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1.
AAPS PharmSciTech ; 21(5): 194, 2020 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-32666160

RESUMO

For polymer-based controlled release drug products (e.g. microspheres and implants), active pharmaceutical ingredient distribution and microporosity inside the polymer matrix are critical for product performance, particularly drug release kinetics. Due to the decreasing domain size and increasing complexity of such products, conventional characterization and release test techniques are limited by their resolution and speed. In this study, samples of controlled release poly(lactic-co-glycolic acid) microspheres in the diameter range of 30-80 µm are investigated with focused ion beam scanning electron microscope imaging at 20 nm or higher resolution. Image data is quantified with artificial intelligence-based image analytics to provide size distributions of drug particles and pores within the microsphere sample. With an innovative image-based numerical simulation method, release profiles are predicted in a matter of days regardless of the designed release time. A mechanistic understanding on the impact of porosity to the interplays of drug, formulation, process, and dissolution was gained.


Assuntos
Preparações de Ação Retardada , Microscopia Eletrônica de Varredura/métodos , Microesferas , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/administração & dosagem , Inteligência Artificial , Composição de Medicamentos , Cinética , Tamanho da Partícula , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Porosidade , Solubilidade
2.
PLoS One ; 15(7): e0236842, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32730363

RESUMO

Cyanobacteria can form biofilms in nature, which have ecological roles and high potential for practical applications. In order to study them we need biofilm models that contain healthy cells and can withstand physical manipulations needed for structural studies. At present, combined studies on the structural and physiological features of axenic cyanobacterial biofilms are limited, mostly due to the shortage of suitable model systems. Here, we present a simple method to establish biofilms using the cyanobacterium Synechocystis PCC6803 under standard laboratory conditions to be directly used for photosynthetic activity measurements and scanning electron microscopy (SEM). We found that glass microfiber filters (GMF) with somewhat coarse surface features provided a suitable skeleton to form Synechocystis PCC6803 biofilms. Being very fragile, untreated GMFs were unable to withstand the processing steps needed for SEM. Therefore, we used polyhydroxybutyrate coating to stabilize the filters. We found that up to five coats resulted in GMF stabilization and made possible to obtain high resolution SEM images of the structure of the surface-attached cells and the extensive exopolysaccharide and pili network, which are essential features of biofilm formation. By using pulse-amplitude modulated variable chlorophyll fluorescence imaging, it was also demonstrated that the biofilms contain photosynthetically active cells. Therefore, the Synechocystis PCC6803 biofilms formed on coated GMFs can be used for both structural and functional investigations. The model presented here is easy to replicate and has a potential for high-throughput studies.


Assuntos
Biofilmes/crescimento & desenvolvimento , Membrana Celular/metabolismo , Microscopia Eletrônica de Varredura/métodos , Polissacarídeos Bacterianos/metabolismo , Synechocystis/crescimento & desenvolvimento , Synechocystis/ultraestrutura , Membrana Celular/ultraestrutura , Polissacarídeos Bacterianos/ultraestrutura , Synechocystis/metabolismo
3.
J Cancer Res Ther ; 16(3): 612-618, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32719276

RESUMO

Objective: The objective of this study is to evaluate the effects of radiotherapy doses on mineral density and percentage mineral volume of human permanent tooth enamel. Materials and Methods: Synchrotron radiation Xray microcomputed tomography (SRµCT) and microhardness testing were carried out on 8 and 20 tooth samples, respectively. Enamel mineral density was derived from SRµCT technique using ImageJ software. Microhardness samples were subjected to Vickers indentations followed by calculation of microhardness and percentage mineral volume values using respective mathematical measures. Data were analyzed using paired t-test at a significance level of 5%. Qualitative analysis of the enamel microstructure was done with two-dimensional projection images and scanned electron micrographs using µCT and field emission scanning electron microscopy, respectively. Results: Vickers microhardness and SRµCT techniques showed a decrease in microhardness and an increase in mineral density, respectively, in postirradiated samples. These changes were related to mineral density variation and alteration of hydroxyapatite crystal lattice in enamel surface. Enamel microstructure showed key features such as microporosities and loss of smooth homogeneous surface. These indicate tribological loss and delamination of enamel which might lead to radiation caries. Conclusions: Tooth surface loss might be a major contributing factor for radiation caries in head-and-neck cancer patients prescribed to radiotherapy. Such direct effects of radiotherapy cause enamel abrasion, delamination, and damage to the dentinoenamel junction. Suitable measures should, therefore, be worked out to protect nontarget oral tissues such as teeth while delivering effective dosages to target regions.


