RESUMO
Both estrogen and hydrogen sulfide (H2S) have been shown to inhibit the development of atherosclerosis. We previously reported that cystathionine γ-lyase knockout (CSE-KO) male mice develop atherosclerosis earlier than male wild-type (WT) mice. The present study investigated the interaction of CSE/H2S pathway and estrogen on the development of atherosclerosis in female mice. Plasma estrogen levels were significantly lower in female CSE-KO mice than in female WT mice. NaHS treatment had no effect on plasma estrogen levels in both WT and CSE-KO female mice. After CSE-KO and WT female mice were fed with atherogenic diet for 12 wk, plasma lipid levels were significantly increased and triglyceride levels decreased compared with those of control diet-fed mice. Atherogenic diet induced more atherosclerotic lesion, oxidative stress, intracellular adhesion molecule-1 (ICAM-1), and NF-κB in CSE-KO mice than in WT mice. Estrogen treatment of atherogenic diet-fed WT mice attenuated hypercholesterolemia, oxidative stress, ICAM-1 expression, and NF-κB in WT mice but not in atherogenic diet-fed CSE-KO mice. Furthermore, H2S production in both the liver and vascular tissues was enhanced by estrogen in WT mice but not in CSE-KO mice. It is concluded that the antiatherosclerotic effect of estrogen is mediated by CSE-generated H2S. This study provides new insights into the interaction of H2S and estrogen signaling pathways on the regulation of cardiovascular functions.NEW & NOTEWORTHY Female cystathionine γ-lyase (CSE)-knockout mice have significantly lower plasma estrogen levels and more severe early atherosclerotic lesion than female wild-type mice. H2S production in liver and vascular tissues is enhanced by estrogen via its stimulatory effect on CSE activity. The antiatherosclerotic effect of estrogen is mediated by CSE-generated H2S.
Assuntos
Aterosclerose/fisiopatologia , Cistationina gama-Liase , Estrogênios , Sulfeto de Hidrogênio , Transdução de Sinais , Animais , Aorta Torácica/patologia , Aterosclerose/genética , Aterosclerose/patologia , Dieta Aterogênica , Estrogênios/sangue , Feminino , Homocisteína/sangue , Proteínas I-kappa B/sangue , Molécula 1 de Adesão Intercelular/metabolismo , Lipídeos/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator de Transcrição RelA/metabolismoRESUMO
Head and neck squamous cell carcinoma (HNSCC) is the major health concern in Indian population. Despite of advanced treatment the mortality rate for this disease has not been improved very much. Current research focused on development of protein marker for the diagnosis and prognosis of HNSCC. The case control study was performed with 125 HNSCC patients and 104 control cases. The level of p50 and IκBα proteins in serum were evaluated at pre and post therapy by label free real time surface plasmon resonance (SPR) and western blot analysis. The serum p50 concentration were significantly (P < 0.0001) higher at the time of diagnosis i.e. pre therapy (Mean ± SD = 27.06 ± 4.88 ng/µl) as compared to controls (Mean ± SD = 16.96 ± 4.04 ng/µl) while it decline at post therapy (Mean ± SD = 21.01 ± 4.98 ng/µl). Similarly, the concentration of IκBα protein in serum were slightly higher at pre therapy (Mean ± SD = 8.33 ± 1.85 ng/µl) as compared to controls (Mean ± SD = 7.27 ± 1.84 ng/µl) and declined at post therapy (Mean ± SD = 7.09 ± 1.24 ng/µl). The level of p50 was also high at the early stage of the disease. The specificity and sensitivity of p50 proteins obtained from ROC analysis revealed the potentiality to be diagnostic protein marker for HNSCC for its accuracy in the study cohort.
Assuntos
Biomarcadores Tumorais/sangue , Carcinoma de Células Escamosas/sangue , Carcinoma de Células Escamosas/diagnóstico , Neoplasias de Cabeça e Pescoço/sangue , Neoplasias de Cabeça e Pescoço/diagnóstico , Subunidade p50 de NF-kappa B/sangue , Adulto , Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Estudos de Casos e Controles , Diagnóstico Precoce , Feminino , Expressão Gênica , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Proteínas I-kappa B/sangue , Proteínas I-kappa B/genética , Masculino , Pessoa de Meia-Idade , Inibidor de NF-kappaB alfa , Subunidade p50 de NF-kappa B/genética , Gradação de Tumores , Estadiamento de Neoplasias , Valor Preditivo dos Testes , Curva ROC , Carcinoma de Células Escamosas de Cabeça e PescoçoRESUMO
Histamine 4 receptor (H4R) is a novel target for the pharmacological modulation of histamine-mediated immune signals during inflammatory diseases. The purpose of this study was to assess the effects of the H4R agonist 4-methylhistamine dihydrochloride (4-MeH) and antagonist JNJ7777120 (JNJ) in the inflamed rat knee. Animals were fasted for 18h before a single dose of 4-MeH or JNJ (30mg/kg) was administered intraperitoneally (i.p.), both followed by intra-articular (i.a.) injection of LPS 2h later. Blood and synovial fluid were collected after a short incubation period and TNF-α, NF-κB, and IkB-α levels were measured via flow cytometry. Additionally, we assessed the effects of H4R engagement on the expression of IL-1ß, TNF-α, and NF-κB mRNAs and the protein levels of TNF-α, NF-κB, JAK-1, and STAT-3 in the inflamed knee tissue. These results revealed increased TNF-α and NF-κB expression and decreased IkB-α levels in both the LPS alone and 4-MeH treated groups in whole blood and synovial fluid. Further, IL-1ß, TNF-α, and NF-κB mRNA levels were significantly increased and western blot analysis confirmed increased expression of TNF-α, NF-κB, JAK-1, and STAT-3 in both LPS and 4-MeH treatment groups. Furthermore, these increases were completely inhibited in the inflamed knee tissue of the JNJ-treated group. Thus, the inhibition of inflammatory mediators and signaling pathways by the H4R antagonist JNJ suggests the anti-arthritic importance of this molecule.
