Modulation of CAT-2B-Mediated l-Arginine Uptake and Nitric Oxide Biosynthesis in HCT116 Cell Line Through Biological Activity of 4'-Geranyloxyferulic Acid Extract from Quinoa Seeds.

Chenopodium quinoa Wild is a "pseudocereal" grain which attracts a lot of attention in the scientific community as it has a positive effect on health. Here, we investigate the presence of biologically active O-prenylated phenylpropanoids in the ethanol extract of commercially available quinoa seeds. We claim that 4'-Geranyloxyferulic acid (GOFA) was the only phytochemical product found that belongs to quinoa's group secondary metabolites. We studied the changes in the oxidative and inflammatory status of the cellular environment in HCT 116 cell line processed with quinoa extract and its component GOFA; the implementation was done through the analysis of the antioxidant enzymes (SOD and CAT), the pro-inflammatory components (iNOS, IL-6 and TNF-α), and the products of intermediary metabolism (ONOO-, O2-). Moreover, the l-arginine uptake was proposed as a target of the tested compounds. We demonstrated that the GOFA, through a decrease of the CAT-2B expression, leads to a reduction of the l-arginine uptake, downregulating the harmful iNOS and restoring the altered redox state. These results propose a new molecular target involved in the reduction of the critical inflammatory process responsible for the cancer progression.

Modulation of the oxidative plasmatic state in gastroesophageal reflux disease with the addition of rich water molecular hydrogen: A new biological vision.

Gastroesophageal reflux disease (GERD), a clinical condition characterized by reflux of gastroduodenal contents in the oesophagus, has proved to demonstrate a strong link between oxidative stress and the development of GERD. Proton pump inhibitors (PPIs) have been universally accepted as first-line therapy for management of GERD. The potential benefits of electrolysed reduced water (ERW), rich in molecular hydrogen, in improving symptoms and systemic oxidative stress associated with GERD was assessed. The study was performed on 84 GERD patients undergoing control treatment (PPI + tap water) or experimental treatment (PPI + ERW) for 3 months. These patients were subjected to the GERD-Health Related Quality of Life Questionnaire as well as derivatives reactive oxigen metabolites (d-ROMs) test, biological antioxidant potential (BAP) test, superoxide anion, nitric oxide and malondialdehyde assays, which were all performed as a proxy for the oxidative/nitrosative stress and the antioxidant potential status. Spearman's correlation coefficient was used to evaluate the correlation between scores and laboratory parameters. Overall results demonstrated that an optimal oxidative balance can be restored and GERD symptoms can be reduced rapidly via the integration of ERW in GERD patients. The relative variation of heartburn and regurgitation score was significantly correlated with laboratory parameters. Thus, in the selected patients, combination treatment with PPI and ERW improves the cellular redox state leading to the improvement of the quality of life as demonstrated by the correlation analysis between laboratory parameters and GERD symptoms.

Biological Effect of Licochalcone C on the Regulation of PI3K/Akt/eNOS and NF-κB/iNOS/NO Signaling Pathways in H9c2 Cells in Response to LPS Stimulation.

Polyphenols compounds are a group molecules present in many plants. They have antioxidant properties and can also be helpful in the management of sepsis. Licochalcone C (LicoC), a constituent of Glycyrrhiza glabra, has various biological and pharmacological properties. In saying this, the effect of LicoC on the inflammatory response that characterizes septic myocardial dysfunction is poorly understood. The aim of this study was to determine whether LicoC exhibits anti-inflammatory properties on H9c2 cells that are stimulated with lipopolysaccharide. Our results have shown that LicoC treatment represses nuclear factor-κB (NF-κB) translocation and several downstream molecules, such as inducible nitric oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Moreover, LicoC has upregulated the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/endothelial nitric oxide synthase (eNOS) signaling pathway. Finally, 2-(4-Morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002), a specific PI3K inhibitor, blocked the protective effects of LicoC. These findings indicate that LicoC plays a pivotal role in cardiac dysfunction in sepsis-induced inflammation.

A Novel Biological Role of α-Mangostin in Modulating Inflammatory Response Through the Activation of SIRT-1 Signaling Pathway.

