Nitrite Oxide Production Research Articles

Enhanced Functionality of Canavalia gladiata Hull by Fermentation with Lactic Acid Bacteria

Abstract Objectives Immature Canavalia Gladiata Hull (ICGH) has been used traditionally as food but the research data on its functionality is insufficient. Antioxidant and anti-inflammation activity of ICGH and fermentation was performed using lactic acid bacteria (LAB) with glycosidase activity to confirm whether the bioconversion affect the functionality. Methods LAB was screened from Kimchi for the glycosidase activity. Antioxidant activity was evaluated by ABTS, DPPH, cellular antioxidant activity (CAA), and nitrite oxide (NO) production was evaluated with RAW264.7 cell treated with lipopolysaccharide (LPS). Active component in ICGH was analyzed using HPLC-PDA, MS/MS, and NMR. Results LABs could grow in medium with ICGH as a sole nutrient source. After fermentation the contents of lactic acid was increased as sugar contents declined and asparagine and γ-aminobutyrate(GABA) were detected as main metabolite, and some flavonoids were identified. After fermentation, higher free radical scavenging activity was confirmed in ABTS and DPPH assay and the tendency to decrease in reactive oxygen species and nitrite production was found in a dose dependent manner. Conclusions The fermentation of ICGH with two selected strains enhanced the antioxidant activity and decreased NO production by increase in the bioavailability of active compounds in ICGH. Funding Sources National Institute of Agricultural Sciences.

Tazarotene-loaded in situ gels for potential management of psoriasis: biocompatibility, anti-inflammatory and analgesic effect

Psoriasis is a chronic autoinflammatory disorder characterized by patches of abnormal skin. For psoriasis management, the application of topical retinoids as Tazarotene is recommended. However, Tazarotene could induce skin irritation limiting its use. Herein, it is evaluated the possible usage of in situ gels for tazarotene skin delivery. The topical in situ gels were developed using thermosensitive poloxamers via cold method. They were examined for their appearance, sol-gel temperature, clarity, pH, viscosity, in vitro release, and stability. Their biocompatibility was evaluated by investigating their cytotoxicity and irritation inducing capacity. The possible anti-inflammatory and analgesic activities were determined by measuring the nitric oxide and prostaglandin E2 levels production in LPS-stimulated RAW264.7 murine macrophage cells. It was revealed that the in situ gels had no cytotoxic effect (∼95–100% cell viability) and nor irritation potential (∼97% cell viability), according to the in vitro EpiDerm™ reconstituted skin irritation test. Additionally, the 10% tazarotene-in situ gels showed possible analgesic activity since the production of prostaglandin E2 (PGE2) was decreased. In further, both concentrations of 5% and 10% tazarotene-in situ gels inhibited significantly the nitrite oxide production at 16% and 19%, respectively. Finally, the prepared in situ gels can act as a potential non-irritant alternative option for tazarotene topical skin delivery.

Design and synthesis of 1,3-benzothiazinone derivatives as potential anti-inflammatory agents

A series of 1,3-benzothiazinone derivatives were designed and synthesized for pharmacological assessments. Among the synthesized 19 compounds, some compounds showed high activities on inhibiting LPS-induced nitrite oxide and TNF-α production, down-regulating COX-2 and increasing IL-10 production in RAW264.7 cells. All the compounds had no obvious cytotoxicity in in vitro assay. LD50 value of compound 25 was greater than 2000 mg/kg, which was safer than meloxicam. Compound 25 significantly inhibited phosphorylation of NF-κB and STAT3 in LPS-induced RAW264.7 cells. Inhibition of synthesized compounds on COX activity was weaker than meloxicam. Compound 25 displayed lower gastrointestinal toxicity than meloxicam. Besides, compound 25 decreased the swelling in carrageenan-induced paw edema models of inflammation and reduced PGE2 level significantly. In summary, 1,3-benzothiazinone derivatives are unique scaffolds with anti-inflammatory activity and low toxicity.

