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Diffractive optical elements that divide an input beam into a set of replicas are used in many optical applications ranging from image processing to communications. Their design requires time-consuming optimization processes, which, for a given number of generated beams, are to be separately treated for one-dimensional and two-dimensional cases because the corresponding optimal efficiencies may be different. After generalizing their Fourier treatment, we prove that, once a particular divider has been designed, its transmission function can be used to generate numberless other dividers through affine transforms that preserve the efficiency of the original element without requiring any further optimization.
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Principal research on energy from thermonuclear fusion uses Deuterium-Tritium plasmas magnetically trapped in toroidal devices. As major scientific problem for an economic (i.e., really feasible) reactor, we must understand how to lead strongly heated plasmas to sustain a high fusion gain while large fraction of current is self-produced via the presence of strong pressure gradient. To suppress turbulent eddies that impair thermal insulation and pressure tight of the plasma, current drive (CD) is necessary. However, tools envisaged so far in ITER (International Thermonuclear Experiment Rector) are unable accomplishing this task that requires efficiently and flexibly matching the natural current profiles of plasma. Consequently, viability of a thermonuclear reactor should be problematic. Multi-megawatt radio-frequency (RF) power coupled to plasma would produce the necessary CD, but modelling results based on previous understanding found difficult the extrapolation of this CD concept to reactor conditions of high temperature plasma, and greater flexibility of method would also be required. Here we present new model results based on standard quasilinear (QL) theory that allow establish conditions to drive efficiently and flexibly the RF-driven current at large radii of the plasma column, as necessary for the goal of a reactor.
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Progress in thermonuclear fusion energy research based on deuterium plasmas magnetically confined in toroidal tokamak devices requires the development of efficient current drive methods. Previous experiments have shown that plasma current can be driven effectively by externally launched radio frequency power coupled to lower hybrid plasma waves. However, at the high plasma densities required for fusion power plants, the coupled radio frequency power does not penetrate into the plasma core, possibly because of strong wave interactions with the plasma edge. Here we show experiments performed on FTU (Frascati Tokamak Upgrade) based on theoretical predictions that nonlinear interactions diminish when the peripheral plasma electron temperature is high, allowing significant wave penetration at high density. The results show that the coupled radio frequency power can penetrate into high-density plasmas due to weaker plasma edge effects, thus extending the effective range of lower hybrid current drive towards the domain relevant for fusion reactors.
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The amyloid cascade hypothesis sustains that beta-amyloid (Abeta) is the main pathogenetic factor of Alzheimer's Disease (AD). Although the direct and indirect neurotoxic role of Abeta are unchallenged, recent findings suggest that the peptide may have so far unforeseen physiological roles. In this regard, the observations showing the ability of Abeta to exert synaptic activities in absence of neurotoxicity are very intriguing. In particular, the peptide is able to affect synaptic transmission of different neurotransmitter systems in key brain areas that regulate executive and cognitive functions, an observation that points Abeta as a new neuromodulator. However, in a pathological context, Abeta may drive functional alterations of several neurotransmitter systems in the early phases of the disease, in turn producing subtle cognitive and behavioural disturbances in addition and before the well known neurodegenerative events. On the other hand, advancing age is the most significant risk factor for the development of AD. In fact, during aging increased Abeta levels have been reported. Moreover, several neurotransmitter systems undergo age-related changes in parallel to a decline of cognitive functions. However, the putative neuromodulatory role of Abeta in the context of aging is nowadays unknown. For these reasons, future studies about the spectrum of action of Abeta (brain areas and neurotransmitter systems affected) are particularly interesting since may suggest new therapeutic targets in order to sustain those functions which may be altered during aging.