Assuntos
Inflamação/patologia , Articulação do Joelho/patologia , NF-kappa B/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Histamínicos/genética , Fator de Necrose Tumoral alfa/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Ativação Enzimática , Feminino , Regulação da Expressão Gênica , Agonistas dos Receptores Histamínicos/farmacologia , Antagonistas dos Receptores Histamínicos/farmacologia , Proteínas I-kappa B/sangue , Indóis/farmacologia , Inflamação/diagnóstico , Inflamação/imunologia , Interleucina-1beta/genética , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Articulação do Joelho/imunologia , Lipopolissacarídeos , Metilistaminas/farmacologia , Inibidor de NF-kappaB alfa , NF-kappa B/sangue , NF-kappa B/genética , Piperazinas/farmacologia , RNA Mensageiro/biossíntese , Ratos , Ratos Wistar , Receptores Histamínicos H4 , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/genéticaRESUMO
The aim was to investigate the metabolic and anti-inflammatory effects of resveratrol alone and when combined with exercise training in skeletal muscle of aged human subjects. Healthy, physically inactive men (60-72 years old) were randomized to either 8 weeks of daily intake of 250 mg resveratrol or placebo or to 8 weeks of high-intensity exercise training with 250 mg resveratrol or placebo. Before and after the interventions, resting blood samples and muscle biopsies were obtained and a one-legged knee-extensor endurance exercise test was performed. Exercise training increased skeletal muscle peroxisome proliferator-activated receptor-γ co-activator-1α mRNA ~1.5-fold, cytochrome c protein ~1.3-fold, cytochrome c oxidase I protein ~1.5-fold, citrate synthase activity ~1.3-fold, 3-hydroxyacyl-CoA dehydrogenase activity ~1.3-fold, inhibitor of κB-α and inhibitor of κB-ß protein content ~1.3-fold and time to exhaustion in the one-legged knee-extensor endurance exercise test by â¼1.2-fold, with no significant additive or adverse effects of resveratrol on these parameters. Despite an overall ~25% reduction in total acetylation level in skeletal muscle with resveratrol, no exclusive resveratrol-mediated metabolic effects were observed on the investigated parameters. Notably, however, resveratrol blunted an exercise training-induced decrease (~20%) in protein carbonylation and decrease (~40%) in tumour necrosis factor α mRNA content in skeletal muscle. In conclusion, resveratrol did not elicit metabolic improvements in healthy aged subjects; in fact, resveratrol even impaired the observed exercise training-induced improvements in markers of oxidative stress and inflammation in skeletal muscle. Collectively, this highlights the metabolic efficacy of exercise training in aged subjects and does not support the contention that resveratrol is a potential exercise mimetic in healthy aged subjects.
Assuntos
Exercício Físico , Músculo Esquelético/efeitos dos fármacos , Estilbenos/farmacologia , Idoso , Citrato (si)-Sintase/sangue , Ciclo-Oxigenase 1/sangue , Método Duplo-Cego , Gliceraldeído-3-Fosfato Desidrogenases/sangue , Humanos , Proteínas I-kappa B/sangue , Masculino , Músculo Esquelético/crescimento & desenvolvimento , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Resveratrol , Fatores de Transcrição/sangue , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND: We investigated the correlation of proinflammatory transcript nuclear factor κB (NF-κB) and antioxidative gene transcript nuclear factor-erythroid 2-related factor 2 (Nrf2) expressions in peripheral blood mononuclear cells (PBMCs) with the tumor necrosis factor α (TNF-α) response after endotoxin stimulation and the clinical outcome of severely injured patients. METHODS: Thirty-two severe blunt trauma patients (injury severity score>16) with systemic inflammatory response syndrome were enrolled. Age- and sex-matched healthy persons were the controls. Patients' blood samples were obtained at 24 and 72 hours after injury. Peripheral blood mononuclear cells were isolated, and measurements for NF-κB p65 translocation, Nrf2 and phosphorylated inhibitory κB-α expressions, and TNF-α levels were assayed after endotoxin stimulation. RESULTS: In the trauma patients, TNF-α hyporesponse, depressed NF-κB p65 translocation, and phosphorylated inhibitory κB-α expression in PBMCs were found at 24 and 72 hours after injury; the Nrf2 expressions in PBMCs were not significantly different between patients and controls. The TNF-α levels had significant correlation with the NF-κB translocation and the trend of negative correlation with Nrf2 expression. Fifteen patients had critical injury (injury severity score≥25). Patients with critical injury had a lower NF-κB signal and a lower TNF-α response than did the counter group. Twelve patients developed organ failure; their Nrf2 expressions were significantly lower than those of patients without organ failure. CONCLUSIONS: The endotoxin hyporesponse associated with NF-κB and Nrf2 signal alternations in PBMCs of injured patients develops early after injury. The hyporesponse of PBMCs with a lower TNF-α level correlates with a lower NF-κB signal and is associated with critical injury, whereas a depressed Nrf2 expression in PBMCs is associated with later organ failure in trauma patients.
