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CUNHA, M. N. PINTO DA. "IMMUNOPROFILE IN EFFUSION CYTOLOGY." Doctoral thesis, Università degli Studi di Milano, 2010. http://hdl.handle.net/2434/150058.
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BACKGROUND: Cytology has a crucial role for diagnosing pleural and abdominal effusions. A prompt accurate diagnosis has both prognostic and therapeutic significance. However, cell morphology alone is not always sufficient to formulate such a diagnosis. In human medicine, immunocytochemistry of effusion cytology has now standardized procedures that provide reliable insights into various diagnostic dilemmas. OBJECTIVE: To describe the method of immunocytochemistry in effusion cytology and to estimate the value of a panel of
markers in identifying cells in canine and feline effusions. MATERIALS AND METHODS: Human, feline and canine mesothelial cells were isolated in culture. Western‐blot (WB) analysis was used to ascertain antibody cross‐reactivity for all the markers, with the exception of HBME‐1. Forty‐four cytospined or smeared effusion specimens from dogs and cats with a cytological diagnosis of reactive
effusion or malignancy of non‐hematopoietic origin were stained with a standard
panel of Vimentin, Cytokeratin (CK) AE1/AE3, CK 5/6 and HBME‐1 as mesothelial cell markers; desmin as mesothelial cell malignancy marker; and CK7/CK20 as a marker of metastasis. Malignancy was confirmed by histologic evaluation; non‐malignant conditions were confirmed by follow‐up. Sensitivities, specificities and predictive values were calculated. RESULTS: The WB analysis confirmed the specific crossreactivity of the human antibodies for canine and feline proteins in mesothelial tissue. No significant differences were found
between canine and feline results. Vimentin/cytokeratin coexpression had a
sensitivity of 79% and a specificity of 92%, HBME‐1 had 89% sensitivity and 23% specificity, and CK5/6 had 26% sensitivity and 100% specificity for mesothelial cells. Desmin had only 20% specificity for benign mesothelial cells, while CK7‐/CK20+ had a specificity of 79% and sensitivity of 30% for metastatic cells on effusions. CONCLUSION: Immunocytochemistry can be applied in effusion samples, and valuable results can be obtained. The most useful marker, with the highest overall accuracy for the identification of mesothelial cells in effusion, is the Vim/CK coexpression, being CK5/6 the more specific and HBME‐1
the more sensitive antibody. Desmin is not useful for discriminating between benign and malignant mesothelial cells. The coordinate expression of CK7‐/CK20+ has not proved to be useful on the identification of metastatic cells on effusions.
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Jonsson, Alexander, and Mena Said. "Metodutveckling av en vätskebaserad cytologisk metod vid preparering av exsudat : En jämförelse med konventionell cytologi." Thesis, Hälsohögskolan, Högskolan i Jönköping, HHJ, Avd. för naturvetenskap och biomedicin, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:hj:diva-31010.
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Två huvudsakliga metodprinciper används inom cytologi för diagnostisering av cellförändringar, nämligen konventionell och vätskebaserad metod. De kan båda appliceras på såväl gynekologiska som icke-gynekologiska prover, där den senare bland annat omfattar olika sorters exsudat. Syftet med den här studien var att utveckla metoden för den vätskebaserade metoden så att etanolfixerade exsudat kunde prepareras och även påvisa bättre resultat än då de preparerats med konventionell metod. För att göra detta har 61 unika prover kategoriserade som exsudat preparerats totalt, varav 61 med konventionell metod, 54 med vätskebaserad metod och 22 med vätskebaserad metod med tillsats av ättiksyra. De färdiga glasen bedömdes sedan i mikroskop och gavs scorevärden utifrån fyra parametrar: mängden celler exklusive inflammatoriska celler; bedömbarheten av cellmorfologin; mängden inflammatorisk komponent samt mängden bakgrundsmaterial. Resultaten visade ingen förbättring mellan de glas som preparerats med konventionell eller vätskebaserad metod. Däremot visade resultaten för de ättiksyrabehandlade proverna på förbättrade scorevärden jämfört med de andra metoderna. Som slutsats drogs att vätskebaserad metod med tillsats av ättiksyra uppnår syftet eftersom det reducerar mängden bakgrundsmaterial, förekomst av ring på objektglasen samt vidhåller en god cellmorfologi, vilket gör proverna lättare att diagnostisera för cytodiagnostikerna.Two main principles is used within cytology in order to diagnose cytological abnormalities; conventional and liquid-based cytology. Both methods can be applied on both gynaecological and non-gynaecological samples of which the later includes samples categorized as exudate. The aim of this study was to develop the method for liquid-based cytology so that exudate fixated with ethanol could be prepared and also achieve better results compared to conventional method. In order to do so, 61 unique samples were prepared of which 61 with conventional method, 54 with liquid-based method and 22 with liquid-based method with added acetic acid. The slides was then examined in microscope and was given score values within four parameters: amount of cells; cell morphology; amount of inflammatory component and amount of background. The results indicated no difference between the slides prepared with conventional or liquid-based method. However, the slides prepared with addition of acetic acid indicated more opportunistic score values when compared. The conclusion was that liquid-based method with the addition of acetic acid did satisfy the aim of this study as it reduces the amount of background, reduces “ring formation” on the slides and preserve the cells morphology well, which makes the samples easier to diagnose.
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Nikkilä, Heikki. "Cerebrospinal fluid cytology in schizophrenia." Helsinki : University of Helsinki, 2000. http://ethesis.helsinki.fi/julkaisut/laa/kliin/vk/nikkila/.
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Santos, Lilia Maria Antunes dos. "Cytology and ultrastructure of Eustigmatophyceae." Thesis, University of Leeds, 1990. http://etheses.whiterose.ac.uk/401/.
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Tvelve-species of the alqa class Eustigmatophyceae were studied by means of light and electron microscopy, with particular reference to structural aspects of the vegetative cells and the flagellar apparatus of the zoospores. The Vegetative cells are shown to have microfibrils (probably cellulose)' in the cell wall (Vischeria stellata), lamellate vesicles in the cytoplasm of all the species observed and a clear connection between the cfiloroplast endoplasmic reticulum and the nuclear envelope only in representatives of the Monodopsidaceae. Microfibrils (probably cellulose) were also found in the cell wall of the tribophycean species Ophiocytium malus. The most significant results on uni- and biflagellate zoospores include the observation of a Golgi body for the first time in a eustigmatophycean zoospore (Vischeria helvetica) and the first reconstruction of the system of flagellar roots in the Eustigmatophyceae (V. stellata). This consists of a rhizoplast and four microtubular roots: roots R1 (3 MTs) and R2 (2 MTs) originate-at basal body B1 and run anteriorly around the flagellar swelling; root R3 (5 MTs) arises between the basal bodies and runs to the posterior end of the cell; root R4 (2 MTs) originates at basal body B2 and curves around the eyespot. For comparison, zoospores of the tribophycean species Heterococcus marietanii and H. protonematoldes were also studied. A system of flagellar roots consisting of a small rhizoplast and three microtubular roots, two directed anteriorly and one posteriorly was confirmed. A double helix was shown to be typical of the transition region of the flagella in-this genus. The few observations on settling cells shoved the withdrawal of the complete flagellar apparatus including the swelling-and the possibility of reformation of the pyrenold in Vischeria from material stored in the spiral vesicles during the motile stage. In preliminary observations on mitosis and cytokinesis it was found that, at early stages, basal bodies appear near the nuclear surface and the chloroplast and the pyrenoid divide. Cytokinesis seems to occur by a cleavage furrow. My reconstruction of the flagellar root system in Eustigmatophyceae shows sufficient similarities with the flagellar, roots of other heterokont algal and fungal classes to justify its, inclusion with them in a single division,the Heterokontophyta. On the basis of this observation and the main ultrastructural features known for these classes,a phylogeny is, constructed for the whole group and the probable characteristics of the, common ancestor are proposed.
