Inhibition Of NO Production Research Articles

New NO production inhibitors from Hyssopus cuspidatus in LPS-induced RAW264.7 cells

Two new sesquiterpenes, including 7-epi-3-eudesmene-1β,11-diol (1) and (7S,10R)-11-hydroxy-4,5-seco-guai-1-ene-4-one (2); one new natural product (11-hydroxy-1-oxo-4β,5β,7β,10β-eremophilane); (3) and nine known compounds were isolated from the aerial parts of Hyssopus cuspidatus Boriss. Their structures as well as their absolute configurations were elucidated by extensive spectroscopic analyses and quantum chemical calculations. Compounds 1 to 9 were tested to evaluate their inhibitory effects on NO production in LPS-induced RAW264.7 cells. Compounds 5 and 8 exhibited inhibitory effects, with IC50 values of 40.43 and 32.56 μM, respectively, compared to the positive control (i.e., dexamethasone, IC50 = 70.35 μM), and compound 4 exhibited a weaker activity, with an IC50 value of 112.56 μM.

Structural characterization and antineuroinflammatory activity of a novel heteropolysaccharide obtained from the fruits of Alpinia oxyphylla

The fruit of Alpinia oxyphylla is often used in traditional Chinese medicine to treat dementia and memory defects. In this study, we isolated a novel acidic polysaccharide (AOP70-2-1) from A. oxyphylla fruit. Structural analysis showed that AOP70-2-1 consists of β-D-GlcAp-(1→, →2,3,6)-α-D-Galp-(1→, α-L-Araf-(1→, →2,5)-α-L-Araf-(1→, →4)-α-D-Glcp-(1→, →3,4)-α-D-Xylp-(1→, →3,6)-β-D-Manp-(1→, and α-L-Rhap-(1→. Morphological analysis indicated that AOP70-2-1 had an irregular sheet structure. The crude polysaccharide (AOP70) from A. oxyphylla significantly improved learning and memory ability of Alzheimer’s disease (AD) mice, and AOP70 exhibited comparable or even better effects than huperzine A. Most important, AOP70 reduced NO, IL-1β, TNF-α, and PGE-2 concentrations to normal levels. AOP70-2-1 significantly inhibited NO production in lipopolysaccharide stimulated BV2 microglial cells. Note that the effect of 2.6 μM AOP70-2-1 was superior to indomethacin. AOP70-2-1 also remarkably decreased the values of IL-6 and TNF-α. AOP70-2-1 may be a bioactive component of AOP70 and has the potential for the treatment of AD.

Studies on active aromatic constituents from rhizomes of Sophora tonkinensis

Aromatic constituents from rhizomes of Sophora tonkinensis were purified by extensive chromatographic techniques including column chromatography over macroporous resin,MCI,silica gel,weak acid cation exchange resin,Sephadex LH-20,ODS,and semi-preparative HPLC. Twelve aromatic compounds were isolated and identified from the water aqueous extract of the rhizomes of S.tonkinensis. Their structures were elucidated as 4-( 3-hydroxypropyl) phenol( 1),( ±)-4-( 2-hydroxypropyl) phenol( 2),benzamide( 3),( ±)-3-( p-methoxyphenyl)-1,2-propanediol( 4),4-methoxybenzamide( 5),3-hydroxy-1-( 4-hydroxy-3-methoxyphenyl) propan-1-one( 6),tyrosol( 7),( ±)-2,3-dihydroxypropyl benzoate( 8),vanillin alcohol( 9),7,3'-dihydroxy-8,4'-dimethoxyisoflavone( 10),7,4'-dihydroxy-3'-methoxyisoflavone( 11),and 7,3'-dihydroxy-5'-methoxyisoflavone( 12). Compounds 1-9 were firstly isolated from the Sophora genus. Compounds 4,5,10 and 11 can remarkably protect Hep G2 cell against APAP-induced damage at the concentration of 10 μmol·L-1. Compounds 1-12 exhibited no significant activities on the assays of inhibition of LPS-induced NO production in RAW cell lines and NF-κB inhibition.