Assuntos
Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos/metabolismo , Fator 2 Relacionado a NF-E2/sangue , NF-kappa B/sangue , Síndrome de Resposta Inflamatória Sistêmica/sangue , Fator de Necrose Tumoral alfa/sangue , Ferimentos não Penetrantes/sangue , Adulto , Biomarcadores/sangue , Western Blotting , Estudos de Casos e Controles , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Proteínas I-kappa B/sangue , Escala de Gravidade do Ferimento , Masculino , Pessoa de Meia-Idade , Insuficiência de Múltiplos Órgãos/sangue , Insuficiência de Múltiplos Órgãos/etiologia , Prognóstico , Transdução de Sinais , Síndrome de Resposta Inflamatória Sistêmica/etiologia , Ferimentos não Penetrantes/complicaçõesRESUMO
SCOPE: Heating during the process of cooking alters the chemical properties of foods and may affect subsequent postprandial inflammation. We tested the effects of four meals rich in different oils subjected to heating on the postprandial inflammatory metabolism of peripheral blood mononuclear cells (PBMCs). METHODS AND RESULTS: Twenty obese participants received four breakfasts following a randomized crossover design, consisting of milk and muffins made with different oils (virgin olive oil (VOO), sunflower oil (SFO), and a mixture of seeds oil (SFO/canola oil) with added either dimethylpolysiloxane (SOD), or natural antioxidants from olive mill wastewater alperujo (phenols; SOP)), previously subjected to 20 heating cycles. Postprandial inflammatory status in PBMCs was assessed by the activation of nuclear NF-κB, the concentration in cytoplasm of the NF-κB inhibitor (IκB-α), the mRNA levels of NF-κB subunits and activators (p65, IKKß, and IKKα) and other inflammatory molecules (TNF-α, IL-1ß, IL-6, MIF, and JNK), and lipopolysaccharide (LPS) levels. VOO and SOP breakfasts reduced NF-κB activation, increased IκB-α, and decreased LPS plasma concentration. SFO increased IKKα, IKKß, p65, IL-1b, IL-6, MIF, and JNK mRNA levels, and plasma LPS. CONCLUSION: Oils rich in phenols, whether natural (VOO) or artificially added (SOP), reduce postprandial inflammation, compared with seed oil (sunflower).
Assuntos
Antioxidantes/administração & dosagem , Ácidos Graxos Monoinsaturados/administração & dosagem , Obesidade/metabolismo , Fenol/administração & dosagem , Óleos de Plantas/administração & dosagem , Estudos Cross-Over , Dimetilpolisiloxanos/metabolismo , Ácidos Graxos Monoinsaturados/química , Manipulação de Alimentos/métodos , Temperatura Alta , Humanos , Quinase I-kappa B/sangue , Proteínas I-kappa B/sangue , Inflamação/tratamento farmacológico , Interleucina-1beta/sangue , Interleucina-6/sangue , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos/sangue , Pessoa de Meia-Idade , Inibidor de NF-kappaB alfa , NF-kappa B/sangue , Obesidade/fisiopatologia , Azeite de Oliva , Óleos de Plantas/química , Período Pós-Prandial , Óleo de Brassica napus , Óleo de Girassol , Fator de Necrose Tumoral alfa/metabolismoRESUMO
AIMS: Chronic inflammation is linked to type 2 diabetes (T2DM), so we investigated correlations between obesity, blood glucose levels, and inflammation in T2DM patients. METHODS: Peripheral blood mononuclear cells (PBMCs) were collected from 40 T2DM patients (27 men, 13 women; mean age 49.63 years), and 10 non-diabetic controls (all men; mean age 38.60 years). Inflammation was measured as DNA-binding activity of nuclear factor kappaB (NF-kappaB), a key transcription factor in inflammation. Protein levels of NF-kappaB subunit p65, and NF-kappaB inhibitor IkappaBalpha were assessed by Western blot. Transcript levels for p65, IkappaBalpha, and the NF-kappaB target genes TNF-alpha, MMP-9, IL-6, IL-8, and IL-18 were measured by real-time PCR. Body mass index (BMI) and glycohemoglobin were measured for all the subjects. RESULTS: NF-kappaB DNA-binding activity, p65 and IkappaBalpha protein levels, and expression of IL-6, TNFalpha and MMP-9 were significantly higher in PBMCs from T2DM patients, than from non-diabetic controls. NF-kappaB binding was significantly positively associated with both BMI and homeostasis model assessment of insulin resistance (HOMA-IR). CONCLUSIONS: Inflammation was observed in PBMCs in T2DM patients in a Chinese population, and correlated independently with obesity and blood glucose levels. Lack of correlation with glycohemoglobin suggested that moderate-term blood glucose control did not mitigate inflammation.
