1,720,974 research outputs found
Evaluation of nanobodies against the selected protein biomarkers of glioblastoma and attempt of their delivery with exosomes
Full text linkGlioblastom (GBM) je najpogostejši primarni možganski tumor, ki se pojavlja s pogostnostjo 3,2 primera na 100 000 prebivalcev. Kljub uveljavljenem zdravljenju, ki obsega kirurško odstranitev tumorja, kemoterapijo s temozolomidom in radioterapijo, večina bolnikov ne preživi več kot 18 mesecev po postavljeni diagnozi. Eden izmed sodobnih možnih načinov zdravljenja GBM je uporaba nanoteles, antigen-prepoznavnih delov težkoverižnih protiteles, ki jih proizvajajo le nekatere živali, npr. lame. Nanotelesa imajo namreč v primerjavi s klasičnimi protiteles precej prednosti, kot so visoka stabilnost, možnost proizvodnje z bakterijo E. coli in hitrejše prehajanje v tumor.
V okviru doktorskega dela smo proučili vpliv nanoteles na preživetje celic, proti osmim možnim označevalcem glioblastoma, njihovo migracijo in tvorjenje kolonij. Najprej smo z uporabo imunohistokemije ugotovili, da na osnovi navzočnosti biooznačevalec oziroma razlik v izražanju, vimentin lahko razlikuje med glioblastomom, gliomi nižje stopnje in normalno možganovino, medtem ko biooznačevalci TUFM, DPYSL1 in CRMP1 razlikujejo med glioblastomom in normalno možganovino. Rezultati proučevanja citotoksičnega delovanja nanoteles kažejo, da nanotelesa Nb79 (anti-vimentin), Nb179 (anti-NAP1L1), Nb225 (anti-TUFM) in Nb314 (anti-DPYSL2) delujejo citotoksično na glioblastomske celice. Posebno velik vpliv na citotoksičnost glioblastomskih matičnih celic ima nanotelo anti-TUFM (Nb225). Na migracijo glioblastomskih celic pa najbolj vpliva nanotelo anti-vimentin (Nb79), ki je popolnoma inhibiralo migracijo celic glioblastomske celične linije U87MG.
V drugem delu raziskave smo razvili dostavni sistem, ki temelji na zunajceličnih veziklih eksosomih, v katere smo zapakirali nanotelesa, da bi izboljšali njihovo dostavo in povečali učinkovitost. Eksosomi so najmanjši zunajcelični vezikli, ki jih izločajo celice, in naj bi, v primerjavi s primerljivimi dostavljalci, liposomi, hitreje prehajali v tarčne celice, obenem pa naj bi tudi imeli daljši razpolovni čas. Eksosome smo izolirali iz glioblastomske celične linije U251MG in jih opredelili z uporabo prenosa western za detekcijo eksosomalnih označevalcev, z uporabo metode sledenja nanodelcem za določitev števila in velikosti eksosomov, ter z elektronsko mikroskopijo za ugotavljanje njihove oblike. V eksosome smo nanotelesa uspešno zapakirali z metodama inkubacije z 0,4 % saponinom in sonikacije, ki sta bili približno enako učinkoviti, medtem ko posredno pakiranje nanoteles v eksosome preko inkubacije celic z nanotelesi ni bilo uspešno.
Eksosomi so poleg tega, da so potencialni dostavljalci oz. komponente sistemov za dostavo zdravil do tkiv in celic, tudi možen vir biooznačevalcev. V naši študiji smo z metodo qPCR analizirali izražanje izbranih mRNA, miRNA in proteinov v eksosomih celic glioblastomskih celičnih linij. Ugotovili smo, da so miR-9-5p, miR-124-3p, mRNA TUFM in mRNA CRMP1 možni označevalci eksosomov glioblastomskih matičnih celic, mRNA VIM pa primeren označevalec eksosomov diferenciranih glioblastomskih celic. Za razliko od omenjenih molekul RNA so bili proteini v eksosomih proučevanih celic slabše zastopani, v njih smo lahko detektirali samo proteinska biooznačevalca ALYREF in DPYSL2.
