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Obesity is frequently accompanied by non-alcoholic fatty liver disease (NAFLD). These two diseases are associated with altered lipid metabolism, in which reverse cholesterol transport (LXRα/ABCA1/ABCG1) and leptin response (leptin receptor (Ob-Rb)/Sam68) are involved. The two pathways were evaluated in peripheral blood mononuclear cells (PBMCs) from 86 patients with morbid obesity (MO) before and six months after Roux-en-Y gastric bypass (RYGB) and 38 non-obese subjects. In the LXRα pathway, LXRα, ABCA1, and ABCG1 mRNA expressions were decreased in MO compared to non-obese subjects (p < 0.001, respectively). Ob-Rb was decreased (p < 0.001), whereas Sam68 was increased (p < 0.001) in MO. RYGB did not change mRNA gene expressions. In the MO group, the LXRα pathway (LXRα/ABCA1/ABCG1) negatively correlated with obesity-related variables (weight, body mass index, and hip), inflammation (C-reactive protein), and liver function (alanine-aminotransferase, alkaline phosphatase, and fatty liver index), and positively with serum albumin. In the Ob-R pathway, Ob-Rb and Sam68 negatively correlated with alanine-aminotransferase and positively with albumin. The alteration of LXRα and Ob-R pathways may play an important role in NAFLD development in MO. It is possible that MO patients may require more than 6 months following RYBGB to normalize gene expression related to reverse cholesterol transport or leptin responsiveness.
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Transportador 1 de Casete de Unión a ATP , Colesterol , Leucocitos Mononucleares , Receptores X del Hígado , Hígado , Obesidad Mórbida , Receptores de Leptina , Humanos , Obesidad Mórbida/metabolismo , Obesidad Mórbida/cirugía , Obesidad Mórbida/genética , Masculino , Leucocitos Mononucleares/metabolismo , Femenino , Receptores de Leptina/genética , Receptores de Leptina/metabolismo , Adulto , Colesterol/metabolismo , Receptores X del Hígado/metabolismo , Receptores X del Hígado/genética , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Persona de Mediana Edad , Hígado/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/genética , Transducción de Señal , Transporte Biológico , Regulación de la Expresión Génica , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genéticaRESUMEN
The present work investigates the modulation of grapefruit flavonoid naringenin over liver X receptor alpha (LXRα) and its target genes in THP-1 macrophages, focusing on AMP-activated protein kinase (AMPK) implication. Naringenin induced LXRα at mRNA and protein levels besides influencing the expression of LXRα target genes ABCA1, ABCG1 (ATP-binding cassette A1 and G1), and SREBP1c (sterol response element binding protein 1c) in THP-1 macrophages. The increased LXRα mRNA and protein expression was reverted when AMPK was inhibited by its chemical inhibitor, compound C or by transfection with AMPK α1 and α2 siRNA. Naringenin treatments were also able to promote reverse cholesterol transport in THP-1 cells, which is in line with the increase in the ABCA1 and ABCG1 expression found. Treatments with this flavonoid also inhibited cell migration in THP-1 cells. In conclusion, LXRα and its target genes are up-regulated by naringenin in an AMPK dependent manner in human macrophages. The enhancement in the expression of genes involved in cholesterol efflux may reveal a new mechanism by which this polyphenol can prevent atherosclerosis and foam cell progression.