Cordyceps cicadae Mycelia Ameliorate Cisplatin-Induced Acute Kidney Injury by Suppressing the TLR4/NF-κB/MAPK and Activating the HO-1/Nrf2 and Sirt-1/AMPK Pathways in Mice.

Acute kidney injury (AKI) is a common clinical problem, characterized by a sudden loss of renal function, a high risk of death, and the eventual development of renal fibrosis and renal failure. Cordyceps cicadae is a traditional Chinese medicine with the potential function of kidney protection. We analyze two sputum extracts, a water extract (WCC), and an ethanol extract (ECC), to assess the potential of treating AKI in an animal model of kidney injury induced by cisplatin. A nephrotoxic mouse model was first established by intraperitoneal injection of cisplatin. Subsequently, WCC and ECC were orally administered in these mice. The results show that WCC and ECC significantly alleviated cisplatin-induced AKI renal histological changes, serum creatinine (CRE) and blood urea nitrogen (BUN) production, and the levels of NO, TNF-α, IL-1β, and IL-6. The levels of malondialdehyde (MDA) and glutathione (GSH) were suppressed by administration of WCC and ECC. However, WCC treatment prevented these changes significantly better than ECC treatment. In addition, Western blot data showed that WCC attenuated the cisplatin-induced protein expression of cyclooxygenase-2 (COX-2) and inducible NO synthase (iNOS), as well as inhibiting nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) activation in the kidney tissues. Furthermore, WCC greatly inhibited the expression of Toll-like receptor 4 (TLR4) and cisplatin-induced NF-κB activation, as well as dramatically increasing the production of antioxidative enzymes (i.e., superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase, nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase 1 (HO-1)), silent information regulator T1 (Sirt1), and p-AMP-activated protein kinase (AMPK) in the kidney tissues. In addition, we found that WCC increased the expression levels of the autophagy-related proteins LC3B and Beclin-1; proapoptotic proteins, including cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP) 1; and organic anion transporters 1 (OAT1) and 3 (OAT3) in the kidney tissues. Finally, WCC, ECC, and two bioactive compounds—adenosine and N6-(2-hydroxyethyl) adenosine (HEA)—inhibited the production of nitrite oxide (NO) and intracellular reactive oxygen species (ROS) triggered by lipopolysaccharide- (LPS-) stimulated RAW264.7 macrophages in vitro. Collectively, WCC could provide a potential therapeutic candidate for the prevention of cisplatin-induced kidney injury through the inhibition of oxidative stress and inflammation.

Centipeda minima extract exerts antineuroinflammatory effects via the inhibition of NF-κB signaling pathway

BackgroundCentipeda minima (L.) A.Br. (C. minima) has been used in traditional Chinese herbal medicine to treat nasal allergy, diarrhea, asthma and malaria for centuries. Recent pharmacological studies have demonstrated that the ethanol extract of C. minima (ECM) and several active components possess anti-bacterial, anti-arthritis and anti-inflammatory properties. However, the effects of ECM on neuroinflammation and the underlying mechanisms have never been reported. PurposeThe study aimed to examine the potential inhibitory effects of ECM on neuroinflammation and illustrate the underlying mechanisms. MethodsHigh performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was performed to qualify the major components of ECM; BV2 and primary microglial cells were used to examine the anti-inflammatory activity of ECM in vitro. To evaluate the anti-inflammatory effects of ECM in vivo, the mice were orally administrated with ECM (100, 200 mg•kg−1•d−1) for 2 days before cotreatment with LPS (2 mg•kg−1•d−1, ip) for an additional 3 days. The mice were sacrificed the day after the last treatment and the hippocampus was dissected for further experiments. The expression of inflammatory proteins and the activation of microglia were respectively detected by real-time PCR, ELISA, Western blotting and immunofluorescence. ResultsHPLC-MS/MS analysis confirmed and quantified seven chemicals in ECM. In BV2 and primary microglial cells, ECM inhibited the LPS-induced production of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), thus protecting HT22 neuronal cells from inflammatory damage. Furthermore, ECM inhibited the LPS-induced activation of NF-κB signaling pathway and subsequently attenuated the induction of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2), NADPH oxidase 2 (NOX2) and NADPH oxidase 4 (NOX4), leading to the decreased production of nitrite oxide, prostaglandin E2 (PGE2) and reactive oxygen species (ROS). In an LPS-induced neuroinflammatory mouse model, ECM was found to exert anti-inflammatory activity by decreasing the production of proinflammatory mediators, inhibiting the phosphorylation of NF-κB, and reducing the expression of COX2, iNOS, NOX2 and NOX4 in the hippocampal tissue. Moreover, LPS-induced microglial activation was markedly attenuated in the hippocampus, while ECM at a high dose possesses a stronger anti-inflammatory activity than the positive drug dexamethansone (DEX). ConclusionThese findings demonstrate that ECM exerts antineuroinflammatory effects via attenuating the activation of NF-κB signaling pathway and inhibiting the production of proinflammatory mediators both in vitro and in vivo. C. minima might become a novel phytomedicine to treat neuroinflammatory diseases.

