Placental growth factor-1 and epithelial haemato-retinal barrier breakdown: potential implication in the pathogenesis of diabetic retinopathy.

AIMS/HYPOTHESIS: Disruption of the retinal pigment epithelial (RPE) barrier contributes to sub-retinal fluid and retinal oedema as observed in diabetic retinopathy. High placental growth factor (PLGF) vitreous levels have been found in diabetic patients. This work aimed to elucidate the influence of PLGF-1 on a human RPE cell line (ARPE-19) barrier in vitro and on normal rat eyes in vivo. METHODS: ARPE-19 permeability was measured using transepithelial resistance and inulin flux under stimulation of PLGF-1, vascular endothelial growth factor (VEGF)-E and VEGF 165. Using RT-PCR, we evaluated the effect of hypoxic conditions or insulin on transepithelial resistance and on PLGF-1 and VEGF receptors. The involvement of mitogen-activated protein kinase (MEK, also known as MAPK)/extracellular signal-regulated kinase (ERK, also known as EPHB2) signalling pathways under PLGF-1 stimulation was evaluated by western blot analysis and specific inhibitors. The effect of PLGF-1 on the external haemato-retinal barrier was evaluated after intravitreous injection of PLGF-1 in the rat eye; evaluation was by semi-thin analysis and zonula occludens-1 immunolocalisation on flat-mounted RPE. RESULTS: In vitro, PLGF-1 induced a reversible decrease of transepithelial resistance and enhanced tritiated inulin flux. These effects were specifically abolished by an antisense oligonucleotide directed at VEGF receptor 1. Exposure of ARPE-19 cells to hypoxic conditions or to insulin induced an upregulation of PLGF-1 expression along with increased transcellular permeability. The PLGF-1-induced RPE cell permeability involved the MEK signalling pathway. Injection of PLGF-1 in the rat eye vitreous induced an opening of the RPE tight junctions with subsequent sub-retinal fluid accumulation, retinal oedema and cytoplasm translocation of junction proteins. CONCLUSIONS/INTERPRETATION: Our results indicate that PLGF-1 may be a potential regulation target for the control of diabetic retinal and macular oedema.

Of great tits and fleas: sleep baby sleep

Many bird parasites reduce their hosts' fitness and, as a consequence, anti-parasite behaviour such as preening and nest sanitation has evolved. These activities are time consuming and, during the day, compete directly with time devoted to foraging and food provisioning to nestlings. Moreover, infested hosts may have to allocate extra time to foraging in order to compensate for the energy loss that ectoparasites impose on the nestlings and parents. Alternatively, brooding females could, at the expense of sleeping, allocate more time to preening and nest sanitation at night. If sleeping has a short-term restoring function, one may then expect a reduction in feeding efficiency of sleep-deprived females. In this study, the effect of a haematophagous ectoparasite, the hen flea, on the activity budgets of breeding female great tits during the day and at night was investigated experimentally. Time allocated to nest sanitation increased only slightly from 0.6 % of daytime in ectoparasite-free nests to 2.8% of daytime in infested nests, thus demonstrating the higher priority given to food provisioning than parasite control. Females in infested nests reduced their sleeping time significantly (73.5% of night-time in parasite-free nests versus 48.1% in infested nests). The time freed from the reduction of sleeping time was mainly used for nest sanitation (8.3% of night-time in parasite-free nests versus 27.1% in infested nests). Despite this strong decrease in sleeping time, there was no effect of ectoparasites on the females' rate of food provisioning to nestlings.

Image-Based Coral Reef Classification and Thematic Mapping

This paper presents a novel image classification scheme for benthic coral reef images that can be applied to both single image and composite mosaic datasets. The proposed method can be configured to the characteristics (e.g., the size of the dataset, number of classes, resolution of the samples, color information availability, class types, etc.) of individual datasets. The proposed method uses completed local binary pattern (CLBP), grey level co-occurrence matrix (GLCM), Gabor filter response, and opponent angle and hue channel color histograms as feature descriptors. For classification, either k-nearest neighbor (KNN), neural network (NN), support vector machine (SVM) or probability density weighted mean distance (PDWMD) is used. The combination of features and classifiers that attains the best results is presented together with the guidelines for selection. The accuracy and efficiency of our proposed method are compared with other state-of-the-art techniques using three benthic and three texture datasets. The proposed method achieves the highest overall classification accuracy of any of the tested methods and has moderate execution time. Finally, the proposed classification scheme is applied to a large-scale image mosaic of the Red Sea to create a completely classified thematic map of the reef benthos

Miniaturized bacterial biosensor system for arsenic detection holds great promise for making integrated measurement device.

