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1.
J Enzyme Inhib Med Chem ; 35(1): 96-108, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31690133

RESUMO

A series of analogues of Amb639752, a novel diacylglycerol kinase (DGK) inhibitor recently discovered by us via virtual screening, have been tested. The compounds were evaluated as DGK inhibitors on α, θ, and ζ isoforms, and as antagonists on serotonin receptors. From these assays emerged two novel compounds, namely 11 and 20, which with an IC50 respectively of 1.6 and 1.8 µM are the most potent inhibitors of DGKα discovered to date. Both compounds demonstrated the ability to restore apoptosis in a cellular model of X-linked lymphoproliferative disease as well as the capacity to reduce the migration of cancer cells, suggesting their potential utility in preventing metastasis. Finally, relying on experimental biological data, molecular modelling studies allow us to set a three-point pharmacophore model for DGK inhibitors.


Assuntos
Indóis/farmacologia , Lipase Lipoproteica/antagonistas & inibidores , Piperazinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células HEK293 , Humanos , Indóis/síntese química , Indóis/química , Lipase Lipoproteica/metabolismo , Linfócitos/efeitos dos fármacos , Células MCF-7 , Modelos Moleculares , Estrutura Molecular , Monócitos/efeitos dos fármacos , Piperazinas/síntese química , Piperazinas/química , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Relação Estrutura-Atividade , Linfócitos T/efeitos dos fármacos
2.
Pulm Pharmacol Ther ; 59: 101851, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31563516

RESUMO

BACKGROUND: Methylxanthines are important pharmacological agents in the treatment of asthma and of chronic obstructive pulmonary diseases. The present study was designed to compare the ability of doxofylline and theophylline to modulate inflammatory pathways in human monocytes. METHODS: Monocytes isolated from healthy anonymous human buffy coats were treated with doxofylline or theophylline in the presence of phorbol 12-myristate 13-acetate (PMA) or lipopolysaccharide (LPS), and their phenotype, the oxidative burst, cytokine expression and release, cAMP production, and protein kinase C (PKC) activity were evaluated. RESULTS: Doxofylline and theophylline did not have overlapping effects on human monocytes. While sharing some common characteristics, they differed significantly in their selectivity. Theophylline affected LPS- above PMA-induced cellular responsivity, while doxofylline behaved in the opposite manner. Furthermore, when testing PKC activity, we found an inhibitory effect of doxofylline but not of theophylline, at equimolar doses. CONCLUSIONS: In conclusion, our data support the growing hypothesis that doxofylline does not have a superimposable mechanism of action compared to theophylline, and this may both explain some differences in the risk/benefit ratio and may direct studies to tailor therapy for patients.

3.
Int J Mol Sci ; 20(2)2019 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-30642067

RESUMO

Abdominal aortic aneurysm (AAA) is a focal dilatation of the aorta, caused by both genetic and environmental factors. Although vascular endothelium plays a key role in AAA progression, the biological mechanisms underlying the mechanical stress involvement are only partially understood. In this study, we developed an in vitro model to characterize the role of mechanical stress as a potential trigger of endothelial deregulation in terms of inflammatory response bridging between endothelial cells (ECs), inflammatory cells, and matrix remodeling. In AAA patients, data revealed different degrees of calcification, inversely correlated with wall stretching and also with inflammation and extracellular matrix degradation. In order to study the role of mechanical stimulation, endothelial cell line (EA.hy926) has been cultured in healthy (10% strain) and pathological (5% strain) dynamic conditions using a bioreactor. In presence of tumor necrosis factor alpha (TNF-α), high levels of matrix metalloproteinase-9 (MMP-9) expression and inflammation are obtained, while mechanical stimulation significantly counteracts the TNF-α effects. Moreover, physiological deformation also plays a significant role in the control of the oxidative stress. Overall our findings indicate that, due to wall calcification, in AAA there is a significant change in terms of decreased wall stretching.


