The release behavior of doxorubicin hydrochloride from medicated fibers prepared by emulsion-electrospinning

The release behavior of a water-soluble small molecule drug from the drug-loaded nanofibers prepared by emulsion-electrospinning was investigated. Doxorubicin hydrochloride (Dox), a water-soluble anticancer agent, was used as the model drug. The laser scanning confocal microscopic images indicated that the drug was well incorporated into amphiphilic poly(ethylene glycol)-poly(L-lactic acid) (PEG-PLA) diblock copolymer nanofibers, forming "core-sheath" structured drug-loaded nanofibers.

Handheld laser profilometry of certain medieval inscribed stones

Higher Education Authority (PRTLI as part of National Development Plan)

CONTRIBUTION OF A SPERM PROTEIN, PAWP, TO THE SIGNAL TRANSDUCT PATHWAY DURING VERTEBRATE FERTILIZATION

PAWP, postacrosomal sheath WW domain binding protein, is a novel sperm protein identified as a candidate sperm borne, oocyte-activating factor (SOAF). PAWP induces both early and later egg activation events including meiotic resumption, pronuclear formation and egg cleavage. Based on the fact that calcium increase is universally accepted as the sole requirement for egg activation, we hypothesized that PAWP is an upstream regulator of the calcium signaling pathway during fertilization. Intracellular calcium increase was detected by two-photon laser scanning fluorescence microscopy following microinjection of recombinant PAWP into Xenopus oocytes, bolstering our hypothesis and suggesting the involvement of a novel PAWP-mediated signaling pathway during fertilization. The N-terminal of PAWP shares a high homology to WW domain binding protein while the C-terminal half contains a functional PPXY motif, which allows it to interact with group I WW domain proteins. These structural considerations together with published data indicating that PPXY synthetic peptide derived from PAWP inhibits ICSI-induced fertilization led to the hypothesis that PAWP triggers egg activation by binding to a group I WW domain protein in the oocyte. By far-Western analysis of oocyte cytoplasmic fraction, PAWP was found to bind to a 52 kDa protein. The competitive inhibition studies with PPXY synthetic peptide, WW domain constructs, and their point mutants demonstrated that the interaction between PAWP and its binding partner is specifically via the PPXY-WW domain module. The 52 kDa protein band crossreacted with antibodies against group I WW domain protein YAP in Western blot assay, indicating that this 52 kDa PAWP binding partner is either YAP or a YAP-related protein. In addition, the far-Western competitive inhibition studies with recombinant GST fusion protein YAP and another WW domain-containing protein, TAZ, demonstrated that the binding of PAWP to its binding partner was significantly reduced by TAZ, providing evidence that TAZ could be the 52 kDa protein candidate. Mass spectrometry was employed to identify this PAWP binding partner candidate. However, due to the low abundance of the candidate protein and the complexity of the sample, several strategies are still needed to enrich this protein. This study correlates PAWP induced meiotic resumption and calcium efflux at fertilization and uncovers a 52 kDa candidate WW domain protein in the oocyte cytoplasm that most likely interacts with PAWP to trigger egg activation.

cAMP/PKA-dependent increases in Ca Sparks, oscillations and SR Ca stores in retinal arteriolar myocytes after exposure to vasopressin.

PURPOSE: To investigate the effects of arginine vasopressin (AVP) on Ca(2+) sparks and oscillations and on sarcoplasmic reticulum (SR) Ca(2+) content in retinal arteriolar myocytes. METHODS: Fluo-4-loaded smooth muscle in intact segments of freshly isolated porcine retinal arteriole was imaged by confocal laser microscopy. SR Ca(2+) store content was assessed by recording caffeine-induced Ca(2+) transients with microfluorimetry and fura-2. RESULTS: The frequencies of Ca(2+) sparks and oscillations were increased both during exposure to, and 10 minutes after washout of AVP (10 nM). Caffeine transients were increased in amplitude 10 and 90 minutes after a 3-minute application of AVP. Both AVP-induced Ca(2+) transients and the enhancement of caffeine responses after AVP washout were inhibited by SR 49059, a V(1a) receptor blocker. Forskolin, an activator of adenylyl cyclase, also persistently enhanced caffeine transients. Rp-8-HA-cAMPS, a membrane-permeant PKA inhibitor, prevented enhancement of caffeine transients by both AVP and forskolin. Forskolin, but not AVP, produced a reversible, Rp-8-HA-cAMPS insensitive reduction in basal [Ca(2+)](i). CONCLUSIONS: AVP activates a cAMP/PKA-dependent pathway via V(1a) receptors in retinal arteriolar smooth muscle. This effect persistently increases SR Ca(2+) loading, upregulating Ca(2+) sparks and oscillations, and may favor prolonged agonist activity despite receptor desensitization.

