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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
91

The effects of bleomycin, mitomycin C, and cytoskeletal-disrupting drugs on angiogenesis in vitro and haemangioma development in vivo

Mabeta, Peaceful Lucy 22 January 2009 (has links)
Angiogenesis, the process of new vessel formation, appears to be a central mechanism that underlies the development of haemangiomas. Recently, intralesional bleomycin injection was used to treat paediatric haemangiomas with very good results. The purpose of this study was to determine whether there was significant systemic circulatory spill-over of bleomycin in haemangioma patients treated with intralesional bleomycin to determine safety of use. Furthermore, in order to elucidate bleomycin’s mechanism of action in inducing haemangioma regression, this study aimed at determining the effects of bleomycin on aspects of angiogenesis, namely, endothelial cell migration, growth and apoptosis, and comparing these effects with those of drugs previously reported to inhibit various aspects of the angiogenic process (mitomycin C, 2-methoxyestradiol, taxol, vincristine, vinblastine, colchicine, nocodazole and cytochalasin D). Lastly, the effects of bleomycin, mitomycin C, 2-methoxyestradiol, taxol, vincristine, vinblastine, colchicine, nocodazole and cytochalasin D were studied in an animal haemangioma model. A rapid and highly sensitive high performance liquid chromatographic (HPLC) method was developed. Blood samples were collected from four haemangioma patients before and after (over a 24 hour period) intralesional bleomycin (IB) therapy. As a control, blood samples were also collected at identical time intervals from four patients undergoing intravenous (IV) bleomycin chemotherapy for various malignant tumours. The HPLC method was used to quantitate bleomycin fractions in patient samples. The mean bleomycin concentration detected in plasma samples obtained from IB treated patients was 0.00 ìg/ml for both bleomycin A<Sub>2 and B2 over the 24-hour period following therapy. Plasma bleomycin A2 and B2 levels of 360.79 and 158.85 ìg/ml respectively were detected in samples obtained from cancer patients treated with bleomycin IV. These findings indicate that the low levels detected may translate to a significantly lesser risk of pulmonary fibrosis following IBI. The effect of drugs on endothelial cell migration was analyzed by wounding a confluent monolayer of cells and determining the number of cells that had migrated from the wound edge. Endothelial cell growth was determined in cells treated with various drug concentrations while apoptosis was examined using hematoxylin and eosin staining, DNA fragmentation assay and acridine orange staining. The effect of test drugs on in vitro angiogenesis was determined on endothelial cells induced to form capillary-like tubes in collagen gel. Test drugs were then evaluated for antitumour activity in an animal haemangioma model. Data demonstrated that test drugs inhibited endothelial cell migration, with the exception of mitomycin C. All test drugs induced a reduction in the percentage of viable endothelial cell in a dose-dependant manner, and also induced endothelial cell apoptosis. The drugs inhibited angiogenesis in vitro and inhibited tumour development in vivo with varying potency. In general, results from this study indicated that there was negligible systemic spill-over of bleomycin following IB administration in patients with haemangiomas, suggesting a much lesser risk of developing bleomycin-induced pulmonary fibrosis. This study also showed that test drugs inhibited angiogenesis in vitro and haemangioma development in vivo in a mouse model. Taken together, these observations demonstrate that bleomycin may inhibit haemangioma growth by inhibiting angiogenesis. In addition, mitomycin C, 2-methoxyestradiol, taxol, vincristine, vinblastine, colchicine, nocodazole and cytochalasin D may have potential in the treatment of haemangiomas of infancy, and should be investigated further in a murine haemangioma model to determine effective dose schedules. / Thesis (PhD)--University of Pretoria, 2009. / Physiology / unrestricted
92