V doktorski nalogi smo pokazali, da so nanotelesa primerno sredstvo za doseganje citotoksičnega učinka in zmanjševanje migracije glioblastomskih celic. Uspešno smo razvili dostavno sredstvo, eksosome, ki vsebujejo nanotelesa, in te bi lahko v prihodnosti uporabili za povečanje učinkovitosti samih nanoteles. Naši rezultati kažejo, da so citotoksična nanotelesa in eksosomi, kot dostavni sistem, obetavna učinkovita in specifična oblika terapije za zdravljenje GBM. Vzporedno smo v naši študiji tudi določili možne eksosomalne označevalce glioblastomskih celic, za katere predlagamo ovrednotenje pri nadaljnjem proučevanju eksosomov, izoliranih iz telesnih tekočin bolnikov. Ti bi v prihodosti lahko služili kot potencialni biomarkerji GBM iz krvi ali cerebrospinalne tekočine bolnikov z GBM.Glioblastoma is the most common primary brain tumor, occurring with a frequency of 3.2 cases per 100 000 population. Despite established treatment, which includes surgical removal of the tumor, chemotherapy with temozolomide, and radiotherapy, most patients do not survive more than 18 months after diagnosis. One of the modern possible ways of treating GBM is the use of nanobodies, antigen-recognizing parts of heavy chain-only antibodies produced by only some animals, e.g. llamas. Namely, nanobodies have many advantages over classical antibodies, such as high stability, possibility of production with E. coli and faster transition and penetration to the tumor.
In the doctoral thesis, we studied the influence of nanobodies against eight possible biomarkers of glioblastoma on cell survival, migration and colony formation. First, using immunohistochemistry, we found that, based on differences in expression or presence, the biomarker vimentin could distinguish between glioblastoma, lower-grade gliomas, and normal brain, while biomarkers TUFM, DPYSL1, and CRMP1 distinguished between glioblastoma and normal brain. The results of the study of the cytotoxic action of nanobodies showed that nanobodies Nb79 (anti-vimentin), Nb179 (anti-NAP1L1), Nb225 (anti-TUFM) and Nb314 (anti-DPYSL2) have a cytotoxic effect on glioblastoma cells. The anti-TUFM nanobody (Nb225) has a particularly large effect on the cytotoxicity of glioblastoma stem cells. The migration of glioblastoma cells is mostly affected by the anti-vimentin nanobody (Nb79), which completely inhibited the migration of cells of the glioblastoma cell line U87MG.
In the second part of the study, we developed a delivery system based on extracellular vesicles, exosomes, into which we packaged nanobodies to improve their delivery and increase efficiency. Exosomes are the smallest extracellular vesicles secreted by cells and are thought to pass more rapidly into target cells than comparable delivery systems, while also having a longer half-life. Exosomes were isolated from the U251MG glioblastoma cell line cells and characterized by detection of exosomal markers using Western blot, a nanoparticle tracking analysis method to determine their number and size, and by electron microscopy to determine their shape. Nanobodies were successfully packaged into exosomes by 0.4% saponin incubation and sonication methods, which were approximately equally effective, while indirect packaging by incubating cells with nanobodies was not successful.
In addition to being carriers of a potential delivery system, exosomes are also a possible source of biomarkers. In our study, we analyzed the expression of selected mRNAs, miRNAs, and proteins in the exosomes of glioblastoma cell lines using the qPCR method. We found that miR-9-5p, miR-124-3p, TUFM mRNA, and CRMP1 mRNA are possible markers of glioblastoma stem cell exosomes, and VIM mRNA is a suitable marker of differentiated glioblastoma cell exosomes. In contrast to the mentioned RNA molecules, proteins were less represented in the exosomes of the studied cells, in which only the protein biomarkers ALYREF and DPYSL2 could be detected.
In our doctoral dissertation, we showed that nanobodies are a suitable means of achieving a cytotoxic effect and reducing the migration of glioblastoma cells. We have successfully developed a delivery vehicle, exosomes that contain nanobodies, and these could be used in the future to increase the efficiency of the nanobodies themselves. Our results suggest that cytotoxic nanobodies and exosomes, as a delivery system, are a promising form of efficient and specific therapy for the treatment of GBM. In parallel, in this study we also identified possible exosomal markers of glioblastoma cells, for which we propose evaluation in the further study of exosomes isolated from patients’ body fluids. These could in the future serve as potential biomarkers of GBM from the blood or cerebrospinal fluid of patients with GBM
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
Nanotechnology meets oncology
No full textAdvances in technology of the past decades led to development of new nanometer scale diagnosis and treatment approaches in cancer medicine leading to establishment of nanooncology. Inorganic and organic nanomaterials have been shown to improve bioimaging techniques and targeted drug delivery systems. Their favorable physico-chemical characteristics, like small sizes, large surface area compared to volume, specific structural characteristics, and possibility to attach different molecules on their surface transform them into excellent transport vehicles able to cross cell and/or tissue barriers, including the blood-brain barrier. The latter is one of the greatest challenges in diagnosis and treatment of brain cancers. Application of nanomaterials can prolong the circulation time of the drugs and contrasting agents in the brain, posing an excellent opportunity for advancing the treatment of the most aggressive form of the brain cancer-glioblastomas. However, possible unwanted side-effects and toxicity issues must be considered before final clinical translation of nanoparticles
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
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
Author-wise bibliometric analysis based on entropy.
No full textAuthor-wise bibliometric analysis based on entropy.</p
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