Assuntos
Envelhecimento/metabolismo , Peptídeos beta-Amiloides/metabolismo , Peptídeos beta-Amiloides/fisiologia , Sistemas de Liberação de Medicamentos/tendências , Neurotransmissores/metabolismo , Sinapses/fisiologia , Envelhecimento/efeitos dos fármacos , Envelhecimento/fisiologia , Animais , Humanos , Preparações Farmacêuticas/administração & dosagem , Preparações Farmacêuticas/metabolismo , Sinapses/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Transmissão Sináptica/fisiologiaRESUMO
In recent years, minimally invasive surgical techniques for lumbar fusion and fixation procedures gained worldwide popularity. Herein we describe a personal technique for percutaneous lumbar interbody fusion associated with minimally invasive posterior fixation for patients affected by degenerative disc disease and lumbar instability. The procedure is described in a step-by-step way and early results are presented. Although the present data reflect only an early experience, we believe that this is a straightforward procedure which may be more advantageous in terms of surgical invasiveness, potentially saving operative and recovery time and reducing risks compared to posterior or anterior approaches for lumbar interbody fusion.
Assuntos
Parafusos Ósseos , Degeneração do Disco Intervertebral/cirurgia , Instabilidade Articular/cirurgia , Vértebras Lombares/cirurgia , Procedimentos Cirúrgicos Minimamente Invasivos/métodos , Fusão Vertebral/métodos , Adulto , Idoso , Feminino , Humanos , Fixadores Internos , Masculino , Pessoa de Meia-Idade , Procedimentos Cirúrgicos Minimamente Invasivos/instrumentação , Procedimentos Neurocirúrgicos/instrumentação , Procedimentos Neurocirúrgicos/métodos , Estudos Retrospectivos , Fusão Vertebral/instrumentação , Resultado do TratamentoRESUMO
The transition of prion protein from a mainly alpha-structured isoform (PrPC) to a beta sheet-containing protein (PrPSc) represents a major pathogenetic mechanism in prion diseases. To study the role of PrP structural conformation in prion-dependent neurodegeneration, we analysed the neurotoxicity of PrP in alpha and beta conformations, using a recombinant protein encompassing amino acids 90-231 of the human PrP (hPrP90-231). Using controlled thermal denaturation (53 degrees C, 1h) we converted hPrP90-231 in a structural isoform displaying PrPSc-related characteristics: high beta sheet content, increased aggregability and a slight increase in the resistance to protease K. In virtue of these structural changes, hPrP90-231 powerfully affected the survival of SH-SY5Y cells, inducing a caspase-3 and p38- dependent apoptosis. Conversely, in the native alpha-helix-rich conformation, hPrP90-231 did not show significant cell toxicity. The relationship between the structural state of hPrP90-231 and its neurotoxicity was demonstrated, inducing the thermal denaturation of the peptide in the presence of Congo red that prevented both the transition of hPrP90-231 into a beta-rich isoform and the acquisition of toxic properties. In conclusion, we report that the toxicity of hPrP90-231 is dependent on its three-dimensional structure, as is supposed to occur for the pathogen PrP during TSE.
Assuntos
Apoptose/efeitos dos fármacos , Proteínas PrPC/química , Proteínas PrPC/farmacologia , Amiloide/biossíntese , Benzotiazóis , Caspase 3 , Caspase 7 , Caspases/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Dicroísmo Circular , Endopeptidase K/química , Corantes Fluorescentes , Humanos , Hidrólise , Immunoblotting , L-Lactato Desidrogenase/metabolismo , Microscopia Eletrônica , Necrose , Conformação Proteica , Desnaturação Proteica , Estrutura Secundária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacologia , Relação Estrutura-Atividade , Sais de Tetrazólio , Tiazóis/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: Docetaxel has a proven significant activity against breast, non-small cell lung, ovarian, head and neck, and hormone refractory prostate cancer. Preclinical pharmacokinetic studies have shown that hepatobiliary extraction is the major route of elimination. We conducted this study to elucidate the feasibility and safety of the use of docetaxel in hemodialysis patients. PATIENT AND METHODS: In a 72-year-old hormone refractory prostate cancer patient on hemodialysis for diabetic nephropathy for 3 years, a first dose (35 mg/m(2) iv) of docetaxel was completed 30 min before starting dialysis, while a second dose was administered 30 min after completion of a different hemodialysis session. Pharmacokinetic analysis was performed following both infusions. RESULTS: No apparent differences could be seen in the plasma concentration-time curves of docetaxel administered before or after dialysis. The patient experienced no significant toxicity after either administration of docetaxel. CONCLUSIONS: Docetaxel is safe in dialysis patients and does not require dose reduction. Dialysis does not remove this drug from blood.