Assuntos
Citocinas/sangue , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/fisiopatologia , Mediadores da Inflamação/sangue , NF-kappa B/sangue , Obesidade/sangue , Obesidade/fisiopatologia , Adulto , Idoso , Glicemia/análise , Índice de Massa Corporal , China , Diabetes Mellitus Tipo 2/complicações , Feminino , Hemoglobinas Glicadas/análise , Hospitais Universitários , Humanos , Proteínas I-kappa B/sangue , Proteínas I-kappa B/genética , Resistência à Insulina , Interleucina-6/sangue , Interleucina-6/genética , Leucócitos Mononucleares/metabolismo , Masculino , Metaloproteinase 9 da Matriz/sangue , Metaloproteinase 9 da Matriz/genética , Pessoa de Meia-Idade , Inibidor de NF-kappaB alfa , Obesidade/complicações , RNA Mensageiro/metabolismo , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/genéticaRESUMO
AIM: The mechanism of hyperbaric oxygen (HBO) therapy for acute pancreatitis has not been fully clarified yet. The main purpose of this study was to investigate the effect of HBO on nuclear factor kappaB (NF-kappaB) activation and the inflammatory response in rats with acute necrotizing pancreatitis (ANP). METHODS: A total of 120 male Sprague-Dawley rats were randomly divided into 3 groups (40 in each): control, ANP and ANP + HBO. ANP rat models were established by a retrograde injection of 5% sodium taurocholate into the pancreatic duct. HBO treatment was performed at 2.5-fold absolute atmospheric pressure in 90% oxygen for 1, 3, 5, and 7 h. The activation of NF-kappaB and its inhibitor IkappaBalpha in peripheral blood neutrophilic granulocytes was measured by electrophoretic mobility shift assay and Western blot, respectively. The inflammatory cytokines [interleukin (IL)-2, IL-6, tumor necrosis factor-alpha (TNF-alpha), and intercellular adhesion molecule 1] in the blood were measured by enzyme-linked immunosorbent assay. RESULTS: The blood levels of inflammatory cytokines and NF-kappaB activation were significantly increased in ANP rats compared to control rats, but IkappaBalpha activation was suppressed. The levels of the elevated inflammatory cytokines were positively correlated with the changes in NF-kappaB activation. After HBO treatment, the blood levels of inflammatory cytokines and NF-kappaB activation were significantly decreased in the ANP + HBO group in a time-dependent manner, but IkappaBalpha activation was increased. CONCLUSION: Our findings suggest that acute pancreatitis is associated with the upregulation of cytokines in blood as well as upregulation of NF-kappaB levels and downregulation of IkappaBalpha activation in peripheral blood neutrophilic granulocytes. In contrast, HBO plays a role in acute pancreatitis treatment by normalizing these changes.
Assuntos
Biomarcadores/sangue , Oxigenoterapia Hiperbárica , NF-kappa B/sangue , Pancreatite Necrosante Aguda/terapia , Amilases/urina , Animais , Ascite/enzimologia , Ativação Enzimática , Proteínas I-kappa B/sangue , Molécula 1 de Adesão Intercelular/sangue , Interleucina-2/sangue , Interleucina-6/sangue , Masculino , Inibidor de NF-kappaB alfa , Neutrófilos/enzimologia , Pâncreas/patologia , Pancreatite Necrosante Aguda/sangue , Pancreatite Necrosante Aguda/patologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/sangueRESUMO
To determine whether insulin administration modulates the systemic inflammatory response in infants undergoing cardiac surgery with cardiopulmonary bypass, 60 infants undergoing cardiopulmonary bypass were randomly assigned into a routine therapy group or to an intensive insulin therapy group with 30 infants in each group. Plasma IL-1beta, IL-6, IL-10, and TNF-alpha levels were determined before anesthesia, at the initiation of cardiopulmonary bypass, and at 0, 6, 12, 24, and 48 h after cardiopulmonary bypass. Nuclear factor-kappaBp65 expression and IkappaB expression in peripheral blood mononuclear cells were also measured by Western blot analysis. TNF-alpha, IL-1beta, IL-6, and IL-10 levels were all elevated after the initiation of cardiopulmonary bypass. However, TNF-alpha, IL-1beta, and IL-6 levels were significantly attenuated in the intensive insulin therapy group compared to those in the routine therapy group after initiation of cardiopulmonary bypass (p<0.05 or <0.01). Meanwhile, plasma IL-10 levels were significantly higher in the intensive insulin therapy group than in the routine therapy group after initiation of cardiopulmonary bypass (p<0.05 or <0.01). Accordingly, Nuclear factor-kappaBp65 expression and IkappaB expression were significantly increased after initiation of cardiopulmonary bypass in both groups (p<0.05 or <0.01). The expression of Nuclear factor-kappaBp65, which induces the transcription of pro-inflammatory cytokines was significantly attenuated in the intensive insulin therapy group (p<0.05 or <0.01). Meanwhile, the expression of IkappaB, an inhibitor of NF-kappaB, was significantly higher in the intensive insulin therapy group (p<0.05 or <0.01). These results suggested that intensive insulin therapy may attenuate the systemic inflammatory response in infants undergoing cardiopulmonary bypass.
Assuntos
Ponte Cardiopulmonar , Mediadores da Inflamação/sangue , Insulina/uso terapêutico , Glicemia/metabolismo , Ponte Cardiopulmonar/efeitos adversos , Feminino , Cardiopatias Congênitas/cirurgia , Humanos , Proteínas I-kappa B/sangue , Lactente , Inflamação/tratamento farmacológico , Insulina/sangue , Interleucina-10/sangue , Interleucina-1beta/sangue , Interleucina-6/sangue , Período Intraoperatório , Masculino , Fator de Transcrição RelA/sangue , Fator de Necrose Tumoral alfa/sangueRESUMO
In order to investigate whether weight loss can lead to improvement of the mononuclear cell (MNC) proinflammatory state, 21 nondiabetic obese women with mean age 34+/-2 years (mean+/-s.e.m.) and BMI 32.5+/-1.2 kg/m2 were enrolled in a 12-week caloric restriction and light exercise-based weight loss program. Ten lean women served as controls. Reverse transcription-PCR of proinflammatory cytokines and adipocytokines as well as homeostasis model assessment of insulin resistance (HOMA-IR) were determined before and after weight reduction. Nuclear factor kappaB (NF-kappaB) binding to DNA and inhibitors of NF-kappaB (IkappaB-alpha and IkappaB-beta) obtained from peripheral MNCs were measured. Overall, subjects lost a mean of 4.0+/-0.4 kg (5.0+/-0.3% of their initial body weight) (P<0.01). In addition to significant reductions in BMI, fasting glucose and insulin concentrations, mean serum high-sensitivity C-reactive protein (hs-CRP), migration inhibitor factor (MIF), leptin and visfatin levels decreased by 49.0, 66.6, 17.2, and 50.2%, respectively (all P<0.05), while adiponectin concentrations rose by 33.9% (P<0.05). The DNA binding of the transcriptionally active NF-kappaB from (p65/p50) decreased by 38.1% (P<0.05). Elevated levels of mRNA of NF-kappaB related proinflammatory genes, tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), MIF, and matrix metalloproteinase-9 (MMP-9), decreased significantly after weight loss. Although mRNA expression of Rel-A, p105, IkappaB-alpha, IkappaB-beta decreased significantly, their protein levels did not change after weight loss. As a group, NF-kappaB binding activity correlated with HOMA-IR (r=0.332, P=0.049) and marginally with values of BMI (r=0.308, P=0.059). In conclusion, weight loss by 5% of initial weight in nondiabetic obese women led to significant improvement in activated intranuclear NF-kappaB binding as well as several transcriptions of proinflammatory genes regulated by NF-kappaB.
