LPS-induced NO Production In Macrophages Research Articles

Trichinella spiralis dipeptidyl peptidase 1 suppressed macrophage cytotoxicity by promoting M2 polarization via the STAT6/PPARγ pathway

Trichinella spiralis dipeptidyl peptidase 1 (TsDPP1), or cysteine cathepsin C, is a secretory protein that is highly expressed during the infective larvae and adult worm stages in the intestines. The aim of this study was to investigate the mechanism by which recombinant TsDPP1 (rTsDPP1) activates macrophages M2 polarization and decreases macrophage cytotoxicity to kill newborn larvae via ADCC. RAW264.7 macrophages and murine peritoneal macrophages were used in this study. The results of the immunofluorescence test (IFT) and confocal microscopy showed that rTsDPP1 specifically bound to macrophages, and the binding site was localized on the cell membrane. rTsDPP1 activated macrophage M2 polarization, as demonstrated by high expression levels of Arg1 (M2 marker) and M2-related genes (IL-10, TGF-β, CD206 and Arg1) and high numbers of CD206+ macrophages. Furthermore, the expression levels of p-STAT6, STAT6 and PPARγ were obviously increased in rTsDPP1-treated macrophages, which were evidently abrogated by using a STAT6 inhibitor (AS1517499) and PPARγ antagonist (GW9662). The results indicated that rTsDPP1 promoted macrophage M2 polarization through the STAT6/PPARγ pathway. Griess reaction results revealed that rTsDPP1 suppressed LPS-induced NO production in macrophages. qPCR and flow cytometry results showed that rTsDPP1 downregulated the expression of FcγR I (CD64) in macrophages. The ability of ADCC to kill newborn larvae was significantly decreased in rTsDPP1-treated macrophages, but AS1517499 and GW9662 restored its killing capacity. Our results demonstrated that rTsDPP1 induced macrophage M2 polarization, upregulated the expression of anti-inflammatory cytokines, and inhibited macrophage-mediated ADCC via activation of the STAT6/PPARγ pathway, which is beneficial to the parasitism and immune evasion of this nematode.

Xenoacremones D-H, Bioactive Tyrosine-decahydrofluorene Analogues from the Plant-Derived Fungus Xenoacremonium sinensis.

Five novel tyrosine-decahydrofluorene analogues, xenoacremones D–H (1–5), each bearing a fused 6/5/6 tricarbocyclic core and a 13-membered para-cyclophane ring system, were isolated from the endophytic fungus Xenoacremonium sinensis. Compound 1 was a novel polyketide synthase–nonribosomal peptide synthetase (PKS–NRPS) tyrosine-decahydrofluorene hybrid containing a 6/5/6/6/5 ring system. Their structures were elucidated from comprehensive spectroscopic analysis and electronic circular dichroism (ECD) calculations. All compounds were evaluated for their inhibitory activities on LPS-induced NO production in macrophages and their cytotoxicities against the NB4 and U937 cell lines. Compounds 3 and 5 exhibited potent anti-inflammatory activities in vitro. Compounds 1 and 3–5 displayed significant antiproliferative activity against the tumor cell lines (IC50 < 20 µM).

Phenolic glycosides from the stems of Homalium stenophyllum.

The phytochemical investigation of the stems of Homalium stenophyllum afforded seven new phenolic glycosides (1-5 and 8-9) and two known compounds (6 and 7). Their structures were elucidated by comprehensive analyses of NMR spectroscopic, mass spectrometric data and chemical hydrolysis. Additionally, their anti-inflammatory activities against the NO production in LPS-induced macrophages were evaluated.

Two new furofuran lignan glycosides from Forsythia suspensa leaves

Abstract Phytochemical investigation into the 70 % ethanol extract of Forsythia suspensa leaves afforded the isolation and identification of two new lignan glycosides, Phillyrigeninside B (1) and Phillyrigeninside C (2). The structure of compounds isolated were elucidated via extensive spectroscopy techniques, including ultraviolet (UV), infrared (IR) and Nuclear Magnetic Resonance (NMR). The inhibitory effect against LPS-induced NO production in macrophages and the antibacterial activity were tested for compounds 1–2. The study showed that the two isolated compounds were furofuran lignan glycosides, and exhibited moderate antibacterial and moderate anti-inflammatory activities. The work further confirmed that the leaves of F. suspensa are also a source of furofuran lignan glycosides with important development value and wide application prospect.