Assuntos
Esmalte Dentário/efeitos da radiação , Neoplasias de Cabeça e Pescoço/radioterapia , Lesões por Radiação/etiologia , Desmineralização do Dente/etiologia , Esmalte Dentário/ultraestrutura , Dureza/efeitos da radiação , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Técnicas In Vitro , Microscopia Eletrônica de Varredura/métodos , Lesões por Radiação/patologia , Propriedades de Superfície , Desmineralização do Dente/patologia , Microtomografia por Raio-X/métodos
4.
Sci Rep ; 10(1): 9395, 2020 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-32523030

RESUMO

Aquatic insects living in fast-flowing streams have developed various types of attachment systems to resist being carried away by strong currents. Combinations of various attachment devices offer aquatic insects advantages in underwater adhesion on substrates with different surface properties. In this study, the net-winged midge (Blepharicera sp.) larvae were investigated to understand micro-/nano-structural attachment mechanisms. The hierarchical structure of insect adhesive surfaces was characterized using Optical Microscopy (OM), Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM). Centrifugal measurements were also conducted to measure the critical rotational velocity at which the larvae of Blepharicera sp. can adhere to substrates with varying roughness. Commercial suckers require smooth substrate surface to maintain a pressure that is lower than the surrounding pressure for adhesion under the sucker cup while the suckers of net-winged midge larvae possess hierarchical micro-/nano-structures, which attach closely to rough surfaces underwater. Furthermore, the functions of microstructures observed on the sucker, including wrinkled surface, inward setae, outer fibers, and nick were explored and may contribute to underwater adhesion. The aligned C-shaped suckers can attach and detach effectively by closing or opening the gap. The unique microstructure and adhesion capability of such suckers could shed light on the design and synthesis of novel bio-inspired devices for reversible underwater adhesion.


Assuntos
Chironomidae/fisiologia , Extremidades/fisiologia , Larva/fisiologia , Adesividade , Animais , Insetos/fisiologia , Microscopia Eletrônica de Varredura/métodos , Sensilas/fisiologia , Propriedades de Superfície
5.
PLoS One ; 15(6): e0233536, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32479553

RESUMO

BACKGROUND: Removal of cement-retained implant fixed restorations when needed, can be challenging. Conventional methods of crown removal are time consuming and costly for patients and practitioners. This research explored the use of two different types of pulsed erbium lasers as a non-invasive tool to retrieve cemented zirconia crowns from zirconia implant abutments. MATERIALS AND METHODS: Twenty identical zirconia crowns were cemented onto 20 identical zirconia prefabricated abutments using self-adhesive resin cement. The specimens were divided into two groups for laser assisted crown removal; G1 for erbium-doped yttrium aluminum garnet laser (Er:YAG), and G2 for erbium, chromium-doped yttrium, scandium, gallium and garnet (Er,Cr:YSGG). For the G1, after the first crown removal, the specimens were re-cemented and removed again using the Er:YAG laser. Times needed to remove the crowns were recorded and analyzed using ANOVA (α = 0.05). The surfaces of the crown and the abutment were further examined using scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS) analyses. RESULTS: The average times of zirconia crown removal from zirconia abutments were 5 min 20 sec and 5 min 15 sec for the Er:YAG laser of first and second experiments (G1), and 5 min 55 sec for the Er,Cr:YSGG laser experiment (G2). No statistical differences were observed among the groups. SEM and EDS examinations of the materials showed no visual surface damaging or material alteration from the two pulsed erbium lasers. CONCLUSIONS: Both types of pulsed erbium lasers can be viable alternatives for retrieving a zirconia crown from a zirconia implant abutment. Despite operating at different wavelengths, the Er:YAG and Er,Cr:YSGG lasers, perform similarly in removing a zirconia crown from a zirconia implant abutment with similar parameters. There are no visual and elemental composition damages as a result of irradiation with pulsed erbium lasers.


