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Determining the reactivity of 2D materials by testing the inhibition and adsorption of the isolated enzyme AChE
No full textRazvoj nanotehnologije je privedel do postopnega izboljšanja lastnosti nanomaterialov (NM), ki se že uporabljajo za številne aplikacije v elektrotehniki, medicini, gradbeništvu in kmetijstvu. Zato je pomembno, da poznamo in razumemo biološko reaktivnost in morebitno strupenost ter potencialna tveganja NM. V okviru magistrskega dela smo testirali biološko reaktivnost 12 različnih izbranih grafenovih oksidov (GO), ki jih uvrščamo med 2D nanomateriale. Biološko reaktivnost izbranih GO smo ugotavljali s testiranjem inhibicije in adsorpcije na izoliranem encimu acetilholinesterazi (AChE) po Ellmanovi metodi, prilagojenim na mikrotitrske plošče. Dva GO, ki sta se najbolj razlikovala v reaktivnosti, izmerjeni s testiranjem aktivnosti AChE, smo testirali še s testi citotoksičnosti na celičnih linijah A549 in Beas-2B, in sicer s testom privzema barvila nevtralno rdeče in testom z resazurinom. Ugotovili smo, da so GO zelo reaktivni materiali, saj dobimo visok delež inhibicije in adsorpcije že pri zelo nizkih koncentracijah materiala. Reaktivnost je bila odvisna od koncentracije GO, pri višjih koncentracijah GO smo izmerili manjšo aktivnost encima AChE. Iz podatkov lahko sklepamo, da je velik delež inhibicije encima AChE posledica adsorpcije GO na encim. Rezultati testov citotoksičnosti in testov inhibicije dveh izbranih GO so se ujemali, biološko bolj reaktiven GO je bil bolj citotoksičen. Rezultati korelacijske analize so pokazali, da je reaktivnost GO odvisna od razmerja med kisikom in ogljikom, ki gradita GO. Naši rezultati so pokazali, da so GO biološko reaktivni materiali, stopnja njihove reaktivnosti pa je odvisna od njihovih fizikalno-kemijskih lastnosti.The development of nanotechnology has led to a gradual improvement in the properties of nanomaterials (NMs), which are already used for a wide range of applications in electrical engineering, medicine, construction and agriculture. Therefore, it is important to know and understand the biological reactivity and potential toxicity and risks of NMs. In this MSc thesis, the biological reactivity of 12 different selected graphene oxides (GOs), classified as 2D nanomaterials, was tested. The biological reactivity of the selected GOs was determined by testing inhibition and adsorption on the isolated acetylcholinesterase (AChE) enzyme using the Ellman method adapted to microtitre plates. The two GOs that differed most in reactivity as measured by AChE activity assays were further tested by cytotoxicity assays on the A549 and Beas-2B cell lines, namely the neutral red dye uptake assay and the resazurin assay. We found that GOs are highly reactive materials, with high inhibition and adsorption rates even at very low material concentrations. The reactivity was dependent on the concentration of GO, with lower AChE activity measured at higher GO concentrations. From the data it can be concluded that much of the inhibition of the AChE is due to adsorption of GO onto the enzyme. The cytotoxicity and reactivity results of the two GOs tested in the AChE assay and cytotoxicity assays were in agreement, the more biologically reactive GO being more cytotoxic. The results of the correlation analysis showed that the reactivity of GO depends on the ratio of oxygen to carbon. Our results showed that GOs are biologically reactive materials, and that the degree of their reactivity depends on their physicochemical properties
Influence of nanomaterials on in vitro phospholipidosis induction on selected cell lines
Full text linkV doktorskem delu nas je zanimal biološki vpliv izbranih nanomaterialov, ki se lahko uporabljajo za medicinske namene. Vpliv nanomaterialov smo testirali s pasjimi ledvičnimi celicami MDCK in s človeškimi pljučnimi celicami A549 z lastnostmi alveolarnih celic tipa II. Glavni namen doktorskega dela je bil preveriti, ali lahko nanomateriali v izbranih celicah sprožijo nastanek fosfolipidoze (prekomerno znotrajcelično kopičenje fosfolipidov v lamelarnih telesih) kot enega zgodnjih znakov škodljivega vpliva nanomaterialov. Poleg tega smo želeli preveriti, ali so biološki vplivi nanomaterialov odvisni od njihove velikosti ter od njihovega raztapljanja. Na celicah MDCK, ki smo jih izpostavili necitotoksičnim koncentracijam nanodelcev ZnO, mikrodelcev ZnO in cinkove soli ZnCl2, smo zaznali poškodbe DNA le pri nanodelcih ZnO. Z merjenjem raztapljanja ZnO smo ugotovili, da sproščanje cinkovih ionov v gojišče za celice ni zadostno, da bi z njim lahko razložili škodljive vplive nanodelcev ZnO na celice. Z barvilom LipidTOX™ phospholipidosis, ki v celici obarva s fosfolipidi bogate organele, in ki se uporablja za testiranje potenciala za sprožitev fosfolipidoze, smo ugotovili, da kljub kvarnemu delovanju nanodelci ZnO ne sprožijo fosfolipidoze v celicah MDCK, kakor tudi ne v človeških pljučnih celicah A549. V celicah MDCK ni nobeden od izbranih nanomaterialov sprožil povečane količine s fosfolipidi bogatih organelov. V celicah A549 pa so nanodelci γ-Fe2O3+SiO2 sprožili povečano vsebnost s fosfolipidi bogatih organelov, povečano vsebnost fosfatidilholina in povečano vsebnost organelov, ki sodelujejo pri biogenezi lamelarnih teles. Vendar pa se je količina lamelarnih teles v