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Antineoplásicos Fitogênicos/farmacocinética , Neoplasias da Próstata/metabolismo , Diálise Renal , Taxoides/farmacocinética , Idoso , Antineoplásicos Fitogênicos/administração & dosagem , Cromatografia Líquida de Alta Pressão , Nefropatias Diabéticas/complicações , Nefropatias Diabéticas/terapia , Docetaxel , Esquema de Medicação , Soluções para Hemodiálise/análise , Humanos , Masculino , Neoplasias da Próstata/complicações , Taxoides/administração & dosagemRESUMO
Peptides corresponding to three alpha helices present in the C-terminal region of the human prion protein have been synthesized and their structural autonomy analyzed by circular dichroism (CD) and NMR spectroscopy. The results obtained indicate that the protein fragment corresponding to the alpha 3-helix, in contrast to alpha 1 and alpha 2 peptides, shows a complete structural autonomy. The chemical shifts values found for NH and CHalpha resonance of the isolated alpha 3 peptide, formed by 30 aminoacid residues, were markedly and surprisingly similar to the corresponding values of the alpha 3-helix in the protein. The structural autonomy of the alpha 3-helix is profoundly determined by the presence of the conserved capping box and, in part, by the ionic bond formed between Glu200 and Lys204. On the basis of these observations a novel PrP consensus pattern, centered on the alpha 3-helix region, has been defined. The data indicate that this autonomous and highly conserved region of the PrPc likely plays a critical role in folding and stability. This gives an explanation of why many of pathogenic mutations occur in this part of the molecule, sharing relevant effects on the overall protein conformation. In particular the D202N capping mutation almost completely destabilizes the isolated alpha 3 peptide. While it is well known that the D202N substitution is associated with a GSS disease, the possible structural basis of this fatal pathology has never been investigated. We propose that a lower alpha 3-helical propensity leading to a major destabilization of the PrPc molecule initiates the pathogenic process associated with D202N capping mutation.
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Mutação/fisiologia , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Príons/genética , Príons/metabolismo , Sequência de Aminoácidos , Animais , Dicroísmo Circular , Humanos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Conformação Molecular , Dados de Sequência Molecular , Fragmentos de Peptídeos/síntese química , Príons/síntese química , Conformação ProteicaRESUMO
Low levels of lignans, namely enterolactone, have been reported to be associated with an increased risk of breast cancer in the general female population. We assessed, retrospectively, the relationship between serum enterolactone concentrations and the occurrence of breast cancer in women with palpable cysts. The levels of enterolactone in cryopreserved serum aliquots, obtained from 383 women with palpable cysts at the time of their first cyst aspiration, were measured using a time-resolved fluoroimmunoassay (TR-FIA). After a median follow-up time of 6.5 years (range 0.5-12.75 years), 18 women were found to have developed an invasive breast cancer. Median values of serum enterolactone were significantly lower in women who subsequently developed breast cancer: 8.5 nM/l versus 16.0 nM/l: P=0.04. Odd Ratios (OR) for breast cancer were: 0.36 (P=0.03), 0.57 (P=0.3) and 0.38 (P=0.25) for 25th (8 nM/l), 50th (16 nM/l) and 75th (24 nM/l) percentile values, respectively. The receiver operating characteristic (ROC) analysis showed a satisfactory accuracy for enterolactone as a breast cancer risk indicator (area under the curve (AUC)=0.64: P=0.04). Logistic regression analysis confirmed that the enterolactone concentration had a strong protective effect on the breast cancer risk. These findings may have important clinical implications with regard to interventional diet-focused chemo-preventive trials.