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Tsun, Ka-lai Obe. "Cervical cytology screening in pregnant women /." View the Table of Contents & Abstract, 2006. http://sunzi.lib.hku.hk/hkuto/record/B36586547.
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Tsun, Ka-lai Obe, and 秦家麗. "Cervical cytology screening in pregnant women." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B4501100X.
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Palmer, Ann. "Population coverage in cervical cytology programmes." Thesis, University of Edinburgh, 1988. http://hdl.handle.net/1842/19212.
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Tamar-Agha, Shawqi Yousif. "The cytology of Solenophrya micraster penard, 1914." Thesis, University of Salford, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308259.
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Haciwa, Coy Hibwaanga. "Cytology, morphology and pathogenicity of peridermium pini." Thesis, University of Cambridge, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.317680.
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Santos, Carla Susana Lima Matos Alves dos. "Cervical cytology use in portuguese urban women." Master's thesis, Faculdade de Medicina da Universidade do Porto, 2009. http://hdl.handle.net/10216/22154.
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Santos, Carla Susana Lima Matos Alves dos. "Cervical cytology use in portuguese urban women." Dissertação, Faculdade de Medicina da Universidade do Porto, 2009. http://hdl.handle.net/10216/22154.
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Lachance, Daniel. "The cytology of a Haliclona oculata (Demospongiae, Haplosclerida) /." Thesis, McGill University, 1985. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=63362.
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Seriburi, Pahnit. "Using electric cell-substrate impedance sensing (ECIS) to measure properties of an individual adherent cell /." Thesis, Connect to this title online; UW restricted, 2008. http://hdl.handle.net/1773/15465.
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Edwards, Louise Jane. "The cytology and metabolism of the rabbit oviduct epithelium." Thesis, University of York, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.358586.
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Stokes-Lampard, Helen Jayne. "Variation in NHS utilisation of vault cytology post-hysterectomy." Thesis, University of Birmingham, 2010. http://etheses.bham.ac.uk//id/eprint/825/.
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Hysterectomy is commonly performed but there is scant evidence concerning appropriate follow-up by vaginal vault cytology testing. This observational, retrospective cohort study, using routinely collected data, linked women’s entire cervical screening histories with their operation details and subsequent vault cytology test results, to establish: Which women are having hysterectomies? What was the indication? Which were followed-up? How did they differ from those who were not? 6,141 women underwent hysterectomy; an incidence of 23/10,000 women/pa. 11.61% had malignancy, 3% had CIN and 82.9% had benign disease. Median age was 48years, women were of greater deprivation and different ethnicity from the background population. Post-operatively 1,016 (16.5%) had vault cytology testing. Those having CIN at total hysterectomy should have vault cytology but only 63% had any, of these less than 10% had it according to protocol. Many factors were associated with having vault cytology (younger, less deprived, non-benign diagnosis and abnormal index cytology) but few clinically meaningful. Only 2.9% of vault cytology tests were abnormal. Efforts to identify and eradicate inappropriate use of vault tests should swiftly lead to savings. Although national guidelines are targeting the right women, it is recommended that all vaginal vault cytology should be undertaken in secondary care hereafter.
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Kirby, Simon. "Statistical discrimination in the automation of cytogenetics and cytology." Thesis, University of Edinburgh, 1990. http://hdl.handle.net/1842/15181.
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The thesis considers two topics in the automation of cytogenetics and cytology: the automated allocation of human chromosomes to the twenty-four classes which humans possess; and the detection of abnormal cervical smear specimens. For chromosome allocation, the following work is presented and evaluated on a number of data sets derived from chromosome preparations of different quality:1. Three new procedures for modelling between-cell variation.2. Six ways of combining class information on variability in multivariate Normal discrimination.3. Covariance selection models for individual chromosome classes and an assumed common covariance structure for a number of classes.4. Some two-stage procedures for the calculation of discriminant scores in multivariate Normal discrimination.5. The application of some non-parametric and semi-parametric methods.6. The modelling of band-transition sequence probabilities. For the detection of abnormal cervical smear specimens, the use of a consensus probability of a specimen being abnormal, derived from a number of cytologists' assessments, is considered. The sequential use of multiple regression equations to try to predict the logit transformations of these consensus probabilities is described. Finally, the sequential use of features in multivariate discrimination is considered mainly for the case of two known multivariate Normal populations with equal covariance matrices.
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Rechcigl, Nancy A. "Ultrastructural cytology of peanut infected with peanut stripe virus." Thesis, Virginia Polytechnic Institute and State University, 1986. http://hdl.handle.net/10919/91063.
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Two isolates of peanut stripe virus (PStV), stripe and blotch, were compared ultrastructurally in peanut (Arachis hypogaea L. 'Florigiant') at several stages of leaf expansion. Ultrathin sections of young leaves infected with either isolate of PStV revealed pinwheel inclusions attached to the cell wall near plasmodesmata. The cytoplasm of infected cells were highly vesiculated. Virus particles amassed in crystalline arrays were observed in blotch infected cells. Virus particles were observed along the arms of pinwheel inclusions. Scroll inclusions appeared in PStV infected cells at a later stage of leaf expansion. In more mature leaves, pinwheel and scroll inclusions occurred in the cytoplasm in association with mitochondria. Virus particles were observed free in the cytoplasm as well as concentrated in linear arrays along the inner surface of the tonoplast. Membrane and organelle degradation was evident in cells infected with either isolate of the virus. Numerous cytoplasmic inclusions and virus particles were observed in cells from light green areas of the leaf. Cells from dark green areas did not contain cytoplasmic inclusions and contained few if any virus particles. Particle measurements show stripe and blotch isolates to have a mean length of 753 nm and 747 nm for leaf dip preparations and 746 nm and 745 nm for partially purified preparations, respectively. Both isolates had a modal length of 750 nm, regardless of the extraction procedure.
The relative virus titer of each isolate was determined in peanut leaves at five stages of leaf expansion and in dark green and light green areas of infected leaves. Virus titer increased significantly from the closed to the fully expanded stage, at which time the virus titer peaked and then decreased slightly. Virus titer was consistently higher in leaves infected with the blotch isolate at all expansion stages. Virus titer was also higher in cells from light green areas of the leaf than from dark green areas of the leaf, regardless of isolate.M.S.
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Reale, Virginia Danielle Hikel. "Engineering a heterologous transforming growth factor-alpha promoter that is glucose-responsive in 293T cells /." View abstract, 2002. http://wilson.ccsu.edu/theses/etd-2002-15/ThesisTitlePage.html.
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Thesis (M.A.)--Central Connecticut State University, 2002.Thesis advisor: Thomas King. " ... in partial fulfillment of the requirements for the degree of Master of Arts in Biological Sciences." Includes bibliographical references (leaf 35). Also available via the World Wide Web.
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NAGASAKA, TETSURO, TOYOHARU YOKOI, TOYONORI TSUZUKI, NAGAKO MAEDA, YOSHIHIRO TOMINAGA, MAKOTO KATO, AYUMI MORIMOTO, and KATSUNORI HASHIMOTO. "Immunocytochemical Analysis for Differential Diagnosis of Thyroid Lesions Using Liquid-Based Cytology." Nagoya University School of Medicine, 2011. http://hdl.handle.net/2237/14911.
