Navegando por Palavras-chave "Confocal microscopy"
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- ItemAcesso aberto (Open Access)Confocal fluorescence microscopy: a powerful tool in the study of Chagas' disease(Sociedade Brasileira de Medicina Tropical - SBMT, 2000-02-01) Mortara, Renato Arruda [UNIFESP]; Silva, Solange Da [UNIFESP]; Taniwaki, Noemi Nosomi [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Instituto Adolfo Lutz Seção de Microscopia EletrônicaConfocal scanning fluorescence microscopy has become widely used in cell biology and pathology. In conjunction with monoclonal antibodies it may turn out to be a powerful diagnostic tool that also enables detailed studies of tissue forms of Trypanosoma cruzi.
- ItemAcesso aberto (Open Access)Development of experimental in vitro burn model(Sociedade Brasileira para o Desenvolvimento da Pesquisa em Cirurgia, 2014-01-01) Fernandes, Ana Carolina Morais; França, Jerônimo Pereira de [UNIFESP]; Gaiba, Silvana; Aloise, Antonio Carlos [UNIFESP]; Oliveira, Andrea Fernandes de; Moraes, Andrea Aparecida Fátima Souza [UNIFESP]; França, Lucimar Pereira de; Ferreira, Lydia Masako [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Universidade Estadual de Santa Cruz Department of Biological SciencesPURPOSE:To propose an experimental burn model in NIH-3T3 cell line.METHODS: Induction of thermal injury in cultures of mouse fibroblast - NIH-3T3- cell line and determination of cell viability by MTT and imunofluorescence.RESULTS: The heating of the Petri dish increased proportionally to the temperature of the base and the time of exposure to microwave. In this in vitro burn model, using the cell line NIH-3T3 was observed drastic cellular injury with significant changes in cell viability and activity. It showed drastically modified cell morphology with altered membrane, cytoskeleton and nucleus, and low cellularity compared to the control group.CONCLUSION: The burn model in vitro using the cell line NIH-3T3 was reproductive and efficient. This burn model was possible to determine significant changes in cell activity and decreased viability, with drastic change in morphology, cell lysis and death.
- ItemSomente MetadadadosDistribution of Trypanosoma cruzi stage-specific epitopes in cardiac muscle of Calomys callosus, BALB/c mice, and cultured cells infected with different infective forms(Elsevier B.V., 2007-07-01) Taniwaki, Noerni N.; Silva, Claudio Vieira da; Silva, Solange da; Mortara, Renato A.; Universidade Federal de São Paulo (UNIFESP); Secao Microscopia Eletron Inst Adolfo LutzTo examine whether distinct parasite infective forms or the mammalian host could affect the distribution of Trypanosoma cruzi stage-specific epitopes defined by monoclonal antibodies (Mabs) raised against mammalian-stage parasite forms, immunofluorescence studies followed the intracellular life cycle of the parasite in the cardiac muscle of Calomys callosus and BALB/c mice in the acute phase of the disease and in LLC-MK(2) Cultured cells. Animals and cells were infected either with tissue-culture derived trypomastigotes (TCT) or bloodstream trypornastigotes (BT) from the Y strain of T cruzi. Samples were examined under confocal fluorescence microscopy after labeling with Mabs 2C2, 1D9, 2B7, 3G8, 3B9, and 4B9 that react with carbohydrate epitopes on Ssp-4, a major amastigote surface glycoprotein; Mab 4B5 that identifies a noncarbohydrate epitope on all intracellular parasites stages, and Mab 3B2 that also recognizes a noncarbohydrate epitope expressed only in flagellated forms. Samples were double labeled with DAPI to visualize parasites' kinetoplasts and nuclei. Most of the Mabs used in this work displayed a surface labeling pattern on amastigotes present in Calomys and mice hearts, and in LLC-MK2 cultured cells infected with BT or TCT. Mab 2B7, however, displayed a marked polymorphic distribution in antigen expression between both mammalian hosts, independent on the infective form. Beyond the polymorphic distribution of amastigote surface epitopes, Calomys, and mice heart sections presented several inflammatory cells around amastigotes and trypornastigotes nests. (c) 2007 Elsevier B.V. All rights reserved.