Anti-inflammatory evaluation and structure-activity relationships of diterpenoids isolated from Euphorbia hylonoma

A phytochemical investigation to obtain chemical components with potential anti-inflammatory activity from E. hylonoma led to the isolation of nine new ent-isopimarane diterpenoids (1 and 3–10), a new ent-rosane diterpenoid (11), along with eight known ones (2 and 12–18) using various chromatographic techniques. Compounds 3, 4, 5, and 10 were rare examples of the epoxy-ent-isopimarane. The structures of these new compounds were confirmed by extensive spectroscopic data, crystal X-ray diffraction analysis, and electronic circular dichroism. And the isolates were evaluated for their inhibitory effects on nitric oxide production induced by lipopolysaccharide in RAW 264.7 cells. The results showed that compounds 2 and 12 exhibited noteworthy inhibitory effects against NO production with IC50 values of 7.12 and 12.73 μM, respectively, which were better than positive control (IC50 = 41.41 μM). The possible mechanism that compounds 2 and 12 could inhibit NO production was investigated by the Western blotting experiments.

Chemical Constituents of Euscaphis konishii and Their Inhibitory Activities

A new megastigmane glycoside, named koniside (1), with five known compounds (2–6) was isolated from the pericarp of Euscaphis konishii. The structure of the new compound was determined by the HR-ESI-MS, IR, 1D, and 2D NMR spectra. Compounds 1 and 5 inhibited NO production of RAW264.7 and show moderate activities.

Anti-inflammatory flavonoids from root bark of Broussonetia papyrifera in LPS-stimulated RAW264.7 cells

Broussonetia papyrifera has been used as a diuretic, tonic and suppressor of edema. Bioactivity-guided fractionation and metabolite investigation of root bark extracts of this plant resulted in the isolation and identification of six 1,3-diphenylpropanes (1, 2, 8, 10, 17, 20), flavanone (3), two chalcones (4, 5), five flavans (6, 11, 14–16), dihydroflavonol (7) and five flavonols (9, 12, 13, 18, 19), including five new compounds (5, 7, 8, 19, 20) that inhibit NO production in LPS-induced RAW264.7 cells. The structures of compounds 1–20 were elucidated on the basis of spectroscopic data (1D and 2D NMR, MS, MS/MS, and HRMS). In particular, compounds 3, 5, 7, 12, and 20 exhibited significant inhibitory effects on the NO, iNOS, and pro-inflammatory cytokine (TNF-α and IL-6) production. Therefore, this study suggests that the flavonoid-rich products of B. papyrifera, including the new compounds, could be valuable candidates for the development of pharmaceuticals or functional foods in the prevention and treatment of anti-inflammatory disease.

Antioxidant, Anti-Inflammatory, and Multidrug Resistance Modulation Activity of Silychristin Derivatives.

Silychristin A is the second most abundant compound of silymarin. Silymarin complex was previously described as an antioxidant with multidrug resistance modulation activity. Here, the results of a classical biochemical antioxidant assay (ORAC) were compared with a cellular assay evaluating the antioxidant capacity of pure silychristin A and its derivatives (anhydrosilychristin, isosilychristin and 2,3-dehydrosilychristin A). All the tested compounds acted as antioxidants within the cells, but 2,3-dehydro- and anhydro derivatives were almost twice as potent as the other tested compounds. Similar results were obtained in LPS-stimulated macrophages, where 2,3-dehydro- and anhydrosilychristin inhibited NO production nearly twice as efficiently as silychristin A. The inhibition of P-glycoprotein (P-gp) was determined in vitro, and the respective sensitization of doxorubicin-resistant ovarian carcinoma overproducing P-gp was detected. Despite the fact that the inhibition of P-gp was demonstrated in a concentration-dependent manner for each tested compound, the sensitization of the resistant cell line was observed predominantly for silychristin A and 2,3-dehydrosilychristin A. However, anhydrosilychristin and isosilychristin affected the expression of both the P-gp (ABCB1) and ABCG2 genes. This is the first report showing that silychristin A and its 2,3-dehydro-derivative modulate multidrug resistance by the direct inhibition of P-gp, in contrast to anhydrosilychristin and isosilychristin modulating multidrug resistance by downregulating the expression of the dominant transmembrane efflux pumps.