Combining bacterial bioreporters with microfluidics systems holds great promise for in-field detection of chemical or toxicity targets. Recently we showed how Escherichia coli cells engineered to produce a variant of green fluorescent protein after contact to arsenite and arsenate can be encapsulated in agarose beads and incorporated into a microfluidic chip to create a device for in-field detection of arsenic, a contaminant of well known toxicity and carcinogenicity in potable water both in industrialized and developing countries. Cell-beads stored in the microfluidics chip at -20°C retained inducibility up to one month and we were able to reproducibly discriminate concentrations of 10 and 50 μg arsenite per L (the drinking water standards for European countries and the United States, and for the developing countries, respectively) from the blank in less than 200 minutes. We discuss here the reasons for decreasing bioreporter signal development upon increased storage of cell beads but also show how this decrease can be reduced, leading to a faster detection and a longer lifetime of the device.

The blood-brain barrier: cellular basis.

Perfusion experiments with horseradish peroxidase have established that the morphological substrate of the blood-brain barrier is represented by microvascular endothelial cells. They are characterized by complexly arranged tight junctions and a very low rate of transcytotic vesicular transport. They express transport enzymes, carrier systems and brain endothelial cell-specific molecules of unknown function not expressed by any other endothelial cell population. These blood-brain barrier properties are not intrinsic to these cells but are inducible by the surrounding brain tissue. Type I astrocytes injected into the anterior eye chamber of the rat or onto the chick chorioallantoic membrane are able to induce a host-derived angiogenesis and some blood-brain barrier properties in endothelial cells of non-neural origin. Recently we have shown that this cellular interaction is due to the secretion of a soluble astrocyte derived factor(s). Astrocytes are also implicated in the maintenance, functional regulation and the repair of the blood-brain barrier. Complex interactions between other constituents of the microenvironment surrounding the endothelial cells, such as the basement membrane, pericytes, nerve endings, microglial cells and the extracellular fluid, take place and are required for the proper functioning of the blood-brain barrier, which in addition is regionally different as reflected by endothelial cell heterogeneity.

Multiplicação in vitro dos porta-enxertos de Prunus sp. 'Barrier' e 'Cadaman'

Este trabalho foi realizado com o objetivo de estabelecer a melhor concentração de sais do meio MS e da citocinina BAP para a multiplicação dos porta-enxertos de Prunus sp. 'Barrier' e 'Cadaman'. Segmentos nodais foram introduzidos em tubos de ensaio contendo 10 mL de meio de cultura com variações na concentração de sais (MS; ½MS; e 2/3MS) combinadas com cinco concentrações de BAP (0; 1,5; 2,5; 3,5 e 4,5 miM). Utilizou-se um fatorial 2x3x5, distribuído em blocos casualizados, compostos por quatro repetições contendo cinco tubos de ensaio cada uma, sendo inoculado um segmento nodal por tubo. As avaliações foram realizadas após cinco semanas de cultivo em ambiente com intensidade luminosa de 20 miE m-2 s-1, fotoperíodo de 16 horas e temperatura de 24 ± 4ºC. Verificou-se maior número médio de gemas e de brotações para a cultivar Barrier. À medida que se reduziu a concentração de sais do meio de cultura, obteve-se maior número de brotações, porém com menor tamanho. As regressões polinomiais das variáveis número de gemas, brotações por explante e comprimento das brotações apresentaram um ajustamento quadrático para níveis de BAP, atingindo os pontos de máximo 31,2 gemas/explante; 4,6 brotações por explante, e 8,1 mm de comprimento nas concentrações 3,3; 3,1, e 3,1 miM de BAP, respectivamente.