Assuntos
Colagem Dentária/métodos , Prótese Dentária/métodos , Lasers de Estado Sólido/uso terapêutico , Cromo , Coroas , Érbio , Gálio , Humanos , Microscopia Eletrônica de Varredura/métodos , Cimentos de Resina , Coroa do Dente/fisiologia , Ítrio , Zircônio
6.
Medicine (Baltimore) ; 99(19): e19972, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32384447

RESUMO

The objective of this proof-of-concept study was to demonstrate the targeted delivery of erythropoietin (EPO) using magnetically guided magnetic nanoparticles (MNPs).MNPs consisting of a ferric-ferrous mixture (FeCl3·6H2O and FeCl2·4H2O) were prepared using a co-precipitation method. The drug delivery system (DDS) was manufactured via the spray-drying technique using a nanospray-dryer. The DDS comprised 7.5 mg sodium alginate, 150 mg MNPs, and 1000 IU EPO.Scanning electron microscopy revealed DDS particles no more than 500 nm in size. Tiny particles on the rough surfaces of the DDS particles were composed of MNPs and/or EPO, unlike the smooth surfaces of the only alginate particles. Transmission electron microscopy showed the tiny particles from 5 to 20 nm in diameter. Fourier-transform infrared spectroscopy revealed DDS peaks characteristic of MNPs as well as of alginate. Thermal gravimetric analysis presented that 50% of DDS weight was lost in a single step around 500°C. The mode size of the DDS particles was approximately 850 nm under in vivo conditions. Standard soft lithography was applied to DDS particles prepared with fluorescent beads using a microchannel fabricated to have one inlet and two outlets in a Y-shape. The fluorescent DDS particles reached only one outlet reservoir in the presence of a neodymium magnet. The neurotoxicity was evaluated by treating SH-SY5Y cells in 48-well plates (1 × 10 cells/well) with 2 µL of a solution containing sodium alginate (0.075 mg/mL), MNPs (1.5 mg/mL), or sodium alginate + MNPs. A cell viability assay kit was used to identify a 93% cell viability after MNP treatment and a 94% viability after sodium alginate + MNP treatment, compared with the control. As for the DDS particle neurotoxicity, a 95% cell viability was noticed after alginate-encapsulated MNPs treatment and a 93% cell viability after DDS treatment, compared with the control.The DDS-EPO construct developed here can be small under in vivo conditions enough to pass through the lung capillaries with showing the high coating efficiency. It can be guided using magnetic control without displaying significant neurotoxicity in the form of solution or particles.


Assuntos
Portadores de Fármacos/farmacologia , Sistemas de Liberação de Medicamentos/métodos , Eritropoetina/farmacologia , Nanopartículas de Magnetita , Materiais Revestidos Biocompatíveis/farmacologia , Meios de Contraste , Fármacos Hematológicos/farmacologia , Humanos , Nanopartículas de Magnetita/química , Nanopartículas de Magnetita/uso terapêutico , Teste de Materiais , Microscopia Eletrônica de Varredura/métodos , Tamanho da Partícula , Propriedades de Superfície , Traumatismos do Sistema Nervoso/terapia
7.
PLoS One ; 15(5): e0232992, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32392263

RESUMO

Although scanning electron microscopy (SEM) can generate high-resolution images of nanosized objects, it requires a high vacuum to do so, which precludes direct observations of living organisms and often produces unwanted structural changes. It has previously been reported that a simple surface modification gives rise to a nanoscale layer, termed the "NanoSuit", which can keep small animals alive under the high vacuum required for field-emission scanning electron microscopy (FE-SEM). We have previously applied this technique to plants, and successfully observed healthy petals in a fully hydrated state using SEM. The flower petals protected with the NanoSuit appeared intact, although we still lack a fundamental understanding of the images of other plants observed using FE-SEM. This report presents and evaluates a rich set of images, acquired using the NanoSuit, for a taxonomically diverse set of plant species. This dataset of images allows the surface features of various plants to be analyzed and thus provides a further complementary morphological profile. Image data can be accessed and viewed through Figshare (https://doi.org/10.6084/m9.figshare.c.4446026.v1).