Assuntos
Inflamação/fisiopatologia , Leucócitos Mononucleares/fisiologia , Obesidade/fisiopatologia , Redução de Peso/fisiologia , Adulto , Glicemia/metabolismo , Proteína C-Reativa/metabolismo , Estudos de Casos e Controles , Feminino , Humanos , Proteínas I-kappa B/sangue , Inflamação/sangue , Inflamação/patologia , Insulina/sangue , Interleucina-6/sangue , Oxirredutases Intramoleculares/sangue , Leptina/sangue , Leucócitos Mononucleares/patologia , Fatores Inibidores da Migração de Macrófagos/sangue , Metaloproteinase 9 da Matriz/sangue , Inibidor de NF-kappaB alfa , NF-kappa B/sangue , Nicotinamida Fosforribosiltransferase/sangue , Obesidade/sangue , Obesidade/patologia , Fator de Necrose Tumoral alfa/sangueRESUMO
Crohn's disease (CD) is an inflammatory disease characterized by the activation of the immune system in the gut. Since tumor necrosis factor (TNF-alpha) plays an important role in the initiation and perpetuation of intestinal inflammation in CD, we investigated whether TX 527 [19-nor-14,20-bisepi-23-yne-1,25(OH)(2)D(3)], a Vitamin D analogue, could affect peripheral blood mononuclear cells (PBMC) proliferation and exert an immunosuppressive effect on TNF-alpha production in CD patients, and whether this immunosuppressive action could be mediated by NF-kappaB down-regulation. TX 527 significantly decreased cell proliferation and TNF-alpha levels. On activation, NF-kappaB, rapidly released from its cytoplasmatic inhibitor (IKB-alpha), transmigrates into the nucleus and binds to DNA response elements in gene promoter regions. The activation of NF-kappaB, stimulated by TNF-alpha, and its nuclear translocation together with the degradation of IKB-alpha were blocked by TX 527. At the same time, NF-kappaB protein levels present in cytoplasmic extracts decreased in the presence of TNF-alpha and increased when PBMC were incubated with TX 527. The results of our studies indicate that TX 527 inhibits TNF-alpha mediated effects on PBMC and the activation of NF-kappaB and that its action is mediated by Vitamin D receptor (VDR), which is activated when the cells are stimulated with TX 527.
Assuntos
Alcinos/sangue , Colecalciferol/sangue , Doença de Crohn/sangue , NF-kappa B/sangue , Vitamina D/análogos & derivados , Adulto , Idoso , Alcinos/uso terapêutico , Estudos de Casos e Controles , Proliferação de Células , Células Cultivadas , Colecalciferol/uso terapêutico , Doença de Crohn/tratamento farmacológico , Interações Medicamentosas , Feminino , Humanos , Proteínas I-kappa B/sangue , Terapia de Imunossupressão , Masculino , Pessoa de Meia-Idade , Estrutura Molecular , Receptores de Calcitriol/sangue , Fator de Necrose Tumoral alfa/farmacologia , VitaminasRESUMO
Calcitriol has important immunomodulation action and can prolong recipient survival after organ transplantation. The data in this study demonstrated that treatment of liver allograft recipient with calcitriol can protect allograft from acute rejection and prolong recipient's survival. Calcitriol inhibited expression of proinflammatory cytokine such as Interleukin-2 and Interferon-gamma intragraft. It also inhibited expression of nuclear factor kappaB (NF-kappaB) significantly as a result of enhancing its inhibitory protein I kappa B (IkappaB). As well, expression of zinc-finger protein A20 (A20) was enhanced significantly. The results suggest that calcitriol exerts its immunosuppression action in part through inducement of the A20, IkB, inhibition of NF-kB, and resultant proinflammatory expression pathway.
Assuntos
Calcitriol/uso terapêutico , Agonistas dos Canais de Cálcio/uso terapêutico , Proteínas de Ligação a DNA/biossíntese , Rejeição de Enxerto/mortalidade , Rejeição de Enxerto/prevenção & controle , Transplante de Fígado/fisiologia , Animais , Proteínas de Ligação a DNA/genética , Rejeição de Enxerto/genética , Proteínas I-kappa B/sangue , Interferon gama/sangue , Interleucina-2/sangue , Fígado/patologia , Masculino , NF-kappa B/sangue , Ratos , Ratos Sprague-Dawley , Análise de Sobrevida , Proteína 3 Induzida por Fator de Necrose Tumoral alfaRESUMO
Macrophages/monocytes and the proinflammatory mediators, such as tumour necrosis factor (TNF)-alpha, prostaglandin E(2) (PGE(2)), macrophage inflammatory protein (MIP)-1alpha and MIP-1alpha, play a critical role in the progression of immunological disorders including rheumatoid arthritis, Behçet's disease and Crohn's disease. In addition, the nicotinic acetylcholine receptor-alpha7 (alpha7nAChR) subunit is an essential regulator of inflammation. In this study, we evaluated the expression of the alpha7nAChR subunit on human peripheral monocytes and the effect of nicotine on the production of these proinflammatory mediators by activated monocytes. Fluorescein isothiocyanate (FITC)-labelled alpha-bungarotoxin demonstrated the cell surface expression of the alpha7nAchR subunit. Pretreatment with low-dose nicotine caused inhibition of TNF-alpha, PGE(2), MIP-1alpha and MIP-1alpha production, and mRNA expression of TNF-alpha, MIP-1alpha and MIP-1alpha and COX-2 in lipopolysaccharide (LPS)-activated monocytes. These suppressive effects of nicotine were caused at the transcriptional level and were mediated through alpha7nAChR. Nicotine suppressed the phosphorylation of I-kappaB, and then inhibited the transcriptional activity of nuclear factor-kappaB. These immunosuppressive effects of nicotine may contribute to the regulation of some immune diseases.