Several studies have shown that xanthones obtained from Garcinia Mangostana (GM) have remarkable biological activities. α-mangostin (α-MG) is the main constituent of the fruit hull of the GM. Several findings have suggested that SIRT-1, a nuclear histone deacetylase, could influence cellular function by the inhibition of NF-kB signaling. ROS can inhibit SIRT-1 activity by initiating oxidative modifications on its cysteine residues, and suppression of SIRT-1 enhances the NF-κB signaling resulting in inflammatory responses. The goals of the present study were to evaluate the quantity of α-MG in the methanolic extract of GM (Vithagroup Spa) and to investigate the activity of this xanthone in U937 cell line and in human monocytes from responsive to inflammatory insult analyzing the possible changes on the activation of SIRT-1 protein via NF-Kb. Cells were treated with the methanolic extract of GM and/or LPS. The chromatographic separation of α-MG was performed by an HPLC analysis. EX 527, a specific SIRT-1 inhibitor, was used to determine if SIRT-1/NfkB signaling pathway might be involved in α-MG action on cells. Our results show that α-MG inhibits p65 acetylation and down-regulates the pro-inflammatory gene products as COX-2, iNOS via SIRT-1 activation. Cells treated with EX 527 showed an up-regulation of NFkB acetylation and an over expression of inducible enzymes and their product of catalysis (NO and PGE2). These results suggest that α-MG may be useful for the development of alternative pharmacological strategies aimed at reducing the inflammatory process. J. Cell. Physiol. 231: 2439-2451, 2016. © 2016 Wiley Periodicals, Inc.

New Approach in Translational Medicine: Effects of Electrolyzed Reduced Water (ERW) on NF-κB/iNOS Pathway in U937 Cell Line under Altered Redox State.

It is known that increased levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS) can exert harmful effects, altering the cellular redox state. Electrolyzed Reduced Water (ERW) produced near the cathode during water electrolysis exhibits high pH, high concentration of dissolved hydrogen and an extremely negative redox potential. Several findings indicate that ERW had the ability of a scavenger free radical, which results from hydrogen molecules with a high reducing ability and may participate in the redox regulation of cellular function. We investigated the effect of ERW on H2O2-induced U937 damage by evaluating the modulation of redox cellular state. Western blotting and spectrophotometrical analysis showed that ERW inhibited oxidative stress by restoring the antioxidant capacity of superoxide dismutase, catalase and glutathione peroxidase. Consequently, ERW restores the ability of the glutathione reductase to supply the cell of an important endogenous antioxidant, such as GSH, reversing the inhibitory effect of H2O2 on redox balance of U937 cells. Therefore, this means a reduction of cytotoxicity induced by peroxynitrite via a downregulation of the NF-κB/iNOS pathway and could be used as an antioxidant for preventive and therapeutic application. In conclusion, ERW can protect the cellular redox balance, reducing the risk of several diseases with altered cellular homeostasis such as inflammation.

Verbascoside down-regulates some pro-inflammatory signal transduction pathways by increasing the activity of tyrosine phosphatase SHP-1 in the U937 cell line.

Polyphenols are the major components of many traditional herbal remedies, which exhibit several beneficial effects including anti-inflammation and antioxidant properties. Src homology region 2 domain-containing phosphatase-1 (SHP-1) is a redox sensitive protein tyrosine phosphatase that negatively influences downstream signalling molecules, such as mitogen-activated protein kinases, thereby inhibiting inflammatory signalling induced by lipopolysaccharide (LPS). Because a role of transforming growth factor ß-activated kinase-1 (TAK1) in the upstream regulation of JNK molecule has been well demonstrated, we conjectured that SHP-1 could mediate the anti-inflammatory effect of verbascoside through the regulation of TAK-1/JNK/AP-1 signalling in the U937 cell line. Our results demonstrate that verbascoside increased the phosphorylation of SHP-1, by attenuating the activation of TAK-1/JNK/AP-1 signalling. This leads to a reduction in the expression and activity of both COX and NOS. Moreover, SHP-1 depletion deletes verbascoside inhibitory effects on pro-inflammatory molecules induced by LPS. Our data confirm that SHP-1 plays a critical role in restoring the physiological mechanisms of inducible proteins such as COX2 and iNOS, and that the down-regulation of TAK-1/JNK/AP-1 signalling by targeting SHP-1 should be considered as a new therapeutic strategy for the treatment of inflammatory diseases.

Emotions, immunity and sport: Winner and loser athlete's profile of fighting sport.