Assuntos
Microscopia Eletrônica de Varredura/métodos , Nanoestruturas , Plantas/ultraestrutura , Animais , Flores/ultraestrutura , Microscopia Eletrônica de Varredura/instrumentação , Microscopia Eletrônica de Transmissão/métodos , Nanotecnologia , Propriedades de Superfície , Vácuo
8.
AAPS PharmSciTech ; 21(4): 125, 2020 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350635

RESUMO

Sunlight is important to health, but higher exposure to radiation causes early aging of the skin and skin damage that can lead to skin cancers. This study aimed at producing a stable octyl p-methoxycinnamate (OMC)-loaded nanostructured lipid carrier (NLC) sunscreen, which can help in the photoprotective effect. NLC was produced by emulsification-sonication method and these systems were composed of myristyl myristate (MM), caprylic capric triglyceride (CCT), Tween® 80 (TW), and soybean phosphatidylcholine (SP) and characterized by dynamic light scattering (DLS), zeta potential (ZP) measurement, atomic force microscopy (AFM), scanning electron microscopy (SEM), differential scanning calorimetry (DSC), and in vitro release studies. Pre-formulation studies were performed changing TW concentrations and no differences were found at concentrations of 1.0 and 2.0%. Two selected formulations were designed and showed an average size of 91.5-131.7, polydispersity index > 0.2, and a negative value of ZP. AFM presented a sphere-like morphology and SEM showed ability to form a thin film. DSC exhibited that the incorporation of OMC promoted reduction of enthalpy due to formation of a more amorphous structure. Drug release shows up to 55.74% and 30.57%, and this difference could be related to the presence of SP in this formulation that promoted a more amorphous structure; the release mechanism study indicated Fickian diffusion and relaxation. Sun protection factor (SPF) evaluation was performed using NLC and presented values around 40, considerably higher than those observed in the literature. The developed formulations provide a beneficial alternative to conventional sunscreen formulations.


Assuntos
Cinamatos/síntese química , Portadores de Fármacos/síntese química , Lipídeos/síntese química , Nanoestruturas/química , Fator de Proteção Solar/métodos , Protetores Solares/síntese química , Varredura Diferencial de Calorimetria/métodos , Cinamatos/farmacocinética , Portadores de Fármacos/farmacocinética , Liberação Controlada de Fármacos , Lipídeos/farmacocinética , Microscopia de Força Atômica/métodos , Microscopia Eletrônica de Varredura/métodos , Tamanho da Partícula , Protetores Solares/farmacocinética
9.
PLoS One ; 15(4): e0231486, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32324746

RESUMO

Vom Rath's organ, located at the distal end of the third segment of the labial palp, is one of the recognized synapomorphies of Lepidoptera (Insecta). Information about the structural and histological morphology of this organ is sparse. The structure of vom Rath's organ in four species of Nymphalidae, three frugivorous: Fountainea ryphea (Charaxinae: Anaeini), Morpho helenor achillaena (Satyrinae: Morphini) and Hamadryas epinome (Biblidinae: Ageroniini), and the nectarivorous species Aeria olena (Danainae: Ithomiini) is described by means of scanning electron microscopy and histology. The species showed significant differences in the cavity shape, setal morphology and arrangement, opening shape and location, associated with the organization of cell groups, type of axon, and degree of development. These differences do not seem to be related to feeding habit. No cell groups were found in Actinote thalia (Heliconiinae: Acraeini) and Heliconius erato phyllis (Heliconiinae: Heliconiini), and for the first time the absence of vom Rath's organ is documented in the clade Ditrysia. A terminology is proposed to improve understanding of the organ morphology, with an extensive analysis of the previous descriptions.


Assuntos
Borboletas/crescimento & desenvolvimento , Animais , Axônios/fisiologia , Microscopia Eletrônica de Varredura/métodos , Filogenia
10.
Molecules ; 25(3)2020 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-32046080

RESUMO

Alkali was used to adjust the pH and neutralize the excess acids of dough in the processing of Chinese northern steamed bread (CNSB). However, extra alkali addition generally resulted in alkalic flavor and poor appearance. The aim of this work was to investigate the role of proofed dough pH on the texture of CNSB. Correlation analysis demonstrated that the pH value of proofed dough has a significant effect on the textural properties of CNSB. The mechanism studies found that gradual acidification of dough by lactic acid bacteria is a critical factor affecting the process. Conversely, chemical acidification weakened the texture property of products and reduced the dough rheology. Scanning electron microscope (SEM) analysis showed that fermentation with starter for 12 h produced a continuous and extensional protein network in the proofed dough. Furthermore, the decreasing pH of proofed dough increased the extractability of protein in a sodium dodecyl sulfate (SDS)-containing medium and the content of free sulfhydryl (SH). The structure and content of gluten, especially influenced by gradual acidification level, change the quality of the final product. It is a novel approach to obtain an alkali-free CNSB with excellent quality by moderate gluten adjustment.


