RESUMO
BACKGROUND: Neuroblastoma is the most common extracranial solid malignancy in childhood, responsible for 15% of all pediatric cancer deaths. It is an heterogeneous disease that does not always respond to classical therapy; so the identification of new and specific molecular targets to improve existing therapy is needed. We have previously demonstrated the involvement of the Notch pathway in the onset and progression of neuroblastoma. In this study we further investigated the role of Notch signaling and identified Delta-like 1 (DLL1) as a novel molecular target in neuroblastoma cells with a high degree of MYCN amplification, which is a major oncogenic driver in neuroblastoma. The possibility to act on DLL1 expression levels by using microRNAs (miRNAs) was assessed. METHODS: DLL1 mRNA and protein expression levels were measured in three different neuroblastoma cell lines using quantitative real-time PCR and Western Blot analysis, respectively. Activation of the Notch pathway as a result of increased levels of DLL1 was analyzed by Immunofluorescence and Western Blot methods. In silico tools revealed the possibility to act on DLL1 expression levels with miRNAs, in particular with the miRNA-34 family. Neuroblastoma cells were transfected with miRNA-34 family members, and the effect of miRNAs transfection on DLL1 mRNA expression levels, on cell differentiation, proliferation and apoptosis was measured. RESULTS: In this study, the DLL1 ligand was identified as the Notch pathway component highly expressed in neuroblastoma cells with MYCN amplification. In silico analysis demonstrated that DLL1 is one of the targets of miRNA-34 family members that maps on chromosome regions that are frequently deregulated or deleted in neuroblastoma. We studied the possibility to use miRNAs to target DLL1. Among all miRNA-34 family members, miRNA-34b is able to significantly downregulate DLL1 mRNA expression levels, to arrest cell proliferation and to induce neuronal differentiation in malignant neuroblastoma cells. CONCLUSIONS: Targeted therapies have emerged as new strategies for cancer treatment. This study identified the Notch ligand DLL1 as a novel and attractive molecular target in childhood neuroblastoma and its results could help to devise a targeted therapy using miRNAs.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteínas de Membrana/genética , MicroRNAs/genética , Terapia de Alvo Molecular , Neuroblastoma/genética , Proteínas de Ligação ao Cálcio , Diferenciação Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Criança , Pré-Escolar , Regulação Neoplásica da Expressão Gênica , Humanos , Ligantes , Neuroblastoma/patologia , Neuroblastoma/terapia , Receptores Notch/genética , Transdução de Sinais/genéticaRESUMO
The beneficial effects of hot springs have been known for centuries and treatments with sulphurous thermal waters are recommended in a number of chronic pathologies as well as acute recurrent infections. However, the positive effects of the therapy are often evaluated in terms of subjective sense of wellbeing and symptomatic clinical improvements. Here, the effects of an S-based compound (NaSH) and of a specific sulphurous thermal water characterized by additional ions such as sodium chloride, bromine and iodine (STW) were investigated in terms of cytokine release and anti-oxidant enzyme activity in primary human monocytes and in saliva from 50 airway disease patients subjected to thermal treatments. In vitro, NaSH efficiently blocked the induction of pro-inflammatory cytokines and counterbalanced the formation of ROS. Despite STW not recapitulating these results, possibly due to the low concentration of S-based compounds reached at the minimum non-toxic dilution, we found that it enhanced the release of IL-10, a potent anti-inflammatory cytokine. Notably, higher levels of IL-10 were also observed in patients' saliva following STW treatment and this increase correlated positively with salivary catalase activity (r2 = 0.19, *p less than 0.01). To our knowledge, these results represent the first evidence suggesting that S-based compounds and STW may prove useful in facing chronic inflammatory and age-related illness due to combined anti-inflammatory and anti-oxidant properties.