- ItemSomente MetadadadosKinin B-1 receptor participates in the control of cardiac function in mice(Elsevier B.V., 2007-08-16) Lauton-Santos, Sandra; Guatimosim, Silvia; Castro, Carlos H.; Oliveira, Fernando A.; Almeida, Alvair P.; Dias-Peixoto, Marco Fabricio; Gomes, Maria Aparecida; Pessoa, Phillipe; Pesquero, Jorge L.; Pesquero, Joao B.; Bader, Michael; Cruz, Jader S.; Universidade Federal de Minas Gerais (UFMG); Universidade Federal de São Paulo (UNIFESP); Max Delbruck Ctr Mol MedThe kinins have an important role in control of the cardiovascular system. They have been associated with protective effects in the heart tissue. Kinins act through stimulation of two 7-transmembrane G protein-coupled receptors, denoted B-1 and 13, receptors. However, the physiological relevance of B receptor in the heart has not been clearly established. Using B-1 kinin receptor gene knock-out mice we tested the hypothesis that the B-1 receptor plays an important role in the control of baseline cardiac function. We examined the functional aspects of the intact heart and also in the isolated cardiomyocytes to study intracellular Ca2+ cycling by using confocal microscopy and whole-cell voltage clamp techniques. We measured heart rate, diastolic and systolic tension, contraction and relaxation rates and, coronary perfusion pressure. Whole-cell voltage clamp was performed to measure L-type Ca2+ current (I-Ca,I-L) the hearts from B-1(-/-) mice showed smaller systolic tension. the average values for WT and B-1(-/-) mice were 2.6 +/- 0.04 g vs. 1.6 +/- 0.08 g, respectively. This result can be explained, at least in part, by the decrease in the Ca2+ transient (3.1 +/- 0.06 vs. 3.4 +/- 0.09 for B and WT, respectively). There was an increase in I-Ca,I-L at depolarized membrane potentials. Interestingly, the inactivation kinetics of I-Ca,I-L was statistically different between the groups. the coronary perfusion pressure was higher in the hearts from B-1(-/-) mice indicating an increase in coronary resistance. This result can be explained by the significant reduction of eNOS (NOS-3) expression in the aorta of B-1(-/-) mice. Collectively, our results demonstrate that B-1 receptor exerts a fundamental role in the mammalian cardiac function. (c) 2007 Elsevier Inc. All rights reserved.
- ItemSomente MetadadadosTooth Bleaching Increases Dentinal Protease Activity(Sage Publications Inc, 2013-02-01) Sato, Claudio Tadaaki; Rodrigues, F. A.; Garcia, Daniel Moreno; Vidal, Cristina de Mattos Pimenta; Pashley, David H.; Tjaderhane, Leo; Carrilho, Marcela Rocha de Oliveira; Nascimento, Fábio Dupart; Tersariol, Ivarne Luis dos Santos [UNIFESP]; Univ Mogi das Cruzes; Universidade Estadual de Campinas (UNICAMP); Georgia Hlth Sci Univ; Univ Oulu; Univ Western Ontario; Universidade Federal de São Paulo (UNIFESP)Hydrogen peroxide is an oxidative agent commonly used for dental bleaching procedures. the structural and biochemical responses of enamel, dentin, and pulp tissues to the in vivo bleaching of human (n = 20) premolars were investigated in this study. Atomic force microscopy (AFM) was used to observe enamel nanostructure. the chemical composition of enamel and dentin was analyzed by infrared spectroscopy (FTIR). the enzymatic activities of dental cathepsin B and matrix metalloproteinases (MMPs) were monitored with fluorogenic substrates. the amount of collagen in dentin was measured by emission of collagen autofluorescence with confocal fluorescence microscopy. the presence of Reactive Oxygen Species (ROS) in the pulp was evaluated with a fluorogenic 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) probe. Vital bleaching of teeth significantly altered all tested parameters: AFM images revealed a corrosion of surface enamel nanostructure; FTIR analysis showed a loss of carbonate and proteins from enamel and dentin, along with an increase in the proteolytic activity of cathepsin-B and MMPs; and there was a reduction in the autofluorescence of collagen and an increase in both cathepsin-B activity and ROS in pulp tissues. Together, these results indicate that 35% hydrogen peroxide used in clinical bleaching protocols dramatically alters the structural and biochemical properties of dental hard and soft pulp tissue.
