Anti-Migratory Effects of 4'-Geranyloxyferulic Acid on LPS-Stimulated U937 and HCT116 Cells via MMP-9 Down-Regulation: Involvement of ROS/ERK Signaling Pathway.

Matrix metalloproteinases (MMPs) play a crucial role in tumor angiogenesis, and metastasis. 4'-geranyloxyferulic acid (GOFA) has anti-tumor and anti-inflammatory proprieties. Herein, we aimed to determine whether this compound affects cell survival, invasion, and migration through reactive oxygen species (ROS)-mediated MMPs activation of extracellular signal-regulated kinases (ERKs) and p38 signaling in lymphocytic histiocytoma (U937) and colorectal cancer (HCT116) cells. We observed that lipopolysaccharide (LPS) stimulated U937 and HCT116 cells presented abnormal cell proliferation and increased metalloproteinase (MMP-9) activity and expression. Non-cytotoxic doses of GOFA blunted matrix invasive potential by reducing LPS-induced MMP-9 expression and cell migration via inhibiting ROS/ ERK pathway. GOFA also attenuated apoptosis and cell senescence. Our findings indicate that GOFA, inhibiting cancer cell proliferation and migration, could be therapeutically beneficial to prevent tumor metastasis.

Erythropoietin induces miRNA-210 by JAK2/STAT5 signaling in PBMCs of End-stage Renal Disease patients.

Anemia of chronic kidney disease is associated with blunted response/resistance to erythropoietin-stimulating agents (ESAs) in hemodialysis (HD) patients. Several molecules have been successfully associated with ESA responsiveness; however, none of them is now considered a valid therapeutic biomarker of erythropoietin resistance in these patients. We performed an evaluation of the level of specific plasma circulating miRNAs in blood samples of HD patients, in relation to ESA treatment, with a follow-up of 1 year (T0-T3). We found significantly lower circulating levels of all miRNAs analyzed at baseline (T0) in HD patients vs. healthy control (HC). The plasmatic levels of miRNA-210 resulted significantly and negatively associated with Erythropoietin Resistance Index (ERI), and the variance of ΔmiRNA-210 (miRNA-210T3 minus miRNA-210T0 ) explained significant percentage of ΔERI (ERIT3 minus ERIT0 ) variance. The receiver operating characteristic analysis at T0 showed that the plasmatic level of miRNA-210 could distinguish HD patients with positive or negative trend in ERI at T3. In vitro, recombinant human erythropoietin (EPO) induced significant release of miRNA-210 from cultured peripheral blood mononuclear cells, through the activation of Janus kinase 2 (JAK2)/ signal transducer and activator of transcription 5 (STAT5) signaling, but not by the activation of the MAPK protein 38α and extracellular signal-regulated kinase ½. Accordingly, HD patients with negative ΔERI showed higher level of phosphor-Janus kinase 2 and nuclear translocation of phosphor-signal transducer and activator of transcription 5. vs. patients with positive ΔERI or HC. Our data highlighted that chronic HD significantly reduces the circulating level of the miRNAs evaluated; within the targets analyzed, the miRNA-210 could be considered as a prognostic indicator of ESA responsiveness and index for anemia management.

Modulation of Apoptotic Cell Death and Neuroprotective Effects of Glutathione-L-Dopa Codrug Against H2O2-Induced Cellular Toxicity.

The L-3,4-dihydroxyphenylalanine (LD) is the gold standard drug currently used to manage Parkinson's disease (PD) and to control its symptoms. However, LD could cause disease neurotoxicity due to the generation of pro-oxidant intermediates deriving from its autoxidation. In order to overcome this limitation, we have conjugated LD to the natural antioxidant glutathione (GSH) to form a codrug (GSH-LD). Here we investigated the effect of GSH-LD on H2O2-induced cellular toxicity in undifferentiated and differentiated lymphoma U-937 and dopaminergic neuroblastoma SH-SY5Y cell lines, used respectively as models to study the involvement of macrophages/microglia and dopaminergic neurons in PD. We analyzed the effect of GSH-LD on apoptosis and cellular oxidative stress, both considered strategic targets for the prevention and treatment of neurodegenerative diseases. Compared to LD and GSH, GSH-LD had a stronger effect in preventing hydrogen peroxide (H2O2) induced apoptosis in both cell lines. Moreover, GSH-LD was able to preserve cell viability, cellular redox status, gluthation metabolism and prevent reactive oxygen species (ROS) formation, in a phosphinositide 3-kinase (PI3K)/kinase B (Akt)-dependent manner, in a neurotoxicity cellular model. Our findings indicate that the GSH-LD codrug offers advantages deriving from the additive effect of LD and GSH and it could represent a promising candidate for PD treatment.

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.

Aging-Related Oxidative Stress: Positive Effect of Memory Training.