Micro and Nanostructuring of Polymers by Femtosecond Laser Pulses

Alshehri, Ali January 2016 (has links)
Micro/Nanostructuring of polymers by femtosecond pulses is of extreme importance because it drives applications in photonics and biomedicine. A femtosecond pulse, with an intensity of ∼ 10^13 W/cm^2, is capable of causing an optical breakdown and inducing permanent modification in the material. With such high intensity, and considering the fact that polymers possess high band gaps, the interaction nature is completely nonlinear, and the material can be modified locally on the surface and in bulk. The irradiated regions exhibit fluorescence, and they display new wetting properties as a consequence of the optical breakdown of a material. The optical breakdown can be investigated by studying the nonlinear absorption. In this thesis, we discuss the nonlinear absorption of fs-laser pulses inside polymers using transmission measurements. We show a step– function–like behaviour of the transmission, dropping abruptly to ∼ 20% at the optical breakdown threshold with a ∼ 40 % reduction in the band gap. Utilizing spectroscopy, we show that the laser-modified regions contain randomly distributed nanoclusters. The presence of localized nanoclusters is responsible for exhibiting fluorescence, within ∼ 10 µm3 for a single pulse. This feature was exploited to demonstrate high-density data storage in Polymethyl methacrylate (PMMA) without any special material preparation. We demonstrate up to 20 layers of embedded data that can be stored in a standard 120 mm disc. Storage capacity of 0.2 TBytes/disc can be achieved by adjusting read laser parameters. Besides the fluorescence capability induced in the bulk of polymers, the hydrophilicity shown by the fs–laser modified surface is utilized to study selective cell growth on the micro-structured Polydimethylsiloxane (PDMS) surface. We show that the C2C12 cells and rabbit anti-mouse protein attach preferentially to the modified regions when the surface is modified with low pulse energies. However, in the high pulse energy regime, the laser-modified regions exhibit superhydrophobicity inhibiting cell adhesion.
93

Studie av celltillväxt hos kondrocyter och mesenkymala stromaceller från benmärg- en utvärdering av optimala odlingsförhållanden / Study of cell growth in chondrocytes and mesenchymal stroma cells from bone marrow - an evaluation of optimal culture conditions

Emilsson, Sara January 2023 (has links)
Introduktion: Hyalint brosk består av kondrocyter inbäddade i ett extracellulärt matrix som täcker alla kroppens synoviala leder. I dag är skador på hyalint brosk ett folkhälsoproblem med begränsade behandlingsmöjligheter. Mesenkymala stamceller har i flertalet studier visat sig vara en lovande kandidat för framtida behandling av broskskador, eftersom dessa kan differentiera till broskproducerande kondrocyter. Modern behandling av broskskador innefattar odling av patientens egna kondrocyter eller stamceller för att sedan återföra dem till broskskadan. För att göra detta effektivt behöver optimering av odlingsförhållanden för aktuella celler utföras. Syfte: Att undersöka tillväxt hos kondrocyter samt mesenkymala stromaceller från benmärg i varierande odlingsförhållanden, detta med avseende att utvärdera vilka tillsatser som mest gynnar cellernas viabilitet och proliferation. Material &amp; Metod: Framtagning av kondrocyter och mesenkymala stromaceller från benmärg i denna studie utfördes på överblivet ortopediskt material från en individ. Cellerna kollagenasbehandlades innan de odlades ut i ett standardmedium under tre veckor. Därefter odlades de i närvaro av olika tillsatser under sex dagar. Cellernas viabilitet mättes genom relativ fluorescens med CellTiter-Blue® reagens medan proliferationen mättes genom cellräkning med flödescytometri. Resultat: Det fanns en signifikant skillnad (ANOVA, p&lt;0,05) mellan minst två tillsatser inom grupperna för viabilitetsanalys samt cellräkning för både kondrocyter och benmärgderiverade mesenkymala stromaceller (BMMSC). Kondrocyterna som odlades i närvaro av fetalt bovint serum (FBS) hade högre proliferation och viabilitet än de celler som odlades i frånvaro av FBS. FBS i kombination med insulin-transferrin-selen (ITS) och kondrocytfaktorerna (KF) dexametason, prolin, insulin samt transforming growth factor beta hade störst positiv effekt på proliferationen. Vidare visade Minimum Essential Medium α med tillsats av FBS 10 % ha störst effekt, av den panel som undersöktes, på både viabilitet och proliferation hos BMMSC. Slutsats: FBS i kombination med ITS och KF hade störst stimulerande effekt på proliferation hos kondrocyter. Val av standardmedium hade större betydelse än tillsats av faktorer vid odling av BMMSC in vitro. / Introduction: Articular cartilage consists of chondrocytes and covers all the synovial joints in the body. Articular cartilage injuries are today a public health issue with limited orthopedic treatments. Mesenchymal stem cells have shown to be a promising candidate for future treatment of articular cartilage injuries due to their many abilities. Modern treatment of cartilage damage involves cultivation of the patient´s own chondrocytes and stem cells. To make this treatment as effective as possible the culture conditions for chondrocytes and stem cells must be optimized. Aim: To study cell growth in chondrocytes and mesenchymal stromal cells from bone marrow in varying culture conditions, this regarding to evaluate the most optimal culture conditions for cell viability and proliferation. Method: Cell extraction in this study was performed on leftover orthopedic material. The cells were treated with collagenase and then cultured in a standard medium for three weeks before they were cultured in presence of various factors for six days. Cell viability was measured by relative fluorescence while proliferation was measured by cell counting. Result: There was a statistically significant difference (ANOVA, p&lt;0,05) between at least two additives within the groups for viability analysis and cell count for both chondrocytes and bone marrow-derived mesenchymal stromal cells (BMMSC). The chondrocytes cultured in presence of fetal bovine serum (FBS) had a higher proliferation and viability compared to the cells cultured in the absence of FBS. FBS in combination with insulin-transferrin-selenium (ITS) and the chondrocyte factors (CF) dexamethasone, proline, insulin and transforming growth factor beta had the biggest impact on proliferation. Minimum Essential Medium α with FBS 10% additive showed positive effects on both viability and proliferation of BMMSCs. Conclusion: FBS in combination with ITS and CF had a good effect on the proliferation of chondrocytes. The choice of standard medium seems to have a more important role than growth factor additive when studying BMMSCs expansion in vitro.
94