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Chiu, Man-kin, and 趙文健. "Comparison of the efficacy of sample collection for cervical cytology between the application of Cervex-Brush and Clover Brush in ThinPrepliquid-based cervical cytology." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2010. http://hub.hku.hk/bib/B45153449.
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Chen, Xiuyun. "Fine needle aspiration cytology in the study of neck mass." Thesis, Click to view the E-thesis via HKUTO, 2003. http://sunzi.lib.hku.hk/hkuto/record/B31970928.
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Harvey, Timothy John. "The development of vibrational spectroscopic cytology for prostate cancer diagnosis." Thesis, University of Manchester, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.489023.
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Brunet, Camille. "Sparse and discriminative clustering for complex data : application to cytology." Thesis, Evry-Val d'Essonne, 2011. http://www.theses.fr/2011EVRY0018/document.
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Les thèmes principaux de ce mémoire sont la parcimonie et la discrimination pour la modélisation de données complexes. Dans un première partie de ce mémoire, nous nous plaçons dans un contexte de modèle de mélanges gaussiens: nous introduisons une nouvelle famille de modèles probabilistes qui simultanément classent et trouvent un espace discriminant tel que cet espace discrimine au mieux les groupes. Une famille de 12 modèles est introduite et se base sur deux idées clefs: tout d'abord, les données réelles vivent dans un sous-espace latent de dimension intrinsèque plus petite que celle de l'espace observé; deuxièmement, un sous-espace de dimensions K-1 est suffisant pour discriminer K groupes; enfin, l'espace observé et celui latent sont liés par une transformation linéaire. Une procédure d'estimation, appelée Fisher-EM, est proposée et améliore la plupart du temps les performances de clustering grâce à l'utilisation du sous-espace discriminant. Puisque chaque axe engendrant le sous-espace discriminant est une combinaison linéaire des variables d'origine, nous avons proposé trois méthodes différentes basées sur des critères pénalisés afin de faciliter l'interprétation des résultats. En particulier, ces méthodes permettent d'introduire de la parcimonie directement dans les composantes de la matrice de projection et peut se traduite comme une étape de sélection de variables discriminantes pour la classification. Dans une seconde partie, nous nous plaçons dans le contexte de la sériation. Nous proposons une mesure de dissimilarités basée sur le voisinage commun qui permet d'introduire de la parcimonie dans les données. Une procédure algorithmique appelée l'algorithme PB-Clus est introduite et permet d'obtenir une représentation diagonale par blocs des données. Cet outil permet de révéler la structure intrinsèque des données même dans le cas de données fortement bruitées ou de recouvrement de groupes. Ces deux méthodes ont été validées dans le cadre d'une application biologique basée sur la détection de cellules cancéreusesThe main topics of this manuscript are sparsity and discrimination for modeling complex data. In a first part, we focus on the GMM context: we introduce a new family of probabilistic models which both clusters and finds a discriminative subspace chosen such as it best discriminates the groups. A family of 12 DLM models is introduced and is based on two three-ideas: firstly, the actual data live in a latent subspace with an intrinsic dimension lower than the dimension of the observed space; secondly, a subspace of K-1 dimensions is theoretically sufficient to discriminate K groups; thirdly, the observation and the latent spaces are linked by a linear transformation. An estimation procedure, named Fisher-EM is proposed and improves, most of the time, clustering performances owing to the use of a discriminative subspace. As each axis, spanning the discriminative subspace, is a linear combination of all original variables, we therefore proposed 3 different methods based on a penalized criterion in order to ease the interpretation results. In particular, it allows to introduce sparsity directly in the loadings of the projection matrix which enables also to make variable selection for clustering. In a second part, we deal with the seriation context. We propose a dissimilarity measure based on a common neighborhood which allows to deal with noisy data and overlapping groups. A forward stepwise seriation algorithm, called the PB-Clus algorithm, is introduced and allows to obtain a block representation form of the data. This tool enables to reveal the intrinsic structure of data even in the case of noisy data, outliers, overlapping and non-Gaussian groups. Both methods has been validated on a biological application based on the cancer cell detection
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Palumbo, Michael O. "Identification, isolation and characterization of proinsulin producing thymic cells." Thesis, McGill University, 2007. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=103277.
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The finding that more than 152 tissue-restricted antigens are expressed by thymic medullary epithelial cells is redefining the importance of thymic central tolerance induction in the prevention of autoimmune diseases. One of the tissue-restricted antigens in the thymus is proinsulin, and in both mice and humans, reduced thymic proinsulin levels have been shown to predispose to Type 1 diabetes. Using transgenic mice expressing a functional beta-Galactosidase gene under the regulation of the Ins2 promoter we have determined that between 1-3% of all medullary thymic epithelial cells express proinsulin and that these cells are frequently part of the Hassall's Corpuscles like structures in mice. Using a cross between the beta-Galactosidase expressing mice and Immortomice (expressing SV40 large T Antigen under the regulation of the MHC I promoter), we have isolated and cultured two proinsulin and two non-proinsulin producing medullary epithelial cell lines. Microarray analysis and RT-PCR analysis of the cell lines revealed the over-expression of approximately 50 genes (>4 fold or more) in the proinsulin producing lineage, versus the non proinsulin producing lineage, and approximately half the over-expressed genes can be considered tissue-restricted antigens. We do not find any evidence for chromosomal clustering of the over-expressed genes nor do we report the expression of any other pancreatic n-cell antigens or specific pancreatic proinsulin regulatory proteins (Pdx-1, Glut-2 or GCK) within the proinsulin producing cell lines but we do detect their expression in whole thymus. Our results suggest that chromosomal clustering is not a phenomenon associated with thymic tissue-restricted antigen expression and that the mechanisms allowing for thymic tissue-restricted antigen expression are not related to the expression mechanisms of such antigens in peripheral tissues.
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Josef, Keith. "The nonlinear phototaxis signaling network of Chlamydomonas investigated by observing ciliary responses of individual cells to green and red light." Related electronic resource: Current Research at SU : database of SU dissertations, recent titles available full text, 2005. http://wwwlib.umi.com/cr/syr/main.
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Gee, Gretchen V. "Mechanisms restricting the cellular tropism of the human Polyomavirus JCV /." View online version; access limited to Brown University users, 2005. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:3174607.
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O'Shaughnessy, Margaret Clare. "Nitric oxide mediated effects on bone cells." Thesis, Imperial College London, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.367610.
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Braim, Shwana. "Thermoresponsive magnetic colloidal gels for in vitro cell expansion." Thesis, University of Nottingham, 2016. http://eprints.nottingham.ac.uk/36832/.