Assuntos
Aneurisma da Aorta Abdominal/fisiopatologia , Técnicas de Cultura de Células/instrumentação , Células Endoteliais/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Aneurisma da Aorta Abdominal/imunologia , Aneurisma da Aorta Abdominal/metabolismo , Reatores Biológicos , Linhagem Celular , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Redes Reguladoras de Genes , Humanos , Metaloproteinase 9 da Matriz/metabolismo , Modelos Biológicos , Estresse Oxidativo , Estresse Mecânico
4.
Eur J Med Chem ; 164: 378-390, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30611057

RESUMO

As part of an effort to identify druggable diacylglycerol kinase alpha (DGKα) inhibitors, we used an in-silico approach based on chemical homology with the two commercially available DGKα inhibitors R59022 and R59949. Ritanserin and compound AMB639752 emerged from the screening of 127 compounds, showing an inhibitory activity superior to the two commercial inhibitors, being furthermore specific for the alpha isoform of diacylglycerol kinase. Interestingly, AMB639752 was also devoid of serotoninergic activity. The ability of both ritanserin and AMB639752, by inhibiting DGKα in intact cells, to restore restimulation induced cell death (RICD) in SAP deficient lymphocytes was also tested. Both compounds restored RICD at concentrations lower than the two previously available inhibitors, indicating their potential use for the treatment of X-linked lymphoproliferative disease 1 (XLP-1), a rare genetic disorder in which DGKα activity is deregulated.


Assuntos
Diacilglicerol Quinase/antagonistas & inibidores , Avaliação Pré-Clínica de Medicamentos/métodos , Transtornos Linfoproliferativos/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Morte Celular/efeitos dos fármacos , Simulação por Computador , Humanos , Piperidinas , Pirimidinonas , Quinazolinonas , Ritanserina , Tiazóis
5.
Fitoterapia ; 127: 252-256, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29499239

RESUMO

The discovery of taste receptors hTAS2Rs expression in extra oral tissue, especially in the gastrointestinal tract and in the respiratory system, has endowed bitter receptors of functionalities that exceed the simple perception of taste and flavour. In particular, stimulation of hTAS2Rs by bitter agents in the airway smooth muscle triggers bronchodilation of possible pharmacological relevance. To study the receptor localization in pulmonary smooth muscle cells and to investigate their biological response to hTAS2R38 activation, we have developed a fluorescent probe for hTAS2R38 starting from the sesquiterpene lactone costunolide, available in multigram amounts from Artemisia umbelliformis Lam. The N-methylanthranilate-containing probe demonstrated a very low cytotoxicity compared to the natural product toward human airway smooth muscle cells and epithelial bronchial cells, but fully retained its binding to hTAS2R38, making it possible the fluorescent detection of cells expressing this bitter receptor.


Assuntos
Artemisia/química , Corantes Fluorescentes/química , Miócitos de Músculo Liso/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Sesquiterpenos/química , Brônquios/citologia , Células Cultivadas , Células Epiteliais/metabolismo , Humanos , Pulmão/citologia , Paladar
6.
Curr Med Chem ; 25(10): 1160-1185, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-28799497

RESUMO

BACKGROUND: Although Cannabis sativa L. is one of the most versatile plant species with multipurpose use both as medical, alimentary source and as psychoactive abuse, its biomedical relevance focused the attention on major cannabinoids. Phytochemical characterization of cannabis highlights the presence of various non-cannabinoids constituents including flavonoids, spiroindans, dihyrostilbenes, dihydrophenanthrenes, lignanamides, steroids and alkaloids. This review aims to identify polyphenols present in this plant, their biosynthesis, their bioactivities and their synthesis, when this occurred. METHODS: We undertook a systematic research focused on bibliographic databases including all noncannabinoids phenolics in various C. sativa strains from their isolation, structural elucidation, their biological activity to their synthesis. RESULT: Nevertheless, attention has so far been focused only on cannabinoids (more than one hundred isolated), cannabis is a complex plant able to produce more than 480 chemical entities that represent almost all of the different biogenetic classes. Regarding phenolic compounds, the plant biosynthesises a plethora of unique non-cannabinoids second metabolites, such as prenylated flavonoids, stilbenoids derivatives and lignanammides. CONCLUSION: Cannabis is a plant with high pharmacological and nutrition values, its potentialities and applications are not only circumscribed to cannabinoids biological activities, but also defined by noncannabinoid compounds. The combination of other cannabinoids together with noncannabinoid components could enhance the beneficial effects of THC and could reduce undesirable side effects.