Assuntos
Ácidos/química , Álcalis/química , Pão/microbiologia , Fermentação/fisiologia , Farinha/microbiologia , Tecnologia de Alimentos/métodos , Glutens/química , Concentração de Íons de Hidrogênio , Lactobacillales/metabolismo , Microscopia Eletrônica de Varredura/métodos , Proteínas/química , Reologia , Vapor , Triticum/química
11.
Sci Rep ; 10(1): 2362, 2020 02 11.
Artigo em Inglês | MEDLINE | ID: mdl-32047210

RESUMO

Phenotypic heterogeneity is an important trait for the development and survival of many microorganisms including the yeast Cryptococcus spp., a deadly pathogen spread worldwide. Here, we have applied scanning electron microscopy (SEM) to define four Cryptococcus spp. capsule morphotypes, namely Regular, Spiky, Bald, and Phantom. These morphotypes were persistently observed in varying proportions among yeast isolates. To assess the distribution of such morphotypes we implemented an automated pipeline capable of (1) identifying potentially cell-associated objects in the SEM-derived images; (2) computing object-level features; and (3) classifying these objects into their corresponding classes. The machine learning approach used a Random Forest (RF) classifier whose overall accuracy reached 85% on the test dataset, with per-class specificity above 90%, and sensitivity between 66 and 94%. Additionally, the RF model indicates that structural and texture features, e.g., object area, eccentricity, and contrast, are most relevant for classification. The RF results agree with the observed variation in these features, consistently also with visual inspection of SEM images. Finally, our work introduces morphological variants of Cryptococcus spp. capsule. These can be promptly identified and characterized using computational models so that future work may unveil morphological associations with yeast virulence.


Assuntos
Variação Anatômica , Cryptococcus/ultraestrutura , Cápsulas Fúngicas/ultraestrutura , Aprendizado de Máquina , Microscopia Eletrônica de Varredura/métodos , Cryptococcus/genética , Fenótipo
12.
Sci Rep ; 10(1): 2368, 2020 02 11.
Artigo em Inglês | MEDLINE | ID: mdl-32047250

RESUMO

The demand for rapid, consistent and easy-to-use techniques for detecting and identifying pathogens in various areas, such as clinical diagnosis, the pharmaceutical industry, environmental science and food inspection, is very important. In this study, the reference strains of six food-borne pathogens, namely, Escherichia coli 0157: H7 ATCC 43890, Cronobacter sakazakii ATCC 29004, Salmonella Typhimurium ATCC 43971, Staphylococcus aureus KCCM 40050, Bacillus subtilis ATCC 14579, and Listeria monocytogenes ATCC 19115, were chosen for scanning electron microscopy (SEM) and energy dispersive X-ray (EDX) analysis. In our study, the time-consuming sample preparation step for the microbial analysis under SEM was avoided, which makes this detection process notably rapid. Samples were loaded onto a 0.01-µm-thick silver (Ag) foil surface to avoid any charging effect. Two different excitation voltages, 10 kV and 5 kV, were used to determine the elemental information. Information obtained from SEM-EDX can distinguish individual single cells and detect viable and nonviable microorganisms. This work demonstrates that the combination of morphological and elemental information obtained from SEM-EDX analysis with the help of principal component analysis (PCA) enables the rapid identification of single microbial cells without following time-consuming microbiological cultivation methods.


Assuntos
Bactérias/ultraestrutura , Microanálise por Sonda Eletrônica/métodos , Microscopia Eletrônica de Varredura/métodos , Análise de Célula Única/métodos
13.
Sci Rep ; 10(1): 1955, 2020 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-32029811

RESUMO

This paper reports the application of a carbon paste electrode modified with magnetite nanoparticles and the ionic liquid 1-butyl-3-methylimidazolium hexafluorophosphate in the electroanalytical determination of 17ß-estradiol and estriol. These estrogens are potential endocrine disruptors and thus it is relevant the development of devices for their monitoring. Transmission electron microscopy, scanning electron microscopy and zeta potential techniques were applied to characterization of the modifier materials. In cyclic voltammetry experiments, irreversible oxidation peaks were observed for 17ß-estradiol and estriol at +0.320 V and +0.400 V, respectively. The anodic currents obtained were approximately three times greater than those provided by the unmodified electrode due to the presence of magnetic nanoparticles and the ionic liquid, which improved the sensitivity of modified electrode. For the analysis, the parameters of the square-wave voltammetry (scan increment, amplitude and frequency) were optimized by Box-Behnken factorial design for each estrogen. For 17ß-estradiol in B-R buffer pH 12.0, the calibration plot was linear from 0.10 to 1.0 µmol L-1, with a detection limit of 50.0 nmol L-1. For estriol in B-R buffer pH 11.0, the linear range was 1.0 to 10.0 µmol L-1, with a detection limit of 300.0 nmol L-1. The modified electrode was applied in the determination of 17ß-estradiol and estriol in pharmaceutical formulations and the results were comparable to those obtained using UV/VIS spectrometry. Statistical tests were applied to evaluate the results and it was concluded that there was no significant difference regarding the precision and accuracy of the data provided by the two methods.