izpostavljenih celicah A549 zmanjšala, povečala pa se je vsebnost avtofagnih vakuol, ki so pogosto vsebovale poškodovana lamelarna telesa, namenjena razgradnji. Rezultati na celicah A549 so pokazali, da nanodelci γ-Fe2O3+SiO2 vplivajo na metabolizem lipidov, na biogenezo lamelarnih teles in na zniževanje površinske napetosti kapljevine nad celicami. Kljub temu, da so naši rezultati pokazali, da pojav fosfolipidoze ni ustrezen indikator zgodnjih kvarnih vplivov nanomaterialov v celicah MDCK in A549, smo pokazali, da lahko nanomateriali pri necitotoksičnih koncentracijah sprožijo poškodbe DNA ter da lahko ovirajo pomembne funkcije celic.We investigated the biological effects of selected nanomaterials that have potential for medical use. Effects of nanomaterials were evaluated on the canine kidney MDCK cells and on the human lung A549 cells with properties of alveolar type II cells. The main purpose of the doctoral work was to determine whether nanomaterials can induce phospholipidosis (excessive intracellular accumulation of phospholipids in lamellar bodies) as early sign of adverse effects of nanomaterials. In addition, we wanted to check if biological effects of nanomaterials depend on their size and on their dissolution. On MDCK cells that were exposed to sub-cytotoxic concentrations of ZnO nanoparticles, ZnO microparticles and to zinc salt ZnCl2, we observed increased DNA damage only in cells exposed to nanoparticles. By measuring the ZnO nanoparticle dissolution, we found that the zinc ion release in the cell culture medium is insufficient to be the main cause of ZnO nanoparticle toxicity. By staining cells with LipidTOX™ phospholipidosis dye that is used to test potential for phospholipidosis induction, we found that ZnO nanoparticles as well as all other selected nanomaterials did not increased the amount of phospholipid rich organelles in MDCK cells. In contrast, in A549 cells, we observed that silica coated maghemite nanoparticles (γ-Fe2O3+SiO2) caused increased cellular content of phospholipids rich organelles, increased content of phosphatidylcholine (PC) and increased content of organelles that are involved in lamellar body biogenesis. Despite these results, indicating intensive lamelar body biogenesis in the exposed A549 cells, the amount of lamellar bodies had been decreased. In γ-Fe2O3+SiO2 treated A549 cells, we observed an increased amount of autophagic vacuoles that often contained defective lamellar bodies destined for degradation. Results on A549 cells showed that γ-Fe2O3+SiO2 affect lipid metabolism, interfered with the lamellar body biogenesis and reduced cell capacity to lower the surface tension of hypophase. Our results show that phospholipidosis is not good indicator of nanotoxicity in MDCK and A549 cells. Nevertheless we have shown that nanomaterials can induce DNA damage and may interfere with important functions of the cell, even at non-cytotoxic concentrations
Prilagoditev kometnega testa s kvasovkami Saccharomyces cerevisiae in izopodi Porcellio scaber za testiranje genotoksičnosti nanodelcev
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Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
In vitro cytotoxicity evaluation of the Magnéli phase titanium suboxides (TiO) on A549 human lung cells
No full textThe use of titanium suboxides, known as Magnéli phase TiO, is expected to increase in the near future due to their desirable properties. In order to use Magnéli phase TiO nanoparticles safely, it is necessary to know how nanoparticles interact with biological systems. In this study, the cytotoxicity of three different Magnéli TiO nanoparticles was evaluated using human lung A549 cells and the results were compared with hazard data on two different TiO nanoparticles whose biological interactions have already been extensively studied. After A549 cells were exposed to nanoparticles, the metabolic activity was measured by the Resazurin assay, the amount of cellular proteins was measured by the Coomassie Blue assay, and lysosomal integrity was measured by the Neutral Red Uptake assay. In order to investigate possible modes of particle actions, intracellular Ca level, reactive oxygen species (ROS) production, and photo-oxidative disruptions of lysosomal membranes were assessed. All experiments were performed in serum-containing and in serum-deprived cell culture mediums. In addition, the photocatalytic activity of Magnéli TiO and TiO nanoparticles was measured. The results show that Magnéli TiO nanoparticles increase intracellular Ca but not ROS levels. In contrast, TiO nanoparticles increase ROS levels, resulting in a higher cytotoxicity. Although Magnéli TiO nanoparticles showed a lower UV-A photocatalytic activity, the photo-stability of the lysosomal membranes was decreased by a greater extent, possibly due to particle accumulation inside lysosomes. We provide evidence that Magnéli TiO nanoparticles have lower overall biological activity when compared with the two TiO formulations. However, some unique cellular interactions were detected and should be further studied in line with possible Magnéli TiO application. We conclude that Magnéli phase nanoparticles could be considered as low toxic material same as other forms of titanium oxide particles
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
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