Assuntos
Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Balneologia , Enzimas/metabolismo , Fontes Termais , Inflamação/terapia , Interleucina-10/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Águas Minerais , Doenças Respiratórias/terapia , Adulto , Idoso , Catalase/metabolismo , Células Cultivadas , Feminino , Glutationa Peroxidase/metabolismo , Humanos , Inflamação/enzimologia , Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Itália , Leucócitos Mononucleares/enzimologia , Leucócitos Mononucleares/imunologia , Masculino , Pessoa de Meia-Idade , NADPH Oxidases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Doenças Respiratórias/enzimologia , Doenças Respiratórias/imunologia , Saliva/enzimologia , Saliva/imunologia , Sulfetos/farmacologia , Superóxido Dismutase/metabolismo , Fatores de TempoRESUMO
Methylglyoxal (MG), a potent glycotoxin that can be found in the diet, is one of the main precursors of Advanced glycation end products (AGEs). It is well known that modifications in lifestyle such as nutritional interventions can be of great value for preventing brain deterioration. This study aimed to evaluate in vivo how an oral MG treatment, that mimics a high MG dietary intake, could affect brain health. From our results, we demonstrated that MG administration affected working memory, and induced neuroinflammation and oxidative stress by modulating the Receptor for Advanced glycation end products (RAGE). The gene and protein expressions of RAGE were increased in the hippocampus of MG mice, an area where the activity of glyoxalase 1, one of the main enzymes involved in MG detoxification, was found reduced. Furthermore, at hippocampus level, MG mice showed increased expression of proinflammatory cytokines and increased activities of NADPH oxidase and catalase. MG administration also increased the gene and protein expressions of Presenilin-1, a subunit of the gamma-secretase protein complex linked to Alzheimer's disease. These findings suggest that high MG oral intake induces alteration directly in the brain and might establish an environment predisposing to AD-like pathological conditions.
Assuntos
Encéfalo/efeitos dos fármacos , Cognição/efeitos dos fármacos , Dieta , Produtos Finais de Glicação Avançada/toxicidade , Presenilina-1/metabolismo , Aldeído Pirúvico/toxicidade , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Envelhecimento , Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Encéfalo/metabolismo , Catalase/metabolismo , Citocinas/metabolismo , Feminino , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Inflamação/etiologia , Inflamação/metabolismo , Lactoilglutationa Liase/metabolismo , Masculino , Memória/efeitos dos fármacos , Camundongos , NADPH Oxidases/metabolismo , Doenças Neuroinflamatórias/etiologia , Doenças Neuroinflamatórias/metabolismo , Doenças Neuroinflamatórias/patologia , Estresse OxidativoRESUMO
In this study, we demonstrated for the first time an increased CD44 gene expression in lymphocytes derived from Alzheimer's disease (AD) patients in comparison with healthy subjects. CD44 is a surface antigen expressed by cells of the immune and central nervous system as well as in a variety of other tissues. Functioning as adhesion molecule, CD44 is furthermore involved in driving immune response into infected tissues, including the CNS. We also found that lymphocytes of the same patients expressed significant levels of unfolded p53 isoform, confirming what we already demonstrated in fibroblasts and lymphocytes derived from other cohorts of AD patients. A correlation between p53 and CD44 expression has been well demonstrated in cancer cells, suggesting that CD44 could be a target gene of mutant p53, or either mutant p53 could lack its ability to negatively regulate CD44 expression. The contemporaneous increased expression of unfolded p53 and CD44 in AD lymphocytes may suggest that these two molecules cross-talk together participating in peripheral immune response during the development of the disease.
Assuntos
Doença de Alzheimer/patologia , Regulação da Expressão Gênica/fisiologia , Receptores de Hialuronatos/metabolismo , Linfócitos/metabolismo , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Receptores de Hialuronatos/genética , Masculino , Pessoa de Meia-IdadeRESUMO
Plants are not just passive living beings that exist in nature. They are complex and highly adaptable species that react sensitively to environmental forces/stimuli with movement, morphological changes and through the communication via volatile molecules. In a way, plants mimic some traits of animal and human behaviour; they compete for limited resources by gaining more area for more sunlight and spread their roots underground. Furthermore, in order to survive and thrive, they evolve and 'learn' to control various environmental stress factors in order to increase the yield of flowering, fertilization and germination processes. The concept of associating complex behaviour, such as intelligence, with plants is still a highly debatable topic among researchers worldwide. Recent studies have shown that plants are able to discriminate between positive and negative experiences and 'learn' from them. Some botanists have interpreted these behavioural data as a form of primitive cognitive processes. Others have evaluated these responses as biological automatisms of plants determined by adaptation to the environment and absence of intelligence. This review aims to explore adaptive behavioural aspects of various plant species distributed in different ecosystems by emphasizing their biological complexity and survival instincts.