Assuntos
Mediadores da Inflamação/sangue , Monócitos/efeitos dos fármacos , Nicotina/farmacologia , Receptores Nicotínicos/fisiologia , Células Cultivadas , Relação Dose-Resposta a Droga , Humanos , Proteínas I-kappa B/sangue , Lipopolissacarídeos/farmacologia , Monócitos/metabolismo , NF-kappa B/sangue , NF-kappa B/genética , Fosforilação/efeitos dos fármacos , Receptores Nicotínicos/sangue , Receptores Nicotínicos/efeitos dos fármacos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Transcrição Gênica/efeitos dos fármacos , Receptor Nicotínico de Acetilcolina alfa7RESUMO
Rheumatoid arthritis (RA) is an autoimmune disease of unknown etiology, characterized by the presence of inflammatory synovitis accompanied by destruction of joint cartilage and bone. Treatment with vasoactive intestinal peptide (VIP) prevents experimental arthritis in animal models by downregulation of both autoimmune and inflammatory components of the disease. The aim of this study was to characterize the protective effect of VIP on bone erosion in collagen-induced arthritis (CIA) in mice. We have studied the expression of different mediators implicated in bone homeostasis, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), receptor activator of nuclear factor-kappaB (RANK), receptor activator of nuclear factor-kappaB ligand (RANKL), osteoprotegerin (OPG), IL-1, IL-4, IL-6, IL-10, IL-11 and IL-17. Circulating cytokine levels were assessed by ELISA and the local expression of mediators were determined by RT-PCR in mRNA extracts from joints. VIP treatment resulted in decreased levels of circulating IL-6, IL-1beta and TNFalpha, and increased levels of IL-4 and IL-10. CIA-mice treated with VIP presented a decrease in mRNA expression of IL-17, IL-11 in the joints. The ratio of RANKL to OPG decreased drastically in the joint after VIP treatment, which correlated with an increase in levels of circulating OPG in CIA mice treated with VIP. In addition, VIP treatment decreased the expression of mRNA for RANK, iNOS and COX-2. To investigate the molecular mechanisms involved, we tested the activity of NFkappaB and AP-1, two transcriptional factors closely related to joint erosion, by EMSA in synovial cells from CIA mice. VIP treatment in vivo was able to affect the transcriptional activity of both factors. Our data indicate that VIP is a viable candidate for the development of treatments for RA.
Assuntos
Artrite Experimental/tratamento farmacológico , Osso e Ossos/efeitos dos fármacos , Peptídeo Intestinal Vasoativo/uso terapêutico , Animais , Artrite Experimental/genética , Artrite Experimental/metabolismo , Osso e Ossos/metabolismo , Ciclo-Oxigenase 2/metabolismo , Citocinas/biossíntese , Citocinas/sangue , Citocinas/genética , Modelos Animais de Doenças , Avaliação de Medicamentos , Perfilação da Expressão Gênica , Glicoproteínas/sangue , Proteínas I-kappa B/sangue , Mediadores da Inflamação/sangue , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos DBA , Inibidor de NF-kappaB alfa , NF-kappa B/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/metabolismo , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Osteólise/etiologia , Osteólise/metabolismo , Osteólise/prevenção & controle , Osteoprotegerina , Transporte Proteico/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-jun/sangue , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores Citoplasmáticos e Nucleares/sangue , Receptores do Fator de Necrose Tumoral/sangue , Fator de Transcrição AP-1/metabolismo , Transcrição Gênica/efeitos dos fármacos , Peptídeo Intestinal Vasoativo/farmacologiaRESUMO
BACKGROUND: Agonists of the peroxisome proliferator-activated receptor-gamma may help to regulate inflammation by modulating the production of inflammatory mediators and adhesion molecules. The purpose of this study was to determine the protective effects of rosiglitazone on renal injury in a sepsis model and to explore the mechanism. METHODS: In lipopolysaccharide (LPS)-induced mouse sepsis, we examined the effect of rosiglitazone on LPS-induced overproduction of inflammatory mediators, on the expression of adhesion molecules in renal tubular epithelial cells and on renal function. The mechanism of the protective effect was investigated in vitro using human renal tubular epithelial cells. RESULTS: Rosiglitazone significantly decreased serum tumour necrosis factor (TNF)-alpha and interleukin (IL)-1beta levels during sepsis. The levels of blood urea nitrogen and creatinine were significantly lower in mice pre-treated with rosiglitazone than that in LPS-treated mice. Rosiglitazone reduced the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in tubular epithelial cells and interstitium of LPS-treated mice. Pre-treatment with rosiglitazone reduced the infiltration of macrophages/monocytes in renal tissue. In cultured tubular epithelial cells, rosiglitazone significantly decreased the expression of ICAM-1 and VCAM-1 induced by TNF-alpha or IL-1beta, inhibited the degradation of inhibitor kappaBalpha (IkappaBalpha) and blocked the activation of the p65 subunit of nuclear factor (NF)-kappaB. CONCLUSIONS: These results indicate that pre-treatment with rosiglitazone attenuated the production of TNF-alpha and IL-1beta and reduced adhesion molecule expression in renal tubular epithelial cells of LPS-treated mice. Rosiglitazone has an anti-inflammatory effect in renal tubular epithelial cells through the inhibition of NF-kappaB activation.
