19 resultados para Cell-based

em Repositório Institucional UNESP - Universidade Estadual Paulista "Julio de Mesquita Filho"


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The construction of a flow-through cell incorporating an array of gold microelectrodes is described and its application to flow injection analysis with amperometric detection is presented, Simple modification of almost any conventional integrated circuit chip, used as an inexpensive source of pre-assembled gold micro-wires, leads to the rapid and successful preparation of arrays of 8-48 elements, the polymeric encapsulation material from the top face of the chip is removed by abrasion until the gold micro-mires (used to interconnect the silicon circuit to the external contact pins of the chip) are disrupted and their transversal (elliptical) sections become exposed. Once polished, the flat and smooth top surface of the gold microelectrode-array chip (MEAC) is provided with a spacer and fitted under pressure against an acrylic block with the reference and auxiliary electrodes, to form the electrochemical (thin-layer) flow cell, while the contact pins are plugged into a standard IC socket, This design ensures autonomous electric contact with each electrode and allows fast dismantling for polishing or substitution, the performance of flow cells with MEACs was investigated utilizing the technique of reverse pulse amperometry without oxygen removal, A method was established for the determination of the copper concentration in sugar cane spirit, regulated by law for beverages, Samples from industrial producers and small-scale (alembic) brewers were compared, With a 24 MEAC, a detection limit of 30 mu g I-l of copper (4.7 x 10(-7) mol l(-1) of Cu-II for 100 mu l injections) was calculated, Routine operation was established at a frequency of 60-90 determinations per hour, Intercomparison with atomic absorption spectrometric determinations resulted in excellent agreement.

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Tendon and ligament injuries have proved difficult to treat effectively. Cell-based therapies offer the potential to harness the complex protein synthetic machinery of the cell to induce a regenerative response rather than fibrous scarring. This article reviews the current state of play with respect to the clinically used cell preparations for the treatment of tendon and ligaments overstrain injuries.

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In the present work, a biosensor was built with smart material based on polymer brushes. The biosensor demonstrated a pH-sensitive on-off property, and it was further used to control or modulate the electrochemical responses of the biosensor. This property could be used to realize pH-controlled electrochemical reaction of hydrogen peroxide and HRP immobilized on polymer brushes. The composite film also showed excellent amperometric i-t response toward hydrogen peroxide in the concentration range of 0-13 μM. In future, this platform might be used for self-regulating targeted diagnostic, drug delivery and biofuel cell based on controllable bioelectrocatalysis. © 2013 Elsevier B.V.

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Chikungunya virus (CHIKV) is a mosquito-borne arthrogenic alphavirus that causes acute febrile illness in humans accompanied by joint pains and in many cases, persistent arthralgia lasting weeks to years. The re-emergence of CHIKV has resulted in numerous outbreaks in the eastern hemisphere, and threatens to expand in the foreseeable future. Unfortunately, no effective treatment is currently available. The present study reports the use of resazurin in a cell-based high-throughput assay, and an image-based high-content assay to identify and characterize inhibitors of CHIKV-infection in vitro. CHIKV is a highly cytopathic virus that rapidly kills infected cells. Thus, cell viability of HuH-7 cells infected with CHIKV in the presence of compounds was determined by measuring metabolic reduction of resazurin to identify inhibitors of CHIKV-associated cell death. A kinase inhibitor library of 4,000 compounds was screened against CHIKV infection of HuH-7 cells using the resazurin reduction assay, and the cell toxicity was also measured in non-infected cells. Seventy-two compounds showing >= 50% inhibition property against CHIKV at 10 mu M were selected as primary hits. Four compounds having a benzofuran core scaffold (CND0335, CND0364, CND0366 and CND0415), one pyrrolopyridine (CND0545) and one thiazol-carboxamide (CND3514) inhibited CHIKV-associated cell death in a dose-dependent manner, with EC50 values between 2.2 mu M and 7.1 mu M. Based on image analysis, these 6 hit compounds did not inhibit CHIKV replication in the host cell. However, CHIKV-infected cells manifested less prominent apoptotic blebs typical of CHIKV cytopathic effect compared with the control infection. Moreover, treatment with these compounds reduced viral titers in the medium of CHIKV-infected cells by up to 100-fold. In conclusion, this cell-based high-throughput screening assay using resazurin, combined with the image-based high content assay approach identified compounds against CHIKV having a novel antiviral activity -inhibition of virus-induced CPE - likely by targeting kinases involved in apoptosis.