Differences in TOR and Yak1 Gene Expression in the Mold and Yeast Phases of Penicillium marneffei

Sethi, Sumedha 06 October 2011 (has links)
No description available.
95

Μελέτη της μοριακής στόχευσης κυττάρων του πλειόμορφου γλοιοβλαστώματος με αναστολείς της αγγειογένεσης και μορίων του μονοπατιού HER

Δημητρόπουλος, Κωνσταντίνος 20 February 2014 (has links)
Το γλοιοβλάστωμα αποτελεί έναν από τους πιο θανατηφόρους τύπους καρκίνου για τον άνθρωπο δεδομένου ότι ο μέσος όρος επιβίωσης είναι 12-15 μήνες. Η συμβατική θεραπεία με τα κλασσικά χημειοθεραπευτικά σχήματα δεν έχει αποδώσει ιδιαιτέρως θετικά αποτελέσματα. Η εξέλιξη της μοριακής βιολογίας έχει «ρίξει φως» στην διερεύνηση και κατανόηση αρκετών μηχανισμών της κυτταρικής λειτουργίας που αφορούν τη μετανάστευση, τον πολλαπλασιασμό και την απόπτωση των κυττάρων. Στο γλοιοβλάστωμα έχει παρατηρηθεί αυξημένη δραστηριότητα πολλών υποδοχέων αυξητικών παραγόντων στην επιφάνεια των κυττάρων όπως ο EGFR, ο PDGFR και ο VEGFR. Ταυτόχρονα, υπάρχουν και άλλα μόρια επιφανείας με ιδιαίτερο ενδιαφέρον όπως οι ιντεγκρίνες λόγω της ιδιότητας τους να καθορίζουν τη μετανάστευση. Στη νέα εποχή των αναστολέων κινάσης τυροσίνης οι οποίοι δρουν έναντι των υποδοχέων των αυξητικών παραγόντων και θεωρούνται πολλά υποσχόμενα μικρά μόρια, πρέπει να διερευνηθεί αν μπορούν να έχουν θέση στην αντιμετώπιση του γλοιοβλαστώματος. Αρχικά έγινε εκτίμηση της επίδρασης των αναστολέων sunitinib και lapatinib ξεχωριστά αλλά και σε συνδυασμό, στο πολλαπλασιασμό και στο κυτταρικό θάνατο των κυττάρων γλοιβλαστώματος U87 και M059K σε συγκεντρώσεις 0,01, 0,1, 1 και 10μΜ. Για το σκοπό αυτό χρησιμοποιήθηκε η μέθοδος του 3-[4,5-dimethylthiazol-2-yl]-2,5 διμεθυλθειαζόλο βρωμιδίου και το ολοκληρωμένο σύστημα ανίχνευσης αννεξίνης V/προπιδίου (annexin V/propidium iodide), αντίστοιχα. Για τη διαδικασία της μετανάστευσης εφαρμόστηκε η μέθοδος μελέτης του χημειοτακτισμού. Η έκφραση μεταλλοπρωτεϊνασών MM-2 και MMP-9 εκτιμήθηκε με τη μέθοδο του ζυμογραφήματος. Περαιτέρω έγινε διερεύνηση της πιθανής εμπλοκής των φαρμάκων στο σχηματισμό συμπλόκων EGFR-υπομονάδα β1 ιντεγκρινών, PDGFR- υπομονάδα β3 ιντεγκρινών και VEGFR- υπομονάδα β3 ιντεγκρινών. Η μελέτη της δημιουργίας των συμπλόκων των β υπομονάδων των ιντεγκρινών με τους υποδοχείς αυξητικών παραγόντων έγινε χρησιμοποιώντας τη μέθοδο της ανοσοκατακρήμνισης και της ανάλυσης κατά Western. Τα αποτελέσματα επαληθεύτηκαν και με τη μέθοδο του ανοσοφθορισμού. Με την ίδια μέθοδο μελετήθηκε και η επίδραση των φαρμάκων στη φωσφορυλιωμένη μορφή της FAK. Τα πειραματικά δεδομένα έδειξαν ότι και οι δύο αναστολείς sunitinib και lapatinib, μείωσαν τον πολλαπλασιασμό με δοσοεξαρτώμενο τρόπο 48 ώρες μετά την προσθήκη τους στα κύτταρα είτε μεμονωμένα είτε σε συνδυασμό. Τα αποτελέσματα της αναστολής του πολλαπλασιασμού συμβάδιζαν με τα αποτελέσματα της απόπτωσης όπου το ποσοστό των αποπτωτικών κυττάρων