Assuntos
Cannabis/química , Extratos Vegetais/química , Polifenóis/química , Polifenóis/farmacologia , Canabinoides/química , Canabinoides/farmacologia , Humanos , Estrutura Molecular , Polifenóis/biossíntese , Relação Estrutura-Atividade
7.
Br J Pharmacol ; 175(1): 113-124, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29057467

RESUMO

BACKGROUND AND PURPOSE: A crosstalk between the immune system and depression has been postulated, with monocytes/macrophages and cytokines having a key role in this interaction. In this study, we examined whether vortioxetine, a multimodal anti-depressive drug, was endowed with anti-inflammatory and antioxidative activity, leading to immunomodulatory effects on human monocytes and macrophages. EXPERIMENTAL APPROACH: Human monocytes were isolated from buffy coats and used as such or differentiated into M1 and M2 macrophages. Cells were treated with vortioxetine before or after differentiation, and their responsiveness was evaluated. This included oxy-radical and TNFα production, TNFα and PPARγ gene expression and NF-κB translocation. KEY RESULTS: Vortioxetine significantly reduced the PMA-induced oxidative burst in monocytes and in macrophages (M1 and M2), causing a concomitant shift of macrophages from the M1 to the M2 phenotype, demonstrated by a significant decrease in the expression of the surface marker CD86 and an increase in CD206. Moreover, treatment of monocytes with vortioxetine rendered macrophages derived from this population less sensitive to PMA, as it reduced the oxidative burst, NF-kB translocation, TNFα release and expression while inducing PPARγ gene expression. FACS analysis showed a significant decrease in the CD14+ /CD16+ /CD86+ M1 population. CONCLUSIONS AND IMPLICATIONS: These results demonstrate that in human monocytes/macrophages, vortioxetine has antioxidant activity and anti-inflammatory effects driving the polarization of macrophages towards their alternative phenotype. These findings suggest that vortioxetine, alongside its antidepressive effect, may have immunomodulatory properties.


Assuntos
Anti-Inflamatórios/farmacologia , Fatores Imunológicos/farmacologia , Macrófagos/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Piperazinas/farmacologia , Sulfetos/farmacologia , Antioxidantes/farmacologia , Células Cultivadas , Relação Dose-Resposta a Droga , Humanos , Macrófagos/imunologia , Monócitos/imunologia , Vortioxetina
8.
Pharmacol Res ; 107: 308-314, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-27045818

RESUMO

Circulating human monocytes, a functionally and phenotypically heterogeneous population, are emerging as fundamental cell types in rheumatoid arthritis (RA). The aim of this pilot study was to assess the correlation, if any, among anti-rheumatic drug therapy, circulating CD14(+)CD16(+) monocytes and validated clinical scales (e.g., DAS28 score and ultrasonography US7 score) of disease severity in RA. Thirty consecutive RA patients, either naïve or under disease-modifying anti-rheumatic drugs (DMARDs) or biological therapy, and 10 age-matched healthy volunteers, were enrolled. Monocytes were prepared from heparinized blood samples; surface expression of CD14 and CD16 was determined by flow cytometry. RA patients presented a significantly higher percentage of CD14(+)CD16(+) monocytes, as compared to healthy subjects. There was a good correlation between DAS28 clinical score and the ultrasound composite score US7 (r=0.66), as well as between both scores and the percentage of CD14(+)CD16(+) monocytes (r=0.43 and 0.47, respectively). Naïve RA patients had the highest expression (19.2±3.2%) of CD14(+)CD16(+) monocytes and elevated DAS28 score; patients on DMARDs presented a 7-fold increased expression of CD14(+)CD16(+) monocytes, relatively to healthy volunteers (2.1±1.4%), and an intermediate disease severity. The RA patients treated with biological therapy had a low percentage of CD14(+)CD16(+) monocytes (5.1±3.6%; p<0.01 vs naïve and DMARDs groups), similar to the one detected in healthy controls, and reduced US7 and DAS28 scores. Interestingly, for the same DAS28 score, monocytes isolated from RA patients on biological therapy had a lower CD16 expression than patients on DMARDs. Therefore, CD14(+)CD16(+) circulating blood monocytes may represent an appropriate biomarker to assess RA disease activity along with DAS28 and US7 scores. Together, these three parameters may represent a better indicator for evaluating therapy efficacy.