Endothelin 1 Stimulates Ca2+-Sparks and Oscillations in Retinal Arteriolar Myocytes via IP3R and RyR-Dependent Ca2+ Release

PURPOSE:
To investigate endothelin 1 (Et1)-dependent Ca(2+)-signaling at the cellular and subcellular levels in retinal arteriolar myocytes.
METHODS:
Et1 responses were imaged from Fluo-4-loaded smooth muscle in isolated segments of rat retinal arteriole using confocal laser microscopy.
RESULTS:
Basal [Ca(2+)](i), subcellular Ca(2+)-sparks, and cellular Ca(2+)-oscillations were all increased during exposure to Et1 (10 nM). Ca(2+)-spark frequency was also increased by 90% by 10 nM Et1. The increase in oscillation frequency was concentration dependent and was inhibited by the EtA receptor (Et(A)R) blocker BQ123 but not by the EtB receptor antagonist BQ788. Stimulation of Ca(2+)-oscillations by Et1 was inhibited by a phospholipase C blocker (U73122; 10 µM), two inhibitors of inositol 1,4,5-trisphosphate receptors (IP(3)Rs), xestospongin C (10 µM), 2-aminoethoxydiphenyl borate (100 µM), and tetracaine (100 µM), a blocker of ryanodine receptors (RyRs).
CONCLUSIONS:
Et1 stimulates Ca(2+)-sparks and oscillations through Et(A)Rs. The underlying mechanism involves the activation of phospholipase C and both IP(3)Rs and RyRs, suggesting crosstalk between these Ca(2+)-release channels. These findings suggest that phasic Ca(2+)-oscillations play an important role in the smooth muscle response to Et1 within the retinal microvasculature and support an excitatory, proconstrictor role for Ca(2+)-sparks in these vessels.

Ca(2+) sparks promote myogenic tone in retinal arterioles

Background and Purpose: Ca(2+) imaging reveals subcellular Ca(2+) sparks and global Ca(2+) waves/oscillations in vascular smooth muscle. It is well established that Ca(2+) sparks can relax arteries, but we have previously reported that sparks can summate to generate Ca(2+) waves/oscillations in unpressurized retinal arterioles, leading to constriction. We have extended these studies to test the functional significance of Ca(2+) sparks in the generation of myogenic tone in pressurized arterioles.

Experimental Approach: Isolated retinal arterioles (25-40 μm external diameter) were pressurized to 70 mmHg, leading to active constriction. Ca(2+) signals were imaged from arteriolar smooth muscle in the same vessels using Fluo4 and confocal laser microscopy.

Key Results: Tone development was associated with an increased frequency of Ca(2+) sparks and oscillations. Vasomotion was observed in 40% of arterioles and was associated with synchronization of Ca(2+) oscillations, quantifiable as an increased cross-correlation coefficient. Inhibition of Ca(2+) sparks with ryanodine, tetracaine, cyclopiazonic acid or nimodipine, or following removal of extracellular Ca(2+) , resulted in arteriolar relaxation. Cyclopiazonic acid-induced dilatation was associated with decreased Ca(2+) sparks and oscillations but with a sustained rise in the mean global cytoplasmic [Ca(2+) ] ([Ca(2+) ]c ), as measured using Fura2 and microfluorimetry.