Assuntos
Evolução Biológica , Ecossistema , Fenômenos Fisiológicos Vegetais , Adaptação Fisiológica , Animais , Germinação/fisiologia , Raízes de Plantas/fisiologia , Plantas/classificação , Plantas/metabolismoRESUMO
Several theories have been postulated, trying to explain why and how living organisms age. Despite some controversies and still huge open questions, a growing body of evidence suggest alterations of mitochondrial functionality and redox-homeostasis occur during the ageing process. Oxidative damage and mitochondrial dysfunction do not represent the cause of ageing per se but they have to be analyzed within the complexity of those series of processes occurring during lifespan. The establishment of a crosstalk among them is a shared common feature of many chronic age-related diseases, including neurodegenerative disorders, for which ageing is a major risk factor. The challenge is to understand when and how the interplay between these two systems move towards from normal ageing process to a pathological phenotype. Here in this review, we discuss the crosstalk between mitochondria and cytosolic-ROS. Furthermore, through a visual data mining approach, we attempt to describe the dynamic interplay between mitochondria and cellular redox state on the route from ageing to an AD phenotype.
Assuntos
Envelhecimento/fisiologia , Doença de Alzheimer/metabolismo , Homeostase/fisiologia , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Humanos , Oxirredução , Estresse OxidativoRESUMO
The identification of biological markers of Alzheimer's disease (AD) can be extremely useful to improve diagnostic accuracy and/or to monitor the efficacy of putative therapies. In this regard, peripheral cells may be of great importance, because of their easy accessibility. After subjects were grouped according to diagnosis, the expression of conformationally mutant p53 in blood cells was compared by immunoprecipitation or by a cytofluorimetric assay. In total, 104 patients with AD, 92 age-matched controls, 15 patients with Parkinson's disease and 9 with other types of dementia were analyzed. Two independent methods to evaluate the differential expression of a conformational mutant p53 were developed. Mononuclear cells were analyzed by immunoprecipitation or by flow-cytometric analysis, following incubation with a conformation-specific p53 antibody, which discriminates unfolded p53 tertiary structure. Mononuclear cells from AD patients express a higher amount of mutant-like p53 compared to non-AD subjects, thus supporting the study of conformational mutant p53 as a new putative marker to discriminate AD from non-AD patients. We also observed a strong positive correlation between the expression of p53 and the age of patients. The expression of p53 was independent from the length of illness and from the Mini Mental State Examination value.
Assuntos
Doença de Alzheimer/genética , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética , Idoso , Idoso de 80 Anos ou mais , Envelhecimento/genética , DNA/sangue , DNA/genética , DNA/isolamento & purificação , Demência/sangue , Demência/genética , Citometria de Fluxo , Humanos , Itália , Leucócitos , Doença de Parkinson/sangue , Doença de Parkinson/genética , Polônia , Conformação Proteica , Valores de Referência , Proteína Supressora de Tumor p53/sangueRESUMO
Aspirin (acetylsalicylic acid) is a commonly prescribed drug with a wide pharmacological spectrum. At concentrations compatible with amounts in plasma during chronic anti-inflammatory therapy, acetylsalicylic acid and its metabolite sodium salicylate were found to be protective against neurotoxicity elicited by the excitatory amino acid glutamate in rat primary neuronal cultures and hippocampal slices. The site of action of the drugs appeared to be downstream of glutamate receptors and to involve specific inhibition of glutamate-mediated induction of nuclear factor kappa B. These results may contribute to the emerging theme of anti-inflammatory drugs and neurodegeneration.