Antioxidant and Anti-Inflammatory Activities of the Crude Extracts of Moringa oleifera from Kenya and Their Correlations with Flavonoids.

Moringa oleifera Lam. (M. oleifera) is commonly distributed and utilized in tropical and sub-tropical areas. There has been a large number of reports on the antioxidant and anti-inflammatory activity of its leaves, but only a few about its seeds and roots. Hence, in this work we aimed to systematically compare the antioxidant and anti-inflammatory activities of the ethanol crude extracts of leaves, seeds, and roots of M. oleifera from Kenya, and further correlate the differential activities with the chemical constituents from these three parts. The antioxidant activities were measured by using three different assays (DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonic acid) and FRAP (Ferric-Reducing Antioxidant Power), respectively). Results showed that the leaf extracts displayed the highest DPPH radical scavenging and FRAP total reducing power activities with IC50 values of 1.02 ± 0.13 mg/mL and 0.99 ± 0.06 mM Fe2+/g, respectively; the leaf and root extracts exhibited potential ABTS radical scavenging activities with the IC50 values of 1.36 ± 0.02 and 1.24 ± 0.03 mg/mL. Meanwhile, the leaf and seed extracts (11.1–100 µg/mL) also exerted obvious anti-inflammatory activities, as indicated by the inhibition of NO production. To further reveal correlations between these differential activities with the chemical constituents in the three organs, the total flavonoids content (TFC) of the three different extracts were evaluated, and the TFC of leaves, seeds and roots were found to be 192.36 ± 2.96, 5.89 ± 0.65 and 106.79 ± 2.12 mg rutin equivalent (RE)/g, respectively. These findings indicated the important impacts of the total flavonoid contents on antioxidant and anti-inflammatory activities. Additionally, we further determined the phytochemical profiles of M. oleifera by HPLC-UV/ESI-MS/MS, and identified most of the chemical constituents of leaves as flavonoids. In summary, the leaves of M. oleifera are a better potential natural source of antioxidants and anti-inflammatory agents, and very promising for development into the health promoting dietary supplements.

Three Novel Monoterpenoid Glycosides From Fruits Of Eleutherococcus Henryi

: The phytochemical investigation on the fruits of Eleutherococcus henryi (Araliaceae) resulted in the discovery of three novel monoterpene glycosides, eleuhenryiside A (1), eleuhenryiside B (2), and eleuhenryiside C (3), as well as a known lignan, (-)-kobusin (4). Their chemical structures were elucidated by mass, 1 D- and 2 D-NMR spectroscopy. The chemical structures of new compounds 1-3 were determined to be (2E,6R)-6-hydroxy-2,6-dimethyl-2,7-octadien-1-yl-(6’-O-acetyl)-O-β-glucopyranoside, (2Z,6R)-6-hydroxy-2,6-dimethyl-2,7-octadien-1-yl-(6’-O-acetyl)-O-β-glucopyranoside, and (-)-(4 R)-4,7-dihydroxy-1-menthene 7-O-β-glucopyranoside, respectively. The anti-neuroinflammatory and anti-inflammatory activities of these compounds were evaluated with LPS-stimulated BV2 microglia and RAW264.7 macrophage, respectively. The results showed that new compounds 1 and 3 have inhibitory effects of NO production with IC50 values of 32.50 ± 1.60 and 3.54 ± 0.20 μM in LPS-stimulated BV2 microglia. Also, (-)-kobusin (4) has abilities to inhibit NO production with the IC50 values of 14.25 ± 2.69 and 36.35 ± 6.27 μM in BV2 and RAW264.7 cells, respectively, which indicated that it may possess the potential anti-neuroinflammatory and anti-inflammatory activities.