Conclusions and Implications: This study provides direct evidence that Ca(2+) sparks can play an excitatory role in pressurized arterioles, promoting myogenic tone. This contrasts with the generally accepted model in which sparks promote relaxation of vascular smooth muscle. Changes in vessel tone in the presence of cyclopiazonic acid correlated more closely with changes in spark and oscillation frequency than global [Ca(2+) ]c , underlining the importance of frequency-modulated signalling in vascular smooth muscle.

Pressing technique and its effect on the quality of Dhaka cheese

Dhaka cheese is a semihard artisanal variety made mainly from bovine milk, using very simple pressing methods. Experimental cheeses were pressed at gauge pressures up to 31 kPa for 12 h at 24 °C and 70% RH. These cheeses were subsequently examined for their compositional, textural and rheological properties plus their microstructures investigated by confocal laser microscopy. The cheese pressed at 15.6 kPa was found to have the best compositional and structural properties.

A multi-level analytical approach for detection and visualization of intracellular NO production and nitrosation events using diaminofluoresceins

Diaminofluoresceins are widely used probes for detection and intracellular localization of NO formation in cultured/isolated cells and intact tissues. The fluorinated derivative, 4-amino-5-methylamino-2′,7′-difluorofluorescein (DAF-FM), has gained increasing popularity in recent years due to its improved NO-sensitivity, pH-stability, and resistance to photo-bleaching compared to the first-generation compound, DAF-2. Detection of NO production by either reagent relies on conversion of the parent compound into a fluorescent triazole, DAF-FM-T and DAF-2-T, respectively. While this reaction is specific for NO and/or reactive nitrosating species, it is also affected by the presence of oxidants/antioxidants. Moreover, the reaction with other molecules can lead to the formation of fluorescent products other than the expected triazole. Thus additional controls and structural confirmation of the reaction products are essential. Using human red blood cells as an exemplary cellular system we here describe robust protocols for the analysis of intracellular DAF-FM-T formation using an array of fluorescence-based methods (laser-scanning fluorescence microscopy, flow cytometry and fluorimetry) and analytical separation techniques (reversed-phase HPLC and LC-MS/MS). When used in combination, these assays afford unequivocal identification of the fluorescent signal as being derived from NO and are applicable to most other cellular systems without or with only minor modifications.

Evidence that nematodes may vector the soft rot-causing enterobacterial phytopathogens

Bacterial soft rot is a globally significant plant disease that causes major losses in the production of many popular crops, such as potato. Little is known about the dispersal and ecology of soft-rot enterobacteria, and few animals have been identified as vectors for these pathogens. This study investigates whether soil-living and bacterial-feeding nematodes could act as vectors for the dispersal of soft-rot enterobacteria to plants. Soft-rot enterobacteria associated with nematodes were quantified and visualized through bacterial enumeration, GFP-tagging, and confocal and electron scanning microscopy. Soft-rot enterobacteria were able to withstand nematode grazing, colonize the gut of Caenorhabditis elegans and subsequently disperse to plant material while remaining virulent. Two nematode species were also isolated from a rotten potato sample obtained from a potato storage facility in Finland. Furthermore, one of these isolates (Pristionchus sp. FIN-1) was shown to be able to disperse soft-rot enterobacteria to plant material. The interaction of nematodes and soft-rot enterobacteria seems to be more mutualistic rather than pathogenic, but more research is needed to explain how soft-rot enterobacteria remain viable inside nematodes.

Effect of hydrophobic plasticizers on functional properties of gelatin-based films

The aim of this study was the production and characterization of gelatin-based films using hydrophobic plasticizers derived from citric acid and soy lecithin as emulsifier. The films were characterized as to their mechanic properties, permeability to water vapor, opacity, morphology and possible interactions using Fourier transform infrared spectroscopy. Tensile strength values (TS) varied from 36 to 103 MPa, how-ever, the increase in the concentration of plasticizers (acetyltributyl citrate and tributyl citrate) reduced TS by 57% and no relation was observed between plasticizer quantities and the elongation in the quantities tested. Permeability to water vapor varied between 0.17 and 0.34 (g mm/m(2) h kPa), slightly increasing with the increase in concentration of plasticizers. The effectiveness in the use of soy lecithin emulsifier in the homogenization between the compounds could be proven by microscopic observation using confocal laser microscopy. (C) 2009 Elsevier Ltd. All rights reserved.

