,5′-cyclic adenosine monophosphate (cAMP) level and subsequently activating protein kinase A. Molecular docking along with fluorescence polarization and enzyme inhibition assays revealed that amlexanox bound directly to phosphodiesterase (PDE) 4B to inhibit its activity. The anti-inflammatory effects of amlexanox could be abolished by the application of cAMP antagonist or PDE4B siRNA. In addition to PDE4B, the activities of PDE1C, 3A, and 3B were directly inhibited by amlexanox. Our results provide mechanistic insight into the clinical utility of amlexanox for the treatment of inflammatory disorders and might contribute to extending the clinical indications of amlexanox.     

Tranilast, an orally active anti-allergic drug, up-regulates the anti-inflammatory heme oxygenase-1 expression but down-regulates the pro-inflammatory cyclooxygenase-2 and inducible nitric oxide synthase expression in RAW264.7 macrophages

Tranilast (N-[3′,4′-dimethoxycinnamonyl] anthranilic acid), an orally active anti-allergic drug, is reported to exert the anti-inflammatory effects, but the underlying mechanisms that could explain the anti-inflammatory actions of tranilast remain largely unknown. Here, we found that tranilast induces heme oxygenase-1 (HO-1) expression through the extracellular signal-regulated kinase-1/2 (ERK1/2) pathway in RAW264.7 macrophages. Tranilast suppressed cyclooxygenase-2 (COX-2) and inducible nitric oxide (NO) synthase (iNOS) expression, and thereby reduced COX-2-derived prostaglandin E₂ (PGE₂) and iNOS-derived NO production in lipopolysaccharide (LPS)-stimulated macrophages. Similarly, tranilast diminished tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) production. Interestingly, the effects of tranilast on LPS-induced PGE₂, NO, TNF-α, and IL-1β production were partially reversed by the HO-1 inhibitor tin protoporphyrin, suggesting that tranilast-induced HO-1 expression is at least partly responsible for the resulting anti-inflammatory effects of the drug. Thus, HO-1 expression via ERK1/2 activation may be at least one of the possible mechanisms explaining the anti-inflammatory actions of tranilast.     

Identification of a novel compound that inhibits iNOS and COX-2 expression in LPS-stimulated macrophages from Schisandra chinensis

A novel α-iso-cubebenol, which has anti-inflammatory effects in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages, was isolated from the fruits of Schisandra chinensis. α-iso-cubebenol inhibited LPS-induced nitric oxide (NO) and prostaglandin E₂ (PGE₂) production. Consistent with these findings, α-iso-cubebenol also reduced the LPS-induced expression of inducible nitric oxide synthase and cyclooxygenase-2 at the protein and mRNA levels in a concentration-dependent manner. α-iso-cubebenol also inhibited LPS-induced nuclear translocation of the NF-κB p65 subunit. Furthermore, α-iso-cubebenol suppressed the phosphorylation of ERK, JNK, and p38 kinase induced by LPS. Since the novel α-iso-cubebenol blocked the production of several pro-inflammatory mediators induced by LPS in macrophages, the molecule can be useful material for the development of anti-inflammatory agents against bacterial infections or endotoxin.     

The anti-inflammatory potential of neuropeptide FF in vitro and in vivo

Neuropeptide FF (NPFF) has many functions in regulating various biological processes. However, little attention has been focused on the anti-inflammatory effect of this peptide. In the present study, the in vitro anti-inflammatory activity of NPFF in both primary peritoneal macrophages and RAW 264.7 macrophages was investigated. Our data showed that NPFF suppressed the nitric oxide (NO) production of macrophages in the inflammation process. RF9, a reported antagonist of NPFF receptors, completely blocked the NPFF-induced NO suppression, suggesting a NPFF receptors-mediated pathway is mainly involved. Down-regulation of the nitric oxide synthases significantly inhibited the NPFF-induced NO reduction, indicating the involvement of nitric oxide synthases. However, the nitric oxide synthases were not the only route by which NPFF modulated the NO levels of macrophages. Pharmacological antagonists of the NF-κB signal pathway also completely suppressed the NPFF-induced NO decline. Moreover, we also observed that NPFF is capable of blocking the LPS-induced nuclear translocation of p65 in macrophages, implying the involvement of the NF-κB signal pathway. Finally, we observed that NPFF markedly attenuated the carrageenan-induced mouse paw edema, indicating that NPFF is capable of exerting anti-inflammatory potency in vivo. Collectively, our findings reveal the potential role of NPFF in the anti-inflammatory field both in vitro and in vivo, which will be helpful for the further exploitation of NPFF utility therapeutically.