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4-Butirolactona/análogos & derivados , 4-Butirolactona/sangue , Biomarcadores Tumorais/sangue , Neoplasias da Mama/etiologia , Doença da Mama Fibrocística/sangue , Lignanas/sangue , Adulto , Idoso , Neoplasias da Mama/sangue , Feminino , Seguimentos , Humanos , Pessoa de Meia-Idade , Análise de Regressão , Estudos Retrospectivos , Fatores de Risco , SoroRESUMO
The present work shows that alpha-glycerylphosphorylethanolamine (alpha-GPE) is effective in recovering astrocytes from mitochondrial membrane integrity and potential derangement and cellular oxidative stress that occur under amyloid beta-peptides-induced reactive gliosis.alpha-Glycerylphosphorylethanolamine (alpha-GPE), a new compound with nootropic properties, known to improve in vivo the learning and memory processes, has been tested for its protective properties on an in vitro model of degeneration. Rat primary astrocytic cultures treated with two amyloid-derived peptides, Abeta((1-40)) and Abeta(3(pE)-42), showed a marked reduction of the mitochondrial redox activity and membrane potential, together with an increase of oxidative species production. Plasma membrane lipid peroxidation (LPO) as well as generation of peroxides is greatly increased under Abeta-peptides toxicity. These features, typical of the reactive gliosis that accompanies neuronal degeneration, were readily recovered by pretreatment with alpha-GPE. alpha-GPE, likely improving the fluidity of cell membrane, has the potential to recover astrocytes from the general redox derangement induced by different amyloid fragments and possibly to protect from inflammation, gliosis and neurodegeneration. This is the first evidence of an antioxidant effect of the ethanolamine derivative on a rat model of chronic gliosis.
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Peptídeos beta-Amiloides/antagonistas & inibidores , Peptídeos beta-Amiloides/toxicidade , Astrócitos/efeitos dos fármacos , Mitocôndrias/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Fragmentos de Peptídeos/antagonistas & inibidores , Fragmentos de Peptídeos/toxicidade , Fosfatidiletanolaminas/farmacologia , Animais , Membrana Celular/efeitos dos fármacos , Membrana Celular/ultraestrutura , Células Cultivadas , Gliose/induzido quimicamente , Gliose/patologia , Peroxidação de Lipídeos/efeitos dos fármacos , Fluidez de Membrana/efeitos dos fármacos , Potenciais da Membrana/efeitos dos fármacos , Microscopia Confocal , Mitocôndrias/efeitos dos fármacos , Degeneração Neural/patologia , Ratos , Sais de Tetrazólio , TiazóisRESUMO
The Src homology 2-containing tyrosine phosphatase, Shp-2, is a crucial enzyme that mediates intracellular signaling and is implicated in cell proliferation and differentiation. Here we investigated the involvement of the Shp-2 tyrosine phosphatase in determining the downstream signaling pathways initiated by the Ret oncogene, carrying either the cysteine 634 to tyrosine or the methionine 918 to threonine substitutions. These mutations convert the receptor tyrosine kinase, Ret, into a dominant transforming protein and induce constitutive activation of its intrinsic tyrosine kinase activity leading to congenital and sporadic cancers in neuroendocrine organs. Using the PC12, rat pheochromocytoma cell line, as model system, we show that Shp-2 mediates immediate-early gene expression if induced by either of the mutant alleles. Furthermore, we show that Shp-2 activity is required for RetM918T-induced Akt activation. The results indicate that Shp-2 is a downstream mediator of the mutated receptors RetC634Y and RetM918T, thus suggesting that it may act as a limiting factor in Ret-associated endocrine tumors, in the neoplastic syndromes multiple endocrine neoplasia types 2A and 2B.