αυξήθηκε. Περαιτέρω η μετανάστευση των κυττάρων εμφανίστηκε μειωμένη μετά την προσθήκη των φαρμάκων είτε μεμονωμένα είτε σε συνδυασμό. Η έκφραση των μεταλλοπρωτεϊνασών MMP-2 και MMP-9 δεν επηρεάστηκε στα κύτταρα U87 μετά την προσθήκη και των 2 φαρμάκων. Αντίθετα, στα κύτταρα M059K το sunitinib αλλά και ο συνδυασμός του με το lapatinib ελαττώσαν την έκφραση των MMPs 48 ώρες μετά τη προσθήκη τους στο θρεπτικό μέσο. Στη συνέχεια βρέθηκε ότι το lapatinib μπορεί να αναστείλει την δημιουργία συμπλόκου του EGFR με την υπομονάδα β1 των ιντεγκρινών, έως και 30 λεπτά μετά την προσθήκη του φαρμάκου στα κύτταρα. Αντίστοιχα το sunitinib μπορεί να αναστέλλει το σχηματισμό συμπλόκου της υπομονάδας β3 των ιντεγκρινών με τον VEGFR εντός δύο ωρών από την προσθήκη των φαρμάκων, χωρίς επίδραση όμως στον σχηματισμό των συμπλόκων PDGFR – υπομονάδα β3 ιντεγκρίνης. Αυτά τα αποτελέσματα επιβεβαιώθηκαν και με την χρήση ανοσοφθορισμού. Συνοψίζοντας, οι δύο αναστολείς κινάσης τυροσίνης, sunitinib και lapatinib κατάφεραν να επιδείξουν αξιόλογα αποτελέσματα στις παραμέτρους του πολλαπλασιασμού και της μετανάστευσης των κυττάρων γλοιοβλαστώματος. Τα αποτελέσματα της έρευνας είναι πρωτοποριακά διότι, αναφορικά με την μετανάστευση στο γλοίωμα, αυτή φαίνεται να αναστέλλεται αποτελεσματικότερα με τον συνδυασμό των δύο φαρμάκων πιθανά μέσω του μηχανισμού διαταραχής της δημιουργίας του συμπλόκου ιντεγκρίνης- υποδοχέας αυξητικού παράγοντα. / Glioblastoma is considered to be one of the most fatal among the malignancies in human with median survival between 12-15 months. The conventional chemotherapy cannot change the fate of these patients. Recent advances in molecular biology have shed light in the various mechanisms recruited by malignant cells in order to proliferate, metastasize and escape apoptosis. Studies in glioblastoma have already shown up-regulation in the function of tyrosine kinase receptors such as EGFR, PDGFR and VEGFR. Besides this, in the more recent years, research supports the significant role of integrins in the migration process of glioblastoma cells. In the new era of tyrosine kinase inhibitors (TKIs), where their main mechanism of action is by blocking various growth factor receptors, it has to be determined a possible contribution in the fight against glioblastoma. Initially, it was estimated a possible effect of sunitinib and lapatinib applied either alone or in combination, on U87 and M059K glioma cells’ proliferation and apoptosis using doses of 0,01 μM, 0,1 μM, 1μM and 10 μM. The proliferation was estimated using the 3-[4,5-dimethylthiazol-2-yl]-2,5 dimethyltetrazolium bromide assay and apoptosis using annexin V/propidium iodide detection assay. Migration assay was performed using Boyden chamber assay. The release of MMP-2 and MMP-9 into the culture medium of U87 and M059K cells was measured by zymography. Furthermore, a possible implication