Dihydroagarofuran sesquiterpenoids esterified with organic acids from the leaves of Tripterygium wilfordii

Six new and one known dihydroagarofuran sesquiterpenoids esterified with organic acids were obtained from the leaves of Tripterygium wilfordii. Spectroscopic techniques (UV, IR, MS, NMR, ORD and CD) were used for the structure elucidation of the compounds. The structures of compounds 1 and 2 were confirmed by X-ray single crystallographic analyses. The inhibitory effects on NO production in LPS-induced macrophages of 1–7 were conducted. At 10 μmol/L, compounds 1, 2 and 7 showed moderate inhibitory effects on NO production in LPS-induced macrophages with inhibitory rate at 31.2 ± 3.6, 40.9 ± 4.3, and 66.79 ± 3.1%, respectively.

In vitro approach for evaluation of carob by-products as source bioactive ingredients with potential to attenuate metabolic syndrome (MetS)

The potential bioactivities for alleviating Metabolic Syndrome associated risk factors were evaluated in carob (Ceratonia siliqua L.) fruit by-products, i.e. seed peel, germ and pod. Carob germ and seed peel showed higher phenolic content than pod (99.72, 80.24 and 47.06 μmol GAE g−1, respectively). Pod mostly contained gallic acid and gallotannins; seed peel and germ's showed as most abundant polyphenols quercetin and apigenin derivatives. Carob pod and seed peel revealed stronger antioxidant capacities compared to germ. The strongest antihypertensive activity was found in seed peel, followed by pod and germ. Anti-inflammatory activity showed inhibition of NO production in LPS-induced macrophages, although only pod was able of reducing pro-inflammatory mediators (TNF-α andPGD2). Finally, fat accumulation on mature adipocytes was reduced by carob seed peel and pod extracts. This work shows the potential use of pod carob by-products as food ingredients with special relevance of carob pod for attenuating metabolic syndrome.

Two New Triterpenoid Glycosides from the Roots ofRosa cymosaTratt.

isolation, structure elucidation and inhibitory activity against LPS-induced NO production in macrophages

Sesquiterpenoids from the seeds of Sarcandra glabra and the potential anti-inflammatory effects

Five new sesquiterpenoids, including two linderanes (1–2) and three eudesmanes (3–5) were isolated from the seeds of Sarcandra glabra. Their structures and relative configurations were established by spectroscopic data analysis. 1 was a rare linderane derivative having an 18-membered triester ring which is a common characteristic in linderane dimers. Compounds 1–5 were investigated for their inhibitory effects on NO production in LPS-induced macrophages and 1 showed moderate bioactivity.

Antiinflammatory Activities of Crebanine by Inhibition of NF-κB and AP-1 Activation through Suppressing MAPKs and Akt Signaling in LPS-Induced RAW264.7 Macrophages.

Crebanine, an aporphine alkaloid, displays various biological activities such as anticancer and antimicrobial activities. In this study, we further investigated the suppressive effect of crebanine on lipopolysaccharide (LPS)-induced expression of proinflammatory mediators and the molecular mechanisms underlying these activities in RAW264.7 macrophages. Crebanine inhibited the production of proinflammatory cytokines including interleukin-6 (IL-6) and tumor necrosis factor-alpha in LPS-induced RAW264.7 cells. Moreover, crebanine suppressed LPS-induced inducible nitric oxide (iNO) and prostaglandin E2 and reduced the expression of iNO synthase and cyclooxygenase-2 in RAW264.7 cells. Crebanine suppressed LPS-induced phosphorylation of Akt and mitogen-activated protein kinases (MAPKs), including extracellular signaling-regulated kinase 1/2, c-Jun NH2-terminal kinase, and p38 MAPK signaling. In addition, the specific inhibitor of MAPKs and Akt reduced the expression of IL-6 and NO production in LPS-induced macrophages. Furthermore, crebanine inhibited LPS-induced nuclear factor kappa B (NF-κB) activation by reducing the phosphorylation of p65 at Ser536 but not the p65 translocation to the nucleus and inhibitory factor kappa B alpha degradation. Crebanine also suppressed phosphorylation and nucleus translocation of activator protein-1 (AP-1). These observations suggest that the antiinflammatory properties of crebanine may stem from the inhibition of proinflammatory mediators via suppression of the NF-κB, AP-1, MAPKs, and Akt signaling pathways.