Assuntos
Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/métodos , Estrogênios/química , Líquidos Iônicos/química , Nanopartículas de Magnetita/química , Calibragem , Carbono/química , Eletrodos , Estradiol/química , Estriol/química , Concentração de Íons de Hidrogênio , Limite de Detecção , Microscopia Eletrônica de Varredura/métodos , Microscopia Eletrônica de Transmissão/métodos , Oxirredução
14.
Sci Rep ; 10(1): 2626, 2020 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-32060319

RESUMO

Assessing the structure and function of organelles in living organisms of the primitive unicellular red algae Cyanidioschyzon merolae on three-dimensional sequential images demands a reliable automated technique in the class imbalance among various cellular structures during mitosis. Existing classification networks with commonly used loss functions were focused on larger numbers of cellular structures that lead to the unreliability of the system. Hence, we proposed a balanced deep regularized weighted compound dice loss (RWCDL) network for better localization of cell organelles. Specifically, we introduced two new loss functions, namely compound dice (CD) and RWCD by implementing multi-class variant dice and weighting mechanism, respectively for maximizing weights of peroxisome and nucleus among five classes as the main contribution of this study. We extended the Unet-like convolution neural network (CNN) architecture for evaluating the ability of our proposed loss functions for improved segmentation. The feasibility of the proposed approach is confirmed with three different large scale mitotic cycle data set with different number of occurrences of cell organelles. In addition, we compared the training behavior of our designed architectures with the ground truth segmentation using various performance measures. The proposed balanced RWCDL network generated the highest area under the curve (AUC) value in elevating the small and obscure peroxisome and nucleus, which is 30% higher than the network with commonly used mean square error (MSE) and dice loss (DL) functions. The experimental results indicated that the proposed approach can efficiently identify the cellular structures, even when the contour between the cells is obscure and thus convinced that the balanced deep RWCDL approach is reliable and can be helpful for biologist to accurately identify the relationship between the cell behavior and structures of cell organelles during mitosis.


Assuntos
Processamento de Imagem Assistida por Computador/métodos , Redes Neurais de Computação , Rodófitas/ultraestrutura , Algoritmos , Imageamento Tridimensional/métodos , Microscopia Eletrônica de Varredura/métodos , Mitose , Organelas/ultraestrutura , Rodófitas/citologia
15.
Chem Biol Interact ; 319: 109019, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-32092302

RESUMO

The inhibition of the enzyme acetylcholinesterase (AChE) is a frequently used therapeutic option to treat Alzheimer's disease (AD). By decreasing the levels of acetylcholine degradation in the synaptic space, some cognitive functions of patients suffering from this disease are significantly improved. Rivastigmine is one of the most widely used AChE inhibitors. The objective of this work was to determine the effects of this drug on human erythrocytes, which have a type of AChE in the cell membrane. To that end, human erythrocytes and molecular models of its membrane constituted by dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE) were used. They correspond to classes of phospholipids present in the outer and inner monolayers of the human erythrocyte membrane, respectively. The experimental results obtained by X-ray diffraction and differential scanning calorimetry (DSC) indicated that rivastigmine molecules were able to interact with both phospholipids. Fluorescence spectroscopy results showed that rivastigmine produce a slight change in the acyl chain packing order and a weak displacement of the water molecules of the hydrophobic-hydrophilic membrane interface. On the other hand, observations by scanning electron microscopy (SEM) showed that the drug changed the normal biconcave shape of erythrocytes in stomatocytes (cup-shaped cells) and echinocytes (speculated shaped).