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Recent studies and clinical trials have shown the potential of cell-based therapies for the treatment of a number of diseases and organ/ tissue damages. However, limited availability of some therapeutically important cells (i.e. adult stem cells) still remain as main challenges in the development of tissue engineering through to the clinic. Healthy cells are required in large numbers to form a tissue-engineered construct and primary cells must therefore be expanded in vitro for both scientific and clinical applications. Various strategies have been developed to expand cells in vitro with increasing emphasis on 3D matrices because it can provide microenvironments which more closely mimic in vivo systems. In this way the inherent difficulties associated with 2D culture such as loss of phenotype could be overcome. Moreover, 3D matrices provide higher surface areas to support expansion of larger cell numbers compared to monolayer culture. Although each 3D method has certain advantages, there is no single technique that can be used to produce material assemblies that address all the fundamental problems linked to 3D cell seeding (penetration into the scaffold), passaging (use of enzymes), and harvesting (cell yield). Recently, thermally reversibly-associating particles have been studied for the growth and support of multiple cell types and for delivery of therapeutic cells. But coupling of thermoresponsive properties to magnetic microspheres would enhance the 3D culture and expansion of multiple cell types, and facilitate rapid recovery of the expanded cell population by simple magnetic separation. In this study, it was proposed that the thermoresponsive properties would allow simple cell seeding at temperatures below the LCST of polymer stabiliser when the suspension is flowing and upon heating to above the LCST cells would be encapsulated and cultured within the particle gels (every cells surrounded by a number of particles, as the size of the particles are much smaller than the cells). The magnetic responsive property would allow efficient and scaffold free cell recovery after expansion without the need for using trypsin or enzymatic treatment. The ‘switchable’ component of reversibly associating colloidal microparticles were prepared via two different strategies. In the first strategy, thermoresponsive PDEGMA was physically adsorbed onto the surface of PS microspheres, whereas, in the second strategy, PDEGMA was chemically grafted from functionalised PCMS microspheres via SI-ATRP. The most simple method i.e. physical adsorption is rapid and can be adapted to many microparticle surfaces but has the drawback of possible desorption of polymer chains during extended use. The chemical grafting method i.e. the formation of covalent bonds between the polymer corona and the microparticle core provides robust and well defined materials but is more complex and time-consuming. In both cases, particle aggregation in their suspensions occurred on increasing the temperature to above the LCST of PDEGMA, but could be reversed by cooling the suspensions back to below the LCST. This confirmed the presence of the thermoresponsive polymer on the surface of the microspheres using both methods (adsorption and grafting). Rheological measurements demonstrated that the viscoelasticity of the prepared particle gels can be tuned, enabling these gels to have the mechanical properties that should facilitate their applications as 3D cell scaffolds for in vitro expansion of cells. Cell culture studies showed that these microparticle based scaffolds can support expansion of clinically relevant cell types (human MSC) and allowed efficient cell recovery after proliferation without the need for using trypsin or enzymatic treatment. Overall, those results suggest that the designed scaffolds had great potential for 3D in vitro cell expansion. The new developed materials have excellent biocompatibility, allow simple and rapid cell seeding and cell recovery after expansion, and possess mechanical strength and stability to support cell growth and proliferation. The materials developed and studied in this thesis may represent a significant contribution to the fields of biomaterials, tissue engineering, 3D cell culture and even bio-separation.
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Shan, Jianguo. "The role of hnrnps in a2re-mediated rna trafficking in oligodendrocytes and neurons /." [St. Lucia, Qld.], 2002. http://www.library.uq.edu.au/pdfserve.php?image=thesisabs/absthe16858.pdf.
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Yang, Fan. "Fabrication and characterization of a micro electroporation cell chip /." View abstract or full-text, 2004. http://library.ust.hk/cgi/db/thesis.pl?MECH%202004%20YANG.
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Thesis (M. Phil.)--Hong Kong University of Science and Technology, 2004.Includes bibliographical references (leaves 79-83). Also available in electronic version. Access restricted to campus users.
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Aladdad, Afnan. "Dynamic patterned electrospun fibres for 3D cell culture." Thesis, University of Nottingham, 2016. http://eprints.nottingham.ac.uk/33895/.
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Current culture methods to generate large quantities of cells destined for tissue engineering and regenerative medicine commonly use enzymatic digestion. However, this method is not desirable for subsequent cell transfer to the body due to the destruction of important cell-surface proteins and the risk of enzymatic contamination [1]. Therefore, research has led to the development of thermo-responsive surfaces for the continued culture of mammalian cells, with passaging achieved via a drop in the culture temperature. Recognising that the three-dimensional (3D) culture environment influences the cell phenotype, our aim was to generate a thermo-responsive 3D fibre-based scaffold, using electrospinning, to create an enzyme-free 3D culture surface for mammalian cell expansion that would be suitable for cells destined for the clinic. Thermo-responsive poly (poly (ethylene glycol) methacrylate), poly (PEGMA188), with lower critical solution temperature (LCST) of 26°C has been proposed for use within this thesis. It was used in combination with poly (lactic-co-glycolic acid) (PLGA) and poly (ethylene terephthalate) (PET) polymers in order to create 3D thermo-responsive non-woven electrospun fibrous scaffolds, on which different cell types could be cultured and passaged. Poly (PEGMA188) was prepared by free radical polymerization, and then incorporated with PLGA and PET polymers via four different methods: (i) surface adsorption, (ii) NaOH surface treatment, (iii) surface entrapment and (iv) blend-electrospinning. Blend-electrospinning was chosen over the other methods as it produced nano-PET and micro-PLGA bead-less fibres with responsive behaviour. The biocompatibility was assessed via the adhesion and proliferation of different mammalian cell types, including (i) red fluorescent protein (RFP)-expressing 3T3 fibroblasts, (ii) green fluorescent protein (GFP)-expressing primary immortalized human mesenchymal stem cells (ihMSCs), (iii) human colon adenocarcinoma cells (Caco2) and (iv) primary human corneal stromal stem cells (hCSSCs). The cell viability (Alamar Blue assay) was determined to measure the difference in cell populations adherent to the scaffolds while changing the culture temperature. These thermo-responsive scaffolds were able to support cell adhesion and proliferation at 37°C (hydrophobic surface). Furthermore, it was possible to detach the cells from the scaffolds by decreasing the temperature to 17°C (hydrophilic surface). Irrespective of the concentration of poly (PEGMA188) used, all scaffolds exhibited thermo-responsive proprieties; the cells were viable and proliferated in a similar manner to those cultured on control surfaces (PLGA or PET scaffolds). Finally, the effects of the thermo-responsive polymer and 3D culture environment on the hCSSC phenotype were assessed by quantitative reverse transcription-polymerase chain reaction (RT-qPCR) and immunocytochemistry. The application of 3D environments can promote the reversion of activated corneal stromal cells’ ‘fibroblastic phenotype’ to a desirable quiescent keratocyte phenotype. Therefore, seven thermal and enzymatic passages on responsive 3D scaffolds and 2D TCPS, respectively, were performed. Cell culture on the 3D scaffolds promoted the quiescent keratocyte phenotype, with the increased expression of the keratocyte markers, CD34 and ALDH, and decreased expression of the myofibroblast marker, ACTA2, when compared with cells cultured on the 2D culture flasks. In this thesis, the preparation and application of first generation, biocompatible thermo-responsive fibrous scaffolds are described. The combination of ease of preparation, positive cell response and the expansion of a desirable cell phenotype make the thermo-responsive fibres promising as a new class of materials for application in cell culture. The materials developed and studied in this thesis are believed to represent a significant contribution to the fields of biomaterials and tissue engineering.
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Rabiaa, Entedhar Kadhum Hussain. "An in vitro model of the impact of chemotherapy on neural stem cells and the protection provided by cells in the neurogenic niche." Thesis, University of Nottingham, 2018. http://eprints.nottingham.ac.uk/49274/.