Assuntos
Antirreumáticos/farmacologia , Artrite Reumatoide/imunologia , Receptores de Lipopolissacarídeos/imunologia , Monócitos/efeitos dos fármacos , Receptores de IgG/imunologia , Adulto , Anticorpos/farmacologia , Anticorpos/uso terapêutico , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Biomarcadores/metabolismo , Feminino , Proteínas Ligadas por GPI/imunologia , Humanos , Masculino , Metotrexato/farmacologia , Metotrexato/uso terapêutico , Pessoa de Meia-Idade , Monócitos/imunologia , Projetos Piloto , Prednisona/farmacologia , Prednisona/uso terapêutico , Índice de Gravidade de Doença
9.
Eur J Pharmacol ; 780: 33-7, 2016 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-26997366

RESUMO

Tocilizumab, etanercept and abatacept are biological drugs used in the therapy of Rheumatoid Arthritis (RA). Their mechanism of action is well documented but their direct effects on human monocytes/macrophages have not been fully investigated. The objective of this study was to evaluate in vitro the influence of these drugs on monocytes/macrophages from healthy volunteers. Human monocytes were isolated from healthy anonymous volunteers and cultured as such or differentiated to monocyte-derived macrophages (MDMs). The effect of tocilizumab, etanercept and abatacept (at concentrations similar to those in plasma of patients) on superoxide anion production, matrix metalloproteinase-9 (MMP-9) gene expression and activity, Peroxisome Proliferator-Activated Receptor (PPAR)γ expression and cell phenotype was evaluated. Exposure of monocytes/macrophages to tocilizumab, etanercept or abatacept resulted in a significant decrease of the PMA-induced superoxide anion production. Interestingly, the expression of PPARγ was significantly increased only by tocilizumab, while etanercept was the only one able to significantly reduce MMP-9 gene expression and inhibit the LPS-induced MMP-9 activity in monocytes. When etanercept and abatacept were added to the differentiating medium, both significantly reduced the amount of CD206(+)MDM. This study demonstrates that etanercept, abatacept and tocilizumab affect differently human monocytes/macrophages. In particular, the IL-6 antagonist tocilizumab seems to be more effective in inducing an anti-inflammatory phenotype of monocytes/macrophages compared to etanercept and abatacept, also in light of the up-regulation of PPARγ whose anti-inflammatory effects are well recognised.


Assuntos
Abatacepte/farmacologia , Anticorpos Monoclonais Humanizados/farmacologia , Etanercepte/farmacologia , Macrófagos/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Biomarcadores/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Macrófagos/citologia , Macrófagos/metabolismo , Metaloproteinase 9 da Matriz/genética , Monócitos/citologia , Monócitos/metabolismo , PPAR gama/metabolismo , Fenótipo
10.
J Affect Disord ; 178: 188-92, 2015 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-25841180

RESUMO

BACKGROUND: Neurokinin 1 receptors (NK-1R) have been involved in several psychiatric disorders including major depression, but less is known for bipolar disorder (BD). METHOD: We compared NK-1R expression and Substance P (SP) ability to induce NF-κB activation in monocytes from BD patients and healthy donors (HD), also looking for the effects of tobacco smoke. After informed written consent, 20 euthymic BD patients, either bipolar type 1 (BDI) or type 2 (BDII), and 14 age-matched healthy donors (HD) were enrolled. NK-1R expression in monocytes was evaluated by Western blot and expressed as the ratio between NK-1R and Na(+)/K(+)-ATPase protein expressions. NF-κB activation was assessed by measuring the nuclear content of the p50 subunit (ELISA kit). RESULTS: NK-1R expression was significantly reduced (P<0.001) in monocytes from BD patients as compared to HD, with no major differences between BDI and BDII patients. Tobacco smoke enhanced NK-1R expression in HD, but not in BD patients. Un-stimulated monocytes from BD patients presented a constitutively higher (P<0.05) content of nuclear p50 subunit as compared to HD. SP and an NK-1R agonist induced NF-κB activation, with a higher effect in HD: this effect was receptor-mediated as it was abrogated by an NK-1R antagonist. LIMITATIONS: As a pilot study enrolling 20 BD patients, an obvious limitation is the sample size. CONCLUSIONS: Our results show the existence of a relevant alteration in NK-1R expression in BD patients and further suggest SP involvement in BD, so improving our understanding of the underlying mechanisms of this disease.