Sea bass (Lateolabrax maculatus) accelerates wound healing: A transition from inflammation to proliferation

Ethnopharmacological relevanceSea bass (Lateolabrax maculatus) has been used for dietary therapy practice for wound healing of puerperal or surgery patients in China. Traditional Chinese medicinal books also documented that sea bass can be used to manage inflammation-associated conditions such as wound, miscarriage and cough. Some studies also proved that dietary supplement with fish benefited for treating many inflammatory - associated conditions, such as cardiovascular disease, ulcerative colitis and hyperlipidemia. However, the studies on the pharmacological mechanisms of wound healing efficacy of sea bass remain lack of investigation. Aim of the studyThe aim of this study is to investigate the molecular mechanisms of sea bass on wound healing efficacy. Establishing a further justification for clinical application of aqueous extract of sea bass (ASB) in treating wound healing. Materials and methodsTransition from inflammation to proliferation phase treated as the critical step in wound repair which were investigated via in vitro and in vivo study. A series of inflammatory mediators associated with wound healing and proliferation effects of fibroblasts upon treatments were studied via Western blotting, enzyme-linked immunosorbent assay (ELISA), real time reverse transcription-polymerase chain reaction (RT-PCR) and scratch assay. The cutaneous wound model was applied on skin wound healing study to observe the healing process in C57BL/6 mice upon ASB treatments. Hematological parameters and tumor necrosis factor-α (TNF-α) secretions in serum were determined. Histopathological examinations were conducted by hematoxylin and eosin (H&E) staining and Masson staining. Immunofluorescence were performed to identify infiltrating neutrophils (MPO) and α-smooth muscle actin (α-SMA). ResultsResults showed that ASB significantly reduced the production of inflammatory mediators cyclooxygenase-2 (COX-2), nitrite oxide (NO) production and TNF-α. The phosphorylation and nuclear protein levels of transcription factor nuclear factor-κB (NF-ĸB) in toll-like receptor 4 (TLR4) signaling were decreased by ASB treatment as well. Wound closure rate and cyclin D1 expression level of fibroblasts were significantly increased by ASB treatments. Moreover, cutaneous wound model in C57BL/6 mice presented many similarities in appearance to the process of wound healing. ConclusionsThe in vitro study demonstrated an inhibitory effect of ASB on the inflammatory mediators regulated by TLR4 signaling pathways, providing evidence that ASB treatment potentially accelerate the wound healing through migration and proliferation enhancement. Additionally, the in vivo study suggested that ASB treatment has a potential in accelerating the proliferation phase of wound healing via well-organized abundant collagen deposition, angiogenesis and re-epithelialization in wounds. The present findings can be treated as a pharmacological basis for the folk use of sea bass and further studies in biological and medical fields.