Alginate aerogels carrying calcium, zinc and silver cations for wound care: Fabrication and metal detection

Calcium-alginate aerogels augmented with zinc and silver can provide the next generation superabsorbent medical devices for advanced wound care with high cell and tissue tolerability and controlled compound release rates during wound healing. In this study, we addressed the fabrication and metal detection of alginate-based aerogels with these immunoactive cations. All three cations were incorporated into the alginate aerogel, albeit in different quantities, due to different magnitudes of leaching during solvent exchange step; an effect not reported previously. Silver addition led to an increase in aerogel zinc content without influencing calcium incorporation. All three metal ions were released into supernatants upon swelling aerogels in aqueous solutions, comparable to those measured for common wound dressings in human and animal injury models. The in vitro bioactivity studies showed that Zn-enriched swelling supernatants were able to suppress NO production in stimulated RAW 264.7 macrophages, indicating an effective anti-inflammatory activity.

Selenium-Rich Yeast Mitigates Aluminum-Mediated Testicular Toxicity by Blocking Oxidative Stress, Inhibiting NO Production, and Disturbing Ionic Homeostasis.

Aluminum (Al) poisoning has been linked to the development of several reproductive system dysfunctions. Dietary supplementation with selenium-rich yeast (SeY) has been shown to prevent a variety of pathologic conditions. In the present study, the potential protect role of SeY on Al-induced testicular toxicity was evaluated, and the possible underlying mechanisms were discussed. Mice were treated with SeY (0.1mg/kg) and/or Al (10mg/kg) by oral gavage for 4weeks. Histopathologic changes were observed in the testes of Al-treated mice. Oxidative stress, ionic disturbances, and the generation of NO systems are believed to have resulted in the observed pathology. Interestingly, SeY supplementation significantly inhibited the Al-induced histopathological and molecular changes and restored these indicators to levels observed in the control animals. These results suggest that SeY exerts a testis-protective effect against Al-induced toxicity through the reduction of oxidative stress, NO production, and the maintenance of ionic homeostasis.

Anti-inflammatory effect of hydrolyzed celery leaves extract in murine primary splenocyte.

Apium graveolens (celery) has been used in traditional medicine for the treatment of various ailments. However, its molecular mechanism of action in inflammatory response is unclear. In this study, we sought to investigate the anti-inflammatory properties of celery leaves. We prepared an acid-hydrolyzed extract of celery leaves (HCE) and studied its effects on concanavalin A (ConA)-stimulated primary splenocytes. HCE at noncytotoxic concentrations, inhibited ConA-induced proliferation of splenocytes. HCE treatment reduced CD4+ T cell population and decreased expressions and production of cytokines in stimulated splenocytes. In addition, HCE significantly inhibited NO production and reduced the expression of COX-2 mRNA in the stimulated splenocytes. The effects seen were probably due to HCE's downregulation of NF-κB/p65 activation in splenocytes. By providing the anti-inflammatory mechanism of action of HCE, these findings are potentially important for future studies that may, ultimately result in the potential use of celery for the treatment/prevention of inflammatory diseases. PRACTICAL APPLICATIONS: Apium graveolens is a well-known edible plant with high concentrations of bioactive compounds such as apigenin, luteolin, and kaempferol. The research investigated the effects of A. graveolens leaves in splenocyte proliferation, and production of inflammatory mediators and cytokines. The data obtained from this study shed light on the use of plant extracts and plant-based bioactives in nutraceutical industries as potential functional food materials for preventing inflammatory diseases.

Flavonoids from Chionanthus retusus (Oleaceae) Flowers and Their Protective Effects against Glutamate-Induced Cell Toxicity in HT22 Cells.

The dried flowers of Chionanthus retusus were extracted with 80% MeOH, and the concentrate was divided into EtOAc, n-BuOH, and H2O fractions. Repeated SiO2, octadecyl SiO2 (ODS), and Sephadex LH-20 column chromatography of the EtOAc fraction led to the isolation of four flavonols (1–4), three flavones (5–7), four flavanonols (8–11), and one flavanone (12), which were identified based on extensive analysis of various spectroscopic data. Flavonoids 4–6 and 8–11 were isolated from the flowers of C. retusus for the first time in this study. Flavonoids 1, 2, 5, 6, 8, and 10–12 significantly inhibited NO production in RAW 264.7 cells stimulated by lipopolysaccharide (LPS) and glutamate-induced cell toxicity and effectively increased HO-1 protein expression in mouse hippocampal HT22 cells. Flavonoids with significant neuroprotective activity were also found to recover oxidative-stress-induced cell damage by increasing HO-1 protein expression. This article demonstrates that flavonoids from C. retusus flowers have significant potential as therapeutic materials in inflammation and neurodisease.