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Membranas Intracelulares/fisiologia , Mutação/fisiologia , Proteínas Oncogênicas/genética , Proteínas Tirosina Fosfatases/metabolismo , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais/fisiologia , Animais , Diferenciação Celular/fisiologia , Linhagem Celular , Sobrevivência Celular/fisiologia , Fator Neurotrófico Derivado de Linhagem de Célula Glial , Peptídeos e Proteínas de Sinalização Intracelular , Fatores de Crescimento Neural/metabolismo , Proteínas Oncogênicas/metabolismo , Células PC12/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 11 , Proteínas Proto-Oncogênicas c-ret , Ratos , Receptores Proteína Tirosina Quinases/metabolismoRESUMO
Brain deposition of the amyloid-beta protein (Abeta) is a frequent complication of Down's syndrome (DS) patients. Abeta peptide is generated by endoproteolytic processing of Abeta precursor protein by gamma and beta secretases. Recently a transmembrane aspartyl protease, BACE, has been identified as the beta-secretase, and its homologous BACE-2 has also been described. BACE-2 gene resides on chromosome 21 in the obligate DS region. It cleaves Abeta precursor protein at its beta site and more efficiently at a different site within Abeta. In the present study we characterized the BACE-2 gene and protein expression in the DS patients and healthy control. We analyzed, by using a nonradioactive ribonuclease protection assay, the levels of BACE-2 mRNA expression in primary skin fibroblasts. The analysis revealed a 2.6-fold increase in BACE-2 mRNA levels in the DS group compared to the levels observed in the control group. Western blot analysis revealed no difference between DS and control in BACE-2 protein levels in the intracellular compartment. In the medium conditioned by fibroblast, we revealed an evident secretion of BACE-2 protein, represented by two different molecular weights, remarkably increased in DS fibroblasts. BACE-2 overexpression was also confirmed in the DS fetal brains and human neural embryonic DS stem cells in which conditioned media BACE-2 was secreted. These data highlight the importance of the extracellular compartment where BACE-2 overexpression could play a role in plaque formation in DS patients.
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Ácido Aspártico Endopeptidases/biossíntese , Encéfalo/enzimologia , Síndrome de Down/enzimologia , Fibroblastos/enzimologia , Células-Tronco/enzimologia , Adulto , Secretases da Proteína Precursora do Amiloide , Ácido Aspártico Endopeptidases/genética , Ácido Aspártico Endopeptidases/metabolismo , Western Blotting , Encéfalo/embriologia , Química Encefálica , Células Cultivadas , Meios de Cultivo Condicionados/química , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Peso Molecular , Neurônios/citologia , Neurônios/enzimologia , RNA Mensageiro/análise , RNA Mensageiro/biossíntese , Valores de Referência , Células-Tronco/citologia , Células-Tronco/metabolismoRESUMO
The authors review the available literature on the preclinical and clinical studies involving GABAergic neurotransmission in mood disorders. Gamma-aminobutyric acid (GABA) is an inhibitory neurotransmitter present almost exclusively in the central nervous system (CNS), distributed across almost all brain regions, and expressed in interneurons modulating local circuits. The role of GABAergic dysfunction in mood disorders was first proposed 20 years ago. Preclinical studies have suggested that GABA levels may be decreased in animal models of depression, and clinical studies reported low plasma and CSF GABA levels in mood disorder patients. Also, antidepressants, mood stabilizers, electroconvulsive therapy, and GABA agonists have been shown to reverse the depression-like behavior in animal models and to be effective in unipolar and bipolar patients by increasing brain GABAergic activity. The hypothesis of reduced GABAergic activity in mood disorders may complement the monoaminergic and serotonergic theories, proposing that the balance between multiple neurotransmitter systems may be altered in these disorders. However, low GABAergic cortical function may probably be a feature of a subset of mood disorder patients, representing a genetic susceptibility. In this paper, we discuss the status of GABAergic hypothesis of mood disorders and suggest possible directions for future preclinical and clinical research in this area.