The Candida albicans arginase family encodes enzymes with diverse catabolic activities that differentially influence host–fungus interactions

In the blood stream, arginine is an essential amino acid that is required by phagocytes to synthesize iNOS. Previously we showed that the fungus Candida albicans induces host arginase production that diverts arginine from the pathway that leads to the production of nitrite oxide. We therefore investigated whether C. albicans arginase activity also contributed to the protection of the fungus by competing for arginine during infections. Three C. albicans genes had been annotated as putative arginase encoding genes. Heterologous expression of these genes suggested all three had some arginase activity and one gene product (Car1) encoded a bone fide arginase that was required for growth on arginine. However, single and double mutations in the two other genes (AGM1 and GBU1) did not affect growth on arginine as a single nitrogen source and were found instead to encode agmatinase and guanidinobutyrase respectively that participate in two other pathways related to arginine metabolism. This family of three enzymes therefore exhibits mixed biochemical activities and collectively participate in the catabolism of exogenous and endogenous sources of arginine. Virulence of the triple mutant lacking all three genes was reduced in a Galleria infection model, but single or double mutants were fully virulent. None of the single or multiple mutants affected host NO production suggesting they do not influence the oxidative burst of phagocytes. In addition, CAR1 expression was required for hyphal growth. This family of enzymes therefore represent a novel enzyme set that is essential for growth in vivo and indirectly for fungal virulence.

Lanceoleins A–G, hydroxychalcones, from the flowers of Coreopsis lanceolata and their chemopreventive effects against human colon cancer cells

Seven new chalcones, lanceolein A–G (compounds 5 and 7–12), as well as five known chalcones (1–4 and 6), were isolated from the methanolic extract of Coreopsis lanceolata flowers. The chemical structures of 5 and 7–12 were determined on the basis of spectroscopic data interpretation. All compounds inhibited the production of nitrite oxide (NO) induced by LPS in RAW264.7 macrophage cells. Also, compounds 1–6 showed moderated cytotoxicity against human colon cancer cell lines, while compounds 7–12 hardly showed the cytotoxicity. Especially, compounds 2, 5, and 6 exhibited a little higher cytotoxicity on HCT15 cells, with IC50 values of 43.7 ± 2.17 μM, 35.6 ± 0.24 μM, and 47.9 ± 1.18 μM, respectively. In the Tali assay, compounds 2 and 5 increased the numeral of apoptotic cells. These compounds also significantly promoted the expression of apoptotic proteins including PARP and caspase-3.

The silkrose of Bombyx mori effectively prevents vibriosis in penaeid prawns via the activation of innate immunity

We previously identified novel bioactive polysaccharides from Bactrocera cucurbitae and Antheraea yamamai that activate innate immunity in RAW264 murine macrophages. However, in terms of potential applications in the cultivation of prawns, there were problems with the availability of these insects. However, we have now identified a polysaccharide from Bombyx mori that activates innate immunity in RAW264 cells and penaeid prawns. This purified polysaccharide, termed silkrose of B. mori (silkrose-BM), has a molecular weight of 1,150,000 and produces a single symmetrical peak on HPLC. Eight of nine constitutive monosaccharides of silkrose-BM are concomitant with dipterose of B. cucurbitae (dipterose-BC) and silkrose of A. yamamai (silkrose-AY). The major differences are found in the molar ratios of the monosaccharides. Silkrose-BM is approximately 500-fold less potent than silkrose-AY (EC50: 2.5 and 0.0043 μg/mL, respectively) in a nitrite oxide (NO) production assay using RAW264 cells. However, the maximum NO production for silkrose-BM and AY were comparable and higher than that of the lipopolysaccharide of Escherichia coli. The survival of penaeid prawns (Litopenaeus vannamei and Marsupenaeus japonicus) after infection with Vibrio penaecida was significantly improved by both dietary silkrose-BM and B. mori pupae. This suggests that silkrose-BM effectively prevents vibriosis in penaeid prawns via the activation of innate immunity.