The cognitive impairment characterizing the phenotype of older adults has been related to the efficiency of the antioxidant system. This study aimed at investigating the effect of memory training (MT) on memory, global cognitive functioning, and the oxidant and antioxidant capacity of plasma. We recruited 52 healthy subjects aged over 60. Twenty-nine subjects were submitted to 6-months of MT (Experimental Group, EG), and 23 were used as a Control Group (CG). Global cognitive functioning was assessed by the Mini-Mental State Examination (MMSE) and Short- and Long-Term Memory (STM and LTM, respectively) by the Rey Auditory Verbal Learning Test (RAVLT) at baseline (T0) and after 6-months (T1). Meanwhile, Reactive Oxygen Metabolites derivative compounds (d-ROMs), Biological Antioxidant Potential (BAP), and their ratio were evaluated on plasma. Results showed that the MMSE and RAVLT scores improved in EG at T1. At the same time, the d-ROMs levels significantly decreased, while the BAP and BAP/d-ROMs ratio showed an opposite trend. In both groups, the MMSE and LTM scores were negatively associated with d-ROMs levels, and positively correlated with BAP levels and the BAP/d-ROMs ratio. When we considered the Δvalue (Δvariable = variable post-MT minus variable pre-MT) in EG, the ΔMMSE and ΔLTM scores were negatively associated to Δd-ROMs, and positively to ΔBAP and ΔBAP/dROM. In conclusion, our results suggest that MT improves memory and global cognitive functioning. These processes were significantly associated to increase in resistance against oxidative stress at the plasma level in healthy older adults.

Memory Training Program Decreases the Circulating Level of Cortisol and Pro-inflammatory Cytokines in Healthy Older Adults.

Aging cognitive decline has been associated to impairment of the Hypothalamus Pituitary Adrenals (HPA) axis activity and a higher level of the systemic inflammation. However, little is known about the molecules driving this process at peripheral level. In addition, the cognitive function is to some extent modifiable with Memory Training (MT) programs, even among older adults and beyond. The study aims to evaluate whether MT could contribute to ameliorate cognitive performance and modulate the HPA axis activity as well the low level inflammation in the aging phenotype. Whether the phosphatase WIP-1, a negative regulator for inflammation, is involved in this process was also investigated. We recruited 31 young adults (19-28, years of age) and 62 older adults aged over 60. Thirty-two older adults were submitted to 6-months of MT program (EG), and 28 older adults were no treated and used as Control Group (CG). Global cognitive functioning (MMSE score), verbal and visual memory, and attention were assessed at baseline (T0) and after 6-months (T1). At the same time, plasmatic level of Cortisol (C), IL-1ß, IL-18, IL-6, and the expression of WIP-1 mRNA and protein in ex vivo Peripheral Blood Mononuclear Cells were analyzed in young adults at T0, as well in older adults at T0 and T1. Together, the results suggest that MT improves the global cognitive functionality, verbal and visual memory, as well as the level of attention. At the same time we observed a decrease of the plasmatic level of C, of the cytokines, and an increase of the expression of mRNA and protein of WIP-1. The analysis of correlations highlighted that the level of the mRNA of WIP-1 was positively associated to the MMSE score, and negatively to the C and cytokine levels. In conclusion, we purpose the MT as tool that could help support successful aging through the improving of memory, attention and global cognitive function performance. Furthermore, this approach could participate to maintain lower the peripheral levels of the C and pro-inflammatory cytokines. The WIP-1 as a potential new target of the pathophysiology of aging is theorized.

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.

The NF-kB regulates the SHP-1 expression in monocytes in congestive heart failure.

It has been shown that functional recovery of patients with acute congestive heart failure (ACHF) after treatment with conventional drugs (CD) is mediated by suppression of inflammation in peripheral blood mononuclear cells. Here, we analyzed gene expression profiles of monocytes from symptomatic ACHF patients (NYHA Class III-IV) before and after pharmacological treatment with CD. The treatment was associated with selective down-regulation of "TNFR signaling" and pro-inflammatory mediators CCL5, MIP-1α receptor, CD14, ITGAM, and significant up-regulation of "TNFR signaling" as evidenced by increase in anti-inflammatory factors including NF-kBIA, TNFAIP3 and SHP-1. In monocyte TNF-alpha-stimulated there is a down-regulation of the phosphatase SHP-1 which induces a significant activation of TAK-1/IKK/NF-kB signaling. These findings suggest that the therapeutic impact of CD treatment in symptomatic ACHF includes negative regulation of the NF-kB signaling in monocytes and the improvement of the SHP-1 activity.

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.

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.

Development of glycine-α-methyl-proline-containing tripeptides with neuroprotective properties.