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Proteínas Quinasas Activadas por AMP/metabolismo , Citrus paradisi/química , Flavanonas/farmacología , Flavonoides/farmacología , Receptores X del Hígado/metabolismo , Macrófagos/efectos de los fármacos , Transportador 1 de Casete de Unión a ATP/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/metabolismo , Aterosclerosis/tratamiento farmacológico , Aterosclerosis/metabolismo , Transporte Biológico/efectos de los fármacos , Línea Celular , Movimiento Celular/efectos de los fármacos , Colesterol/metabolismo , Células Espumosas/efectos de los fármacos , Células Espumosas/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Macrófagos/metabolismo , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Transducción de Señal/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacosRESUMEN
PURPOSE: Oxysterols are cholesterol-oxygenated derivatives generated in the organism and also present in foods because of cholesterol oxidation during processing and storage. They are the natural ligands of liver X receptors (LXRs) and are generally recognized as hypocholesterolemic and anti-inflammatory molecules although this latter property is still controversial. Most oxysterol studies have been performed in macrophages, whereas the effects of oxysterols in neutrophils are poorly known. In this study, human neutrophils were exposed to two different oxysterols, 7-keto-cholesterol (7-k-chol) and 25-hydroxy-cholesterol (25-OH-chol), and their possible participation in inflammatory process was evaluated. METHODS: Human neutrophils were incubated with 7-k-chol and 25-OH-chol, and ROS production, translocation of the NADPH oxidase cytosolic components, hemoxygenase-1 (HO-1) expression and lysozyme secretion were analyzed. RESULTS: An increase in ROS production was observed within a short period of time (minutes) with both molecules. These oxysterols also stimulated the cellular membrane translocation of the NADPH oxidase cytosolic components, p47phox and p67phox. On the other hand, HO-1 expression, a cytoprotector enzyme, is inhibited in human neutrophils upon oxysterols treatment. Moreover, both oxysterols were associated with high lysozyme enzyme secretion at 5 and 18 h of incubation. CONCLUSIONS: The present paper describes for the first time that two oxysterols (7-k-chol and 25-OH-chol) enhance the ROS production within a short period of time in human neutrophils, stimulate the translocation of the cytosolic components of NADPH oxidase to the cellular membrane and increase lysozyme secretion. These data suggest that both oxysterols are able to activate pro-inflammatory effects in human neutrophils which contrasts with the role assigned to the oxysterols when they act through LXR at long time of incubation.
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Hidroxicolesteroles/farmacología , Cetocolesteroles/farmacología , Neutrófilos/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Membrana Celular/metabolismo , Hemo-Oxigenasa 1/genética , Hemo-Oxigenasa 1/metabolismo , Humanos , Muramidasa/metabolismo , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Neutrófilos/citología , Fosfoproteínas/genética , Fosfoproteínas/metabolismoRESUMEN
PURPOSE: Regulation of liver X receptors (LXRs) is essential for cholesterol homeostasis and inflammation. The present study was conducted to determine whether oleic acid (OA) could regulate mRNA expression of LXRα and LXRα-regulated genes and to assess the potential promotion of oxidative stress by OA in neutrophils. METHODS: Human neutrophils were treated with OA at different doses and LXR target gene expression, oxidative stress production, lipid efflux and inflammation state were analyzed. RESULTS: We describe that mRNA synthesis of both LXRα and ABCA1 (a reverse cholesterol transporter) was induced by OA in human neutrophils. This fatty acid enhanced the effects of LXR ligands on ABCA1 and LXR expression, but it decreased the mRNA levels of sterol regulatory element-binding protein 1c (a transcription factor that regulates the synthesis of triglycerides). Although OA elicited a slight oxidative stress in the short term (15-30 min) in neutrophils, it is unlikely that this is relevant for the modulation of transcription in our experimental conditions, which involve longer incubation time (i.e., 6 h). Of physiological importance is our finding that OA depresses intracellular lipid levels and that markers of inflammation, such as ERK1/2 and p38 mitogen-activated protein kinase phosphorylation, were decreased by OA treatment. In addition, 200 µM OA reduced the migration of human neutrophils, another marker of the inflammatory state. However, OA did not affect lipid peroxidation induced by pro-oxidant agents. CONCLUSIONS: This work presents for the first time evidence that human neutrophils are highly sensitive to OA and provides novel data in support of a protective role of this monounsaturated acid against the activation of neutrophils during inflammation.
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Transportador 1 de Casete de Unión a ATP/genética , Neutrófilos/efectos de los fármacos , Ácido Oléico/farmacología , Receptores Nucleares Huérfanos/genética , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Receptores X del Hígado , Neutrófilos/metabolismo , Receptores Nucleares Huérfanos/metabolismo , Estrés Oxidativo/efectos de los fármacos , Fosforilación , ARN Mensajero/genética , ARN Mensajero/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Factores de Transcripción/metabolismo , Triglicéridos/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
KEY POINTS: T-cell activation in patients with chronic rhinosinusitis with nasal polyps (CRSwNP) is enriched by late cytotoxic T cells. The proportion of early and intermediate activated cytotoxic T cells decreases in nasal polyps of patients with CRSwNP. Our results identify late activated cytotoxic T cells as potential biomarkers or therapeutic targets for patients with CRSwNP.