Sesquiterpene dimers esterified with diverse small organic acids from the seeds of Sarcandra glabra

11 new sesquiterpene dimers, sarglabolides A–K (1–11), and five known ones were isolated from the seeds of Sarcandra glabra. Their structures were elucidated by spectroscopic data analysis and chemical evidence. Sarglabolide A (1) was verified to exclusively possess a seventeen-membered macrocyclic ester ring formed by the scaffold of the sesquiterpene dimer and small organic acids, different from the eighteen-membered rings of the other reported analogues. The chiral small organic acid moieties were assigned to l-malic acid, d-malic acid, and d-tartaric acid based on the combination of spectroscopy, chemical derivatization and HPLC analysis. Dimers 1, 12 and 13 can significantly inhibit NO production in LPS-induced macrophages with IC50 values at 3.04, 4.65 and 2.33 μmol/L, respectively.

Oleuropein suppresses LPS-induced inflammatory responses in RAW 264.7 cell and zebrafish.

Oleuropein is one of the primary phenolic compounds present in olive leaf. In this study, the anti-inflammatory effect of oleuropein was investigated using lipopolysaccharide (LPS)-stimulated RAW 264.7 and a zebrafish model. The inhibitory effect of oleuropein on LPS-induced NO production in macrophages was supported by the suppression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). In addition, our enzyme immunoassay showed that oleuropein suppressed the release of pro-inflammatory cytokines such as interleukin-1β (IL-1β) and interleukin-6 (IL-6). Oleuropein inhibited the translocation of p65 by suppressing phosphorylation of inhibitory kappa B-α (IκB-α). Oleuropein also decreased activation of ERK1/2 and JNK, which are associated with LPS-induced inflammation, and its downstream gene of AP-1. Furthermore, oleuropein inhibited LPS-stimulated NO generation in a zebrafish model. Taken together, our results demonstrated that oleuropein could reduce inflammatory responses by inhibiting TLR and MAPK signaling, and may be used as an anti-inflammatory agent.

Se improves indole glucosinolate hydrolysis products content, Se-methylselenocysteine content, antioxidant capacity and potential anti-inflammatory properties of sauerkraut

Selenium (Se) has a well-known role in prevention of chronic diseases associated with oxidative stress and inflammation. The objective was to produce Se-enriched sauerkraut and to study the effect of selenite addition on indole glucosinolate hydrolysis products, vitamin C, Se biotransformation and microbial quality. The antioxidant activity of sauerkraut and its ability to inhibit NO production in LPS-induced macrophages were also examined. White cabbage was naturally fermented with 0.3mg of Na2SeO3/kg fresh cabbage (NFSe) or without Se addition (NF). Total Se content reached up to 1.29μg/gd.m. (0.11μg/gf.m.). Selenomethylselenocysteine was the major Se form found in NFSe cabbage. Se addition caused a slight reduction of ascorbigen (6%) and vitamin C (5%) contents in sauerkraut (P⩽0.05); however, lactic acid bacteria increased (3%), and the formations of indole-3-carbinol and indole-3-acetonitrile were markedly enhanced (74% and 13%, respectively). NFSe cabbage extracts showed higher (P⩽0.05) antioxidant activity (163μmoltrolox/gd.m.) and inhibition of NO production in LPS-induced macrophages (IC50=44.0μg/ml) than did NF cabbage extracts. Consequently, Se-enriched sauerkraut can be considered as a health-promoting food.

The Stem Bark ofKalopanax pictusExhibits Anti-inflammatory Effect through Heme Oxygenase-1 Induction and NF-κB Suppression

BackgroundThe stem bark of Kalopanax pictus (KP) has been used in traditional medicine to treat rheumatoidal arthritis, neurotic pain and diabetes mellitus in China and Korea. In this study, the mechanism responsible for anti-inflammatory effects of KP was investigated.MethodsWe examined the effects of KP on NO production, nitric oxide synthase (iNOS) and HO-1 expression, NF-κB, Nrf2 and MAPK activation in mouse peritoneal macrophages.ResultsThe aqueous extract of KP inhibited LPS-induced NO secretion as well as inducible iNOS expression, without affecting cell viability. KP suppressed LPS-induced NF-κB activation, phosphorylation and degradation of IκB-α, phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK). Furthermore, KP induced HO-1 expression and Nrf2 nuclear translocation.ConclusionThese results suggest that KP has the inhibitory effects on LPS-induced NO production in macrophages through NF-κB suppression and HO-1 induction.