Assuntos
Transtorno Bipolar/metabolismo , Monócitos/metabolismo , NF-kappa B/metabolismo , Receptores da Neurocinina-1/metabolismo , Substância P/metabolismo , Adulto , Western Blotting , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Transdução de Sinais , ATPase Trocadora de Sódio-Potássio/metabolismo
11.
Life Sci ; 126: 28-36, 2015 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-25711428

RESUMO

AIMS: Cyclooxygenase (COX)-inhibiting nitric oxide donors (CINODs) are a new class of drugs that structurally combine a COX inhibitor with a nitric oxide (NO) donating moiety. This combination reduces potential toxicity of the non-steroidal anti-inflammatory drugs (NSAIDs) whilst maintaining the analgesic and anti-inflammatory effects. The present study was undertaken to investigate the anti-inflammatory effects of NCX 429, a naproxen-based CINOD, and to assess the additional properties of NO donation beyond those related to naproxen. MAIN METHODS: We evaluated the in vitro effects of NCX 429 on oxy-radical production, phagocytosis, cytokine release, MMP-9, PPARγ expression and NF-κB activation in human monocytes/MDM and compared to naproxen. Moreover, we compared the in vivo efficacy of NCX 429 and naproxen in a murine model of peritonitis. KEY FINDINGS: In all the experiments performed in vitro, NCX 429 reduced the inflammatory responses with equal or higher efficacy compared to naproxen. Moreover, in in vivo experiments, NCX 429, at the lowest dose tested, was able to significantly inhibit cell influx in response to IL-1ß administration although naproxen was found to be more potent than NCX 429 at reducing PGE2 in inflammatory exudates. SIGNIFICANCE: These results demonstrate that both in vitro and in vivo--in a murine model of peritonitis--NCX 429 elicits significant anti-inflammatory activity, beyond the simple COX inhibition or pure NO release. Therefore, NO donation along with COX inhibition may represent a strategy for investigating inflammatory diseases in which pain and function are not fully resolved by analgesics/anti-inflammatory drugs.


Assuntos
Anti-Inflamatórios não Esteroides , Naproxeno/análogos & derivados , Nitratos , Doadores de Óxido Nítrico , Óxido Nítrico/farmacocinética , Peritonite , Animais , Anti-Inflamatórios não Esteroides/farmacocinética , Anti-Inflamatórios não Esteroides/farmacologia , Dinoprostona/metabolismo , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Humanos , Interleucina-1beta/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , NF-kappa B/metabolismo , Naproxeno/farmacocinética , Naproxeno/farmacologia , Nitratos/farmacocinética , Nitratos/farmacologia , Doadores de Óxido Nítrico/farmacocinética , Doadores de Óxido Nítrico/farmacologia , Peritonite/tratamento farmacológico , Peritonite/metabolismo , Peritonite/patologia , Fagocitose/efeitos dos fármacos
12.
Pharmacol Res ; 68(1): 24-30, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23142211

RESUMO

Increasing evidence suggests that tachykinins are involved in the control of different pathological conditions, including psychiatric disorders. In this study we evaluated the expression of NK(1) and NK(2) receptors (NK-1R and NK-2R), as well as the effects of substance P (SP) and neurokinin A (NKA), in monocytes isolated from 15 healthy subjects and 15 patients with recurrent major depressive disorder (RMDD), under stable antidepressant therapy. NK-1R expression in monocytes from RMDD patients was significantly decreased as compared to healthy subjects, whereas NK-2R expression was markedly increased. Both NK-1R and NK-2R expression correlated with HAM-D, but not HAM-A, score. SP, NKA and selective NK-1R and NK-2R agonists stimulated TNF-α release in monocytes of both groups, with a significant higher effect observed in RMDD. Moreover they induced NF-κB activation, which was reversed by selective NK-1R and NK-2R antagonists, so demonstrating that it was receptor-mediated. The occurrence of a profound alteration in NK receptor expression in RMDD is a novel finding that suggests NK-1R and NK-2R pathways as possible relevant players in major depressive disorder, so improving our understanding of the complex pathogenesis of the disease.