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Inmunofenotipificación , Activación de Linfocitos , Pólipos Nasales , Rinitis , Sinusitis , Humanos , Pólipos Nasales/inmunología , Sinusitis/inmunología , Rinitis/inmunología , Enfermedad Crónica , Activación de Linfocitos/inmunología , Masculino , Adulto , Persona de Mediana Edad , Femenino , Linfocitos T Citotóxicos/inmunología , Anciano , RinosinusitisRESUMEN
Dendritic cells (DCs) serve as professional antigen-presenting cells (APC) bridging innate and adaptive immunity, playing an essential role in triggering specific cellular and humoral responses against tumor and infectious antigens. Consequently, various DC-based antitumor therapeutic strategies have been developed, particularly vaccines, and have been intensively investigated specifically in the context of acute myeloid leukemia (AML). This hematological malignancy mainly affects the elderly population (those aged over 65), which usually presents a high rate of therapeutic failure and an unfavorable prognosis. In this review, we examine the current state of development and progress of vaccines in AML. The findings evidence the possible administration of DC-based vaccines as an adjuvant treatment in AML following initial therapy. Furthermore, the therapy demonstrates promising outcomes in preventing or delaying tumor relapse and exhibits synergistic effects when combined with other treatments during relapses or disease progression. On the other hand, the remarkable success observed with RNA vaccines for COVID-19, delivered in lipid nanoparticles, has revealed the efficacy and effectiveness of these types of vectors, prompting further exploration and their potential application in AML, as well as other neoplasms, loading them with tumor RNA.
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PURPOSE: Arterial hypertension is associated with a high production of reactive oxygen species and a decrease in the antioxidant defense systems. Based on the lack of toxicity of L-carnitine (LC) and previous studies reporting beneficial effects of this compound in experimental models of hypertension, the aim of this work was to test the hypothesis that LC might protect the kidney against hypertension-induced oxidative damage, as well as to investigate the mechanisms involved in this effect. To this end, specific activities and protein/mRNA expression of the antioxidant enzymes (glutathione peroxidase, glutathione reductase, and superoxide dismutase), and those of NADPH oxidase (the main responsible for superoxide anion production in renal tissue) have been measured in renal cortex homogenates from NG-nitro-L-arginine methyl ester (L-NAME)-treated rats and control normotensive rats. In addition, components of the renin-angiotensin system (RAS) and redox-sensitive transcription factors (NF-κB, Nrf2, and PPARα) have also been evaluated. METHODS: Male Wistar rats aged 6-8 weeks were divided into four groups of six animals each: (1) control, normotensive Wistar rats (with free access to tap water); (2) Wistar rats subjected to treatment with 25 mg of L-NAME/kg body weight/day dissolved in the drinking water, in order to develop L-NAME-induced hypertension; (3) Wistar rats subjected to treatment with 400 mg of LC/kg body weight/day (also dissolved in the drinking water); and (4) L-NAME-treated rats subjected to simultaneous treatment with LC at the indicated doses. RESULTS: The beneficial effect of LC supplementation on oxidative damage in the renal cortex of hypertensive rats reversed hypertension-associated renal function damage and produced an upregulation of both antioxidant enzymes and eNOS, and with a downregulation of both NADPH oxidase and RAS components. LC improves the oxidative stress response through a specific modulation of NF-κB, Nrf2, and PPARα transcription factors. Thus, the low production of superoxide anions, subsequent to NADPH oxidase inhibition, might act by increasing the expression of Nrf2 and PPARα and by decreasing that of NF-κB, which, in turn, would enhance the antioxidant defense systems. CONCLUSIONS: Our results might support the use of LC to prevent hypertension-induced renal damage.