Several studies have focused on the relationship between hormonal changes and affective states in sporting contexts relating to an agonistic outcome. More recently, pro-inflammatory cytokines have also been successfully associated with affective state modulation. The aim of this study was to investigate whether athletes who won or lost show different levels of steroid hormones (testosterone and cortisol), pro-inflammatory cytokine IL-1ß, or expressions of anger and anxiety during six training fights in seasonal competitions down to the main seasonal competition. In 25 male kick-boxing athletes (age±SD, 28.68±5.34), anger states (RS score) and anxiety states (AS score) were assessed by STAXI-2 and STAI-Y, respectively. Cortisol (C), testosterone (T) and IL-1ß salivary levels were measured by the ELISA method. The saliva samples were taken in the afternoon, 30min prior to the start and 30min from the end of both simulated and official competitions. The results showed that the RS score, T, T/C ratio salivary levels increased during the season, whereas the AS score, C and IL-1ß suggested an opposite trend. Close to an official competition, the RS score, T, T/C ratio and IL-1ß salivary concentrations were significantly higher, and then decreased during competition. By contrast, the AS score and C levels significantly increased throughout the official competition. In addition, significant differences were found for hormones and IL-1ß concentrations as well as psychometric assessment close to the outcome of an official match. Athletes who lost showed an higher AS score and C level, while those who won were characterized by an higher level during the pre-competition RS score, T, T/C ratio, and IL-1ß. Note that these factors were positively and significantly correlated at the pre-official competition time, while in a linear regression analysis, IL-1ß, T and T/C ratio concentrations explained 43% of the variance in the RS score observed at the same time (adjusted R(2)=0.43, ANOVA P<.05). Our data suggest that the beginning of an agonistic event could trigger emotional responses which correspond to different biological processes instead that of a simulated fight. In particular, IL-1ß could be a potential new biological marker of anger and the combined measurement of these factors may be a useful way of understanding athletes' change in relation to their performance.

Synthesis of a novel cyclic prodrug of S-allyl-glutathione able to attenuate LPS-induced ROS production through the inhibition of MAPK pathways in U937 cells.

A novel cyclic prodrug of S-allyl-glutathione (CP11), obtained by using an acyloxy-alkoxy linker, was estimated for its pharmacokinetic and biological properties. The stability of CP11 was evaluated at pH 1.2, 7.4, in simulated fluids with different concentrations of enzymes, and in human plasma. The anti-inflammatory ability of CP11 was assessed in U937 cells, an immortalized human monocyte cell line. Results showed that CP11 is stable at acidic pH showing a possible advantage for oral delivery due to the longer permanence in the stomach. Having a permeability coefficient of 2.49 × 10(-6) cm s(-1), it was classified as discrete BBB-permeable compound. Biological studies revealed that CP11 is able to modulate inflammation mediated by LPS in U937 cells preventing the increase of ROS intracellular levels through interaction with the MAPK pathway.

Novel aminobenzyl-acetamidine derivative modulate the differential regulation of NOSs in LPS induced inflammatory response: role of PI3K/Akt pathway.

BACKGROUND: Previous reports suggest that NO may contribute to the pathophysiology of septic shock. Recently, we have synthesized and characterized a series of benzyl- and dibenzyl derivative of N-(3-aminobenzyl)acetamidine, a potent and selective inhibitor of iNOS, in vitro assay. We evaluated the molecular mechanisms by which these compounds are involved in the regulation of NOSs expression. METHODS: H9c2 cells were stimulated with lipopolysaccharide (LPS) in the presence or absence of acetamidine-derivative. The NOSs mRNA and protein, and activation of signaling pathways (Akt and NF-κB) were assayed. RESULTS: The induction of endotoxic shock in H9c2 with LPS caused an increase of inducible NOS and a down-regulation of constitutive NOS. The molecular mechanism involved in the modulation of NOSs expression in H9c2 cells upon LPS stimulation resulted in the modification of the redox state responsible for NF-kB nuclear translocation via NIK -IKKα/ß-IkBα, simultaneously to the inactivation of the PI3K/Akt pathway. The compounds acted as an anti-inflammatory modulator. CONCLUSION: These results suggest that LPS regulates the opposite NOS expression in H9c2 cells by modifying the redox state of these cells responsible for the NF-kB nuclear translocation via NIK-IKKα/ß-IkBα, simultaneous to the inactivation of the PI3K/Akt pathway. The new molecule acts as an anti-inflammatory modulator in LPS-induced inflammation in H9c2 cells by the restoration of eNOS and nNOS expressions, mechanistically involving the PI3K/Akt pathway. GENERAL SIGNIFICANCE: This study delineates the underlying mechanisms of opposite NOSs expression in H9c2 cells stimulated with LPS.