Thein vitroantileishmanial activity of essential oil fromAloysia gratissimaand guaiol, its major sesquiterpene againstLeishmania amazonensis

Leishmaniases is a tropical disease caused by protozoa of the genus Leishmania for which the current treatment is expensive, besides increasing reports of parasite resistance. This study investigated the anti-Leishmania amazonensis activity of the essential oil from Aloysia gratissima (AgEO) and guaiol, the major sesquiterpene constituent in the oil. Our results showed that AgEO killed promastigotes and intracellular amastigotes at an IC50 of 25 and 0·16 µg mL-1, respectively, while guaiol killed amastigotes at an IC50 of 0·01 µg mL-1. Both AgEO and guaiol were safe for macrophages up to 100 µg mL-1, as evaluated by the dehydrogenase activity, membrane integrity and phagocytic capacity. AgEO and guaiol did not induce nitrite oxide (NO) in resting macrophages and inhibited the production of NO in lipopolysaccharide-stimulated macrophages. The ultrastructural analysis suggested that AgEO and guaiol act directly on parasites, affecting promastigotes kinetoplast, mitochondrial matrix and plasma membrane. Together, these results pointed out that AgEO and guaiol could be promising candidates to develop anti-Leishmania drugs.

CD14 is critical for TLR2-mediated M1 macrophage activation triggered by N-glycan recognition

Agonist interaction with Toll-like receptors (TLRs) induces T cell-mediated immunity, which is effective against intracellular pathogens. Consequently, TLR agonists are being tried as immunomodulatory agents. The lectin ArtinM targets TLR2 N-glycans on macrophages, induces cytokines production, and promotes T helper-1 immunity, a process that culminates in resistance to several parasitic and fungal infections in vivo. Because co-receptors influence agonist binding to TLRs, we investigated whether CD14 is required for macrophage activation induced by ArtinM. Macrophages from wild-type mice stimulated by ArtinM not only produced cytokines but also had the following activation profile: (i) expression of M1 polarization markers; (ii) nitrite oxide production; (iii) cellular migration; (iv) enhanced phagocytic and fungicide activity; (v) modulation of TLR2 expression; and (vi) activation of NF-κB pathway. This activation profile induced by ArtinM was evaluated in macrophages lacking CD14 that showed none of the ArtinM effects. We demonstrated by immunoprecipitation and sugar inhibition assays the physical interaction of ArtinM, TLR2, and CD14, which depends on recognition of the trimannoside that constitutes the core of N-glycans. Thus, our study showed that CD14 is critical for ArtinM-induced macrophage activation, providing fundamental insight into the design of anti-infective therapies based on carbohydrate recognition.

Nitrification gene ratio and free ammonia explain nitrite and nitrous oxide production in urea-amended soils

The atmospheric concentration of nitrous oxide (N2O), a potent greenhouse gas and ozone-depleting chemical, continues to increase, due largely to the application of nitrogen (N) fertilizers. While nitrite (NO2−) is a central regulator of N2O production in soil, NO2− and N2O responses to fertilizer addition rates cannot be readily predicted. Our objective was to determine if quantification of multiple chemical variables and structural genes associated with ammonia (NH3)- (AOB, encoded by amoA) and NO2−-oxidizing bacteria (NOB, encoded by nxrA and nxrB) could explain the contrasting responses of eight agricultural soils to five rates of urea addition in aerobic microcosms. Significant differences in NO2− accumulation and N2O production by soil type could not be explained by initial soil properties. Biologically-coherent statistical models, however, accounted for 70–89% of the total variance in NO2− and N2O. Free NH3 concentration accounted for 50–85% of the variance in NO2− which, in turn, explained 62–82% of the variance in N2O. By itself, the time-integrated nxrA:amoA gene ratio explained 78 and 79% of the variance in cumulative NO2− and N2O, respectively. In all soils, nxrA abundances declined above critical urea addition rates, indicating a consistent pattern of suppression of Nitrobacter-associated NOB due to NH3 toxicity. In contrast, Nitrospira-associated nxrB abundances exhibited a broader range of responses, and showed that long-term management practices (e.g., tillage) can induce a shift in dominant NOB populations which subsequently impacts NO2− accumulation and N2O production. These results highlight the challenges of predicting NO2− and N2O responses based solely on static soil properties, and suggest that models that account for dynamic processes following N addition are ultimately needed. The relationships found here provide a basis for incorporating the relevant biological and chemical processes into N cycling and N2O emissions models.