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Carnitina/farmacología , Regulación de la Expresión Génica , Hipertensión/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Animales , Antioxidantes/farmacología , Regulación hacia Abajo , Glutatión Peroxidasa/metabolismo , Glutatión Reductasa/metabolismo , Riñón/efectos de los fármacos , Riñón/patología , Peroxidación de Lípido/efectos de los fármacos , Masculino , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , FN-kappa B/metabolismo , NG-Nitroarginina Metil Éster/efectos adversos , Óxido Nítrico Sintasa de Tipo III/genética , Óxido Nítrico Sintasa de Tipo III/metabolismo , PPAR alfa/genética , PPAR alfa/metabolismo , Ratas , Ratas Wistar , Sistema Renina-Angiotensina , Superóxido Dismutasa/metabolismo , Superóxidos/metabolismo , Regulación hacia ArribaRESUMEN
Nutraceuticals act as cellular and functional modulators, contributing to the homeostasis of physiological processes. In an inflammatory microenvironment, these functional foods can interact with the immune system by modulating or balancing the exacerbated proinflammatory response. In this process, immune cells, such as antigen-presenting cells (APCs), identify danger signals and, after interacting with T lymphocytes, induce a specific effector response. Moreover, this conditions their change of state with phenotypical and functional modifications from the resting state to the activated and effector state, supposing an increase in their energy requirements that affect their intracellular metabolism, with each immune cell showing a unique metabolic signature. Thus, nutraceuticals, such as polyphenols, vitamins, fatty acids, and sulforaphane, represent an active option to use therapeutically for health or the prevention of different pathologies, including obesity, metabolic syndrome, and diabetes. To regulate the inflammation associated with these pathologies, intervention in metabolic pathways through the modulation of metabolic energy with nutraceuticals is an attractive strategy that allows inducing important changes in cellular properties. Thus, we provide an overview of the link between metabolism, immune function, and nutraceuticals in chronic inflammatory processes associated with obesity and diabetes, paying particular attention to nutritional effects on APC and T cell immunometabolism, as well as the mechanisms required in the change in energetic pathways involved after their activation.
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Células Presentadoras de Antígenos , Linfocitos T , Humanos , Células Presentadoras de Antígenos/metabolismo , Macrófagos/metabolismo , Inflamación/metabolismo , Suplementos Dietéticos , Obesidad/metabolismoRESUMEN
Dendritic cells (DCs) are antigen presenting cells that link innate and adaptive immunity. DCs have been historically considered as the most effective and potent cell population to capture, process and present antigens to activate naïve T cells and originate favorable immune responses in many diseases, such as cancer. However, in the last decades, it has been observed that DCs not only promote beneficial responses, but also drive the initiation and progression of some pathologies, including inflammatory bowel disease (IBD). In line with those notions, different therapeutic approaches have been tested to enhance or impair the concentration and role of the different DC subsets. The blockade of inhibitory pathways to promote DCs or DC-based vaccines have been successfully assessed in cancer, whereas the targeting of DCs to inhibit their functionality has proved to be favorable in IBD. In this review, we (a) described the general role of DCs, (b) explained the DC subsets and their role in immunogenicity, (c) analyzed the role of DCs in cancer and therapeutic approaches to promote immunogenic DCs and (d) analyzed the role of DCs in IBD and therapeutic approaches to reduced DC-induced inflammation. Therefore, we aimed to highlight the "yin-yang" role of DCs to improve the understand of this type of cells in disease progression.
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Enfermedades Inflamatorias del Intestino , Neoplasias , Humanos , Células Dendríticas , Inmunidad Adaptativa , Neoplasias/metabolismo , Progresión de la EnfermedadRESUMEN
In 2010, the Mediterranean diet was recognized by UNESCO as an Intangible Cultural Heritage of Humanity. Olive oil is the most characteristic food of this diet due to its high nutraceutical value. The positive effects of olive oil have often been attributed to its minor components; however, its oleic acid (OA) content (70-80%) is responsible for its many health properties. OA is an effective biomolecule, although the mechanism by which OA mediates beneficial physiological effects is not fully understood. OA influences cell membrane fluidity, receptors, intracellular signaling pathways, and gene expression. OA may directly regulate both the synthesis and activities of antioxidant enzymes. The anti-inflammatory effect may be related to the inhibition of proinflammatory cytokines and the activation of anti-inflammatory ones. The best-characterized mechanism highlights OA as a natural activator of sirtuin 1 (SIRT1). Oleoylethanolamide (OEA), derived from OA, is an endogenous ligand of the peroxisome proliferator-activated receptor alpha (PPARα) nuclear receptor. OEA regulates dietary fat intake and energy homeostasis and has therefore been suggested to be a potential therapeutic agent for the treatment of obesity. OEA has anti-inflammatory and antioxidant effects. The beneficial effects of olive oil may be related to the actions of OEA. New evidence suggests that oleic acid may influence epigenetic mechanisms, opening a new avenue in the exploration of therapies based on these mechanisms. OA can exert beneficial anti-inflammatory effects by regulating microRNA expression. In this review, we examine the cellular reactions and intracellular processes triggered by OA in T cells, macrophages, and neutrophils in order to better understand the immune modulation exerted by OA.