Analysis of genomic methylation level using micellar electrokinetic chromatography with UV detection.

Analytical methods for quantification of 5'-methylcytosine in genomes are important tools to investigate epigenetic changes in gene expression during development, differentiation, aging, or cancer. Here, we report a novel genomic methylation content assay based on enzymatic hydrolysis of DNA and MEKC separation of 5'-deoxyribonucleoside monophosphates (dNMP) using the cationic surfactant CTAB as pseudostationary phase. Calf Thymus DNA was used during method development to determine electrophoretic parameters and electrolyte composition for a complete separation between 2'-deoxycytosine-5'-monophosphate and 2'-deoxy-5'-methylcytosine 5'-monophosphate (d5mCMP). Methylated and not methylated oligonucleotides were used to confirm the identity of each peak and evaluate analytical parameters of the method. The LOD of the method was found to be 12.5 pmol/µL for d5mCMP.

Positive correlation between serum interleukin-1ß and state anger in rugby athletes.

Recently, several studies reported a relationship between immune system activation and anger expression. Consequently, the aim of this study was to explore immunitary molecular mechanisms that potentially underlie anger expression. To this end, we applied the Frustration-Aggression Theory in a contact sport model, utilizing the nearing of sporting events to trigger anger feelings. In parallel, we evaluated the activation of immune system at mRNA levels. We enrolled 20 amateur rugby players (age ± SD, 27.2 ± 4.5) who underwent psychological assessment to evaluate anger, with the State-Trait Anger Expression Inventory-2 (STAXI-2), before rugby matches; at the same time blood samples were taken to analyze the variations of gene expression by microarray. During the 2 hr before each game, a significant increase was verified in the Rage State (RS) score compared to the score ascertained 72 hr before. At the same time, we found modulation in expression profile, in particular increased expression of gene that encodes interleukin l-ß (IL-1ß). In a regression analysis, RS score was related to IL-1ß, and the potential risk factors age, body mass index, smoking, and drinking. The levels of cytokine were positively and independently related to RS score. Our results suggest that the nearing of sporting event can trigger anger state feelings and activate immune system in rugby players. We propose the IL-1ß as a potential biological marker of anger. However, further research is necessary to clarify the correlation between cytokine and anger.

Activity of matrix metallo proteinases (MMPs) and the tissue inhibitor of MMP (TIMP)-1 in electromagnetic field-exposed THP-1 cells.

Matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) are the main determinants of tissue remodeling in both physiological and pathological processes. Metabolic processes, which generate oxidants and antioxidants can be influenced by environmental factors such as electromagnetic fields (EMF). We analyzed the effects of EMF on the activity and expression of MMPs in THP-1 cells. Cells were exposed to a 50 Hz, 1 mT EMF for 24 h and incubated with or without LPS. Our data indicate that THP-1 cells exposed to EMF causes a reduction of anti-oxidant enzyme activity and an enhancement of nitrogen intermediates involving the iNOS pathway. We then analyzed the role of nitration of TIMP-1 in increasing the activity of MMPs in EMF exposed cells. Molecular modeling tools were employed to identify the most plausible sites in the active conformation of TIMP-1; at least two protein sites, Y120 and Y38 and/or Y72 were identified. Reactive nitrogen species (RNS) may affect protein targets, such as TIMP-1, which are crucial for the regulation of MMP activities by oxidation of sulfydryl groups, or by nitration of tyrosine residues. These results may suggest a pathway connecting an imbalance of MMPs and their cognate inhibitor TIMP-1.

The role of inducible nitric oxide synthase and haem oxygenase 1 in growth and development of dental tissue'.

In this study, the activity of the antioxidant enzyme network was assessed spectrophotometrically in samples of dental pulp and dental papilla taken from third-molar gem extracts. The production of nitric oxide by the conversion of l-(2,3,4,5)-[3H] arginine to l-(3H) citrulline, the activity of haem oxygenase 1 (HO-1) through bilirubin synthesis and the expression of inducible nitric oxide synthase (iNOS), HO-1 proteins and messenger RNA by Western blot and reverse-transcribed polymerase chain reaction were also tested. The objective of this study was to evaluate the role of two proteins, iNOS and HO-1, which are upregulated by a condition of oxidative stress present during dental tissue differentiation and development. This is fundamental for guaranteeing proper homeostasis favouring a physiological tissue growth. The results revealed an over-expression of iNOS and HO-1 in the papilla, compared with that in the pulp, mediated by the nuclear factor kappa B transcription factor activated by the reactive oxygen species that acts as scavengers for the superoxide radicals. HO-1, a metabolically active enzyme in the papilla, but not in the pulp, seems to inhibit the iNOS enzyme by a crosstalk between the two proteins. We suggest that the probable mechanism through which this happens is the interaction of HO-1 with haem, a cofactor dimer indispensible for iNOS, and the subsequent suppression of its metabolic activity.