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Química Encefálica/fisiologia , Transtornos do Humor/fisiopatologia , Ácido gama-Aminobutírico/fisiologia , HumanosRESUMO
BACKGROUND: Anthracycline cardiotoxicity is increased by the contemporaneous administration of trastuzumab. The mechanism by which it occurs is as yet unknown. The aim of this study was to evaluate whether trastuzumab modifies the pharmacokinetics of epirubicin and its metabolites. PATIENTS AND METHODS: Women with HER2-positive metastatic breast cancer were treated with epirubicin 75 mg/m(2) i.v. bolus followed by docetaxel 75 mg/m(2) in a 1-h infusion, every 3 weeks for six cycles, and trastuzumab (once at 4 mg/m(2), then 2 mg/m(2) weekly thereafter) in a 30-min infusion. Epirubicin pharmacokinetic data of seven patients were evaluated at the first cycle of therapy (baseline, with trastuzumab administered 24 h after epirubicin), and at the sixth cycle (i.e. 15 weeks after baseline, with trastuzumab administered immediately before epirubicin). RESULTS: No pharmacokinetic change in the parent compound epirubicin was detected. The area under the plasma concentration-time curve (AUC(0-24 h)) was 1230 +/- 318 [mean +/- standard deviation (SD)] at the first cycle and 1287 +/- 385 h. micro g/l at the sixth. The mean (+/-SD) maximum plasma concentration (C(max)) and the terminal elimination half-life at the first cycle (1303 +/- 490 micro g/l and 12.5 +/- 3.1 h, respectively) were similar to those obtained at the sixth cycle (1229 +/- 580 micro g/l and 11.5 +/- 2.9 h, respectively). Pharmacokinetic data of epirubicin metabolites evaluated at the first and sixth cycle of chemotherapy were superimposable without any statistical difference. CONCLUSION: Enhanced anthracycline cardiotoxicity related to trastuzumab administration was not linked to pharmacokinetic interferences with epirubicin and its metabolites.
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Anticorpos Monoclonais/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias da Mama/tratamento farmacológico , Doenças Cardiovasculares/induzido quimicamente , Epirubicina/farmacocinética , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/secundário , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Relação Dose-Resposta a Droga , Esquema de Medicação , Interações Medicamentosas , Epirubicina/administração & dosagem , Feminino , Seguimentos , Humanos , Infusões Intravenosas , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/secundário , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Probabilidade , Estudos Prospectivos , Receptor ErbB-2/análise , Medição de Risco , Neoplasias de Tecidos Moles/tratamento farmacológico , Neoplasias de Tecidos Moles/secundário , Estatísticas não Paramétricas , Taxa de Sobrevida , Trastuzumab , Resultado do TratamentoRESUMO
Somatostatin was reported to inhibit Kaposi's sarcoma (KS) cell (KS-Imm) xenografts through an antiangiogenic activity. Here, we show that somatostatin blocks growth of established KS-Imm tumors with the same efficacy as adriamycin, a clinically effective cytotoxic drug. Whereas KS-Imm cells do not express somatostatin receptors (SSTRs), endothelial cells express several SSTRs, in particular SSTR3. We investigated the molecular mechanisms and receptor specificity of somatostatin inhibition of angiogenesis. Somatostatin significantly inhibited angiogenesis in vivo in the matrigel sponge assay; this inhibition was mimicked by the SSTR3 agonist L-796778 and reversed by the SSTR3 antagonist BN81658, demonstrating involvement of SSTR3. In vitro experiments showed that somatostatin directly affected different endothelial cell line proliferation through a block of growth-factor-stimulated MAPK and endothelial nitric oxide (NO) synthase (eNOS) activities. BN81658 reversed somatostatin inhibition of cell proliferation, NO production, and MAPK activity, indicating that SSTR3 activation is required for the effects of somatostatin in vitro. Finally in vivo angiogenesis assays demonstrated that eNOS inhibition was a prerequisite for the antiangiogenic effects of somatostatin, because high concentrations of sodium nitroprusside, an NO donor, abolished the somatostatin effects. In conclusion, we demonstrate that somatostatin is a powerful antitumor agent in vivo that inhibits tumor angiogenesis through SSTR3-mediated inhibition of both eNOS and MAPK activities.