Assuntos
Anti-Inflamatórios/farmacologia , Túbulos Renais/efeitos dos fármacos , Sepse/tratamento farmacológico , Sepse/fisiopatologia , Tiazolidinedionas/farmacologia , Animais , Western Blotting , Modelos Animais de Doenças , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Proteínas I-kappa B/sangue , Imuno-Histoquímica , Molécula 1 de Adesão Intercelular/sangue , Interleucina-1/sangue , Túbulos Renais/citologia , Túbulos Renais/metabolismo , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Inibidor de NF-kappaB alfa , NF-kappa B/antagonistas & inibidores , Rosiglitazona , Sepse/sangue , Fator de Necrose Tumoral alfa/análise , Molécula 1 de Adesão de Célula Vascular/sangueRESUMO
BACKGROUND: In view of the increase in plasma concentrations of proinflammatory mediators tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and C-reactive protein (CRP) in obesity, we investigated whether peripheral blood mononuclear cells (MNC) from obese subjects are in a proinflammatory state. METHODS AND RESULTS: MNC were prepared from fasting blood samples of obese (n=16; body mass index [BMI]=37.7+/-5.0 kg/m2) and normal-weight control (n=16; BMI=23.8+/-1.9 kg/m2) subjects. Nuclear factor kappaB (NF-kappaB) binding to DNA in nuclear extracts was elevated (P<0.05) and the inhibitor of NFkappaB-beta (IkappaB-beta) was significantly lower (P<0.001) in the obese group. Reverse transcription-polymerase chain reaction revealed elevated levels of migration inhibitor factor (MIF), IL-6, TNF-alpha, and matrix metalloproteinase-9 (MMP-9) mRNA expression in the obese subjects (P<0.05). Plasma concentrations of MIF, IL-6, TNF-alpha, MMP-9, and CRP were also significantly higher. Plasma glucose, insulin, and free fatty acids (FFAs) were measured, and homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. Plasma FFA concentration related significantly to BMI, IL-6, and TNF-alpha mRNA expression and plasma CRP levels but not to HOMA-IR. On the other hand, the inflammatory mediators were significantly related to BMI and HOMA-IR. CONCLUSIONS: These data show (1) for the first time that MNC in obesity are in a proinflammatory state with an increase in intranuclear NF-kappaB binding, a decrease in IkappaB-beta, and an increase in the transcription of proinflammatory genes regulated by NF-kappaB; (2) that plasma FFAs are a modulator of inflammation; and (3) that insulin resistance is a function of inflammatory mediators.
Assuntos
Inflamação/sangue , Leucócitos Mononucleares/patologia , Obesidade/sangue , Adulto , Glicemia/análise , Índice de Massa Corporal , Proteína C-Reativa/análise , Proteína C-Reativa/genética , Núcleo Celular/química , DNA/sangue , Jejum/sangue , Ácidos Graxos não Esterificados/sangue , Feminino , Humanos , Proteínas I-kappa B/sangue , Proteínas I-kappa B/genética , Inflamação/etiologia , Inflamação/genética , Resistência à Insulina , Interleucina-6/sangue , Interleucina-6/genética , Leucócitos Mononucleares/química , Leucócitos Mononucleares/ultraestrutura , Fatores Inibidores da Migração de Macrófagos/sangue , Fatores Inibidores da Migração de Macrófagos/genética , Metaloproteinase 9 da Matriz/sangue , Metaloproteinase 9 da Matriz/genética , Pessoa de Meia-Idade , NADPH Oxidases , Inibidor de NF-kappaB alfa , NF-kappa B/biossíntese , NF-kappa B/sangue , Obesidade/complicações , Fosfoproteínas/sangue , RNA Mensageiro/biossíntese , RNA Mensageiro/sangue , Fator de Transcrição RelA , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/genéticaRESUMO
Human peripheral blood mononuclear cells (PBMCs) were activated using anti-CD3/CD28 (HIT3A/CD28.2) resulting in degradation of IkB alpha, an inhibitor of NFkB, relative to unactivated cells. Degradation of IkB alpha began by 30 min and proceeded for at least 5 h. Calpeptin, a calpain inhibitor, inhibited IkB alpha degradation in a time- and dose-dependent manner. Furthermore, calpain inhibition increased IkB alpha levels compared to nonactivated controls. Recombinant IkB alpha was incubated with purified porcine m-calpain in the presence of 0.1% Triton X-100, and the degradation products were monitored by SDS-PAGE and sequenced. Most of the degradation products were peptides derived from calpain, but one was derived from IkB alpha cleaved between amino acids 50 and 51 (glutamine and glutamic acid). The liberated fragment included the entire signal response domain (SRD), a region containing key serine and threonine residues necessary for phosphorylation by the IKKinase complex and sites required for ubiquitination. The results suggest that calpain plays an important role in IkB alpha degradation, a crucial event in T cell activation.