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Stemming from in vitro and in vivo pre-clinical and human models, tissue-engineering-based strategies continue to demonstrate great potential for the regeneration of the pulp-dentin complex, particularly in necrotic, immature permanent teeth. Nanofibrous scaffolds, which closely resemble the native extracellular matrix, have been successfully synthesized by various techniques, including but not limited to electrospinning. A common goal in scaffold synthesis has been the notion of promoting cell guidance through the careful design and use of a collection of biochemical and physical cues capable of governing and stimulating specific events at the cellular and tissue levels. The latest advances in processing technologies allow for the fabrication of scaffolds where selected bioactive molecules can be delivered locally, thus increasing the possibilities for clinical success. Though electrospun scaffolds have not yet been tested in vivo in either human or animal pulpless models in immature permanent teeth, recent studies have highlighted their regenerative potential both from an in vitro and in vivo (i.e., subcutaneous model) standpoint. Possible applications for these bioactive scaffolds continue to evolve, with significant prospects related to the regeneration of both dentin and pulp tissue and, more recently, to root canal disinfection. Nonetheless, no single implantable scaffold can consistently guide the coordinated growth and development of the multiple tissue types involved in the functional regeneration of the pulp-dentin complex. The purpose of this review is to provide a comprehensive perspective on the latest discoveries related to the use of scaffolds and/or stem cells in regenerative endodontics. The authors focused this review on bioactive nanofibrous scaffolds, injectable scaffolds and stem cells, and pre-clinical findings using stem-cell-based strategies. These topics are discussed in detail in an attempt to provide future direction and to shed light on their potential translation to clinical settings.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

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In this work it is introduced a new approach to calculate the density of liquids in terms of the energies of the acoustic signals. This method is compared to other methods in the time domain (peak-to-peak amplitudes) and frequency domain magnitudes at a single frequency. It is used a measurement cell based on a multiple reflection technique, and it is developed an acoustic model for the cell. Simulations and experiments using several liquids are presented, showing that the energy method a less sensitive to noise than the other techniques. The relative errors in the density are smaller than 0.2% when compared to the values measured with a pycnometer.

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Some modifying factors may determine the risk of brain tumors. Until now, it could not be attempted to identify people at risk and also to improve significantly disease progression. Current therapy consists of surgical resection, followed by radiation therapy and chemotherapy. Despite of these treatments, the prognosis for patients is poor. In this review, we highlight general aspects concerning genetic alterations in brain tumors, namely astrocytomas, glioblastomas, oligodendrogliomas, medulloblastomas and ependymomas. The influence of these genetic alterations in patients' prognosis is discussed. Mutagen sensivity is associated with cancer risk. The convincing studies that linked DNA damages and DNA repair alterations with brain tumors are also described. Another important modifying factor is immunity. General immune response against cancer, tumor microenvironment and immune response, mechanisms of tumor escape, CNS tumor immunology, immune defects that impair anti-tumor systemic immunity in brain tumor patients and local immunosuppressive factors within CNS are also reviewed. New hope to treatment perspectives, as dendritic-cell-based vaccines is summarized too. Concluding, it seems well established that there is association between brain tumor risk and mutagen sensivity, which is highly heritable. Primary brain tumors cause depression in systemic host immunity; local immunosuppressive factors and immunological characteristics of tumor cells may explain the poor prognosis and DNA damages responses can alert immune system. However, it is necessary to clarify if individuals with both constitutional defects in immune functions and genetic instability have higher risk of developing brain tumors. Cytogenetic prospective studies and gene copy number variations analysis also must be performed in peripheral lymphocytes from brain tumor patients. © 2011 Bentham Science Publishers Ltd.

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Objectives: The clinical translation of stem cell-based Regenerative Endodontics demands further development of suitable injectable scaffolds. Puramatrix™ is a defined, self-assembling peptide hydrogel which instantaneously polymerizes under normal physiological conditions. Here, we assessed the compatibility of Puramatrix™ with dental pulp stem cell (DPSC) growth and differentiation. Methods: DPSC cells were grown in 0.05-0.25% Puramatrix™. Cell viability was measured colorimetrically using the WST-1 assay. Cell morphology was observed in 3D modeling using confocal microscopy. In addition, we used the human tooth slice model with Puramatrix™ to verify DPSC differentiation into odontoblast-like cells, as measured by expression of DSPP and DMP-1. Results: DPSC survived and proliferated in Puramatrix™ for at least three weeks in culture. Confocal microscopy revealed that cells seeded in Puramatrix™ presented morphological features of healthy cells, and some cells exhibited cytoplasmic elongations. Notably, after 21 days in tooth slices containing Puramatrix™, DPSC cells expressed DMP-1 and DSPP, putative markers of odontoblastic differentiation. Significance: Collectively, these data suggest that self-assembling peptide hydrogels might be useful injectable scaffolds for stem cell-based Regenerative Endodontics. © 2012 Academy of Dental Materials.

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)