Assuntos
Hormônios/farmacologia , Neovascularização Patológica/tratamento farmacológico , Óxido Nítrico Sintase/metabolismo , Receptores de Somatostatina/metabolismo , Somatostatina/farmacologia , Amidas/farmacologia , Animais , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Endotélio Vascular/citologia , Endotélio Vascular/enzimologia , Ativação Enzimática/efeitos dos fármacos , Humanos , Técnicas In Vitro , Camundongos , Camundongos Nus , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Óxido Nítrico Sintase Tipo II , Óxido Nítrico Sintase Tipo III , Nitrobenzenos/farmacologia , Receptores de Somatostatina/agonistas , Receptores de Somatostatina/antagonistas & inibidores , Veias Umbilicais/citologiaRESUMO
Prion diseases are neurodegenerative disorders of the central nervous system of humans and animals, characterized by spongiform degeneration of the central nervous system, astrogliosis, and deposition of amyloid into the brain. The conversion of a cellular glycoprotein (prion protein, PrP(C)) into an altered isoform (PrP(Sc)) has been proposed to represent the causative event responsible for these diseases. The peptide corresponding to the residues 106-126 of PrP sequence (PrP106-126) is largely used to explore the neurotoxic mechanisms underlying the prion diseases. We investigated the intracellular signaling responsible for PrP106-126-dependent cell death in the SH-SY5Y human neuroblastoma cell line. In these cells, PrP106-126 treatment induced apoptotic cell death and the activation of caspase-3. The p38 MAP-kinase blockers (SB203580 and PD169316) prevented the apoptotic cell death evoked by PrP106-126 and Western blot analysis revealed that the exposure of the cells to the peptide induced p38 activation. However, whether the neuronal toxicity of PrP106-126 is caused by a soluble or fibrillar form of this peptide is still unknown. In this study, we correlated the structural state of this peptide with its neurotoxicity. We show that the two conserved glycines in position 114 and 119 prevent the peptide to assume a structured conformation, favoring its aggregation in amyloid fibrils. The substitution of both glycines with alanine residues (PrP106-126AA) generates a soluble nonamyloidogenic peptide, that retained its toxic properties when incubated with neuroblastoma cells. These data show that the amyloid aggregation is not necessary for the induction of the toxic effects of PrP106-126.
Assuntos
Peptídeos beta-Amiloides/biossíntese , Apoptose/fisiologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fragmentos de Peptídeos/farmacologia , Príons/farmacologia , Sequência de Aminoácidos , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Dados de Sequência Molecular , Neuroblastoma/patologia , Fragmentos de Peptídeos/química , Príons/química , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
BACKGROUND: Previously we observed a pharmacokinetic interference of epirubicin elimination when paclitaxel is given in combination in a sequence-dependent manner (i.e. when paclitaxel is administered as first drug). The aim of this study was to determine whether these sequence-dependent pharmacological effects were also evident when epirubicin was combined with docetaxel. PATIENTS AND METHODS: Patients who received epirubicin 75 mg/m2 or 90 mg/m2 as an intravenous bolus followed immediately by docetaxel 70 mg/m2 or 80 mg/m2 over a 1-h infusion, or the opposite sequence, every 3 weeks were eligible for this study. The pharmacokinetics of docetaxel, epirubicin and its metabolites were studied at the first and second cycle of treatment. Pharmacokinetic data were normalised to the lower dose of each drug. Toxicity was recorded at nadir and graded according to National Cancer Institute Common Toxicity Criteria. RESULTS: Twelve consecutive patients, each acting as their own control, entered the study. The sequence of drug administration of docetaxel and epirubicin did not affect the pharmacokinetics of the parent anthracycline. Statistically significant lower glucuronidation metabolism of epirubicin was observed in patients who received docetaxel before epirubicin. The pharmacokinetics of docetaxel were not influenced by the sequence of drug administration. No difference in haematological and non-haematological toxicity was observed in the two sequences of treatment. CONCLUSIONS: The pharmacokinetics of the parent anthracycline and of docetaxel were similar between the two schemes of treatment. The metabolic variations observed, i.e. differences in the plasma levels of epirubicin glucuronides, seem not to have clinical relevance.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Epirubicina/administração & dosagem , Paclitaxel/análogos & derivados , Paclitaxel/administração & dosagem , Taxoides , Docetaxel , Epirubicina/efeitos adversos , Epirubicina/farmacocinética , Feminino , Humanos , Paclitaxel/efeitos adversos , Paclitaxel/farmacocinéticaRESUMO