Bioassay-Guided Isolation of Anti-Inflammatory Components from the Bulbs of Lilium brownii var. viridulum and Identifying the Underlying Mechanism through Acting on the NF-κB/MAPKs Pathway.

The bulbs of Lilium brownii var. viridulum (LB) are commonly used as both traditional Chinese medicines and popular functional food for many centuries in China. Previous studies reported that the extract of lily bulbs exhibited anti-inflammatory activity both in vivo and in vitro, but its active components and associated molecular mechanisms remain elusive. In the present study, using bioassay-guided isolation method, two phenylpropenoid acylglycerols, 1-O-feruloyl-2-O-p-coumaroylglycerol (1) and 1,3-O-diferuloylglycerol (2), were obtained and identified from the chloroform fraction of LB. Both compounds 1 and 2 significantly decreased the production of nitrite oxide (NO) in lipopolysaccharide (LPS)-stimulated mouse macrophage RAW264.7 cells in a dose-dependent manner with half maximal inhibitory concentration (IC50) values of 9.12 ± 0.72 μM and 12.01 ± 1.07 μM, respectively. They also inhibited the production of prostaglandin E2 (PGE2) and several other pro-inflammatory cytokines, such as interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). Furthermore, compounds 1 and 2 downregulated the protein levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). They also inhibited the nuclear translocation of nuclear factor-κB (NF-κB) p65 subunit and suppressed mitogen-activated protein kinases (MAPKs) pathway. Taken these data together, compounds 1 and 2 exhibited anti-inflammatory activities through acting on the NF-κB and MAPKs pathway. This research provides the first evidence on the major bioactive constituents and related molecular mechanisms of LB as an anti-inflammatory agent. Our findings also advanced the understanding of LB as a traditional herbal medicine for the prevention and treatment of inflammation.

Effect of curcumin on acute spinal cord injury in mice via inhibition of inflammation and TAK1 pathway.

Traumatic spinal cord injury (SCI) is damage to the spinal cord that results in damaged spinal cord function. As a natural compound, curcumin has recently been shown to have anti-inflammatory and strong antioxidant activities. To investigate the effect of curcumin against acute spinal cord injury (SCI), we explored its induced effects in SCI mice. Transforming growth factor (TGF)-activated kinase 1 (TAK1) is a member of the MAPKKK family and plays an essential role in TNF, IL-1, and Toll-like receptor (TLR) signaling pathways. One hundred adult female KM mice were randomly divided into 5 groups (Control, Model, Test-L, Test-M, and Test-H). SCI was induced using the method described by Allen's. Motor function of the hindlimbs was evaluated on days 1, 7, 14, 21, and 28 after the injury using the motor rating test on the Basso mouse scale (BMS). 7 days after SCI, the levels of TNF-α, IL-1β, and IL-6 were measured by enzyme-linked immunosorbent assay (ELISA); the level of NO was evaluated by Griess assay; and Western blot was used to verify the levels of proteins in the TAK1 pathway. Expressions of GFAP positive cells in injured spinal cord were detected by immunohistochemical staining. The experiment showed that curcumin markedly inhibited SCI-induced production of inflammatory mediators, including TNF-α, IL-1β, IL-6 (ELISA assay) and nitrite oxide (Griess method) in a concentration-dependent manner. Curcumin decreased the phosphorylation levels of TGF-β-activated kinase 1 (TAK1) protein, leading to decreased phosphorylation levels of MKK6 and p38 MAPKs, key players in the microglia-mediated inflammatory response. Curcumin also significantly down-regulated the expression levels of the NF-κB upstream regulators IκB and IκB kinase (IKK). Additionally, behavior research showed that curcumin-treated mice showed significantly improved functional recovery compared to untreated mice (BMS assay). The expressions of GFAP increased in the injured spinal cord segments, which were decreased by Teat-M and Teat-H at 7d after SCI. Curcumin restores mice hind-limb function that has been reduced by SCI. This occurs by inhibition of TAK1/MKK6/p38MAPK via the TAK1 and NFκB pathways and inflammation. These results suggest the therapeutic potential for curcumin in the treatment of SCI.

