974 resultados para Michelson interference
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Recurrent breast cancer occurring after the initial treatment is associated with poor outcome. A bimodal relapse pattern after surgery for primary tumor has been described with peaks of early and late recurrence occurring at about 2 and 5 years, respectively. Although several clinical and pathological features have been used to discriminate between low- and high-risk patients, the identification of molecular biomarkers with prognostic value remains an unmet need in the current management of breast cancer. Using microarray-based technology, we have performed a microRNA expression analysis in 71 primary breast tumors from patients that either remained disease-free at 5 years post-surgery (group A) or developed early (group B) or late (group C) recurrence. Unsupervised hierarchical clustering of microRNA expression data segregated tumors in two groups, mainly corresponding to patients with early recurrence and those with no recurrence. Microarray data analysis and RT-qPCR validation led to the identification of a set of 5 microRNAs (the 5-miRNA signature) differentially expressed between these two groups: miR-149, miR-10a, miR-20b, miR-30a-3p and miR-342-5p. All five microRNAs were down-regulated in tumors from patients with early recurrence. We show here that the 5-miRNA signature defines a high-risk group of patients with shorter relapse-free survival and has predictive value to discriminate non-relapsing versus early-relapsing patients (AUC = 0.993, p-value<0.05). Network analysis based on miRNA-target interactions curated by public databases suggests that down-regulation of the 5-miRNA signature in the subset of early-relapsing tumors would result in an overall increased proliferative and angiogenic capacity. In summary, we have identified a set of recurrence-related microRNAs with potential prognostic value to identify patients who will likely develop metastasis early after primary breast surgery.
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Stable gene silencing by RNA interference (RNAi) can be achieved by expression of small hairpin RNAs (shRNAs) from RNA polymerase III promoters. We have tested lentiviral vectors expressing shRNAs targetting CCR5 in primary CD4 T cells from donors representing various CCR5 and CCR2 genetic backgrounds covering the full spectrum of CCR5 expression levels and permissiveness for HIV-1 infection. A linear decrease in CCR5 expression resulted in a logarithmic decrease in cellular infection, giving up to three logs protection from HIV-1 infection in vitro. Protection was maintained at very high multiplicity of infection. This and other recent reports on RNAi should open a debate about the use of RNAi gene therapy for HIV infection.
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This randomised, double-blind, multicentre study with children nine-23 months old evaluated the immunogenicity of yellow fever (YF) vaccines prepared with substrains 17DD and 17D-213/77. YF antibodies were tittered before and 30 or more days after vaccination. Seropositivity and seroconversion were analysed according to the maternal serological status and the collaborating centre. A total of 1,966 children were randomised in the municipalities of the states of Mato Grosso do Sul, Minas Gerais and São Paulo and blood samples were collected from 1,714 mothers. Seropositivity was observed in 78.6% of mothers and 8.9% of children before vaccination. After vaccination, seropositivity rates of 81.9% and 83.2%, seroconversion rates of 84.8% and 85.8% and rates of a four-fold increase over the pre-vaccination titre of 77.6% and 81.8% were observed in the 17D-213/77 and 17DD subgroups, respectively. There was no association with maternal immunity. Among children aged 12 months or older, the seroconversion rates of 69% were associated with concomitant vaccination against measles, mumps and rubella. The data were not conclusive regarding the interference of maternal immunity in the immune response to the YF vaccine, but they suggest interference from other vaccines. The failures in seroconversion after vaccination support the recommendation of a booster dose in children within 10 years of the first dose.