Tiivispaperin rasvanläpäisyn luonnehtiminen

Työn tavoitteena oli kehittää Metsä Tissue Oyj:n Mäntän tehtaille uusi testimenetelmä leivinpaperin rasvanläpäisyn mittaamiseen. Uudella testimenetelmällä korvataan kaksi nykyisistä testimenetelmistä, jotka ovat aikaa vieviä ja epäluotettavia. Uudelta menetelmältä vaaditaan lyhyttä kestoa, hyvää toistettavuutta ja luotettavuutta sekä korreloivuutta nykyisiin testimenetelmiin. Nykyisissä testeissä leivinpaperilla paistetaan sikaa ja kanaa uunissa, jonka jälkeen tarkastetaan leivinpaperin läpäisseen rasvan määrää leivinpaperin alla olleen lautasen puhtauden perusteella. Sikatesti on huomattavasti kriittisempi kuin kanatesti, joten tässä työssä keskitytään sikatestin korvaamiseen uudella testimenetelmällä. Työn kirjallisuusosassa perehdyttiin rasvatiiviisiin papereihin, niiden valmistukseen ja ominaisuuksiin. Lähinnä keskityttiin tiivispaperin valmistukseen ja sen silikonointiin. Lisäksi esitettiin rasvanläpäisyn mekanismeja ja niihin vaikuttavia tekijöitä sekä rasvanläpäisyn mittaamiseen kehitettyjä menetelmiä. Kokeellisen osan alussa tehtiin esikokeita olemassa olevilla testeillä, kuten Helga- ja Tappi-rasvatiiveystestillä, DIN 53116 standardin mukaisella menetelmällä sekä DPM menetelmällä. Esikokeiden tarkoituksena oli selvittää missä määrin sikatesti korreloi muiden testimenetelmien kanssa. Näistä menetelmistä ei löydetty korrelaatiota sikatestiin, joten kehitettiin aivan uusi testimenetelmä. Kokeellisessa osassa tutkittiin sikatestissä leivinpaperin pidättämiä ja läpäisemiä aineita. Testeissä huomattiin, että leivinpaperin läpäisseet aineet olivat pääosin vesiliukoisia proteiineja. Leivinpaperin pidättämässä aineessa oli vettä, vesiliukoista proteiineja ja rasvaa, siasta riippuen hyvinkin erilaisissa suhteissa. Täten todettiin sikatestin huono toistettavuus ja luotettavuus. Uudessa testimenetelmässä leivinpaperia ja sen päällä olevaa koeainetta puristetaan määrätyllä paineella määrätyssä lämpötilassa, jonka jälkeen leivinpaperin läpäissyt rasva havainnoidaan leivinpaperin alla olevasta indikaattoripaperista. Lopuksi lasketaan tietynkokoisten läpimenotahrojen lukumäärän perusteella näytteelle rasvan läpäisyindeksi. Uusi menetelmä korreloi sekä sika- että kanatestin kanssa.

Disruption of embryonic blood-CSF barrier in chick embryos reveals the actual importance of this barrier to control E-CSF composition and homeostasis in early brain development

In vertebrates, early brain development takes place at the expanded anterior end of the neural tube. After closure of the anterior neuropore, the brain wall forms a physiologically sealed cavity that encloses embryonic cerebrospinal fluid (E-CSF), a complex and protein-rich fluid that is initially composed of trapped amniotic fluid. E-CSF has several crucial roles in brain anlagen development. Recently, we reported the presence of transient blood-CSF barrier located in the brain stem lateral to the ventral midline, at the mesencephalon and prosencephalon level, in chick and rat embryos by transporting proteins, water, ions and glucose in a selective manner via transcellular routes. To test the actual relevance of the control of E-CSF composition and homeostasis on early brain development by this embryonic blood-CSF barrier, we block the activity of this barrier by treating the embryos with 6-aminonicotinamide gliotoxin (6-AN). We demonstrate that 6-AN treatment in chick embryos blocks protein transport across the embryonic blood-CSF barrier, and that the disruption of the barrier properties is due to the cease transcellular caveolae transport, as detected by CAV-1 expression cease. We also show that the lack of protein transport across the embryonic blood-CSF barrier influences neuroepithelial cell survival, proliferation and neurogenesis, as monitored by neurepithelial progenitor cells survival, proliferation and neurogenesis. The blockage of embryonic blood-CSF transport also disrupts water influx to the E-CSF, as revealed by an abnormal increase in brain anlagen volume. These experiments contribute to delineate the actual extent of this blood-CSF embryonic barrier controlling E-CSF composition and homeostasis and the actual important of this control for early brain development, as well as to elucidate the mechanism by which proteins and water are transported thought transcellular routes across the neuroectoderm, reinforcing the crucial role of E-CSF for brain development.

The embryonic blood-CSF barrier has molecular elements for specific glucose transport and for the general transport of molecules via transcellular routes.