Assuntos
Aspirina/farmacologia , Ácido Glutâmico/farmacologia , Hipocampo/efeitos dos fármacos , NF-kappa B/metabolismo , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Salicilato de Sódio/farmacologia , Animais , Cálcio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cerebelo/citologia , Cerebelo/efeitos dos fármacos , Cerebelo/metabolismo , Giro Denteado/citologia , Giro Denteado/efeitos dos fármacos , Giro Denteado/metabolismo , Hipocampo/citologia , Hipocampo/metabolismo , Técnicas In Vitro , N-Metilaspartato/farmacologia , Neurônios/citologia , Neurônios/metabolismo , Células Piramidais/citologia , Células Piramidais/efeitos dos fármacos , Células Piramidais/metabolismo , Ratos , Ratos Sprague-Dawley , Fator de Transcrição AP-1/metabolismo , Regulação para CimaRESUMO
Sodium fluoride, guanylimidodiphosphate, and the D1 dopamine receptor agonist SKF 38393 elicited a greater activation of adenylate cyclase in homogenates of caudate nucleus in schizophrenic than in nonschizophrenic subjects used as controls. Similarly, a greater activation of adenylate cyclase by sodium fluoride was observed in the nucleus accumbens of schizophrenics. These findings suggest that the coupling of dopamine D1 recognition sites with adenylate cyclase is more efficient in the brain of the schizophrenic, presumably because of an increased affinity of the G/F protein for guanosine 5'-triphosphate.
Assuntos
Adenilil Ciclases/metabolismo , Núcleo Caudado/metabolismo , Núcleo Accumbens/metabolismo , Receptores de Superfície Celular/fisiologia , Receptores Dopaminérgicos/fisiologia , Esquizofrenia/metabolismo , Núcleos Septais/metabolismo , Ativação Enzimática/efeitos dos fármacos , Fluoretos/farmacologia , Proteínas de Ligação ao GTP , Guanilil Imidodifosfato/farmacologia , Humanos , Proteínas de Membrana/metabolismoRESUMO
BACKGROUND: Colorectal adenocarcinoma cells (Caco-2) are a widely used model of intestinal barrier to study cancer development, toxicological assessments, absorption and metabolism in food science or drug discovery. Caco-2 spontaneously differentiate into a monolayer expressing several specific characteristics, typically showed by mature enterocytes. For in vitro experiments, it is crucial to identify non-invasive and non-destructive techniques able to evaluate the integrity and differentiation of the cells monolayer. Thus, we aimed to assess these properties by analyzing electrical impedance measurements. METHODS: Caco-2 cells were differentiated for 21â¯days. The monolayer integrity and differentiation were primarily evaluated by means of morphological, biochemical and molecular data. Impedance measurements in a range of frequencies from 400â¯Hz to 50â¯kHz were performed using a dedicated set up, including customized Aerosol Jet Printed carbon-based sensors. RESULTS: The trends of RI observed at three different frequencies were able to describe cell growth and differentiation. In order to evaluate which frequencies better correlate with cell differentiation, Principal Component Analysis have been employed and the concordance analysis between RI magnitude and morphological, biochemical and molecular data, highlighted 40â¯kHz as the optimal frequency to assess Caco-2 cells differentiation process. CONCLUSION: We demonstrated the feasibility and reliability of applying impedance-based measurements not only to provide information about the monolayer status, but also for cell differentiation monitoring. GENERAL SIGNIFICANCE: This study underlined the possibility to use a dedicated sensor to assess the integrity and differentiation of Caco-2 monolayer, as a reliable non-destructive alternative to conventional approaches.
Assuntos
Diferenciação Celular , Impedância Elétrica , Técnicas Eletroquímicas , Impressão Tridimensional , Células CACO-2 , Proliferação de Células , Eletrodos , HumanosRESUMO
The appropriate level of microtubule stability is fundamental in neurons to assure correct polarity, migration, vesicles transport and to prevent axonal degeneration. In the present study, we have identified Notch pathway as an endogenous microtubule stabilizer. Stimulation of Notch receptors by exposure of mouse cortical neurons to the Notch ligand Jagged1 resulted in increased microtubule stability, as measured by using antibodies against post-translationally modified alpha tubulin, and changes in axonal morphology and branching, with varicosity loss, thicker neurites and enlarged growth cones. Similar effects were found after exposure of the cells to different doses of Taxol. However, contrary to Taxol, Jagged1 induced downregulation of the microtubule severing protein Spastin. We suggest that a fine-tuned manipulation of Notch signaling may represent a novel approach to modulate neuronal cytoskeleton plasticity.