Desenvolvimento de argamassa para restauração utilizando resíduo do polimento do porcelanato

The coatings mortars are essential elements of building structures because they execute an important role in protecting walls and are particularly exposed to aggressive action responsible for its degradation over time. The importance of wall coverings has been the subject of discussion and analysis in the conservation and rehabilitation of old buildings. Are sometimes removed and replaced with inappropriate solutions of constructive point of view or architecture. The most commonly used coatings on walls of old buildings is based on traditional hydraulic lime mortars. The present study aims at the formulation of new lime- based mortars and aerial fine aggregate, in order to contribute to a better field of conservation and restoration mortar coating of old buildings. Residue was used for polishing porcelain as fine aggregate, replacing the aggregate (sand), in percentages 05-30% by mass. We conducted a thorough evaluation of the mortar properties in fresh and hardened state by comparing the performance of the same with a reference mortar. The residue used was characterized as the density, bulk density, and particle size laser, scanning electron microscopy, X-ray diffraction and X-ray fluorescence. Formulations were produced 7, 6 with residue and one commonly used formulation, which served as a reference. In the formulations of lime mortars air (hydrated lime powder CH-I) has been adopted a stroke volume (1:3) with constant binder, was varied and the water / binder and aggregate and waste. For evaluation of mortars fresh, proceeded to consistency analysis, specific gravity, water retention and air content embedded. In the hardened state assays were performed in specific gravity, water retention, modulus of elasticity, tensile strength in bending, compressive strength, water absorption by capillary action, adhesion, tensile strength, resistance to shrinkage and salts by of crystallization trials with resources chloride solution, nitrate and sulfate all sodium in prismatic at 90 days of age, in addition to the micro structural analysis of mortars. Based on the results we can see that the mortar formulated with 10% content of waste and the reference free retraction feature more stable closer to neutrality. The composition of 10% was obtained better performance against the action of the salt crystallization. The mortar with 15% residue obtained better density, lower air content embedded and high capacity for water retention developing good workability. The replacement of 20% of waste generates a satisfactory utilization of resistance to compression, flexion and traction grip the base. And, finally, it can be seen that the mortar with 10, 15 and 20% residual show, in principle, good suitability as coatings, thus enabling a final result consistent with durability, workability and aesthetics developing therefore a material with better performance to repair or replace existing mortars in old buildings

Monitoramento da morfologia costeira em setores da bacia potiguar sob influência da indústria petrolífera utilizando geodésia de alta precisão e laser escâner terrestre

The objective of this Doctoral Thesis was monitoring, in trimestral scale, the coastal morphology of the Northeastern coast sections of Rio Grande do Norte State, in Brazil, which is an area of Potiguar Basin influenced by the oil industry activities. The studied sections compose coastal areas with intense sedimentary erosion and high environmental sensitivity to the oil spill. In order to achieve the general objective of this study, the work has been systematized in four steps. The first one refers to the evaluation of the geomorphological data acquisition methodologies used on Digital Elevation Model (DEM) of sandy beaches. The data has been obtained from Soledade beach, located on the Northeastern coast of Rio Grande Norte. The second step has been centered on the increasing of the reference geodetic infrastructure to accomplish the geodetic survey of the studied area by implanting a station in Corta Cachorro Barrier Island and by conducting monitoring geodetic surveys to understand the beach system based on the Coastline (CL) and on DEM multitemporal analysis. The third phase has been related to the usage of the methodology developed by Santos; Amaro (2011) and Santos et al. (2012) for the surveying, processing, representation, integration and analysis of Coastlines from sandy coast, which have been obtained through geodetic techniques of positioning, morphological change analysis and sediment transport. The fourth stage represents the innovation of surveys in coastal environment by using the Terrestrial Laser Scanning (TLS), based on Light Detection and Ranging (LiDAR), to evaluate a highly eroded section on Soledade beach where the oil industry structures are located. The evaluation has been achieved through high-precision DEM and accuracy during the modeling of the coast morphology changes. The result analysis of the integrated study about the spatial and temporal interrelations of the intense coastal processes in areas of building cycles and destruction of beaches has allowed identifying the causes and consequences of the intense coastal erosion in exposed beach sections and in barrier islands