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BACKGROUND: The Complete Arabidopsis Transcript MicroArray (CATMA) initiative combines the efforts of laboratories in eight European countries 1 to deliver gene-specific sequence tags (GSTs) for the Arabidopsis research community. The CATMA initiative offers the power and flexibility to regularly update the GST collection according to evolving knowledge about the gene repertoire. These GST amplicons can easily be reamplified and shared, subsets can be picked at will to print dedicated arrays, and the GSTs can be cloned and used for other functional studies. This ongoing initiative has already produced approximately 24,000 GSTs that have been made publicly available for spotted microarray printing and RNA interference. RESULTS: GSTs from the CATMA version 2 repertoire (CATMAv2, created in 2002) were mapped onto the gene models from two independent Arabidopsis nuclear genome annotation efforts, TIGR5 and PSB-EuGène, to consolidate a list of genes that were targeted by previously designed CATMA tags. A total of 9,027 gene models were not tagged by any amplified CATMAv2 GST, and 2,533 amplified GSTs were no longer predicted to tag an updated gene model. To validate the efficacy of GST mapping criteria and design rules, the predicted and experimentally observed hybridization characteristics associated to GST features were correlated in transcript profiling datasets obtained with the CATMAv2 microarray, confirming the reliability of this platform. To complete the CATMA repertoire, all 9,027 gene models for which no GST had yet been designed were processed with an adjusted version of the Specific Primer and Amplicon Design Software (SPADS). A total of 5,756 novel GSTs were designed and amplified by PCR from genomic DNA. Together with the pre-existing GST collection, this new addition constitutes the CATMAv3 repertoire. It comprises 30,343 unique amplified sequences that tag 24,202 and 23,009 protein-encoding nuclear gene models in the TAIR6 and EuGène genome annotations, respectively. To cover the remaining untagged genes, we identified 543 additional GSTs using less stringent design criteria and designed 990 sequence tags matching multiple members of gene families (Gene Family Tags or GFTs) to cover any remaining untagged genes. These latter 1,533 features constitute the CATMAv4 addition. CONCLUSION: To update the CATMA GST repertoire, we designed 7,289 additional sequence tags, bringing the total number of tagged TAIR6-annotated Arabidopsis nuclear protein-coding genes to 26,173. This resource is used both for the production of spotted microarrays and the large-scale cloning of hairpin RNA silencing vectors. All information about the resulting updated CATMA repertoire is available through the CATMA database http://www.catma.org.
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Background: The glycosylated hemoglobin (HbA1c) is used to help monitor the degree of a diabetic’s hyperglycemia. Security and accuracy of the methods used in its detection are affected by variants forms of Hb or elevations in levels of Fetal Hb (HbF). These interference are the result of a change in the haemoglobin total net charge of the variant due of a substitution of one amino acid in the remaining amino terminal of the beta chain. International Standardization for HbA1c values (NGSP) not include interference assessment as part of the certification program. Therefore, the effect of each variant or the lifting of the HbF on HbA1c result should be examined in each sample depending on the detected variant and the method used for the detection of the same. The objectives were: to describe the possible variants of Hb and their interference in HbA1c measurement by our method, after the implementation of a computer program for their detection. To identify some variants detected by chromatography liquid ion exchange high resolution (HPLC) with DNA molecular sequencing.
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The prefrontal (PFC) and orbitofrontal cortex (OFC) appear to be associated with both executive functions and olfaction. However, there is little data relating olfactory processing and executive functions in humans. The present study aimed at exploring the role of olfaction on executive functioning, making a distinction between primary and more cognitive aspects of olfaction. Three executive tasks of similar difficulty were used. One was used to assess hot executive functions (Iowa Gambling Task-IGT), and two as a measure of cold executive functioning (Stroop Colour and Word Test-SCWT and Wisconsin Card Sorting Test-WCST). Sixty two healthy participants were included: 31 with normosmia and 31 with hyposmia. Olfactory abilities were assessed using the ''Sniffin' Sticks'' test and the olfactory threshold, odour discrimination and odour identification measures were obtained. All participants were female, aged between 18 and 60. Results showed that participants with hyposmia displayed worse performance in decision making (IGT; Cohen's-d = 0.91) and cognitive flexibility (WCST; Cohen's-d between 0.54 and 0.68) compared to those with normosmia. Multiple regression adjusted by the covariates participants' age and education level showed a positive association between odour identification and the cognitive inhibition response (SCWT-interference; Beta = 0.29; p = .034). The odour discrimination capacity was not a predictor of the cognitive executive performance. Our results suggest that both hot and cold executive functions seem to be associated with higher-order olfactory functioning in humans. These results robustly support the hypothesis that olfaction and executive measures have a common neural substrate in PFC and OFC, and suggest that olfaction might be a reliable cognitive marker in psychiatric and neurologic disorders.