Assuntos
Transtorno Depressivo Maior/metabolismo , Receptores da Neurocinina-1/metabolismo , Receptores da Neurocinina-2/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Monócitos/metabolismo , NF-kappa B/metabolismo , Neurocinina A/farmacologia , Neurotransmissores/farmacologia , Receptores da Neurocinina-1/agonistas , Receptores da Neurocinina-2/agonistas , Substância P/farmacologia , Fator de Necrose Tumoral alfa/metabolismo
13.
Rheumatology (Oxford) ; 51(11): 1942-52, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22829690

RESUMO

OBJECTIVES: Peroxisome proliferator-activated receptor-gamma (PPARγ) is expressed by different cell types in the joints and plays a relevant anti-inflammatory role in various diseases. This pilot study aimed to evaluate PPARγ expression in monocytes/macrophages isolated from RA patients as compared with healthy subjects, the relationships between PPARγ expression, MMP-9 activity and disease, and the influence of therapy with anti-rheumatic drugs on these parameters. METHODS: Thirty RA patients of both sexes (treated with CSs and MTX, mainly) and 15 healthy volunteers were enrolled in this study. Disease severity was evaluated by the 28-joint disease activity score (DAS-28). Monocytes and monocyte-derived macrophages (MDMs) were isolated by standard procedures. PPARγ protein and mRNA expression were assessed by immunoblotting and real-time PCR, respectively; MMP-9 activity was determined by gelatin zymography. Moreover, we checked the ability of 15-deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ, a PPARγ agonist), MTX and methylprednisolone (MP) to affect PPARγ expression and lipopolysaccharide (LPS)-induced MMP-9 activity. RESULTS: Monocytes/MDMs from RA patients have significantly enhanced PPARγ expression (both protein and mRNA) and MMP-9 activity as compared with healthy donors. Interestingly, cells from patients with less active disease (DAS-28 <3.2) present higher PPARγ protein expression and lower MMP-9 activity than RA patients with DAS-28 >3.2. At therapeutic concentrations, MTX and MP increase in vitro PPARγ protein expression and inhibit LPS-induced MMP-9 activity. CONCLUSION: PPARγ expression in human monocytes/MDMs could represent an indicator of disease activity and therapy efficacy in RA because patients with a DAS-28 score <3.2 show the highest expression.


Assuntos
Antirreumáticos/uso terapêutico , Artrite Reumatoide/metabolismo , Macrófagos/metabolismo , Monócitos/metabolismo , PPAR gama/metabolismo , Artrite Reumatoide/tratamento farmacológico , Estudos de Casos e Controles , Quimioterapia Combinada , Feminino , Humanos , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Projetos Piloto , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Resultado do Tratamento , Adulto Jovem
14.
Pharmacol Res ; 62(5): 391-9, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20670683