Antifungal-demelanizing properties and RAW264.7 macrophages stimulation of glucan sulfate from the mycelium of the mushroom Ganoderma lucidum.

Underutilized mycelium of Ganoderma lucidum BCCM 31549 has been a significant source of a glucan sulfate (GS) possessing therapeutic activities. GS have been evaluated for their antifungaldemelanizing properties and nitrite oxide production from stimulated RAW264.7 macrophages. GS exhibited antifungal activity against Aspergillus niger A60 with a minimum inhibitory concentration of 60 mg/mL and a minimum fungicidal concentration of 100 mg/mL. At 60mg/mL (sublethal) and 30mg/mL (subinhibitory) doses of GS, the mycelium of A. niger A60 was successfully demelanized with a conidiophore head and black pigment reduction. Additionally, GS successfully stimulated RAW264.7 macrophage cells at a concentration of 500 μg/mL to produce 0.45 μM of nitric oxide. The GS-stimulated RAW264.7 macrophages were morphologically similar to those treated with lipopolysaccharide. The results highlight a novel bifunctional property of mycelial GS from G. lucidum.

The effect of nano-encapsulated Aliskiren on nitrite oxide production in cardiovascular system in experimental hypertension

The effect of nano-encapsulated Aliskiren on nitrite oxide production in cardiovascular system in experimental hypertension

The Anti-inflammatory Activities of Two Major Withanolides from Physalis minima Via Acting on NF-κB, STAT3, and HO-1 in LPS-Stimulated RAW264.7 Cells.

Physalis minima has been traditionally used as a folk herbal medicine in China for the treatment of many inflammatory diseases. However, little is known about its anti-inflammatory constituents and associated molecular mechanisms. In our study, withaphysalin A (WA) and 2, 3-dihydro-withaphysalin C (WC), two major withanolide-type compounds, were obtained from the anti-inflammatory fraction of P. minima. Both WA and WC significantly inhibited the production of nitrite oxide (NO), prostaglandin E2 (PGE2), and several pro-inflammatory cytokines, such as interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α) in lipopolysaccharide (LPS)-activated RAW264.7 macrophages. Further research indicated that they downregulated the LPS-induced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at the mRNA and protein levels. In addition, they also suppressed nuclear translocation of NF-κB p65, phosphorylation of STAT3, and upregulated HO-1 expression. Intriguingly, the activation of MAPKs was suppressed by WA but was not altered by WC. Taken together, these data provide scientific evidence for elucidating the major bioactive constituents and related molecular mechanisms for the traditional use of P. minima and suggest that WA and WC can be attractive therapeutic candidates for various inflammatory diseases.

Role of transient receptor potential vanilloid 1 in regulating erythropoietin-induced activation of endothelial nitric oxide synthase.

Erythropoietin (EPO), the key hormone involved in erythropoiesis, beneficially affects endothelial cells (ECs), but the detailed mechanisms are yet to be completely understood. In this study, we investigated the role of transient receptor potential vanilloid type 1 (TRPV1), a ligand-gated non-selective calcium (Ca2+ ) channel, in EPO-mediated endothelial nitric oxide synthase (eNOS) activation and angiogenesis. In ECs, EPO time dependently increased intracellular levels of calcium; this increase was abrogated by the Ca2+ chelators and pharmacological inhibitors of TRPV1 in bovine aortic ECs (BAECs) and TRPV1-transfected HEK293 cells. In addition, EPO-induced nitrite oxide (NO) production, phosphorylation of eNOS, Akt and AMP-activated protein kinase (AMPK) and the formation of TRPV1-Akt-AMPK-eNOS complex as well as tube formation were diminished by the pharmacological inhibition of TRPV1 in BAECs. Moreover, EPO time dependently induced the phosphorylation of phospholipase C-γ1 (PLC-γ1). Inhibition of PLC-γ1 activity blunted the EPO-induced Ca2+ influx, eNOS phosphorylation, TRPV1-eNOS complex formation and NO production. The phosphorylated level of eNOS increased in the aortas of EPO-treated wild-type (WT) mice or EPO-transgenic (Tg) mice but not in those of EPO-treated TRPV1-deficient (TRPV1-/- ) mice or EPO-Tg/TRPV1-/- mice. Matrigel plug assay showed that EPO-induced angiogenesis was abrogated in TRPV1 antagonist capsazepine-treated WT mice and TRPV1-/- mice. These findings indicate the EPO-induced Ca2+ influx via the activation of the PLC-γ1 signalling pathway, which leads to TRPV1 activation and consequently increases the association of the TRPV1-Akt-AMPK-eNOS complex, eNOS activation, NO production and angiogenesis.