Astaxanthin treatment reduced oxidative induced pro-inflammatory cytokines secretion in U937: SHP-1 as a novel biological target.

It has been suggested that oxidative stress activates various intracellular signaling pathways leading to secretion of a variety of pro-inflammatory cytokines and chemokines. SHP-1 is a protein tyrosine phosphatase (PTP) which acts as a negative regulator of immune cytokine signaling. However, intracellular hydrogen peroxide (H(2)O(2)), generated endogenously upon stimulation and exogenously from environmental oxidants, has been known to be involved in the process of intracellular signaling through inhibiting various PTPs, including SHP-1. In this study, we investigated the potential role of astaxanthin, an antioxidant marine carotenoid, in re-establishing SHP-1 negative regulation on pro-inflammatory cytokines secretion in U-937 cell line stimulated with oxidative stimulus. ELISA measurement suggested that ASTA treatment (10 µM) reduced pro-inflammatory cytokines secretion (IL-1ß, IL-6 and TNF-α) induced through H(2)O(2), (100 µM). Furthermore, this property is elicited by restoration of basal SHP-1 protein expression level and reduced NF-κB (p65) nuclear expression, as showed by western blotting experiments.

Dysregulation of chemo-cytokine production in schizophrenic patients versus healthy controls.

BACKGROUND: The exact cause of schizophrenia is not known, although several aetiological theories have been proposed for the disease, including developmental or neurodegenerative processes, neurotransmitter abnormalities, viral infection and immune dysfunction or autoimmune mechanisms. Growing evidence suggests that specific cytokines and chemokines play a role in signalling the brain to produce neurochemical, neuroendocrine, neuroimmune and behavioural changes. A relationship between inflammation and schizophrenia was supported by abnormal cytokines production, abnormal concentrations of cytokines and cytokine receptors in the blood and cerebrospinal fluid in schizophrenia. Since the neuropathology of schizophrenia has recently been reported to be closely associated with microglial activation we aimed to determined whether spontaneous or LPS-induced peripheral blood mononuclear cell chemokines and cytokines production is dysregulated in schizophrenic patients compared to healthy subjects. We enrolled 51 untreated first-episode schizophrenics (SC) and 40 healthy subjects (HC) and the levels of MCP-1, MIP-1α, IL-8, IL-18, IFN-γ and RANTES were determined by Elisa method in cell-free supernatants of PBMC cultures. RESULTS: In the simultaneous quantification we found significantly higher levels of constitutively and LPS-induced MCP-1, MIP-1α, IL-8 and IL-18, and lower RANTES and IFNγ levels released by PBMC of SC patients compared with HC. In ten SC patients receiving therapy with risperidone, olanzapine or clozapine basal and LPS-induced production of RANTES and IL-18 was increased, while both basal and LPS-induced MCP-1 production was decreased. No statistically significant differences were detected in serum levels after therapy. CONCLUSION: The observation that in schizophrenic patients the PBMC production of selected chemo-cytokines is dysregulated reinforces the hypothesis that the peripheral cyto-chemokine network is involved in the pathophysiology of schizophrenia. These preliminary, but promising data are supportive of the application of wider profiling approaches to the identification of biomarker as diagnostic tools for the analysis of psychiatric diseases.

The biological effect of pharmacological treatment on dimethylaminohydrolases (DDAH-1) and cationic amino acid transporter-1 (CAT-1) expression in patients with acute congestive heart failure.