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Chemotherapy has been highly successful in treating many forms of cancer; however there are increasing reports that this treatment causes cognitive declines in cancer survivors. These effects have been called “chemobrain” and while not affecting all patients, can persist for many years after the completion of treatment. The symptoms of chemobrain include a decline in concentration, memory and attention which are associated with a lower quality of life and in ability to return to work. Very little is known about the mechanisms behind these changes, or even the brain regions that are affected. Animal studies have found that systemic chemotherapy causes a decrease in the proliferation of neural stem cells (NSCs) in the subgranular zone (SGZ) neurogenic niche of the hippocampus and a decline in spatial memory. As hippocampal neurogenesis is required for a number of memory functions including the consolidation of long term memory, a decline in neurogenesis is likely to be one of the causes of the cognitive decline experienced by patients after chemotherapy. Previous animal work has shown that chronic treatment with the antidepressant fluoxetine prevents the decrease in neurogenesis and the associated cognitive decline. In the absence of fluoxetine chemotherapy spares dividing cells which are in contact with the surface of blood vessels in the SGZ of the hippocampus. This project used in vitro techniques to firstly look at whether fluoxetine has a direct effect on the sensitivity of neural cells to chemotherapy or whether treatment of astrocytes cells with fluoxetine produces an indirect effect on sensitivity. Secondly the effect of contact between NSCs and either astrocytes or endothelial cells was investigated to see if these cell types could provide protection to NSCs from chemotherapy. In the third part of the project, fluorescence activated cell sorting (FACS) and measurements of oxygen consumption after different drug treatments were used to investigate DNA damage, apoptosis and changes in the cell cycle and the metabolic response of different cell types to chemotherapy respectively. To do this we evaluated the relative sensitivity of primary NSCs, neural N2a cells, endothelial cells HBMEC, primary astrocytes, C6 astrocytes and 3T3 cells to the chemotherapy drug 5-fluorouracil (5-FU). NSCs were found to be more sensitive to 5-FU than other cell types. A concentration of 5 μM 5-FU was found to reduce NSC viability by 50% but largely to spare astrocytes and endothelial cells. FACS analysis showed that 5-FU was causing DNA damage and inducing apoptosis with some cells arresting in the G1 stage of the cell cycle. Measurements of oxygen consumption were not conclusive in explaining the differences in sensitivity between different cell types. Treatment with fluoxetine, either directly to neural cell cultures or via conditioned media from fluoxetine treated astrocytes or endothelial cells, was not found to be protective against 5-FU treatment. In contrast co-culture of NSCs with astrocytes or endothelial cells, protected NSCs from chemotherapy treatment. This effect was specific to cells in the neurogenic niche as co-culture with 3T3 fibroblasts did not provide protection. Direct cell contact between neural cells and astrocytes or endothelial cells was required for protection as neither conditioned media nor co-culture separated by a porous membrane was found to be effective. To investigate the mechanism behind astrocyte or endothelial contact dependent protection of neural cells, the gap junction protein Cx34 was stained for and found to be present on neural, astrocytic and endothelial cells. Fluorescent dye loading of marked cells showed that gap junctions were functional and dye could pass between them. Blocking gap junctions with the gap junction inhibitor carbenoxolone (CBX) abolished the protection provided by contact with astrocytes or endothelial cells. I hypothesise that the protection provided in vivo by fluoxetine may be mediated by its anti-inflammatory effect on the brain and is not a direct effect on cells in the stem cell niche. Contact with astrocytes or endothelial cells may provide Ca2+ buffering via gap junctions and in this way protect the more sensitive neural cells from the effects of chemotherapy.
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Amaral, Fabio M. R. "Gene expression network analysis of the routes to pluripotency." Thesis, University of Nottingham, 2018. http://eprints.nottingham.ac.uk/49714/.
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Great progress has been made towards the understanding of the molecular mech- anisms driving factor induced somatic cell reprogramming to pluripotency since the discovery by Takahashi and Yamanaka. However this process remains highly stochastic and inef cient. More study is needed in order to achieve a more de- terministic cell fate conversion which could further improve the quality of stem cells generated, essential for prospective therapeutic applications. The work presented here was developed under the premise that natural embryonic devel- opment can serve as a guide to achieve more ef cient pluripotency induction. It was observed that the histone variant H2A.Z, which has a role in pluripotency in embryonic stem cells, is highly expressed in the oocyte and upon over-expression, together with Pou5f1, Sox2, Klf4 and Myc, was able to increase the ef ciency of somatic cell reprogramming to induced pluripotency. A gene co-expression network analysis of somatic cells being reprogrammed identi ed hub genes as- sociated with H2Af.z and chromatin remodelling related genes which could be tested for further improving the reprogramming ef ciency induced by H2Af.z over-expression. Moreover, the study of genetic networks from pre-implantation embryos identi ed preserved genetic circuits also present during the course of reprogramming. The most preserved network modules are associated with the nal stages of pluripotency induction. However the analysis also identi ed a genetic network associated with the zygotic genome activation in the totipotent embryo stage which is also found in a sub-population of pluripotent stem cells characterised by the expression of genes from the Zscan4 family, Tet1, Etv5 and Mga among other genes. This provocative observation led me to hypothesise that during the course of reprogramming the forced activation of hub genes from such network may help improve its ef ciency, possibly by recapitulating the natural embryonic processes which induces totipotency prior to pluripotency. The identi- cation of preserved network modules and its hub genes presented in this work may serve a platform for further reprogramming studies in a quest for improved cocktails of reprogramming factors capable of more ef ciently generating induced pluripotent stem cells.
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Bailey, John Paul. "Cytology and breeding behaviour of giant alien Polygonum species in Britain." Thesis, University of Leicester, 1989. http://hdl.handle.net/2381/9471.
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An investigation into the cytology and breeding behaviour of the large asiatic Reynoutria species (R. japonica and R.sachalinensis), which were introduced into the British Isles in the last century and are now a significant component of the British Flora. A number of colonies have been examined morphologically and cytologically; hybrids have been identified and artificially re-synthesised in the laboratory. The sex-expression of the Reynoutria taxa has been examined, and has been found to be gynodioecious. R. japonica var. japonica has been found only as male-sterile plants in the British Isles, which has made it particularly susceptible to hybridisation with Fallopia baldschuanica (the commonly grown garden plant Russian Vine). Seed production and viability, and the role of seed production in the colonization of Britain has also been investigated. Chromosome counts and karyotypes have been produced of Reynoutria taxa and of the closely related Fallopia species. 2c DNA amounts have been determined, and Giemsa and fluorescent banding techniques employed. A comprehensive synonomy has been produced, and the relationship between the genera Fallopia and Reynoutria discussed. One conclusion of this research is that the genus Reynoutria should be incorporated into the older genus Fallopia.
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Pipes, L. D. "Cytology and ultrastructure of new, rare and interesting algae from Tasmania." Thesis, University of Leeds, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.383756.
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Al-Jumaily, Rakad Mohammed Khamas. "Molecular cytology of hypoxic cancer and stem cells : an epigenetic approach." Thesis, Keele University, 2018. http://eprints.keele.ac.uk/5582/.