RESUMO

Previous studies have shown that NCX 6550 (NCX), a nitric oxide (NO)-donating pravastatin, induces anti-inflammatory effects in murine macrophage cell lines. Here, we have studied its activity in human monocyte/macrophages, by investigating cytokine release, NF-κB translocation and peroxisome proliferator-activated receptor γ (PPARγ) expression and function. For comparison, pravastatin, isosorbide-5-mononitrate (ISMN), sodium nitroprusside (SNP) and the PPARγ ligand 15-deoxy-Δ(12,14)-prostaglandin J(2) (PGJ) were also tested. Monocytes and macrophages (MDM: monocyte-derived macrophages) were isolated from healthy donors; cytokine release was measured by ELISA, NF-κB by electrophoretic mobility shift assay and PPARγ by Western blot and Real-Time PCR. NCX (1 nM-50 µM) dose-dependently inhibited phorbol 12-myristate 13-acetate (PMA)-induced TNF-α release from monocytes (IC(50)=240 nM) and MDM (IC(50)=52 nM). At 50 µM, it was more effective than pravastatin, ISMN and SNP (P<0.05), but less efficient than PGJ. Similar results were obtained for IL-6. Likewise, NCX was more effective than pravastatin and the other NO donors in inhibiting PMA-induced NF-κB translocation in both cell types, and, at the highest concentration, significantly (P<0.05) enhanced PPARγ protein expression in monocytes. We conclude that NCX 6550 exerts a significant anti-inflammatory activity in human monocyte/macrophages, that is also contributed by its NO donating properties, as the effects exerted by NCX are significantly higher than those evoked by pravastatin in many experimental assays. These data further indicate that the incorporation of a NO-donating moiety into a statin structure confers pharmacological properties which may translate into useful therapeutic benefits.


Assuntos
Citocinas/metabolismo , Macrófagos/efeitos dos fármacos , Monócitos/efeitos dos fármacos , NF-kappa B/metabolismo , Nitratos/farmacologia , PPAR gama/metabolismo , Pravastatina/análogos & derivados , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Relação Dose-Resposta a Droga , Humanos , Interleucina-6/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Monócitos/metabolismo , NF-kappa B/antagonistas & inibidores , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/farmacologia , Pravastatina/farmacologia , Superóxidos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
15.
Atherosclerosis ; 211(1): 242-8, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20206356

RESUMO

OBJECTIVE: Atherosclerosis and restenosis are largely ruled by inflammation. The aim of this study was to test the effects of a short-course, high-dose oral prednisone on the release of interleukin-6 (IL-6) and tumour necrosis factor (TNF)-alpha from circulating monocytes and on the neointimal growth that follows bare metal stent (BMS) implantation. In a sub-group of patients activated NF-kappaB was also evaluated. METHODS: Out of 40 patients with coronary artery disease treated with BMS implantation, 20 were randomly assigned to receive oral prednisone during 40 days according to a standardized protocol. In non-stimulated and stimulated (LPS and PMA) monocytes we evaluated the release of IL-6 and TNF-alpha, and NF-kappaB p50 subunit translocation at baseline, at 10 and 30 days. Late luminal loss (LLL) 9 months after angioplasty was calculated by quantitative coronary angiography. RESULTS: Plasma concentrations of prednisone correlated inversely with IL-6 and TNF-alpha release (R2=0.45, p=0.04 and R2=0.69, p=0.005, respectively) and NF-kappaB activation from monocytes (R2=0.58, p=0.01). The reduction of TNF-alpha release and NF-kappaB activation were significantly related (R2=0.56, p=0.01). Prednisone patients showed a significantly larger reduction of cytokine release and NF-kappaB activation compared to non-treated patients, at 10 days and 30 days. LLL was lower in the prednisone group (0.44+/-0.35 mm versus 0.80+/-0.53 mm, p=0.02) and correlated with reduction of TNF-alpha (R2=0.41, p=0.01). CONCLUSIONS: High doses of oral prednisone reduce NF-kappaB pathway activation and pro-inflammatory cytokine release in circulating activated monocytes of patients treated with coronary stenting. TNF-alpha release reduction correlates with decreased LLL.


Assuntos
Doença da Artéria Coronariana/tratamento farmacológico , Reestenose Coronária/prevenção & controle , Citocinas/metabolismo , Interleucina-6/metabolismo , Monócitos/efeitos dos fármacos , NF-kappa B/metabolismo , Prednisona/uso terapêutico , Fator de Necrose Tumoral alfa/metabolismo , Idoso , Angioplastia Coronária com Balão , Doença da Artéria Coronariana/cirurgia , Feminino , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Stents
16.
Life Sci ; 81(11): 906-15, 2007 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-17765929