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BACKGROUND Measurement of HbA1c is the most important parameter to assess glycemic control in diabetic patients. Different point-of-care devices for HbA1c are available. The aim of this study was to evaluate two point-of-care testing (POCT) analyzers (DCA Vantage from Siemens and Afinion from Axis-Shield). We studied the bias and precision as well as interference from carbamylated hemoglobin. METHODS Bias of the POCT analyzers was obtained by measuring 53 blood samples from diabetic patients with a wide range of HbA1c, 4%-14% (20-130 mmol/mol), and comparing the results with those obtained by the laboratory method: HPLC HA 8160 Menarini. Precision was performed by 20 successive determinations of two samples with low 4.2% (22 mmol/mol) and high 9.5% (80 mmol/mol) HbA1c values. The possible interference from carbamylated hemoglobin was studied using 25 samples from patients with chronic renal failure. RESULTS The means of the differences between measurements performed by each POCT analyzer and the laboratory method (95% confidence interval) were: 0.28% (p<0.005) (0.10-0.44) for DCA and 0.27% (p<0.001) (0.19-0.35) for Afinion. Correlation coefficients were: r=0.973 for DCA, and r=0.991 for Afinion. The mean bias observed by using samples from chronic renal failure patients were 0.2 (range -0.4, 0.4) for DCA and 0.2 (-0.2, 0.5) for Afinion. Imprecision results were: CV=3.1% (high HbA1c) and 2.97% (low HbA1c) for DCA, CV=1.95% (high HbA1c) and 2.66% (low HbA1c) for Afinion. CONCLUSIONS Both POCT analyzers for HbA1c show good correlation with the laboratory method and acceptable precision.
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The inflammatory prostaglandin E2 (PGE2) cytokine plays a key role in the development of colon cancer. Several studies have shown that PGE2 directly induces the growth of colon cancer cells and furthermore promotes tumor angiogenesis by increasing the production of the vascular endothelial growth factor (VEGF). The signaling intermediaries implicated in these processes have however not been fully characterized. In this report, we show that the mechanistic target of rapamycin complex 1 (mTORC1) plays an important role in PGE2-induced colon cancer cell responses. Indeed, stimulation of LS174T cells with PGE2 increased mTORC1 activity as observed by the augmentation of S6 ribosomal protein phosphorylation, a downstream effector of mTORC1. The PGE2 EP4 receptor was responsible for transducing the signal to mTORC1. Moreover, PGE2 increased colon cancer cell proliferation as well as the growth of colon cancer cell colonies grown in matrigel and blocking mTORC1 by rapamycin or ATP-competitive inhibitors of mTOR abrogated these effects. Similarly, the inhibition of mTORC1 by downregulation of its component raptor using RNA interference blocked PGE2-induced LS174T cell growth. Finally, stimulation of LS174T cells with PGE2 increased VEGF production which was also prevented by mTORC1 inhibition. Taken together, these results show that mTORC1 is an important signaling intermediary in PGE2 mediated colon cancer cell growth and VEGF production. They further support a role for mTORC1 in inflammation induced tumor growth.
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AIMS/HYPOTHESIS: Pro-atherogenic and pro-oxidant, oxidised LDL trigger adverse effects on pancreatic beta cells, possibly contributing to diabetes progression. Because oxidised LDL diminish the expression of genes regulated by the inducible cAMP early repressor (ICER), we investigated the involvement of this transcription factor and of oxidative stress in beta cell failure elicited by oxidised LDL. METHODS: Isolated human and rat islets, and insulin-secreting cells were cultured with human native or oxidised LDL or with hydrogen peroxide. The expression of genes was determined by quantitative real-time PCR and western blotting. Insulin secretion was monitored by EIA kit. Cell apoptosis was determined by scoring cells displaying pycnotic nuclei. RESULTS: Exposure of beta cell lines and islets to oxidised LDL, but not to native LDL raised the abundance of ICER. Induction of this repressor by the modified LDL compromised the expression of important beta cell genes, including insulin and anti-apoptotic islet brain 1, as well as of genes coding for key components of the secretory machinery. This led to hampering of insulin production and secretion, and of cell survival. Silencing of this transcription factor by RNA interference restored the expression of its target genes and alleviated beta cell dysfunction and death triggered by oxidised LDL. Induction of ICER was stimulated by oxidative stress, whereas antioxidant treatment with N-acetylcysteine or HDL prevented the rise of ICER elicited by oxidised LDL and restored beta cell functions. CONCLUSIONS/INTERPRETATION: Induction of ICER links oxidative stress to beta cell failure caused by oxidised LDL and can be effectively abrogated by antioxidant treatment.