Assuntos
Doença de Alzheimer/tratamento farmacológico , Inibidores da Colinesterase/farmacologia , Eritrócitos/efeitos dos fármacos , Rivastigmina/farmacologia , Acetilcolinesterase/metabolismo , Doença de Alzheimer/metabolismo , Varredura Diferencial de Calorimetria/métodos , Forma Celular/efeitos dos fármacos , Dimiristoilfosfatidilcolina/metabolismo , Membrana Eritrocítica/efeitos dos fármacos , Membrana Eritrocítica/metabolismo , Eritrócitos/metabolismo , Humanos , Microscopia Eletrônica de Varredura/métodos , Modelos Moleculares , Fosfatidiletanolaminas/metabolismo , Fosfolipídeos/metabolismo , Espectrometria de Fluorescência/métodos , Difração de Raios X/métodos
16.
Biochem Soc Trans ; 48(1): 187-197, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32065234

RESUMO

Membrane contact sites (MCSs) are sites where the membranes of two different organelles come into close apposition (10-30 nm). Different classes of proteins populate MCSs including factors that act as tethers between the two membranes, proteins that use the MCSs for their function (mainly lipid or ion exchange), and regulatory proteins and enzymes that can act in trans across the MCSs. The ER-Golgi MCSs were visualized by electron microscopists early in the sixties but have remained elusive for decades due to a lack of suitable methodological approaches. Here we report recent progress in the study of this class of MCSs that has led to the identification of their main morphological features and of some of their components and roles. Among these, lipid transfer proteins and lipid exchange have been the most studied and understood so far. However, many unknowns remain regarding their regulation and their role in controlling key TGN functions such as sorting and trafficking as well as their relevance in physiological and pathological conditions.


Assuntos
Retículo Endoplasmático/metabolismo , Membranas Intracelulares/metabolismo , Rede trans-Golgi/metabolismo , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Transporte/metabolismo , Proteínas do Olho/metabolismo , Homeostase , Humanos , Proteínas de Membrana/metabolismo , Microscopia Eletrônica de Varredura/métodos , Microscopia de Fluorescência/métodos , Fosfatidilinositóis/metabolismo , Transporte Proteico
17.
J Vis Exp ; (156)2020 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-32065162

RESUMO

This report described sample preparation methods that scanning and transmission electron microscope observations, demonstrated by preparing appendages of the woodboring beetle, Chlorophorus caragana Xie & Wang (2012), for both types of electron microscopy. The scanning electron microscopy (SEM) sample preparation protocol was based on sample chemical fixation, dehydration in a series of ethanol baths, drying, and sputter-coating. By adding Tween 20 (Polyoxyethylene sorbitan laurate) to the fixative and the wash solution, the insect body surface of woodboring beetle was washed more cleanly in SEM. This study's transmission electron microscopy (TEM) sample preparation involved a series of steps including fixation, ethanol dehydration, embedding in resin, positioning using fluorescence microscopy, sectioning, and staining. Fixative with Tween 20 enabled penetrate the insect body wall of woodboring beetle more easily than it would had been without Tween 20, and subsequently better fixed tissues and organs in the body, thus yielded clear transmission electron microscope observations of insect sensilla ultrastructures. The next step of this preparation was determining the positions of insect sensilla in the sample embedded in the resin block by using fluorescence microscopy to increase the precision of target sensilla positioning. This improved slicing accuracy.


Assuntos
Besouros/ultraestrutura , Microscopia Eletrônica de Varredura/métodos , Microscopia Eletrônica de Transmissão/métodos , Manejo de Espécimes , Animais , Sensilas , Fixação de Tecidos
18.
Appl Spectrosc ; 74(5): 544-552, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32031010

RESUMO

Fourier transform infrared (FT-IR) spectroscopic imaging and microscopy of single living cells are established label-free technique for the study of cell biology. The constant driver to improve the spatial resolution of the technique is due to the diffraction limit given by infrared (IR) wavelength making subcellular study challenging. Recently, we have reported, with the use of a prototype zinc sulfide (ZnS) transmission cell made of two hemispheres, that the spatial resolution is improved by the factor of the refractive index of ZnS, achieving a λ/2.7 spatial resolution using the synchrotron-IR microscopy with a 36× objective with numerical aperture of 0.5. To refine and to demonstrate that the ZnS hemisphere transmission device can be translated to standard bench-top FT-IR imaging systems, we have, in this work, modified the device to achieve a more precise path length, which has improved the spectral quality of the living cells, and showed for the first time that the device can be applied to study live cells with three different bench-top FT-IR imaging systems. We applied focal plane array (FPA) imaging, linear array, and a synchrotron radiation single-point scanning method and demonstrated that in all cases, subcellular details of individual living cells can be obtained. Results have shown that imaging with the FPA detector can measure the largest area in a given time, while measurements from the scanning methods produced a smoother image. Synchrotron radiation single-point mapping produced the best quality image and has the flexibility to introduce over sampling to produce images of cells with great details, but it is time consuming in scanning mode. In summary, this work has demonstrated that the ZnS hemispheres can be applied in all three spectroscopic approaches to improve the spatial resolution without any modification to the existing microscopes.