AIM: Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide (NO) which plays an important role in controlling vascular tone and regulates the contractile properties of cardiac myocytes. The aim of this study was to investigate the effect of pharmacological treatment on symmetric dimethylarginine (SDMA), ADMA and arginine plasma concentrations in patients with acute congestive heart failure (ACHF) through the evaluation of type-1 system cationic amino acid transporter-1/type 1 dimethylarginine dimethylaminohydrolases-1 (CAT-1/DDAH-1). METHODS AND RESULTS: 25 hospitalized cardiology patients with symptomatic acute congestive HF (NYHA Class III-IV) and impaired left ventricular (LV) function (ejection fraction<35%) were included in the study. ADMA, SDMA, and arginine plasma concentrations were assessed before and after pharmacological treatment by high performance liquid chromatography. All patients received an adequate pharmacological treatment for ACHF. ADMA and SDMA plasma levels were significantly higher after pharmacological treatment respect to baseline values (pre-treatment) (0.75 vs 0.48; 1.31 vs 1.03; p<0.01). Arginine plasma concentration was significantly lower after therapy respect to baseline values (0.78 vs 0.99; p<0.01). This is associated more with the modulation of DDAH-1 protein than with of CAT-1 system transport. CONCLUSIONS: In patients with ACHF, acute renal impairment function and the modulation of metabolism and extracellular transport by the DDAH-1/CAT-1 system determine high ADMA and SDMA levels after therapy for acute congestive heart failure.

Licocalchone-C extracted from Glycyrrhiza glabra inhibits lipopolysaccharide-interferon-γ inflammation by improving antioxidant conditions and regulating inducible nitric oxide synthase expression.

The genus Glycyrrhiza consists of about 30 species, amoung these, G. glabra is the source of several phenolic compounds, known as flavonoids, such as licoagrodin, licoagrochalcones, licoagroaurone and licochalcone C, kanzonol Y, glyinflanin B and glycyrdione A, which have shown various pharmacological activities, including antitumor, antiparasitic, antileishmanial, anti-ulcer and antioxidative effects. Among these compounds, licochalcone C was isolated but its biology has not been fully examined. In our study we reproduced an inflammatory state by treating THP-1 (human myelomonocytic leukaemia) cells with pro-inflammatory stimuli, such as LPS and IFN-γ and we investigated the possible antioxidant activity of licochalcone C at a concentration of 50 µM. Our results show that treatment with licochalcone C attenuates the LPS-IFN-γ-induced inflammatory response by significantly decreasing the expression and activity of iNOS via NFκB (nuclear factor kappa-B), by influencing extracellular O2⁻ production, and by modulating the antioxidant network activity of SOD (superoxide dismutase), CAT (catalase) and GPx (glutathione peroxidase) activity. Based on these results we hypothesize that Licochalcone C has antioxidant properties since it reduces the production of superoxide radicals and consequently reduces the activity of iNOS.

N-Substituted acetamidines and 2-methylimidazole derivatives as selective inhibitors of neuronal nitric oxide synthase.

A series of N-substituted acetamidines and 2-methylimidazole derivatives structurally related to W1400 were synthesized and evaluated as Nitric Oxide Synthase (NOS) inhibitors. Analogs with sterically hindering isopropyl and phenyl substituents on the benzylic carbon connecting the aromatic core of W1400 to the acetamidine nitrogen, showed good inhibitory potency for nNOS (IC(50)=0.2 and 0.3 µM) and selectivity over eNOS (500 and 1166) and to a lesser extent over iNOS (50 and 100). A molecular modeling study allowed to shed light on the effects of the structural modifications on the selectivity of the designed inhibitors toward the different NOS isoforms.

Antiinflammatory effects in THP-1 cells treated with verbascoside.

Verbascum thapsus commonly known as 'mullein' is part of a large family of Scrophulariaceae consisting of more than 360 species. From antiquity Verbascum thapsus has been used as a medicinal herb, it contains diverse polysaccharides, iroid glycosides, flavonoids, saponins, volatile oils and phenylentanoids. Inducible nitric oxide synthase (iNOS) represents one of the three isoforms that produce nitric oxide using L-arginine as a substrate in response to an increase in superoxide anion activated by NF-kB. It is implicated in different pathophysiological events and its expression increases greatly during an inflammatory process, due to oxidative stress and the activation of the enzymes of the antioxidant network such as SOD, CAT and GPx.In this study an inflammatory state was reproduced by treating THP-1 cells (human myelomonocytic leukaemia) with pro-inflammatory stimuli, such as LPS and IFN-gamma, obtaining an up-regulation both in the expression and in the activity of iNOS. The aim of the work was to investigate the antiinflammatory action of verbascoside using a concentration of 100 mum. The results show a significant decrease of the expression and activity of iNOS, extracellular O(2) (-) production, SOD, CAT and GPx activity when the cells were treated with verbascoside. Based on these results it is hypothesized that verbascoside has antiinflammatory properties since it reduces the production of superoxide radicals and consequently reduces the activity of iNOS.