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El SPION (Super Paramagnetic Iron Oxide Nanoparticles) ha estat estudiat com un nou adsorbent per eliminar l’arsènic d’aigües contaminades. Les condicions òptimes de treball es van assolir per un pH de 3,6 i per concentracions inferiors als 100ppm. No es van trobar interferències significatives produïdes pels cations Cu, Ni i Zn en l’adsorció de l’As, sent el fosfat l’anió que més interfereix. Una esponja de cel·lulosa s’ha utilitzat com a suport del SPION, per disminuir les agregacions de les nanopartícules en suspensió i per proporcionar una material adequat per l’adsorció en continu, experiment amb columnes. Així, es va obtenir un augment de la capacitat d’adsorció del SPION per l’As(V), mentre que per l’As(III) continua sent baixa, per tant s’augmenta la selectivitat per l’As(V). Les interferències aniòniques afecten d’igual manera a l’adsorció de l’As(III) i l’As(V) quan l’adsorció és en continu o en discontinu. Els cations metàl·lics no interfereixen en l’adsorció de l’arsènic, a excepció del coure que és adsorbit i porta a la disminució de l’adsorció d’arsènic.
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El projecte consisteix en el desenvolupament d'un algorisme que millori el posicionament final d'un sistema que adquireix les dades d'una antena de GPS estàndard. Aquest sistema en certs moments té pèrdua total de senyal GPS o rep senyal amb pertorbacions, derivant en un mal posicionament. Nosaltres hem proposat una solució que utilitza les coordenades del GPS, el filtre Kalman per resoldre els problemes de pertorbacions de senyal, bases de dades digitals geogràfiques per garantir la circulació del vehicle per sobre la carretera, i finalment combina la informació temporal de posicions anteriors i la de les bases de dades per posicionar el vehicle quan hi ha pèrdua total de senyal. Els experiments realitzats ens indiquen que s'obté una millora del posicionement.
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The generation of vaccines against HIV/AIDS able to induce long-lasting protective immunity remains a major goal in the HIV field. The modest efficacy (31.2%) against HIV infection observed in the RV144 phase III clinical trial highlighted the need for further improvement of HIV vaccine candidates, formulation, and vaccine regimen. In this study, we have generated two novel NYVAC vectors, expressing HIV-1 clade C gp140(ZM96) (NYVAC-gp140) or Gag(ZM96)-Pol-Nef(CN54) (NYVAC-Gag-Pol-Nef), and defined their virological and immunological characteristics in cultured cells and in mice. The insertion of HIV genes does not affect the replication capacity of NYVAC recombinants in primary chicken embryo fibroblast cells, HIV sequences remain stable after multiple passages, and HIV antigens are correctly expressed and released from cells, with Env as a trimer (NYVAC-gp140), while in NYVAC-Gag-Pol-Nef-infected cells Gag-induced virus-like particles (VLPs) are abundant. Electron microscopy revealed that VLPs accumulated with time at the cell surface, with no interference with NYVAC morphogenesis. Both vectors trigger specific innate responses in human cells and show an attenuation profile in immunocompromised adult BALB/c and newborn CD1 mice after intracranial inoculation. Analysis of the immune responses elicited in mice after homologous NYVAC prime/NYVAC boost immunization shows that recombinant viruses induced polyfunctional Env-specific CD4 or Gag-specific CD8 T cell responses. Antibody responses against gp140 and p17/p24 were elicited. Our findings showed important insights into virus-host cell interactions of NYVAC vectors expressing HIV antigens, with the activation of specific immune parameters which will help to unravel potential correlates of protection against HIV in human clinical trials with these vectors. IMPORTANCE: We have generated two novel NYVAC-based HIV vaccine candidates expressing HIV-1 clade C trimeric soluble gp140 (ZM96) and Gag(ZM96)-Pol-Nef(CN54) as VLPs. These vectors are stable and express high levels of both HIV-1 antigens. Gag-induced VLPs do not interfere with NYVAC morphogenesis, are highly attenuated in immunocompromised and newborn mice after intracranial inoculation, trigger specific innate immune responses in human cells, and activate T (Env-specific CD4 and Gag-specific CD8) and B cell immune responses to the HIV antigens, leading to high antibody titers against gp140. For these reasons, these vectors can be considered vaccine candidates against HIV/AIDS and currently are being tested in macaques and humans.