Assuntos
Espectroscopia de Infravermelho com Transformada de Fourier/instrumentação , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Células A549 , Humanos , Lentes , Microscopia Eletrônica de Varredura/instrumentação , Microscopia Eletrônica de Varredura/métodos , Sulfetos/química , Síncrotrons/instrumentação , Compostos de Zinco/química
19.
Acta Odontol Scand ; 78(5): 321-326, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31909679

RESUMO

Objective: The objective of this study is to evaluate the wall adaptation and apical microleakage values following the application of various irrigation protocols in primary teeth.Material and methods: For the two parts of the study, extracted upper incisor primary teeth were randomly included to the 1% sodium hypochlorite (NaOCl), 10% ethylenediaminetetraacetic acid (EDTA)+1% NaOCl, 6% citric acid (CA)+1% NaOCl and 0.9% physiological saline (PS) groups. Canal wall adaptation and apical microleakage were assessed by scanning electron microscopy (SEM) and stereomicroscope, respectively.Results: 6% CA + 1% NaOCl group was found to be the most successful irrigation protocol in providing strong canal wall adaptation and less apical microleakage, followed by 10% EDTA +1% NaOCl. 6% CA +1% NaOCl was significantly superior regarding apical microleakage (p < .05).Conclusions: Due to the ability to provide appropriate changes in the root canal walls to make a well-adapted and leak-proof canal filling, 6% CA + 1% NaOCl can be recommended as an irrigation protocol in primary teeth.


Assuntos
Infiltração Dentária , Ácido Edético/administração & dosagem , Microscopia Eletrônica de Varredura/métodos , Irrigantes do Canal Radicular/farmacologia , Preparo de Canal Radicular , Camada de Esfregaço , Hipoclorito de Sódio/administração & dosagem , Dente Decíduo/diagnóstico por imagem , Cavidade Pulpar/efeitos dos fármacos , Cavidade Pulpar/ultraestrutura , Humanos , Irrigantes do Canal Radicular/administração & dosagem , Fatores de Tempo
20.
Acta Histochem ; 122(2): 151503, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31955907

RESUMO

The current study aimed to investigate the tongue (lingual) morphometry, histology, and histochemistry of two chiropterans endemic in the Egyptian fauna, and having different feeding preferences. The tongues of nine adult individuals of each species were utilized in our investigation. The tongue of fruit-eating bat was observed relatively longer than the one of insect-eating bat. Grossly, the insect-eating bat had a lingual prominence on the dorsum of lingual body, while the fruit-eating bat had a concave midline over the lingual body. Histologically, numerous forms of lingual papillae were scattered along the dorsal epithelium of the tongue. The lingual papillae of the fruit-eating bat seem to be well adapted for piercing the skin of a fruit and liquid sap retention. The lingual glands of both species were lodged in the muscular layer. Two main sets were identified; the serous von Ebner's gland usually seen accompanied by the circumvallate papillae and Weber's gland with mixed mucoserous secretions. Von Ebner's gland showed more prominent acidic mucins, while Weber's gland expressed neutral mucins. The lingual epithelium of the fruit-eating bat had an outer covering of cornified non-nucleated epithelium. On the other hand, the insect-eating bat had an outer covering of nucleated epithelium. It is for the first time to record the existence of the entoglossal plates of both species which consisted of a bony core in the fruit-eating bat and a cartilaginous element in the insect-eating bat. The current study represents an attempt to shed more light on the tongue evolution among mammalian vertebrates.


Assuntos
Epitélio/patologia , Pele/metabolismo , Papilas Gustativas/patologia , Língua/patologia , Animais , Quirópteros , Egito , Frutas/metabolismo , Microscopia Eletrônica de Varredura/métodos
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