Assuntos
Calpaína/antagonistas & inibidores , Proteínas I-kappa B/sangue , Leucócitos Mononucleares/metabolismo , Linfócitos/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Antígenos CD28/sangue , Complexo CD3/sangue , Calpaína/sangue , Células Cultivadas , Inibidores de Cisteína Proteinase/farmacologia , Dipeptídeos/farmacologia , Humanos , Proteínas I-kappa B/genética , Cinética , Ativação Linfocitária/fisiologia , Linfócitos/efeitos dos fármacos , Dados de Sequência Molecular , Inibidor de NF-kappaB alfa , NF-kappa B/antagonistas & inibidores , NF-kappa B/sangue , Sondas de OligonucleotídeosRESUMO
The risk of infection and cancer increases dramatically beyond middle age, when T-cell function is noticeably altered. Nevertheless, many elderly people remain in apparently good health. To identify immunological adaptations favouring longevity, a pilot study was undertaken to compare peripheral blood T cells from healthy volunteers aged 18-25 years with those >65 years. Instead of preserved immune function in the elderly, there was an emergence of haematopoietic space particularly affecting T- and B-cell numbers, together with early signs of immunoglobulin dysregulation. Age-associated proliferative defects were present irrespective of the stimuli used. A higher constitutive expression of NF-kappaB and I-kappaBalpha in the nuclei of peripheral lymphocytes from the elderly remained unaltered by activation stimuli, despite the presence of exogenous cytokines. Nevertheless, activation resulted in their higher CD95 upregulation and more intracellular bcl-2 (suggesting a survival advantage), but lower CD27, CD28 and CD45Rb expression. The presence of CD45RO(+) CD45Rb(-) populations was unique to the elderly and their lower replicative potential was not due to the presence of CD25(+) regulatory T cells. These data collectively suggest altered gene regulation and the accumulation of terminally differentiated T cells during healthy ageing.
Assuntos
Envelhecimento/imunologia , Proteínas I-kappa B/sangue , NF-kappa B/sangue , Linfócitos T/citologia , Adolescente , Adulto , Idoso , Diferenciação Celular/imunologia , Divisão Celular/imunologia , Humanos , Imunofenotipagem , Antígenos Comuns de Leucócito/sangue , Contagem de Leucócitos , Ativação Linfocitária/imunologia , Inibidor de NF-kappaB alfa , Proteínas Proto-Oncogênicas c-bcl-2/sangue , Subpopulações de Linfócitos T/imunologia , Linfócitos T/imunologia , Receptor fas/sangueRESUMO
Leukocytosis in tobacco smokers has been well recognized; however, the exact cause has not been elucidated. To test the hypothesis that tobacco nicotine stimulates neutrophils in the respiratory tract to produce IL-8, which causes neutrophilia in vivo, we examined whether nicotine induces neutrophil-IL-8 production in vitro; the causative role of NF-kappaB in its production, in association with the possible production of reactive oxygen intermediates that activate NF-kappaB; and the nicotinic acetylcholine receptors (nAChRs) involved in IL-8 production. Nicotine stimulated neutrophils to produce IL-8 in both time- and concentration-dependent manners with a 50% effective concentration of 1.89 mM. A degradation of IkappaB-alpha/beta proteins and an activity of NF-kappaB p65 and p50 were enhanced following nicotine treatment. The synthesis of superoxide and the oxidation of dihydrorhodamine 123 (DHR) were also enhanced. The NOS inhibitor, nomega-Nitro-l-arginine methyl ester, prevented nicotine-induced IL-8 production, with an entire abrogation of DHR oxidation, IkappaB degradation, and NF-kappaB activity. Neutrophils spontaneously produced NO whose production was not increased, but rather decreased by nicotine stimulation, suggesting that superoxide, produced by nicotine, generates peroxynitrite by reacting with preformed NO, which enhances the NF-kappaB activity, thereby producing IL-8. The nAChRs seemed to be involved in IL-8 production. In smokers, blood IL-8 levels were significantly higher than those in nonsmokers. In conclusion, nicotine stimulates neutrophil-IL-8 production via nAChR by generating peroxynitrite and subsequent NF-kappaB activation, and the IL-8 appears to contribute to leukocytosis in tobacco smokers.
Assuntos
Interleucina-8/sangue , NF-kappa B/sangue , Neutrófilos/imunologia , Nicotina/farmacologia , Ácido Peroxinitroso/sangue , Humanos , Proteínas I-kappa B/sangue , Interleucina-8/biossíntese , Cinética , Inibidor de NF-kappaB alfa , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/biossíntese , Peroxidase/sangue , Transporte Proteico , Espécies Reativas de Oxigênio/metabolismo , Fumar/sangue , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Minimal-change nephrotic syndrome (MCNS) is a renal disease characterized by heavy glomerular proteinuria and increased production of cytokines by immune cells. Because of the central role of nuclear factor-kappaB (NF-kappaB) in the regulation of cytokine expression, its activity during the relapse and remission phases of steroid-sensitive MCNS was analyzed. During relapse, nuclear extracts from peripheral blood mononuclear cells displayed high levels of NF-kappaB DNA-binding activity, consisting primarily of p50/RelA (p65) complexes. NF-kappaB p65 and IkappaBalpha proteins were barely detected or not detected in cytosolic fractions during relapse, in contrast to remission. The lack of expression of IkappaBalpha protein was associated with downregulation of IkappaBalpha mRNA and increases in the levels of the mRNA encoding the proteasome alpha2 subunit proteolytic pathway. In addition, inhibition of proteasome activity induced cytosolic accumulation of phosphorylated IkappaBalpha and significant reductions in the NF-kappaB binding activity in nuclear extracts from peripheral blood mononuclear cells from patients experiencing relapses. These results suggest that alterations in the NF-kappaB/IkappaBalpha regulatory feedback loop may contribute to the immunologic abnormalities that occur in steroidsensitive MCNS.