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Viruses have evolved strategies to overcome the antiviral effects of the host at different levels. Besides specific defence mechanisms, the host responds to viral infection via the interferon pathway and also by RNA interference (RNAi). However, several viruses have been identified that suppress RNAi. We addressed the question of whether hepatitis C virus (HCV) suppresses RNAi, using cell lines constitutively expressing green fluorescent protein (GFP) and inducibly expressing HCV proteins. It was found that short interfering RNA-mediated GFP gene silencing was inhibited when the entire HCV polyprotein was expressed. Further studies showed that HCV structural proteins, and in particular envelope protein 2 (E2), were responsible for this inhibition. Co-precipitation assays demonstrated that E2 bound to Argonaute-2 (Ago-2), a member of the RNA-induced silencing complex, RISC. Thus, HCV E2 that interacts with Ago-2 is able to suppress RNAi.
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Background: A developmental dysregulation of glutathione (GSH) synthesis leading to oxidative stress, when combined with environmental risk factors (viral infections) generating reactive oxygen species, can play a critical role in inducing schizophrenia phenotypes. GSH deficit induces morphological, physiological and behavioral anomalies analogous to those reported in schizophrenic patients, including disrupted parvalbumine (PV) inhibitory interneuron's integrity and neuronal synchrony (β/γ-oscillations). Methods: We assessed PV immunoreactivity (PV-IR) and local synchronization in prefrontal cortex of two mouse models: (1) mice with a genetic deficit in GSH (GCLM-/-) and (2) mice with prenatal immune activation at embryonic day17 (PolyI:C). Results: Adults from both mice models display reduced PV-IR in prefrontal cortex. In anterior cingulate (ACC) of GCLM-/-, appearance and maturation of PVI are delayed and worsened with peribubertal stress but not in adult one. This effect is reversed by treatment with the GSH precursor N-acetyl-cysteine. The power of beta and gamma oscillations are decreased in ACC of GCLM-/- while they increased in prelimbic cortex of PolyI:C mice. Conclusions: Despite reduced PV-IR in both models, alteration of the synchronization was different, indicating that the structural/functional disruption of the cortical circuitry was partly different in both models. Novel therapeutic strategies are proposed, based on interference with oxidative stress and inflammatory processes.
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Arteriovenous-lymphatic endothelial cell fates are specified by the master regulators, namely, Notch, COUP-TFII, and Prox1. Whereas Notch is expressed in the arteries and COUP-TFII in the veins, the lymphatics express all 3 cell fate regulators. Previous studies show that lymphatic endothelial cell (LEC) fate is highly plastic and reversible, raising a new concept that all 3 endothelial cell fates may co-reside in LECs and a subtle alteration can result in a reprogramming of LEC fate. We provide a molecular basis verifying this concept by identifying a cross-control mechanism among these cell fate regulators. We found that Notch signal down-regulates Prox1 and COUP-TFII through Hey1 and Hey2 and that activated Notch receptor suppresses the lymphatic phenotypes and induces the arterial cell fate. On the contrary, Prox1 and COUP-TFII attenuate vascular endothelial growth factor signaling, known to induce Notch, by repressing vascular endothelial growth factor receptor-2 and neuropilin-1. We show that previously reported podoplanin-based LEC heterogeneity is associated with differential expression of Notch1 in human cutaneous lymphatics. We propose that the expression of the 3 cell fate regulators is controlled by an exquisite feedback mechanism working in LECs and that LEC fate is a consequence of the Prox1-directed lymphatic equilibrium among the cell fate regulators.