In vertebrates, early brain development takes place at the expanded anterior end of the neural tube, which is filled with embryonic cerebrospinal fluid (E-CSF). We have recently identified a transient blood-CSF barrier that forms between embryonic days E3 and E4 in chick embryos and that is responsible for the transport of proteins and control of E-CSF homeostasis, including osmolarity. Here we examined the presence of glucose transporter GLUT-1 as well the presence of caveolae-structural protein Caveolin1 (CAV-1) in the embryonic blood-CSF barrier which may be involved in the transport of glucose and of proteins, water and ions respectively across the neuroectoderm. In this paper we demonstrate the presence of GLUT-1 and CAV-1 in endothelial cells of blood vessels as well as in adjacent neuroectodermal cells, located in the embryonic blood-CSF barrier. In blood vessels, these proteins were detected as early as E4 in chick embryos and E12.7 in rat embryos, i.e. the point at which the embryonic blood-CSF barrier acquires this function. In the neuroectoderm of the embryonic blood-CSF barrier, GLUT-1 was also detected at E4 and E12.7 respectively, and CAV-1 was detected shortly thereafter in both experimental models. These experiments contribute to delineating the extent to which the blood-CSF embryonic barrier controls E-CSF composition and homeostasis during early stages of brain development in avians and mammals. Our results suggest the regulation of glucose transport to the E-CSF by means of GLUT-1 and also suggest a mechanism by which proteins are transported via transcellular routes across the neuroectoderm, thus reinforcing the crucial role of E-CSF in brain development.

Transmission properties of high barrier coated paperboard packages

High barrier materiaaleilla pyritään pidentämään pakattujen elintarvikkeiden hyllyikää. Barrierin tärkein tehtävä on elintarvikkeen suojaaminen hapelta ja kosteudelta. Alumiinin käyttöä barriermateriaalina pyritään vähentämään korvaamalla alumiini polymeereillä, jotka täyttävät elintarvikkeiden asettamat korkeat säilyvyysvaatimukset. Etyylivinyylialkoholin (EVOH) hapenläpäisevyys on kuivissa olosuhteissa alhaisin kaupallisista muovilaaduista. EVOH tarjoaa myös erinomaisen suojan muita kaasuja, rasvoja, hajuja ja aromeja vastaan ja sitä on helppo prosessoida. Polyamideilla on erinomainen kaasutiiveys sekä hyvä lujuus ja sitkeys. Eri muovilaatuja sekoittamalla voidaan vähentää hapenläpäisyä ja parantaa prosessointia. Polyolefiineja käytetään yleisesti päällystysmateriaaleina, koska ne suojaavat tuotetta erinomaisesti kosteudelta. Hapenläpäisyllä tarkoitetaan hapen kulkeutumista materiaalin läpi joko permeaation kautta tai reikien ja vuotojen läpi. Kaasun permeoitumiseen materiaalin läpi vaikuttavat materiaalin vapaa tilavuus, kiteisyysaste, orientaatio, substituointi, suhteellinen kosteus, lämpötila, barrierkerroksen paksuus, paine-ero ja permeoituvan molekyylin ominaisuudet. Kokeellisessa osassa analysoitiin ja vertailtiin kartonkipohjaisia mehutölkkejä, joissa käytettävät high barrier materiaalit olivat EVOH ja PA. Kartonkipohjaisia alumiinitölkkejä käytettiin referenssinä. Pakkausten hapenläpäisevyysmittauksissa saatiin samasta näytteestä toistettavia tuloksia, vaikka vuotomittauksissa saadut tulokset eivät olleet vertailukelpoisia hapenläpäisytulosten kanssa. Tölkkien valmistus vaikutti oleellisesti pakkausten tiiveysominaisuuksiin. Hapenläpäisy vuotojen ja reikien läpi oli merkittävämpää kuin hapenläpäisy materiaalin läpi. Pakkausten tiiveysominaisuuksia analysoitiin mittaamalla appelsiinimehun askorbiini-happopitoisuus. Askorbiinihapon hajoaminen mitattiin koetölkkeihin pakatusta appelsiinimehusta, ja lämpötilan, valon ja hapen vaikutusta askorbiinihapon hajoamiseen tutkittiin 12 viikon ajan. Lämpötilalla oli suurin vaikutus askorbiinihapon hajoamiseen huolimatta käytetystä pakkausmateriaalista.