- ItemSomente MetadadadosTranscorneal iontophoresis of dendrimers: PAMAM corneal penetration and dexamethasone delivery(Elsevier B.V., 2015-02-28) Souza, Joel G.; Dias, Karina; Silva, Silas A. M. [UNIFESP]; Rezende, Lucas C. D. de; Rocha, Eduardo M.; Emery, Flavio S.; Lopez, Renata F. V.; Universidade de São Paulo (USP); Universidade Federal de São Paulo (UNIFESP)Iontophoresis of nanocarriers in the eye has been proposed to sustain drug delivery and maintain therapeutic concentrations. Fourth generation polyamidoamine (PAMAM) dendrimers are semi-rigid nanoparticles with surface groups that are easily modified. These dendrimers are known to modulate tight junctions, increase paracellular transport of small molecules and be translocated across epithelial barriers, exhibiting high uptake by different cell lines. the first aim of this study was to investigate the effect of iontophoresis on PAMAM penetration and distribution into the cornea. the second aim was to evaluate, ex vivo and in vivo, the effect of these dendrimers in dexamethasone (Dex) transcorneal iontophoresis. Anionic (PAMAM G3.5) and cationic (PAMAM G4) dendrimers were labeled with fluorescein isothiocyanate (FITC), and their distribution in the cornea was investigated using confocal microscopy after ex vivo anodal and cathodal iontophoresis for various application times. the particle size distribution and zeta potential of the dendrimers in an isosmotic solution were determined using dynamic light scattering and Nanoparticle Tracking Analysis (NTA), where the movement of small particles and the formation of large aggregates, from 5 to 100 nm, could be observed. Transcorneal iontophoresis increased the intensity and depth of PAMAM-FITC fluorescence in the cornea, suggesting improved transport of the dendrimers across the epithelium toward the stroma. PAMAM complexes with Dex were characterized by C-13-NMR, H-1-NMR and DOSY. PAMAMG3.5 and PAMAMG4 increased the aqueous solubility of Dex by 10.3 and 3.9-fold, respectively; however, the particle size distribution and zeta potential remained unchanged. PAMAM G3.5 decreased the Dex diffusion coefficient 48-fold compared with PAMAM G4. the ex vivo studies showed that iontophoresis increased the amount of Dex that penetrated into the cornea by 2.9, 5.6 and 3.0-fold for Dex, Dex-PAMAM G4 and Dex-PAMAM G3.5, respectively. in vivo experiments, however, revealed that iontophoresis of Dex-PAMAM-G3.5 increased Dex concentration in the aqueous humor by 6.6-fold, while iontophoresis of Dex-PAMAM G4 and Dex increased it 2.5 and 2-fold, respectively. Therefore, iontophoresis targeted PAMAM to the cornea but it is the sustained delivery of the Dex from PAMAM that prevents its rapid elimination from the aqueous humor. in conclusion, iontophoresis of PAMAM complexes represents a promising strategy for targeted and sustained topical drug delivery to the eye. (C) 2015 Elsevier B.V. All rights reserved.
- ItemAcesso aberto (Open Access)Vesicles with charged domains(Elsevier B.V., 2010-07-01) Vequi-Suplicy, Cíntia Cristina; Riske, Karin Amaral [UNIFESP]; Knorr, Roland L.; Dimova, Rumiana; Max Planck Inst Colloids & Interfaces; Universidade de São Paulo (USP); Universidade Federal de São Paulo (UNIFESP)This work summarizes results obtained on membranes composed of the ternary mixture dioleoylphosphatidylglycerol (DOPG), egg sphingomyelin (eSM) and cholesterol (Chol). the membrane phase state as a function of composition is characterized from data collected with fluorescence microscopy on giant unilamellar vesicles. the results suggest that the presence of the charged DOPG significantly decreases the composition region of coexistence of liquid ordered and liquid disordered phases as compared to that in the ternary mixture of dioleoylphosphatidycholine, sphingomyelin and cholesterol. the addition of calcium chloride to DOPG:eSM:Chol vesicles, and to a lesser extent the addition of sodium chloride, leads to the stabilization of the two-phase coexistence region, which is expressed in an increase in the miscibility temperature. On the other hand, addition of the chelating agent EDTA has the opposite effect, suggesting that impurities of divalent cations in preparations of giant vesicles contribute to the stabilization of charged domains. We also explore the behavior of these membranes in the presence of extruded unilamellar vesicles made of the positively charged lipid dioleoyltrimethylammoniumpropane (DOTAP). the latter can induce domain formation in DOPG:eSM:Chol vesicles with initial composition in the one-phase region. (C) 2010 Elsevier B.V. All rights reserved.