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Oxygen is an essential factor for life in many organisms. Oxygen concentrations vary widely across different human tissues, but in general, are much lower than the 21% oxygen in atmospheric air. In spite of this, most research continues to be based on the culture of cells in an air oxygen (21% O2) environment. This ignores the mounting evidence of the role of physiological oxygen levels in the maintenance of survival, proliferation, stemness, genetic and epigenetic changes. The positive benefits of low oxygen tension on the maintenance of properties of several cells including embryonic and mesenchymal stem cells are well established while data describing negative impacts via genomic stability are few and conflicting. The role of cytosine modifications in cancer cells in response to hypoxia is poorly understood either in vitro or in vivo. In this study, we aimed to determine the role of low oxygen in the regulation of DNA methylation marks (5mC and 5hmC) and their associated genes including DNMT1/3A/3B/3L and TET1/2/3 in different cell lines including human embryonic stem cells, human mesenchymal stem cells and cancer cells. In addition to above, we also sought to corroborate upon and extend previous studies describing the effect of reduced oxygen on a range of cellular aspects including proliferation, metabolic activity, stemness and differentiation. To achieve these aims we cultured cells in either air oxygen, a fully defined 2% O2 environment (Hypoxycool media, tri-gas workstation), or in a multiuser tri-gas incubator with handling in a standard biological safety cabinet. Routine culture of stem cells in physiological normoxia enhanced the functional profile of stem cell populations including proliferation, metabolic activity and stemness. In contrast, culture of cancer cells in reduced oxygen caused significant decreases in growth profiles (vs. air oxygen). Quantitative RT-PCR and Western blots results of cells cultured in reduced oxygen revealed significant transcriptional translational downregulation of DNMT3B and TET1 vs. air oxygen (except for TET1 in cancer cell lines cultured in 2% O2 where it increased significantly), accompanied by significantly reduced in levels of 5mC and 5hmC (again except for 5hmC in cancer cell lines cultured in 2% O2 where significant increases were noted). Noteworthy was that the downregulation in gene expression of DNMT3B was associated with an increase in its CpG promoter methylation. Importantly, these changes observation was associated with increases HIF2A, at protein levels in most cell types investigated. The role of physiological oxygen in these changes was confirmed by transitioning cancer cell lines between 2% O2 and air oxygen and detailing the reversibility of both DNMT3B and DNMT3L expression at mRNA level and promoters CpG island methylation. Together these data suggest that the level of 5mC is reduced by HIF2A, via DNMT3B- mediated methylation. In conclusions, our cells display oxygen-sensitive methylation patterns where de novo methylation is linked to oxygen culture and correlates directly with transcriptional and translational regulation of the de novo methylase DNMT3B. Delineating cancer and stem cell biology under niche-like culture conditions will ultimately enhance our understanding of mechanisms of action promoting improvements to both tumour-targeting medicines and regenerative medicine applications.
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Taukobong, Tshegofatso Martha. "The visual literacy of Grade 10 Life Sciences learners in cytology." Diss., University of Pretoria, 2017. http://hdl.handle.net/2263/65471.
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In Life Sciences Education, the use of educational external representations (ERs) such as diagrams, models and animations are increasingly appearing in learning and teaching resources. However, their effectiveness is limited if learners experience learning difficulties due to lack of visual literacy and spatial ability skills to work with ERs. The study explored the level of visual literacy of 225 Grade 10 Life Sciences learners in cytology across six secondary schools in Pretoria, Gauteng. It was theorised that ERs need to be integrated in the Life Sciences curricula to develop learners’ visual literacy and spatial ability skills. With this theory, the study aimed to explore the visual literacy of Grade 10 Life Sciences learners and the influence of gender and school location on the visual literacy and spatial ability skills of the learners. Through a quantitative research method a Life Sciences visual literacy questionnaire and a spatial ability test were used to collect data. Collected data was analysed descriptively and inferentially through Statistical Package Social Sciences Version 23. The results showed that most Grade 10 Life Sciences learners lack average visual literacy skills. Furthermore, the results showed that gender doesn’t play a role on the learners’ performance in visual literacy skills as both genders performed equally in both tests, On the other hand, the results showed that the location of the school (urban, rural or township) has an effect on the learners’ performance in visual literacy skills. Teachers need to incorporate different ERs that would stimulate different senses and which will also enhance learners’ visual literacy and spatial ability skills in their lessons. A conclusion and some recommendations for future research are given.Dissertation (MEd)--University of Pretoria, 2017.
Science, Mathematics and Technology Education
MEd
Unrestricted
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Nihan, Laura. "Conjunctival Impression Cytology Assessment of Vitamin A Status of Migrant Children." DigitalCommons@USU, 1995. https://digitalcommons.usu.edu/etd/5437.
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Subclinical vitamin A deficiency was assessed in 65 Hispanic children attending four migrant Head Start programs in Utah. Subjects aged 2 to 6 years (median 3 years 10 months) were examined for evidence of vitamin A deficiency by conjunctival impression cytology. Biochemical indices for serum vitamin A, retinol-binding protein, zinc, and iron were performed.
Of eight children (12.5%) with subclinical vitamin A deficiency, one child had a marginal serum vitamin A of 11 μg/dl. Retinol-binding protein concentrations were significantly lower in two subjects with abnormal conjunctival impression cytology. Serum zinc, which when low can mimic signs of ocular vitamin A lesions, was normal for all 65 subjects. Fifteen children (23%) had iron-deficiency anemia.
Logistic regression was the central method of analysis used in this study. The results of the statistical analyses indicated there was a correlation value (0.31) between abnormal conjunctival impression cytology and serum vitamin A, which supports the hypothesis that abnormal conjunctiva! impression cytology is concurrent with decreased serum vitamin A.
Assessment of vitamin A status of Hispanic migrant children by impression cytology was effective in identifying children at risk for hypovitaminosis A. Beyond vitamin A's role in vision and maintenance of epithelium, it is also required for growth and hematopoiesis. The children of migrant workers may be suffering physiologically important consequences of vitamin A and iron deficiency that can be prevented by screening with biochemical and histological testing. Nutrition intervention for deficient children is warranted.
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Machado, Patrícia da Silva. "Epitypification, cytology, genetic and physiological variability, and genomic analysis of Puccinia psidii." Universidade Federal de Viçosa, 2015. http://www.locus.ufv.br/handle/123456789/6474.
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Designou-se, neste trabalho, o epítipo de Puccinia psidii Winter, originário de Psidium guajava para servir como referência para futuros estudos taxonômicos e moleculares do fungo. Esta ferrugem é considerada autoécia e macrocíclica, com a fase de pícnio desconhecida e com poucas informações sobre a formação, condição nuclear e o papel dos basidiósporos no ciclo vital de P. psidii. Assim, a partir de estudos citológicos, encontrou-se que após a fusão nuclear e migração do núcleo diplóide do teliósporo para o metabasídio ocorre meiose e originam-se quatro núcleos haplóides. Formam-se septos entre cada um dos quatro núcleos, os quais podem sofrer divisão mitótica e resultar em quatro basidiósporos predominantemente binucleados. A partir de inoculações de basidiósporos em folhas destacadas de Syzygium jambos, observaram-se apressórios lobados rudimentares, mas sem evidência de penetração e infecção. Em outro estudo, seis marcadores microssatélites foram utilizados para analisar a variabilidade genética de 79 isolados de P. psidii coletados em diferentes hospedeiros da Austrália, Nova Caledônia e China. Até o presente momento os isolados são geneticamente uniformes, exceto para algumas mutações pontuais em quatro isolados da Austrália. Ainda que a correlação seja pequena (p< 0.01, r= 0.061), encontrou-se associação entre variabilidade genética de isolados de P. psidii do Brasil, estimada por microssatélites, e os resultados de inoculação cruzadas em diferentes espécies de mirtáceas. Realizou-se o sequenciamento completo e uma breve análise genômica de cinco isolados P. psidii do viiiBrasil e da Austrália. A montagem de cada isolado foi comparada com o isolado VIC42496. Mais de 70% dos contigs de cada isolado que não estavam presente na montagem do isolado VIC 42496 não exibiram homologia com nenhuma sequência depositada no banco de dados do NCBI. Para aquelas sequências que apresentaram similaridade com outras sequências fúngicas, incluindo P. psidii, apenas três contigs foram similares com genes que codificam produtos envolvidos em patogenicidade.