Two New Spirobiflavonoids fromAbies chensiensiswith Moderate NO Production Inhibitory Activity

Phytochemical investigation of the aerial parts of Abies chensiensis afforded two new (compounds 2 and 3) and 27 known compounds, including the related compound larixinol ( 1). The structures of spirobiflavonoids 1- 3 were established using 1D and 2D NMR spectroscopic techniques. In addition, the structure of larixinol ( 1) was confirmed by X-ray crystallographic analysis. Compounds 1- 3 were evaluated for inhibitory activities against LPS-induced NO production in macrophages. Larixinol ( 1) showed moderate effects, with an IC(50) value of 60.0 microg/mL. In addition, it did not show any cytotoxicity on RAW 264.7 macrophages at 100 microg/mL.

Inhibition of LPS-induced NO production by taurine chloramine in macrophages is mediated though Ras-ERK-NF-κB

Taurine is an abundant free amino acid in inflammatory cells that protects cells from inflammatory damages. Although the protection mechanism remains unclear, taurine chloramine (Tau-Cl) produced by the reaction between taurine and hypochlorous acid in neutrophils plays an important role. In this study, we investigated the mechanism(s) by which Tau-Cl inhibits LPS-induced NO production in macrophages. Tau-Cl inhibited LPS-induced iNOS expression and NO production in RAW 264.7 cells. LPS treatment elevated the level of active Ras-GTP, and Tau-Cl inhibited LPS-induced Ras activation. Tau-Cl also inhibited ERK1/2 activation in a dose-dependent manner in both RAW 264.7 cells and murine peritoneal macrophages, whereas it did not exert any effect on p38 MAPK activation. Furthermore, Tau-Cl inhibited NF-κB activation without affecting AP-1 activity. These results suggest that Tau-Cl suppresses LPS-induced NO production by inhibiting specific signaling pathways. Thus, Tau-Cl protects cells from inflammatory injury resulting from overproduction of NO in a signaling pathway-specific manner.

Elevation of intracellular calcium levels contributes to the inhibition of nitric oxide production by atrial natriuretic peptide.

Atrial natriuretic peptide (ANP) attenuates LPS-induced inducible nitric oxide synthase (iNOS) expression in murine macrophages by destabilizing iNOS mRNA. Because elevated intracellular free Ca2+ levels [Ca2+]i reduce iNOS mRNA stability, the aim of the present study was to determine whether inhibition of iNOS by ANP is due to alterations in intracellular calcium. As determined by fluorescence photometry, ANP (10(-7) and 10(-6) mol/L) was shown to elevate intracellular calcium levels in bone marrow-derived macrophages. This effect seemed to be mediated via the guanylate cyclase-coupled A receptor, because dibutyryl-cGMP mimicked and the A-receptor antagonist HS-142-1 partially abrogated the effect of ANP. Because the Ca2+ increase was also observed in Ca2+-free buffer, it is suggested that the liberation of intracellular calcium pools contributes to the elevation of [Ca2+]i by ANP. The B-receptor ligand C-type natriuretic peptide (CNP), which does not alter iNOS expression, had no effect on [Ca2+]i. The Ca2+-ionophore 4-Br-A23187 and thapsigargin, a compound known to liberate Ca2+ from intracellular stores, were further demonstrated to reduce LPS-induced NO production in macrophages (Griess assay), confirming a functional link for elevated [Ca2+]i and iNOS inhibition. These effects were abrogated by coincubation with extra- as well as intracellular Ca2+ chelators (EGTA, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA)). The inhibitory effect of ANP on NO production was also abrogated by Ca2+ chelation. These findings support a causal relationship between reduced iNOS induction and elevation of [Ca2+]i. Taken together, the data indicate that intracellular Ca2+ elevation by ANP is involved in the inhibition of LPS-induced nitric oxide production in macrophages.