Assuntos
Córtex Cerebral/fisiologia , Microtúbulos/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Receptores Notch/agonistas , Adenosina Trifosfatases/biossíntese , Adenosina Trifosfatases/genética , Secretases da Proteína Precursora do Amiloide/metabolismo , Animais , Antineoplásicos Fitogênicos/farmacologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/fisiologia , Córtex Cerebral/citologia , Córtex Cerebral/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Proteína Jagged-1 , Ligantes , Proteínas de Membrana/genética , Proteínas de Membrana/fisiologia , Camundongos , Microscopia Confocal , Paclitaxel/farmacologia , Ácido Poliglutâmico/metabolismo , RNA/biossíntese , RNA/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Serrate-Jagged , Transdução de Sinais/efeitos dos fármacos , Espastina , Tubulina (Proteína)/metabolismo , Tirosina/metabolismoRESUMO
γ-Oryzanol (ORY) is well known for its antioxidant potential. However, the mechanism by which ORY exerts its antioxidant effect is still unclear. In this paper, the antioxidant properties of ORY were investigated for its potential effects as a reactive oxygen and nitrogen species (ROS/RNS) scavenger and in activating antioxidant-promoting intracellular pathways utilizing the human embryonic kidney cells (HEK-293). The 24 h ORY exposure significantly prevented hydrogen peroxide- (H2O2-) induced ROS/RNS production at 3 h, and this effect was sustained for at least 24 h. ORY pretreatment also enhanced the activity of antioxidant enzymes: superoxide dismutase (SOD) and glutathione peroxidase (GPX). Interestingly, ORY induced the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation and upregulation of Nrf2-dependent defensive genes such as NAD(P)H quinone reductase (NQO1), heme oxygenase-1 (HO-1), and glutathione synthetase (GSS) at mRNA and protein levels in both basal condition and after H2O2 insult. Thus, this study suggested an intriguing effect of ORY in modulating the Nrf2 pathway, which is also involved in regulating longevity as well as age-related diseases.
Assuntos
Fator 2 Relacionado a NF-E2/metabolismo , Fenilpropionatos/metabolismo , Antioxidantes , HumanosRESUMO
Cell-cycle-related proteins, such as cyclins or cyclin-dependent kinases, are re-expressed in neurons committed to death in response to a variety of insults, including excitotoxins, hypoxia and ischemia, loss of trophic support, or beta-amyloid peptide. In some of these conditions events that are typical of the mid-G1 phase, such as cyclin-dependent kinase 4/6 activation, are required for the induction of neuronal death. In other cases, the cycle must proceed further and recruit steps that are typical of the G1/S transition for death to occur. Finally, there are conditions in which cell-cycle proteins might be re-expressed, but do not contribute to neuronal death. We hypothesize that cell-cycle signaling becomes a mandatory component of neuronal demise when other mechanisms are not enough for neurons to reach the threshold for death. Under this scheme, the death threshold is set by the extent of DNA damage. Whenever the extent of DNA damage is below this threshold, a cell-cycle signaling becomes crucial for the induction of neuronal death through p53-dependent or -independent pathways.