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Dieta Mediterránea , Ácido Oléico , Ácido Oléico/farmacología , Ácido Oléico/uso terapéutico , Aceite de Oliva/farmacología , Ácidos Oléicos/farmacología , Antiinflamatorios/farmacologíaRESUMEN
The defense mechanism against harmful stimuli is inflammation. Indeed, neurodegenerative disorders can arise as a result of a persistent neuroinflammation. Beta-amyloid (Aß1-42) is an early trigger in the origination of Alzheimer's disease, leading to synaptic and cognitive impairments. Virgin olive oil (VOO) is correlated with a decreased risk of developing immune-inflammatory disorders, but the potential effects of the phenolic fraction (PF) from VOO in the modulation of neuroinflammatory processes in neutrophils remain unknown. In this study, we investigated the ability of the PF to modulate the activation of Aß1-42-stimulated primary human neutrophils, focusing on the expression of gene and surface markers and the release of pro-inflammatory and chemoattractant mediators. Down-regulation of pro-inflammatory cytokine gene expression in Aß1-42-treated neutrophils, among other changes, was reported. Furthermore, pretreatment with PF prevented neutrophil activation. The beneficial effects in the modulation of inflammatory responses show the relevance of VOO to achieve a healthier diet that can help prevent inflammatory diseases.
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Neutrófilos , Fenoles , Humanos , Aceite de Oliva/farmacología , Fenoles/farmacología , Péptidos beta-Amiloides , DietaRESUMEN
BACKGROUND: Sulforaphane (SFN) is an isothiocyanate of vegetable origin with potent antioxidant and immunomodulatory properties. The characterization of its pleiotropic activity in human dendritic cells (DCs) is poorly summarized. The aim of this work was to study the immunomodulatory power of SFN in response to an inflammatory microenvironment on human monocyte-derived DCs (moDCs). METHODS: We studied the immunological response induced by SFN. Apoptosis and autophagy assays were performed using flow cytometry on moDCs and a cancer cell line (THP-1). These included moDC maturation, lymphocyte proliferation and cytokine production under different experimental conditions. We investigated whether these results were associated with an inflammatory microenvironment induced by lipopolysaccharides (LPSs). RESULTS: Our results demonstrated that SFN could interact with moDCs, significantly reducing the autophagy process and enhancing apoptosis similarly to cancer cell line THP-1 cells in a chronic inflammatory microenvironment. Under chronic inflammation, SFN modulated the phenotypical characteristics of moDCs, reducing the expression of all markers (CD80, CD83, CD86, HLA-DR and PD-L1). SFN significantly reduced the Th2 proliferative response, with a decrease in the IL-9 and IL-13 levels. Although we did not observe any changes in the regulatory proliferative response, we noted an increase in the IL-10 levels. CONCLUSIONS: These findings demonstrate that SFN exerts protective effects against LPS-induced inflammation via the modulation of moDCs/T cells towards a regulatory profile. SFN may be a potential candidate for the treatment of pathologies with an inflammatory profile.
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Citocinas , Isotiocianatos , Humanos , Citocinas/metabolismo , Isotiocianatos/farmacología , Isotiocianatos/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Células Dendríticas , Inmunidad , Monocitos/metabolismo , Diferenciación Celular , Células CultivadasRESUMEN
Dietary fatty acids have been demonstrated to modulate systemic inflammation and induce the postprandial inflammatory response of circulating immune cells. We hypothesized that postprandial triglyceride-rich lipoproteins (TRLs) may have acute effects on immunometabolic homeostasis by modulating dendritic cells (DCs), sentinels of the immunity that link innate and adaptive immune systems. In healthy volunteers, saturated fatty acid (SFA)-enriched meal raised serum levels of granulocyte/macrophage colony-stimulating factor GM-CSF (SFAs > monounsaturated fatty acids (MUFAs) = polyunsaturated fatty acids (PUFAs)) in the postprandial period. Autologous TRL-SFAs upregulated the gene expression of DC maturation (CD123 and CCR7) and DC pro-inflammatory activation (CD80 and CD86) genes while downregulating tolerogenic genes (PD-L1 and PD-L2) in human monocyte-derived DCs (moDCs). These effects were reversed with oleic acid-enriched TRLs. Moreover, postprandial SFAs raised IL-12p70 levels, while TRL-MUFAs and TRL-PUFAs increased IL-10 levels in serum of healthy volunteers and in the medium of TRL-treated moDCs. In conclusion, postprandial TRLs are metabolic entities with DC-related tolerogenic activity, and this function is linked to the type of dietary fat in the meal. This study shows that the intake of meals enriched in MUFAs from olive oil, when compared with meals enriched in SFAs, prevents the postprandial production and priming of circulating pro-inflammatory DCs, and promotes tolerogenic response in healthy subjects. However, functional assays with moDCs generated in the presence of different fatty acids and T cells could increase the knowledge of postprandial TRLs' effects on DC differentiation and function.