Investigation of Some Medicinal Plants Inhibitory Effect on NO Production in Oxidative Stressed PC12 Cells

Background: Nitric oxide and reactive nitrogen species play an important role in various pathological conditions like cancer, inflammation and neurodegeneration. As plants and natural compounds have a great potency of discovering lead compounds which might affect NO production during inflammation and various pathologies, we examined the effects of three medicinal plants native to Iran, on NO production during oxidative stress in PC12 cells. Methods: In this study, cell death and NO levels were measured by MTT and by Griess assay, respectively. Oxidative stress was induced by hydrogen peroxide and extracts of Astragalus jolderensis, Convolvulus commutatus and Salvia multicaulis were used as pretreatment in oxidative stressed PC12 cells. Results: A. jolderensis extract significantly suppressed NO production in 150 and 200 μg/ml concentrations and C. commutatus extract in all concentration inhibited NO production in stressed PC12 cells. In addition, the extract of S. multicaulis inhibited NO production during stress at all concentrations above 50 μg/ml. Besides, the extract of S. multicaulis showed protective effect at lower doses in stressed cells. Conclusion: According to the results, S. multicaulis inhibited NO production and protected cells from oxidative stress. Hence, S. multicaulis is a good candidate for further in vitro and in vivo investigations. A. jolderensis and C. commutatus also suppressed NO production during stress. Therefore, they could be noticed in experiments that centralize on the inhibition of NO production and drug discovery studies in the field of neurodegenerative and chronic inflammatory diseases.

Ergosterols from Ganoderma sinense and their anti-inflammatory activities by inhibiting NO production

Three new ergosterols, ganocalidophins A–C (1‒3), along with four known ones, were isolated from the fruiting bodies of Ganoderma sinense. Their structures were established on the basis of extensive NMR and MS technologies, as well as the single crystal X-ray diffraction. Compounds 1‒3 and 6 exhibited inhibitory activities against NO production with IC50 of 17.7–32.4 μM.

Effect of Anadenanthera colubrina protease inhibitors as an antiinflamatory mediator

This work aimed to obtain and characterize protease inhibitors from A. colubrina leaves and evaluate their potential as inflammatory mediator and cell viability. The protein extract was analyzed and characterized by SDS-PAGE, RP-HPLC-PDA, MALDI-TOF/MS and Zeta potential. Bioassays were conducted in order to evaluate the cell viability in RAW 264.7, in vitro (NO and TNF-α production inhibition) and in vivo anti-inflammatory potential, inhibition rate of trypsin and hemagglutination activity from protein extract. The results revealed the presence of bands at 14, 21 and 30 kDa in SDS-PAGE, the RP-HPLC-PDA analysis showed peaks at 12, 13, 28 and 40 minutes and MALDI-TOF/MS showed peaks with 3.4, 4.7, 5.6, 9.4 and 11.2 kDa. The protein extracts presented enzymatic activity inhibition of trypsin (IC50 59.2 μgmL−1), did not show any cytotoxicity to RAW264.7 cells, hemagglutination 8HU and insignificant reduction in NO and TNF-α production and reduced anti-inflammatory potential in vivo compared to dexamethasone.