In this study, the epitype of Puccinia psidii Winter from Psidium guajava was designed to serve as reference for future taxonomic and molecular studies of the fungus. This rust is it considered autoecious and macrocyclic, but lacking pycnium and with little knowledge about the formation, nuclear condition and the role of basidiospores in the life cycle of P. psidii. Thus, cytological studies showed that after nuclear fusion the diploid nucleus migrated into the metabasidium and underwent meiotic division resulting in four haploid nuclei. Septum was formed between each nucleus, which can undergo mitotic division and result in four basidiospores mainly binucleate. After inoculations on detached leaves of Syzygium jambos, rudimentary lobate apressoria were observed, but with no evidence of penetration and infection. In another study, six microsatellite markers were used to analyze the genetic variability of 79 isolates of P. psidii collected from different hosts in Australia, New Caledonia and China. Up to now it was found that isolates are genetically uniform, except for a few point mutations in four isolates from Australia. Although the correlation is low (p <0.01, r = 0.061), there was an association between the genetic variability of isolates of P. psidii from Brazil, estimated by microsatellites, and the results of cross-inoculation on different Myrtaceae species. Genome sequencing of five isolates of Puccinia psidii from Brazil and Australia and a brief genomic analysis among them was provided. The assembly of each isolate was compared to VIC 42496. The analysis reveled that more than 70% of the contigs of xeach isolate that was not present on assembly of VIC42496 had no significant homology (no hits) to anything currently residing in the fungal genome databases. For those contigs that displayed similar to fungal sequences, including P. psidii, only three contigs matched with genes coding for products that could be involved in pathogenicity.
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Lytvynenko, D. S. "Sir John Bertrand Gurdon and his contribution to histology, cytology and embryology." Thesis, Сумський державний університет, 2013. http://essuir.sumdu.edu.ua/handle/123456789/31965.
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Sir John Bertrand Gurdon (born 2 October 1933) is a British developmental embryologist. He is best known for his pioneering research in nuclear transplantation and cloning. He was awarded the Lasker Award in 2009. In 2012, he and Shinya Yamanaka were awarded the Nobel Prize for Physiology or Medicine for the discovery that mature cells can be reprogrammed to become pluripotent. His Nobel Lecture was called "The Egg and the Nucleus: A Battle for Supremacy".
When you are citing the document, use the following link http://essuir.sumdu.edu.ua/handle/123456789/31965
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GRIBBINS, KEVIN MICHAEL. "THE CYTOLOGY OF SPERMATOGENESIS AND ULTRASTRUCTURE OF THE SEMINIFEROUS EPITHELIUM IN REPTILES." University of Cincinnati / OhioLINK, 2003. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1053715753.
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Kosmacek, Elizabeth Anne Ianzini Fiorenza Mackey Michael A. "Live cell imaging technology development for cancer research." [Iowa City, Iowa] : University of Iowa, 2009. http://ir.uiowa.edu/etd/388.
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Yang, Zhongfa. "The role of GA binding protein (GABP) transcription factor in myeloid differentiation and cell cycle progression /." View online version; access limited to Brown University users, 2005. http://wwwlib.umi.com/dissertations/fullcit/3174701.
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Mackin, Nancy A. "The spatial and temporal regulation of morphogenesis in the budding yeast Saccharomyces cerevisiae." Related electronic resource: Current Research at SU : database of SU dissertations, recent titles available full text, 2006. http://proquest.umi.com/login?COPT=REJTPTU0NWQmSU5UPTAmVkVSPTI=&clientId=3739.
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Iyoho, Anthony E. "Modeling and analysis of nonlinear biological systems." Diss., Columbia, Mo. : University of Missouri-Columbia, 2007. http://hdl.handle.net/10355/6012.
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Thesis (Ph. D.)--University of Missouri-Columbia, 2007.The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed on December 28, 2007) Includes bibliographical references.
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Xie, Xun. "Leukocyte rolling and firm adhesion in the microvasculature : functional significance for leukocyte recruitment in inflammation /." Stockholm, 1998. http://diss.kib.ki.se/1998/91-628-2830-4.
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Camirand, Anne. "Glycosyltransferases from pea membranes : glucose and fucose incorporation into cell wall polysaccharides." Thesis, McGill University, 1986. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=75336.
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Synthesis from UDP-($ sp{14}$C) glucose of charged lipid-linked glucosyl compounds by pea membranes was short-lived, and of very limited magnitude compared to the synthesis of 1,4- and 1,3-linked B-glucans. Lipid-linked monophosphoryl glucose was the only charged lipid formed at initial stages, and had properties similar to that of dolichol-monophosphoryl glucose. It exhibited no turnover during pulse-chase experiments. Lipid-linked pyrophosphoryl-glucose or -oligosaccharides were not detected. Coumarin inhibited the synthesis of SDS-soluble products and glucans, but not of the lipid-P-glucose. Transfer of the label from endogeneous lipid-P-($ sp{14}$C) glucose or from dolichol-P-($ sp3$H) glucose into non-lipid products was minimal. It was concluded that the lipid-linked phosphoryl saccharide formed from UDP-glucose was not an obligate intermediate in the formation of B-glucans in pea membranes.Fucose-containing lipid-linked intermediates were not involved in the biosynthesis of xyloglucans. However, pea microsomal membranes catalysed the transfer of $ lbrack sp{14}{ rm C} rbrack$-fucose from GDP-$ lbrack sp{14}$C) fucose, with or without added unlabelled UDP-glucose, UDP-xylose or UDP-galactose, to an insoluble product with properties characteristic of xyloglucan. After digestion of the ethanol-insoluble pellet with Streptomyces griseus endocellulase, $ lbrack sp{14}$C) fucose residues occurred exclusively in a fragment identified as the xyloglucan nonasaccharide, Glc$ sb4$ Xyl$ sb3$ Gal Fuc. By comparison, in incubations with UDP-$ lbrack sp3$H) xylose, the maximum size of labeled oligosaccharide found following cellulase digestion of products was an octasaccharide. In the presence of both GDP-$ lbrack sp{14}$C) -fucose and UDP-$ lbrack sp3$H) xylose, a nonasaccharide containing both labels was produced. Fucose and xylose residues were transferred rapidly to acceptor molecules of MW up to 300,000. Such products did not elongate detectably over 60 min of incubation. We concluded that the nonasaccharide subunit of xyloglucan was generated in vitro by transfucosylation to preformed acceptor chains, and that its synthesis was dependent on exogenous GDP-fucose.
Microsomal membranes were separated by rate-zonal centrifugation on renografin gradients. Transfer to xyloglucan of labelled fucose and xylose from GDP- ($ sp{14}$C) fucose and UDP- ($ sp{14}$C) xylose occurred mainly in dictyosome-enriched fractions. No transferase activity was detected in secretory vesicle fractions. Pulse-chase experiments using pea stem slices incubated with ($ sp3$H) fucose suggested that xyloglucan chains are fucosylated and their structure completed within the dictyosomes, before being transported to the cell wall by secretory vesicles.
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Thansa, Kwanta. "Novel approaches to the isolation of farm animal embryonic stem cells." Thesis, University of Nottingham, 2009. http://eprints.nottingham.ac.uk/10918/.