Assuntos
Peptídeos beta-Amiloides/metabolismo , Apoptose/fisiologia , Proteínas de Ciclo Celular/metabolismo , Dano ao DNA/fisiologia , Neurônios/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Peptídeos beta-Amiloides/farmacologia , Animais , Apoptose/efeitos dos fármacos , Proteínas de Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Dano ao DNA/efeitos dos fármacos , Humanos , Degeneração Neural/metabolismo , Neurônios/efeitos dos fármacos , Neurotoxinas/farmacologia , Oligonucleotídeos Antissenso/farmacologia , Proteína Supressora de Tumor p53/farmacologiaRESUMO
It is well recognized that mitochondrial dysfunction contributes to neurodegeneration occurring in Alzheimer's disease (AD). However, evidences of mitochondrial defects in AD peripheral cells are still inconclusive. Here, some mitochondrial-encoded and nuclear-encoded proteins, involved in maintaining the correct mitochondria machine, were investigated in terms of protein expression and enzymatic activity in peripheral blood mononuclear cells (PBMCs) isolated from AD and Mild Cognitive Impairment (MCI) patients and healthy subjects. In addition mitochondrial DNA copy number was measured by real time PCR. We found some differences and some similarities between AD and MCI patients when compared with healthy subjects. For example, cytochrome C and cytochrome B were decreased in AD, while MCI showed only a statistical reduction of cytochrome C. On the other hand, both AD and MCI blood cells exhibited highly nitrated MnSOD, index of a prooxidant environment inside the mitochondria. TFAM, a regulator of mitochondrial genome replication and transcription, was decreased in both AD and MCI patients' blood cells. Moreover also the mitochondrial DNA amount was reduced in PBMCs from both patient groups. In conclusion these data confirmed peripheral mitochondria impairment in AD and demonstrated that TFAM and mtDNA amount reduction could be two features of early events occurring in AD pathogenesis.
Assuntos
Doença de Alzheimer/sangue , Disfunção Cognitiva/sangue , Leucócitos Mononucleares/citologia , Mitocôndrias/patologia , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Núcleo Celular/metabolismo , Citocromos b/metabolismo , Citocromos c/metabolismo , DNA Mitocondrial/metabolismo , Feminino , Dosagem de Genes , Humanos , Masculino , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Oxidantes/metabolismo , Oxirredução , Reação em Cadeia da Polimerase em Tempo Real , Superóxido Dismutase/metabolismoRESUMO
Preclinical data indicate a direct anti-tumor effect of zoledronic acid (ZA) outside the skeleton, but its molecular mechanism is still not completely clarified. The aim of this study was to investigate the anti-cancer effects of ZA in human breast cancer cell lines, suggesting that they may in part be mediated via the miR-21/PTEN/Akt signaling pathway. The effect of ZA on cell viability was measured by MTT assay, and cell death induction was analyzed using either a double AO/EtBr staining and M30 ELISA assay. A Proteome Profiler Human Apoptosis Array was executed to evaluate the molecular basis of ZA-induced apoptosis. Cell cycle analysis was executed by flow cytometry. The effect of ZA on miR-21 expression was quantified by qRT-PCR, and the amount of PTEN protein and its targets were analyzed by Western blot. ZA inhibited cell growth in a concentration- and time-dependent manner, through the activation of cell death pathways and arrest of cell cycle progression. ZA downregulated the expression of miR-21, resulting in dephosphorilation of Akt and Bad and in a significant increase of p21 and p27 proteins expression. These results were observed also in MDA-MB-231 cells, commonly used as an experimental model of bone metastasis of breast cancer. This study revealed, for the first time, an involvement of the miR-21/PTEN/Akt signaling pathway in the mechanism of ZA anti-cancer actions in breast cancer cells. We would like to underline that this pathway is present both in the hormone responsive BC cell line (MCF-7) as well as in a triple negative cell line (MDA-MB-231). Taken together these results reinforce the use of ZA in clinical practice, suggesting the role of miR-21 as a possible mediator of its therapeutic efficacy.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/metabolismo , Difosfonatos/farmacologia , Imidazóis/farmacologia , MicroRNAs/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Feminino , Humanos , Transdução de Sinais/efeitos dos fármacos , Ácido ZoledrônicoRESUMO
Apoptosis is now recognized as an important component in many progressive and acute neurodegenerative diseases. Extracellular signals and intracellular mechanisms triggering and regulating apoptosis in neuronal cells are still a matter of investigation. Here we review data from our and other laboratories with the aim to elucidate the nature of some proteins which are known to be involved in cell cycle regulation as well as in promoting degeneration and apoptosis of neurons. The following molecules will be taken into consideration: NF-kappaB, p53, p21 and MSH2. These proteins are activated by neurotoxic experimental conditions which involve the stimulation of selective receptors for the excitatory aminoacid glutamate. Thus, we hypothesize their contribution to an intracellular pathway responsible for the glutamate-induced neuronal death. Identification of such mechanisms could be relevant for understanding the apoptosis associated with various neurodegenerative diseases as well as for developing novel strategies of pharmacological intervention.