DEVELOPMENT OF SEMISOLID PREPARATIONS CONTAINING EXTRACT OF THAI POLYHERBAL RECIPE FOR ANTI-INFLAMMATORY EFFECT

Objective: The objective was to develop the semisolid preparations containing extract of Thai polyherbal recipe with anti-inflammatory effect.
 Methods: Polyherbal ethanolic extract was prepared by maceration and determined for phytochemicals and antioxidant activity. Effects of extract on the production of pro-inflammatory mediator-nitric oxide (NO)-were examined in RAW 264.7 cells. Semisolid preparations, balm, and gel, were prepared and evaluated. In vitro release profiles and mechanisms of phenolic compounds, phytochemical markers, from the preparations were investigated.
 Results: Polyherbal ethanolic extract was dark yellow-green, viscous liquid with the yields of 8.2%. Total phenolic and total flavonoid contents were 121.21±1.60 mg GAE/g and 26.55±1.38 mg QE/g, respectively. Antioxidant assay showed that polyherbal extract can scavenge the radical to a certain extent, with DPPH IC50 of 160.75±3.43 µg/ml and FRAP values of 91.94±4.17 mg FeSO4/g. In vitro anti-inflammatory test revealed that the extract inhibited NO production in a dose-dependent manner, with IC50 of 145.65±3.26 µg/ml. The yellowish-green color, homogenous and suitable for skin application polyherbal balm and gel was obtained. The higher release of phenolics from the gel was observed, with the cumulative release at 8 h of 119.0±4.3 mg GAE, whereas that from the balm was only 39.7±2.0 mg GAE. The phenolic release profile was found to be best fitted with the Higuchi model.
 Conclusion: The semisolid preparations containing Thai polyherbal recipe extract with anti-inflammatory effect were successfully prepared. The proper semisolid base and compositions are crucial for effective skin delivery as they influence the release rates of phytochemical markers.

CGRP signalling inhibits NO production through pannexin-1 channel activation in endothelial cells

Blood flow distribution relies on precise coordinated control of vasomotor tone of resistance arteries by complex signalling interactions between perivascular nerves and endothelial cells. Sympathetic nerves are vasoconstrictors, whereas endothelium-dependent NO production provides a vasodilator component. In addition, resistance vessels are also innervated by sensory nerves, which are activated during inflammation and cause vasodilation by the release of calcitonin gene-related peptide (CGRP). Inflammation leads to superoxide anion (O2• −) formation and endothelial dysfunction, but the involvement of CGRP in this process has not been evaluated. Here we show a novel mechanistic relation between perivascular sensory nerve-derived CGRP and the development of endothelial dysfunction. CGRP receptor stimulation leads to pannexin-1-formed channel opening and the subsequent O2• −-dependent connexin-based hemichannel activation in endothelial cells. The prolonged opening of these channels results in a progressive inhibition of NO production. These findings provide new therapeutic targets for the treatment of the inflammation-initiated endothelial dysfunction.

Design, Synthesis and Biological Activities of New Pyrazole Derivatives Possessing Both Coxib and Combretastatins Pharmacophores.

In our efforts to discover novel multi-target agents having better antitumor activities than celecoxib, 21 new aryl-substituted pyrazole derivatives possessing cis-diphenylethylene scaffold were mostly synthesized by a one-pot approach to ethyl 1,4,5-triaryl-1H-pyrazole-3-carboxylates via an improved Claisen condensation - Knorr reaction sequence. The cytotoxic effects of these compounds against three human cancer cell lines HT-29, Hep-G2, MCF-7 as well as their inhibition of NO production were studied. Results showed that incorporation of the important pharmacophoric groups of two original molecules celecoxib and combretastatin A-4 in a single molecule plays an important role in determining a better biological activities of the new coxib-hybrided compounds.

Inhibitory Effects on NO Production and DPPH Radicals and NBT Superoxide Activities of Diarylheptanoid Isolated from Enzymatically Hydrolyzed Ehthanolic Extract of Alnus sibirica.

Alnus sibirica (AS) is geographically distributed in Korea, Japan, Northeast China, and Russia. Various anti-oxidant, anti-inflammation, anti-atopic dermatitis and anti-cancer biological effects of AS have been reported. Enzymatic hydrolysis decomposes the sugar bond attached to glycoside into aglycone which, generally, has a superior biological activity, compared to glycoside. Enzymatic hydrolysis of the extract (EAS) from AS was processed and the isolated compounds were investigated—hirsutanonol (1), hirsutenone (2), rubranol (3), and muricarpon B (4). The structures of these compounds were elucidated, and the biological activities were assessed. The ability of EAS and the compounds (1–4) to scavenge 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals and Nitroblue tetrazolium (NBT) superoxide, and to inhibit NO production was evaluated in vitro. EAS showed more potent antioxidant and anti-inflammatory activity than AS. All investigated compounds showed excellent antioxidant and anti-inflammatory activities.