RESUMO

Previous observations demonstrated that Peroxisome Proliferator-Activated Receptor-gamma (PPAR-gamma), a key regulator of adipocyte differentiation, is expressed in a large variety of cells, including cells of the monocyte/macrophage lineage. This study was aimed to quantify both the constitutive and ligand-induced PPAR-gamma expression in monocytes and monocyte-derived macrophages (MDM) isolated from healthy smokers and non-smokers, and to evaluate the possible direct effect of nicotine. PPAR-gamma protein was detected by Western blot and quantification was performed by calculating the ratio between PPAR-gamma and beta-actin protein expression. Cytokine release was measured with enzyme-linked immunoassay kits. Constitutive PPAR-gamma protein was detected in human monocytes and its expression was up-regulated along with differentiation to MDM. The endogenous ligand 15-deoxy-delta(12,14)-prostaglandin J(2) and the synthetic agonist ciglitazone enhanced PPAR-gamma expression, the former being effective also at low micromolar concentrations. Both agonists significantly inhibited the basal secretion of pro-inflammatory cytokines (e.g., TNF-alpha, IL-6), ciglitazone being more potent. Monocytes and MDM from healthy smokers presented a significantly enhanced (4-fold and 2.5-fold, respectively) constitutive PPAR-gamma expression, as compared to those from healthy non-smokers. However, ligand-induced PPAR-gamma expression and inhibition of cytokine secretion were similar in healthy smokers and non-smokers. Nicotine dose-dependently enhanced PPAR-gamma expression with a maximum at 10 muM, and inhibited release of pro-inflammatory cytokines; these effects were reversed by alpha-bungarotoxin. Nicotine and PPAR-gamma agonists did not exert synergistic effects. In conclusion, monocytes and MDM from healthy smokers present a constitutively enhanced PPAR-gamma expression; this effect is reproduced, to some extent, by nicotine in vitro.


Assuntos
Regulação Enzimológica da Expressão Gênica , Macrófagos/metabolismo , Monócitos/metabolismo , Nicotina/farmacologia , PPAR gama/metabolismo , Fumar , Actinas/metabolismo , Adulto , Feminino , Humanos , Interleucina-6/metabolismo , Macrófagos/efeitos dos fármacos , Masculino , Monócitos/efeitos dos fármacos , Nicotina/efeitos adversos , Receptores Nicotínicos/metabolismo , Tiazolidinedionas/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima , Receptor Nicotínico de Acetilcolina alfa7
17.
Eur J Pharmacol ; 501(1-3): 199-208, 2004 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-15464079

RESUMO

Inhibition of tumor necrosis factor-alpha (TNF-alpha) represents a relevant target in rheumatoid arthritis therapy. Besides inhibiting cyclooxygenase, anti-inflammatory drugs can affect the activation of transcription factors. We investigated the ability of dexamethasone, indomethacin, and rofecoxib to modulate nuclear factor-kappaB (NF-kappaB) activation and TNF-alpha release from human monocytes challenged with lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA). Both stimuli induced NF-kappaB nuclear translocation and TNF-alpha secretion. Dexamethasone potently inhibited TNF-alpha release, indomethacin inhibited only PMA-evoked release, while rofecoxib had no effect. In the electrophoretic mobility shift assay, dexamethasone and rofecoxib dose-dependently inhibited the DNA binding activity of NF-kappaB in stimulated monocytes, whereas indomethacin failed to inhibit the LPS-evoked one. These results were further confirmed by evaluating the drugs' ability to reduce nuclear NF-kappaB subunits, as well as the amount of phosphorylated IkappaBalpha in cytosolic fractions. In conclusion, these results indicate that anti-inflammatory drugs differ largely in their ability to inhibit NF-kappaB activity and/or TNF-alpha release from human monocytes. These effects can be relevant to rheumatoid arthritis therapy.


Assuntos
Anti-Inflamatórios/farmacologia , Artrite Reumatoide/tratamento farmacológico , Monócitos/efeitos dos fármacos , NF-kappa B/fisiologia , Fator de Necrose Tumoral alfa/biossíntese , Adulto , Anti-Inflamatórios/uso terapêutico , Artrite Reumatoide/metabolismo , Relação Dose-Resposta a Droga , Humanos , Pessoa de Meia-Idade , Monócitos/metabolismo
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