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Diferenciación Celular , Células Dendríticas/inmunología , Grasas de la Dieta/administración & dosificación , Grasas de la Dieta/efectos adversos , Ácidos Grasos/administración & dosificación , Ácidos Grasos/efectos adversos , Lipoproteínas/metabolismo , Monocitos , Periodo Posprandial/inmunología , Triglicéridos/metabolismo , Adulto , Antígeno B7-1/genética , Antígeno B7-1/metabolismo , Células Cultivadas , Células Dendríticas/metabolismo , Células Dendríticas/fisiología , Femenino , Expresión Génica , Factor Estimulante de Colonias de Granulocitos y Macrófagos/sangre , Homeostasis/efectos de los fármacos , Humanos , Subunidad alfa del Receptor de Interleucina-3/genética , Subunidad alfa del Receptor de Interleucina-3/metabolismo , Masculino , Comidas , Aceite de OlivaRESUMEN
Pluripotent stem cells maintain the property of self-renewal and differentiate into all cell types under clear environments. Though the gene regulatory mechanism for pluripotency has been investigated in recent years, it is still not completely understood. Here, we show several signaling pathways involved in the maintenance of pluripotency. To investigate whether AMPK is involved in maintaining the pluripotency in mouse embryonic stem cells (mESCs) and elucidating the possible molecular mechanisms, implicated D3 and R1/E mESC lines were used in this study. Cells were cultured in the absence or presence of LIF and treated with 1 mM and 0.5 mM 5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside (AICAR), 2 mM metformin, compound C, and the PI3K inhibitor LY294002 for 24, 72, and 120 h. The levels of Nanog, Oct3/4, and REX1 and Brachyury, Notch2, and Gata4 mRNAs and Nanog or OCT3/4 protein levels were analyzed. Alkaline phosphatase and the cellular cycle were determined. The pGSK3ß, GSK3ß, p-ß-catenin, and ß-catenin protein levels were also investigated. We found that AMPK activators such as AICAR and metformin increase mRNA expression of pluripotency markers and decrease mRNA expression of differentiation markers in R1/E and D3 ES cells. AICAR increases phosphatase activity and arrests the cellular cycle in the G1 phase in these cells. We describe that AICAR effects were mediated by AMPK activation using a chemical inhibitor or by silencing this gene. AICAR effects were also mediated by PI3K, GSK3ß, and ß-catenin in R1/E ES cells. According to our findings, we provide a mechanism by which AICAR increases and maintains a pluripotency state through enhanced Nanog expression, involving AMPK/PI3K and p-GSK3ß Ser21/9 pathways backing up the AICAR function as a potential target for this drug controlling pluripotency. The highlights of this study are that AICAR (5-aminoimidazole-4-carboxamied-1-b-riboside), an AMP protein kinase (AMPK) activator, blocks the ESC differentiation and AMPK is a key enzyme for pluripotency and shows valuable data to clarify the molecular pluripotency mechanism.