Assuntos
Apoptose/efeitos dos fármacos , Ácido Glutâmico/farmacologia , NF-kappa B/metabolismo , Neurônios/efeitos dos fármacos , Proteína Supressora de Tumor p53/metabolismo , Ciclo Celular/fisiologia , Linhagem Celular , Proteínas do Tecido Nervoso/metabolismo , Doenças Neurodegenerativas/etiologia , Neurônios/metabolismo , Transdução de Sinais/fisiologiaRESUMO
Here we report that a novel member of the TNF-alpha family, TNF-related apoptosis-inducing ligand (TRAIL), contributes substantially to amyloid-induced neurotoxicity in human SH-SY5Y neuronal cell line. Involvement of TRAIL in the amyloid-induced cell death is supported by cDNA array, Northern blot, and Western blot data, demonstrating increased TRAIL expression after treatment of the cells with a neurotoxic fragment of amyloid protein (betaAP). TRAIL was also found to be released in the culture media after betaAP treatment with a time-course overlapping to contents of the intracellular protein. Contribution of TRAIL to betaAP neurotoxicity is demonstrated by data showing that TRAIL-neutralizing monoclonal antibody protects neuronal SH-SY5Y cells from betaAP neurotoxicity. Moreover, exposure of neuronal SH-SY5Y cells to TRAIL leads to cell death, indicating that this substance per se is endowed with neurotoxic properties. We also found that, similarly to betaAP and TRAIL, activation of the death-domain adaptor protein FADD results in neuronal cell death. Lack of FADD function, by overexpression of its dominant negative, rescued cells from either TRAIL- or betaAP-induced neurotoxicity, supporting the hypothesis that these three molecules share common intracellular pathways. Finally, we found that betaAP strongly activated caspase-8, and the cell-permeable, selective caspase-8 inhibitor z-IETD-FMK prevents both betaAP- and TRAIL-induced neurotoxicity. In view of TRAIL's potency in inducing neuronal death, and its role as mediator of betaAP, it is plausible to hypothesize that TRAIL can be regarded as a molecule that provides substantial contribution to betaAP-dependent cell death, which takes part in the progression of the neurodegenerative process and related chronic inflammatory response.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Doença de Alzheimer/tratamento farmacológico , Peptídeos beta-Amiloides/toxicidade , Apoptose/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Glicoproteínas de Membrana/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Anticorpos Monoclonais/farmacologia , Apoptose/fisiologia , Proteínas Reguladoras de Apoptose , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Proteínas de Transporte/agonistas , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Caspase 8 , Caspase 9 , Inibidores de Caspase , Caspases/metabolismo , Inibidores de Cisteína Proteinase/farmacologia , Proteína de Domínio de Morte Associada a Fas , Humanos , Neurônios/metabolismo , Neurônios/patologia , Fármacos Neuroprotetores/imunologia , Fármacos Neuroprotetores/farmacologia , Oligopeptídeos/farmacologia , Ligante Indutor de Apoptose Relacionado a TNF , Células Tumorais CultivadasRESUMO
Demyelinating diseases are high impact neurological disorders. Steroids are regarded as protective molecules in the susceptibility to these diseases. Here, we studied the interactions between tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), a potent proapoptotic molecule toxic to oligodendrocytes, and 17-beta-estradiol (E-17-beta), in human oligodendrocytic MO3.13 cells. Exposure of cells to TRAIL resulted in the upregulation of both death receptors DR4 and DR5 and apoptosis, as well as the activation of caspase-8 and -3, increased phosphorylation of Jun-N-terminal kinase and p38 kinase, and the reduction of bcl-2 and bcl-xL proteins. TRAIL-mediated MO3.13 cell apoptosis was abrogated by the dominant-negative form of the adaptor protein FADD and by caspase inhibitors. Preincubation with E-17-beta completely prevented both TRAIL-induced DR4 and DR5 upregulation and apoptosis. Estrogen-induced cytoprotection was time and concentration dependent and reverted by antiestrogens. Estrogen treatment per se reduced kinase phosphorylation, and upregulated bcl-2 and bcl-xL proteins. In conclusion, our data show that the detrimental role of TRAIL on oligodendrocytes can be effectively counteracted by estrogens, thus suggesting that the underlying molecular interactions can be of potential relevance in characterizing novel targets for therapy of demyelinating disorders.