Herein is described the synthesis of novel glycine-α-methyl-proline-containing tripeptides (GP(Me)X tripeptides namely GP(Me)R, GP(Me)K, and GP(Me)H) with the aim of obtaining derivatives highly stable in human plasma and able to counteract neuroinflammatory processes that are distinctive of neurodegenerative pathologies. The syntheses of GP(Me)R, GP(Me)K, and GP(Me)H were all achieved both by introducing the Pro(Me) residue into the Gly-Pro-Arg (GPR) sequence in place of the native Pro in P2 position and replacing the basic amino acid Arg in P3 position by Lys or His. Results showed that all novel GP(Me)X tripeptides are stable in human plasma (t1/2 > 51 h) and that GP(Me)H - generating stable intramolecular H-bond in a C11-turn by interaction of His imidazole ring and Gly carbonyl group - restored physiological levels of nitric oxide deriving from neuronal NOS (nNOS) activity, thus preventing the inflammatory response by suppression of the NF-kB activity and, consequently, the expression of inflammatory genes such as inducibile NOS (iNOS). Therefore, GP(Me)H could be a lead compound for further development of peptidomimetics able to contrast neuroinflammatory processes.

mTOR Activation by PI3K/Akt and ERK Signaling in Short ELF-EMF Exposed Human Keratinocytes.

Several reports suggest that ELF-EMF exposures interact with biological processes including promotion of cell proliferation. However, the molecular mechanisms by which ELF-EMF controls cell growth are not completely understood. The present study aimed to investigate the effect of ELF-EMF on keratinocytes proliferation and molecular mechanisms involved. Effect of ELF-EMF (50 Hz, 1 mT) on HaCaT cell cycle and cells growth and viability was monitored by FACS analysis and BrdU assay. Gene expression profile by microarray and qRT-PCR validation was performed in HaCaT cells exposed or not to ELF-EMF. mTOR, Akt and MAPKs expressions were evaluated by Western blot analysis. In HaCaT cells, short ELF-EMF exposure modulates distinct patterns of gene expression involved in cell proliferation and in the cell cycle. mTOR activation resulted the main molecular target of ELF-EMF on HaCaT cells. Our data showed the increase of the canonical pathway of mTOR regulation (PI3K/Akt) and activation of ERK signaling pathways. Our results indicate that ELF-EMF selectively modulated the expression of multiple genes related to pivotal biological processes and functions that play a key role in physio-pathological mechanisms such as wound healing.

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.

Effect of erythropoietin on primed leucocyte expression profile.

Resistance to erythropoietin (EPO) affects a significant number of anaemic patients with end-stage renal disease. Previous reports suggest that inflammation is one of the major independent predictors of EPO resistance, and the effects of EPO treatment on inflammatory mediators are not well established. The aim of this study was to investigate EPO-induced modification to gene expression in primary cultured leucocytes. Microarray experiments were performed on primed ex vivo peripheral blood mononuclear cells (PBMCs) and treated with human EPO-α. Data suggested that EPO-α modulated genes involved in cell movement and interaction in primed PBMCs. Of note, EPO-α exerts anti-inflammatory effects inhibiting the expression of pro-inflammatory cytokine IL-8 and its receptor CXCR2; by contrast, EPO-α increases expression of genes relating to promotion of inflammation encoding for IL-1ß and CCL8, and induces de novo synthesis of IL-1α, CXCL1 and CXCL5 in primed cells. The reduction in MAPK p38-α activity is involved in modulating both IL-1ß and IL-8 expression. Unlike the induction of MAPK, Erk1/2 activity leads to upregulation of IL-1ß, but does not affect IL-8 expression and release. Furthermore, EPO-α treatment of primed cells induces the activation of caspase-1 upstream higher secretion of IL-1ß, and this process is not dependent on caspase-8 activation. In conclusion, our findings highlight new potential molecules involved in EPO resistance and confirm the anti-inflammatory role for EPO, but also suggest a plausible in vivo scenario in which the positive correlation found between EPO resistance and elevated levels of some pro-inflammatory mediators is due to treatment with EPO itself.

Astaxanthin treatment confers protection against oxidative stress in U937 cells stimulated with lipopolysaccharide reducing O2- production.

Recently, astaxanthin (ASTA) studies have focused on several biological functions such as radical scavenging, singlet oxygen quenching, anti-carcinogenesis, anti-diabetic, anti-obesity, anti-inflammatory, anti-melanogenesis, and immune enhancement activities. In this study, we investigated the potential role protective of ASTA, an antioxidant marine carotenoid, in restoring physiological conditions in U937 cells stimulated with LPS (10 µg/ml). Our results show that pre-treatment with ASTA (10 µM) for 1 h attenuates the LPS-induced toxicity and ROS production. The beneficial effect of ASTA is associated with a reduction intracellular O2 (-) production by restoring the antioxidant network activity of superoxide dismutase (SOD) and catalase (CAT), which influence HO-1 expression and activity by inhibiting nuclear translocation of Nrf2. We accordingly hypothesize that ASTA has therapeutic properties protecting U937 cells from LPS-induced inflammatory and oxidative stress.

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.