Extração automática de contornos de telhados usando dados de varredura a laser e campos randômicos de Markov

This paper proposes a methodology for automatic extraction of building roof contours from a Digital Elevation Model (DEM), which is generated through the regularization of an available laser point cloud. The methodology is based on two steps. First, in order to detect high objects (buildings, trees etc.), the DEM is segmented through a recursive splitting technique and a Bayesian merging technique. The recursive splitting technique uses the quadtree structure for subdividing the DEM into homogeneous regions. In order to minimize the fragmentation, which is commonly observed in the results of the recursive splitting segmentation, a region merging technique based on the Bayesian framework is applied to the previously segmented data. The high object polygons are extracted by using vectorization and polygonization techniques. Second, the building roof contours are identified among all high objects extracted previously. Taking into account some roof properties and some feature measurements (e. g., area, rectangularity, and angles between principal axes of the roofs), an energy function was developed based on the Markov Random Field (MRF) model. The solution of this function is a polygon set corresponding to building roof contours and is found by using a minimization technique, like the Simulated Annealing (SA) algorithm. Experiments carried out with laser scanning DEM's showed that the methodology works properly, as it delivered roof contours with approximately 90% shape accuracy and no false positive was verified.

Uso de dados fotogramétricos no refinamento geométrico de contornos de telhados de edifícios extraídos de dados laser

In this paper, a methodology is proposed for the geometric refinement of laser scanning building roof contours using high-resolution aerial images and Markov Random Field (MRF) models. The proposed methodology takes for granted that the 3D description of each building roof reconstructed from the laser scanning data (i.e., a polyhedron) is topologically correct and that it is only necessary to improve its accuracy. Since roof ridges are accurately extracted from laser scanning data, our main objective is to use high-resolution aerial images to improve the accuracy of roof outlines. In order to meet this goal, the available roof contours are first projected onto the image-space. After that, the projected polygons and the straight lines extracted from the image are used to establish an MRF description, which is based on relations ( relative length, proximity, and orientation) between the two sets of straight lines. The energy function associated with the MRF is minimized by using a modified version of the brute force algorithm, resulting in the grouping of straight lines for each roof object. Finally, each grouping of straight lines is topologically reconstructed based on the topology of the corresponding laser scanning polygon projected onto the image-space. The preliminary results showed that the proposed methodology is promising, since most sides of the refined polygons are geometrically better than corresponding projected laser scanning straight lines.

Segmentação de dados de perfilamento a laser em áreas urbanas utilizando uma abordagem Bayesiana

In this paper is presented a region-based methodology for Digital Elevation Model segmentation obtained from laser scanning data. The methodology is based on two sequential techniques, i.e., a recursive splitting technique using the quad tree structure followed by a region merging technique using the Markov Random Field model. The recursive splitting technique starts splitting the Digital Elevation Model into homogeneous regions. However, due to slight height differences in the Digital Elevation Model, region fragmentation can be relatively high. In order to minimize the fragmentation, a region merging technique based on the Markov Random Field model is applied to the previously segmented data. The resulting regions are firstly structured by using the so-called Region Adjacency Graph. Each node of the Region Adjacency Graph represents a region of the Digital Elevation Model segmented and two nodes have connectivity between them if corresponding regions share a common boundary. Next it is assumed that the random variable related to each node, follows the Markov Random Field model. This hypothesis allows the derivation of the posteriori probability distribution function whose solution is obtained by the Maximum a Posteriori estimation. Regions presenting high probability of similarity are merged. Experiments carried out with laser scanning data showed that the methodology allows to separate the objects in the Digital Elevation Model with a low amount of fragmentation.