Assuntos
Apoptose/efeitos dos fármacos , Estradiol/farmacologia , Glicoproteínas de Membrana/metabolismo , Oligodendroglia/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Reguladoras de Apoptose , Proteínas de Arabidopsis/metabolismo , Inibidores de Caspase , Caspases/metabolismo , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Ácidos Graxos Dessaturases/metabolismo , Células HeLa , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fosforilação , Receptores do Ligante Indutor de Apoptose Relacionado a TNF , Receptores do Fator de Necrose Tumoral/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
A common arginine to proline polymorphism is harboured at codon 72 of the human p53 gene. In this investigation, we found that fibroblasts and lymphocytes isolated from arginine allele homozygote centenarians and sexagenarians (Arg+) undergo an oxidative-stress-induced apoptosis at a higher extent than cells obtained from proline allele carriers (Pro+). At variance, the difference in apoptosis susceptibility between Arg+ and Pro+ is not significant when cells from 30-year-old people are studied. Further, we found that Arg+ and Pro+ cells from centenarians differ in the constitutive levels of p53 protein and p53/MDM2 complex, as well as in the levels of oxidative stress-induced p53/Bcl-xL complex and mitochondria-localised p53. Consistently, all these differences are less evident in cells from 30-year-old people. Finally, we investigated the in vivo functional relevance of the p53 codon 72 genotype in a group of old patients (66-99 years of age) affected by acute myocardial ischaemia, a clinical condition in which in vivo cell death occurs. We found that Arg+ patients show increased levels of Troponin I and CK-MB, two serum markers that correlate with the extent of the ischaemic damage in comparison to Pro+ patients. In conclusion, these data suggest that p53 codon 72 polymorphism contributes to a genetically determined variability in apoptotic susceptibility among old people, which has a potentially relevant role in the context of an age-related pathologic condition, such as myocardial ischaemia.
Assuntos
Apoptose , Códon , Genes p53 , Isquemia , Proteína Supressora de Tumor p53/fisiologia , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Alelos , Arginina , Western Blotting , Morte Celular , Creatina Quinase/sangue , Creatina Quinase Forma MB , Relação Dose-Resposta a Droga , Feminino , Fibroblastos/metabolismo , Citometria de Fluxo , Genótipo , Homozigoto , Humanos , Imunoprecipitação , Isoenzimas/sangue , Leucócitos/metabolismo , Linfócitos/metabolismo , Masculino , Potenciais da Membrana , Microscopia de Fluorescência , Pessoa de Meia-Idade , Isquemia Miocárdica/patologia , Estresse Oxidativo , Polimorfismo Genético , Prolina , Proteínas Proto-Oncogênicas c-bcl-2 , Análise de Regressão , Serina/química , Fatores de Tempo , Transfecção , Troponina I/sangue , Proteína bcl-XRESUMO
The concept of heterogeneity of Alzheimer's disease is based on molecular, neuropathological, clinical and neuropsychological features, and also supported by the observation that Alzheimer's patients differ in their response to pharmacological interventions. Recent investigations evaluating the therapeutic potential of cholinesterase inhibitors have disclosed the existence of at least two subsets of patients with dementia, defined as 'responders' and 'nonresponders' to this therapy. In this article, Paolo Liberini and colleagues suggest that the cluster of responders to the cholinesterase inhibitors might include a significant number of subjects with a rather selective dysfunction of the cholinergic system, as in the case of Lewy-body dementia. A neuropathological demonstration of this correlation should open up new therapeutic perspectives.