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Liver X receptor alpha (LXRα) is a nuclear receptor involved in cholesterol homeostasis. Curcumin, a traditional Chinese derivative from the rhizomes of Curcuma longa and a well-known AMP-activated protein kinase (AMPK) activator, possess hypocholesterolemic activity, however, the possible link between AMPK and cholesterol is unknown. In this study, we have investigated whether curcumin regulates metabolic changes in cholesterol metabolism via LXRα in THP-1 human macrophages, the cells implicated in atheroma plaques formation. Results showed that curcumin induced AMPK phosphorylation, increased LXRα mRNA and protein expression. Curcumin up-regulated mRNA expression of genes involved in cholesterol transport and metabolism as ATP-binding cassette (ABC) transporters ABCA1 and ABCG1, and the sterol response element binding protein 1c (SREBP1c). On the other hand, this increased LXRα mRNA and protein expression was reverted when AMPK was inhibited by its chemical inhibitor, compound C. Transfection with AMPK α1 and α2 siRNA decreased the LXRα mRNA expression and its target genes. Curcumin treatment inhibited cell migration and was also able to promote reverse cholesterol transport in THP-1 cells. This enhanced reverse cholesterol transport might be related to the up-regulating of ABCA1 and ABCG1 mRNA expression by activating AMPK-LXRα signaling in THP-1 cells. This study describes a possible mechanism for understanding the hypocholesterolemic effects of curcumin and expand knowledge about the LXRα regulation by AMPK.
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Proteínas Quinasas Activadas por AMP/metabolismo , Curcumina/farmacología , Receptores X del Hígado/genética , Macrófagos/efectos de los fármacos , Proteínas Quinasas Activadas por AMP/genética , Transporte Biológico/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Colesterol/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Hidrocarburos Fluorados/farmacología , Receptores X del Hígado/metabolismo , Macrófagos/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Sulfonamidas/farmacología , Células THP-1RESUMEN
The present study aimed to investigate whether l-carnitine (LC) protects the vascular endothelium and tissues against oxidative damage in hypertension. Antioxidant enzyme activities, glutathione and lipid peroxidation were measured in the liver and heart of spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. Nitrite and nitrate levels and total antioxidant status (TAS) were evaluated in plasma, and the expression of endothelial nitric oxide synthase (eNOS) and p22phox subunit of NAD(P)H oxidase was determined in aorta. Glutathione peroxidase activity was lower in SHR than in WKY rats, and LC increased this activity in SHR up to values close to those observed in normotensive animals. Glutathione reductase and catalase activities, which were higher in SHR, tended to increase after LC treatment. No differences were found in the activity of superoxide dismutase among any animal group. The ratio between reduced and oxidized glutathione and the levels of lipid peroxidation were respectively decreased and increased in hypertensive rats, and both parameters were normalized after the treatment. Similarly, LC was able to reverse the reduced plasma nitrite and nitrate levels and TAS observed in SHR. We found no alterations in the expression of aortic eNOS among any group; however, p22phox mRNA levels showed an increase in SHR that was reversed by LC. In conclusion, chronic administration of LC leads to an increase in hepatic and cardiac antioxidant defense and a reduction in the systemic oxidative process in SHR. Therefore, LC might increase NO availability in SHR aorta by a reduction in superoxide anion production.
Asunto(s)
Carnitina/farmacología , Hipertensión/fisiopatología , Estrés Oxidativo/efectos de los fármacos , Animales , Presión Sanguínea/efectos de los fármacos , Glutatión/sangre , Disulfuro de Glutatión/sangre , Glutatión Reductasa/metabolismo , Masculino , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Superóxido Dismutasa/metabolismoRESUMEN
Oxidative stress plays an important role in arterial hypertension and propionyl-L-carnitine (PLC) has been found to protect cells from toxic reactive oxygen species. In this work, we have evaluated the antioxidant capacity of chronic PLC treatment in spontaneously hypertensive rats (SHR) by measuring the activity of antioxidant enzymes and the lipid peroxidation in liver and cardiac tissues. The activity of glutathione peroxidase was decreased in liver and cardiac tissues of SHR when compared with their normotensive controls, Wistar- Kyoto (WKY) rats, this alteration being prevented by PLC treatment. Glutathione reductase activity was increased in hypertensive rats and no effect was observed after the treatment. No significant changes in superoxide dismutase activity were observed among all experimental groups. Liver of hypertensive rats showed higher catalase activity than that of normotensive rats, and PLC enhanced this activity in both rat strains. Thiobarbituric acid reactive substances, determined as a measure of lipid peroxidation, were increased in SHR compared with WKY rats, and PLC treatment decreased these values not only in hypertensive rats but also in normotensive ones. The content of carnitine in serum, liver and heart was higher in PLC-treated rats, but PLC did not prevent the hypertension development in young SHR. In addition, triglyceride levels, which were lower in SHR than WKY rats, were reduced by chronic PLC treatment in both rat strains. These results demonstrate: i) the hypotriglyceridemic effect of PLC and ii) the antioxidant capacity of PLC in SHR and its beneficial use protecting tissues from hypertension-accompanying oxidative damage.