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The establishment of stable immortal ES cell lines using embryos as a source of isolation in domesticated farm animals, in particular for pigs, which are closer to humans than other ungulates, has not been reported; hence this information could contribute to the improvement of regenerative medicine in humans, biotechnology and agriculture. Therefore, the discovery of effective protocols to derive and maintain ES cells and the induction of purified somatic cells from ES cells in pigs is of importance. The objectives of this study were to produce pES-like cells and direct differentiation of the ES-like cells obtained by improving the culture conditions. In vivo-derived porcine blastocysts at day 6-8 were classified into two groups distinguished by the exhibition of ICMs and epiblasts of the embryos. In each group, intact blastocysts and isolated ICMs or epiblasts were designed to culture in either KO4bh or DM40bh medium on mitotically inactivated MEFs under the humidified air of 5%CO2 at 39°C until the primary outgrowth of ES-like cells was observed. Two morphologically distinct pES-like cells, pESA-like and pESB-like cells were isolated from the epiblasts, whereas no cell lines were generated from ICMs. pESA-like cells were observed as individual small round cells containing one or multiple nucleoli along with a high ratio of nucleus to cytoplasm, while pESB-like cells formed dome-like colonies. The pESA-like cells were stained both negative and positive with the alkaline phosphatase enzyme, while pESB-like cells were all stained positive. With immunofluorescence staining of OCT-4 and nanog, the nuclei of pESB-like cells appeared not to be stained positive with these two antibodies, while the designed self-renewing genes such as OCT-4, nanog, SOX-2, REX-1 and DPPA-3 were detectable as similar to mES cells. Regarding the pluripotent abilities of pESB-like cells, they could be induced to form neuronal-like, neuronal supporting-like, smooth muscle-like and hepatic-like cells in a variety of desirable differentiation media under the feeder-free culture system. The cytoplasmic contents of certain induced mature cells were stained positive with nestin, α-smooth muscle actin and α-fetoprotein in association with the expression of differentiated genes specific to each germ layer such as nestin, α-smooth muscle actin, smooth muscle myosin heavy chain, α-cardiac actin, transthyretin, α-fetoprotein, albumin and HGF1β. In conclusion, pESB-like cells obtained in this study may possibly have the potential to be authentic ES cells isolated from early epiblast origin as mES cells.
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Uhiara, Chukwuemeka Obinna. "The role of extracellular signal-regulated kinase in beta-adrenoceptor-mediated vasodilatation." Thesis, University of Nottingham, 2012. http://eprints.nottingham.ac.uk/12695/.
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Beta-Adrenoceptors (B-ARs) mediate vasodilatation by activating various mechanisms that collectively contribute to vascular smooth muscle (VSM) relaxation. It has been shown that B2-AR stimulation in cultured cells results in activation of extracellular signal-regulated kinase (ERK). As the functional relevance of this was not known, the aim of the current investigation was determine the role of ERK in beta-AR-mediated vasodilatation. Isoprenaline-induced relaxation of porcine coronary artery (PCA) segments pre-contracted with the thromboxane mimetic U46619 was significantly enhanced by inhibition of ERK activation. Relaxations to the beta2-AR agonist salbutamol, but not those to the beta1-AR agonist xamoterol or the adenylyl cyclase activator forskolin, were also enhanced. The intermediate-conductance Ca2+-activated K+ (IKCa) channel blocker TRAM-34 prevented the enhancement of beta2-AR-mediated responses. Taken together, the data indicate that ERK inhibits beta2-AR-mediated vasodilatation by interacting with a cyclic 3’, 5’-adenosine monophosphate-independent relaxation pathway involving K+ channels. This may occur through a direct regulatory action on the IKCa channel via phosphorylation. Furthermore, the finding that increased ERK activation in a rat model of Type II diabetes was associated with significantly impaired beta-AR-mediated vasodilatation raises the possibility that ERK may represent a promising therapeutic target in the treatment of disease states characterised by abnormal vascular function.
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Carter, David Andrew. "Sourcing cells for gut tissue engineering : understanding and inducing embryonic stem cell differentiation to the intestinal cell lineage." Thesis, University of Nottingham, 2012. http://eprints.nottingham.ac.uk/12713/.
Full textAbstract:
Tissue engineering of any tissue type requires the combination of a supporting scaffold, a range of biological factors and a suitable source of cells. This source of cells must satisfy a number of criteria:- • The ability to form all of the mature/specialised cell types found in the target tissue. • Readily obtainable. • Readily maintainable in the laboratory without requiring excessive resources or time. For intestinal tissue engineering there are a number of issues associated with the use of tissue derived stem cells particularly quantity of normal tissue available (from the patient) and maintenance and expansion of the cells when cultured in vitro. Using embryonic stem cells offers a potential alternative strategy but methods must be established to efficiently differentiate the cells towards the desired fate. Many strategies for differentiating embryonic stem cells are based upon treatment with growth factors in vitro. There is a (logistical) limit to the degree of complexity that these systems can achieve and therefore a limit to the number of differentiation signals that occur during in vivo development that can be mimicked. In recent years using embryonic tissue to provide signals to undifferentiated cells has proved a successful method of directing the differentiation of naïve cells towards a particular fate (with the choice of tissue determined by the desired target cell type). The aims of this thesis were to explore the potential of differentiating embryonic stem cells towards the intestinal progenitor fate using a combination of in vitro cell culture treatment with the growth factor Activin-A and ex vivo co-culture with embryonic chick gut tissue. Previous studies [Kubo et al 2004, Tada et al 2005, Yasunaga et al 2005, D’Amour et al 2005, MacClean et al 2007] have shown that Activin-A treatment will induce embryonic cells to more efficiently differentiate to definitive endoderm, the germ layer from which the intestines (and other visceral organs) arise. These techniques were applied to the Columnar Epithelial Epiblast murine embryonic stem cell line and cell differentiation was then evaluated at the molecular level using Reverse Transcription-Polymerase Chain Reaction, immunocytochemistry and Western blotting. Activin-A treatment produced an upregulation of definitive endoderm markers at both the mRNA and proteomic levels compared to the control conditions. However the cell population produced retained expression of pluripotent markers and showed some expression of markers of other cell lineages. Further studies [Sugie et al 2005, Fair et al 2003, Van Vranken et al 2005, Coleman et al 2007, Krassowska et al 2006] have shown that co-culture of embryonic stem cells with early stage embryonic tissue can induce the formation of particular tissue types; the tissue must be selected based on proximity to the target cell type during development. This exposes the embryonic stem cells to the signals that prompt differentiation towards the target tissue during normal development. With gut tissue much signalling occurs between the different tissue layers that make up the whole organ both during development and in adult tissue. Ex vivo co-culture of murine embryonic stem cells with embryonic chick gut tissue was used to direct their differentiation to the intestinal epithelial stem cell fate. Before the co-culture was carried out various experiments were carried out to establish if the proposed protocol was viable e.g. defining how long chick gut tissue explants could survive in culture. Once this had been established co-culture experiments were undertaken and cell differentiation was then evaluated at the molecular level using Reverse Transcription-Polymerase Chain Reaction, immunocytochemistry and Western blotting. The cells showed some expression of intestinal epithelial stem cell markers at both the mRNA and proteomic levels following co-culture. The cells were also assessed at a physiological/functional level by evaluating their ability to form a functional intestinal epithelial barrier. This was achieved using an in vitro co-culture model with intestinal subepithelial myofibroblasts by measuring transepithelial resistance, permeability to protein and morphology in a simple tissue model co-culture. The cells did not display the morphological or physiological characteristics associated with intestinal epithelial cells in the model system. Overall this work has shown that co-culturing pluripotent mES cells with embryonic chick gut tissue can induce differentiation towards the ISC fate. Pre-treating the cells with growth factors in vitro did not seem to enhance this differentiation but there was scope to refine these techniques. Following the differentiation protocols the cells did not display the desired physiological characteristics but again there was scope to refine the techniques particularly with regard to selecting cells positive for the expression of the chosen molecular markers. These techniques show promise but do require some further development.