The present findings show that an atypical non-steroidal anti-inflammatory drug, such as acetaminophen, retains the ability to recover amyloid beta-peptides driven neuronal apoptosis through the impairment of oxidative stress. Moreover, this compound reduces the increased NF-kappaB binding activity, which occurs in these degenerative conditions. Therapeutic interventions aimed at reducing the inflammatory response in Alzheimer's disease (AD) recently suggested the application of non-steroidal anti-inflammatory drugs. Although the anti-inflammatory properties of acetaminophen are controversial, it emerged that in an amyloid-driven astrocytoma cell degeneration model acetaminophen proved to be effective. On these bases, we analyzed the role of acetaminophen against the toxicity exerted by different Abeta-peptides on rat primary hippocampal neurons and on a rat pheochromocytoma cell line. We found a consistent protection from amyloid beta-fragments 1-40 and 1-42-induced impairment of mitochondrial redox activity on both cell cultures, associated with a marked reduction of apoptotic nuclear fragmentation. An antioxidant component of the protective activity emerged from the analysis of the reduction of phospholipid peroxidation, and also from a significant reduction of cytoplasmic accumulation of peroxides in the pheochromocytoma cell line. Moreover, activation of NF-kappaB by amyloid-derived peptides was greatly impaired by acetaminophen pre-treatment in hippocampal cells. This evidence points out antioxidant and anti-transcriptional properties of acetaminophen besides the known capability to interfere with inflammation within the central nervous system, and suggests that it can be exploited as a possible therapeutic approach against AD.
Assuntos
Acetaminofen/farmacologia , Peptídeos beta-Amiloides/metabolismo , Hipocampo/efeitos dos fármacos , NF-kappa B/metabolismo , Estresse Oxidativo , Fragmentos de Peptídeos/metabolismo , Animais , Apoptose/efeitos dos fármacos , Ensaio de Desvio de Mobilidade Eletroforética , Hipocampo/citologia , Neurônios/efeitos dos fármacos , Células PC12 , RatosRESUMO
Prion diseases are fatal neurodegenerative disorders of the CNS of men and animals, characterized by spongiform degeneration of the CNS, astrogliosis and deposition of amyloid into the brain. The conversion of a cellular glycoprotein (the prion protein, PrP(C)) into an altered isoform (the prion scrapie, PrP(Sc)), which accumulates within the brain tissue by virtue of its resistance to the intracellular catabolism, is currently believed to represent the etiologic agent responsible for these diseases. Synthetic or recombinant polypeptides are commonly used to elucidate the mechanism of proteins involved in neurodegenerative diseases. Here we describe a procedure, which allows the synthesis and purification in its native folding, of the human prion protein fragment 90-231, corresponding to the protease resistant core of PrP(Sc). We synthesized the polypeptides 90-231 of both the wild type and the E200K mutant isoforms of PrP. Using a gluthatione S-transferase (GST) fusion protein approach, milligram amounts of polypeptides were obtained after expression in E. coli. The recovery of the purified fusion protein was monitored following the evaluation of the GST activity. The PrP fragment was released from the fusion protein immobilized on a glutathione-coupled agarose resin by direct cleavage with thrombin. The recombinant protein was identified by comassie stained acrylamide gel and by immunoblotting employing a monoclonal anti-PrP antibody. The peptide purified by gel filtration chromatography showed mainly an alpha-helix structure, as analysed by circular dichroism (CD) and an intact disulfide bridge. The same procedure was also successfully employed to synthesize and purify the E200K mutant PrP fragment.