Asunto(s)
Antioxidantes/farmacología , Carnitina/análogos & derivados , Ventrículos Cardíacos/efectos de los fármacos , Hipertensión/metabolismo , Hígado/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Animales , Antioxidantes/metabolismo , Antioxidantes/uso terapéutico , Presión Sanguínea/efectos de los fármacos , Carnitina/metabolismo , Carnitina/farmacología , Carnitina/uso terapéutico , Catalasa/metabolismo , Enzimas , Glutatión Peroxidasa/metabolismo , Glutatión Reductasa/metabolismo , Ventrículos Cardíacos/enzimología , Hipertensión/tratamiento farmacológico , Hipertensión/fisiopatología , Peroxidación de Lípido/efectos de los fármacos , Hígado/enzimología , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Triglicéridos/metabolismoRESUMEN
Liver X receptors (LXRs) are ligand-activated nuclear receptors involved mainly in the regulation of cholesterol metabolism in many organs, including liver and intestine, as well as in macrophages and neutrophils. Besides, both anti-inflammatory and pro-inflammatory properties have been ascribed to LXRs. The effect of the inflammatory condition on the expression of LXRα and its target genes has not been previously addressed in human neutrophils. We have described that platelet-activating factor (PAF) and hydrogen peroxide (H2O2) are potent pro-inflammatory mediators that link the haemostatic and innate immune systems. In this work we report that H2O2 at low doses (1 pM-1µM) exerts an inhibitory effect on TO901317-induced mRNA expression of LXRα and of its target genes encoding the ATP-binding cassette (ABC) transporters ABCA1 and ABCG1, and the sterol regulatory element-binding protein 1c (SREBP1c). However, an opposite behaviour, i.e., a transcription-enhancing effect, was found at higher H2O2 doses (100-500µM) on most of these genes. A similar dual effect was observed when the pro-inflammatory molecule PAF was used. Interestingly, H2O2 production separately elicited by 10nM PAF or 1µM H2O2 was similarly low, and analogously, H2O2 production levels elicited by 5µM PAF or 100µM H2O2 were similarly high when they were compared. On the other hand, low doses of PAF or H2O2 induced phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK 1/2) and NF-κB activation, However, PAF or H2O2 at high doses did not produce changes in NF-κB activation levels. In summary, our results show that H2O2, either exogenous or PAF-induced, exerts a dual regulation on mRNA expression of LXRα and its target genes.
Asunto(s)
Proteínas Portadoras/farmacología , Peróxido de Hidrógeno/farmacología , Receptores X del Hígado/metabolismo , Neutrófilos/efectos de los fármacos , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Células Cultivadas , Proteínas de Unión al ADN , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Hidrocarburos Fluorados/farmacología , Inmunidad Innata , Metabolismo de los Lípidos , Receptores X del Hígado/genética , FN-kappa B/metabolismo , Neutrófilos/inmunología , Fosforilación/efectos de los fármacos , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/genética , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Sulfonamidas/farmacología , Activación Transcripcional/efectos de los fármacosRESUMEN
In the present study, we have examined the potential ability of 5'-AMP-activated protein kinase (AMPK) to modulate NADPH oxidase activity in human neutrophils. AMPK activated with either 5'-aminoimidazole-4-carboxamide ribonucleoside (AICAR) or with 5'-AMP significantly attenuated both phorbol 12-myristate 13-acetate (PMA) and formyl methionyl leucyl phenylalanine-stimulated superoxide anion O2- release by human neutrophils, consistently with a reduced translocation to the cell membrane and phosphorylation of a cytosolic component of NADPH oxidase, namely p47phox. AMPK was found to be present in human neutrophils and to become phosphorylated in response to either AICAR or other stimulators of its enzyme activity. Furthermore, AICAR also strongly reduced PMA-dependent H2O2 release, and induced the phosphorylation of c-jun N-terminal kinase 1 (p46), p38 mitogen-activated protein kinase and extracellular signal-regulated kinase. Present data demonstrate for the first time that the activation of AMPK, in states of low cellular energy charge (such as under high levels of 5'-AMP) or other signals, could be a factor contributing to reduce the host defense mechanisms.