Isocyperol, isolated from the rhizomes of Cyperus rotundus, inhibits LPS-induced inflammatory responses via suppression of the NF-κB and STAT3 pathways and ROS stress in LPS-stimulated RAW 264.7 cells

The rhizomes of Cyperus rotundus (cyperaceae) have been used in Korean traditional medicines for treating diverse inflammatory diseases. However, little is known about the biological activities of isocyperol, a sesquiterpene isolated from C. rotundus, and their associated molecular mechanisms. In this study, we found that isocyperol significantly inhibited lipopolysaccharide (LPS)-induced production of nitrite oxide (NO) and prostaglandin E2 (PGE2) and suppressed LPS-induced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at the mRNA and protein levels in RAW 264.7 macrophages. In addition, isocyperol downregulated the LPS-induced expression of several proinflammatory cytokines, such as interleukin-1beta (IL-1β), IL-6, and monocyte chemotactic protein-1 (MCP-1). Isocyperol treatment suppressed the LPS-induced nuclear translocation and transcriptional activation of nuclear factor-kappaB (NF-κB) in macrophages. Moreover, the activation of STAT3, another proinflammatory signal, was suppressed by isocyperol in LPS-stimulated RAW 264.7 cells. Isocyperol pretreatment also induced heme oxygenase-1 (HO-1) expression and reduced LPS-stimulated reactive oxygen species (ROS) accumulation in macrophages. Furthermore, isocyperol significantly increased the survival rate and attenuated serum levels of NO, PGE2, and IL-6 in LPS-induced septic shock mouse model. Taken together, these data indicate that isocyperol suppress septic shock through negative regulation of pro-inflammatory factors through inhibition of the NF-κB and STAT3 pathways and ROS. To our knowledge, this is the first report on the biological activity of isocyperol and its molecular mechanism of action.

The anti-inflammation and pharmacokinetics of a novel alkaloid from Portulaca oleracea L.

This study was to elucidate the pharmacokinetics of a novel alkaloid, 6-acetyl-2,2,5-trimethyl-2,3-dihydrocyclohepta[b]pyrrol-8(1H)-one, named oleracone isolated from Portulaca oleracea L., and to examine the anti-inflammatory ability with lipopolysaccharide (LPS) stimulated macrophages. The novel alkaloid, oleracone, was isolated from Portulaca oleracea L., and its structure was determined by spectroscopic analysis including HRESIMS, 2D NMR spectroscopic data and single-crystal X-ray diffraction. The activity of anti-inflammation was assayed via the test with RAW 264.7 activated by LPS, and the pharmacokinetics of oleracone in rat plasma after intravenous and oral administration at dose of 2.5 mg/kg was, respectively, investigated by a rapid and sensitive ultra high-performance liquid chromatography (UHPLC) method with bergapten as internal standard. Oleracone was a novel alkaloid first isolated from Portulaca oleracea L. and possessed unique structure in natural products, whose anti-inflammatory effecting on nitrite oxide production and several pivotal pro-inflammatory cytokines was found at the concentration of 50 μm, and the pharmacokinetic results indicated that oleracone was rapidly distributed with Tmax of 15.7 min after oral administration and presented a higher oral absolute bioavailability to be 74.91 ± 10.7%. Oleracone as novel alkaloid presented remarkably anti-inflammatory effect, which was rapid distributed in rat with high bioavailability of 74.91 ± 10.7%.