of the tested agents in the formation of EGFR-integrin β1 complex, VEGFR-integrin β3 and PDGFR-integrin β3 complexes formation was studied. Immunoprecipitation and western blot analysis were used for studying the complex formation of EGFR, PDGFR and VEGFR with integrins. Validation of the results was made using immunofluorescence assay. Also, similar experiments were performed for the effect of sunitinib and lapatinib in FAK phosphorylation. The results showed that both tested agents decreased cell proliferation in a dose-dependent manner 48 h after their application in both cell lines either alone or in combination. The results in cell proliferation were in line with the increase in apoptotic cells after their treatment with the tested agents. Furthermore, the ability of U87 and M059K cells to migrate was inhibited either by each agent alone or in combination. Both agents did not affect MMP-2 and MMP-9 levels in U87 cells, however, MMPs levels were decreased in M059K cells, 48h after their treatment with sunitinib either alone or in combination with lapatinib. Additionally, a time course study for the effect of lapatinib on EGFR-integrin β1 complex revealed an inhibition in complex formation up to 30 min after agent application. Likewise, sunitinib inhibited complex formation of VEGFR-integrin β3 complex within 2h after its application without affecting PDGFR-integrin β3 complex. The previously-described interruption of complexes formation was confirmed with an immunofluorescence assay. Summarizing the results, sunitinib and lapatinib exerted significant effects on glioblastoma cell proliferation, apoptosis and migration. The preliminary results of the current study are the first to support the implication of a dual anti-EGFR/HER-2 agent, lapatinib and a multi-targeted agent, sunitinib in glioma cells migration, through a mechanism implying interruption of growth factor receptor - integrin complexes formation.
96

Automates cellulaires pour la modélisation multi-échelle des systèmes biologiques / Cellular automata for multi-scale modeling of biological systems

Louvet, Benjamin 11 July 2014 (has links)
Ce projet de thèse, dans le cadre d’une collaboration entre le LIMOS et le LPC, s’inscrit dans une démarche de recherche permettant la mise en synergie des domaines de la biologie, de la physique et de l’informatique par la proposition d’une démarche de simulation permettant la réalisation d’expériences in silico. Pour cela, nous nous proposons de développer une plateforme logicielle dédiée à la modélisation multiéchelle des systèmes biologiques qui pourra par la suite être interfacée avec les outils de simulation de physique des particules. Nous proposons également un modèle individu-centré de cellule biologique paramétrable à l’aide de données obtenues d’expériences in vitro. Nous présentons l’élaboration de cette plateforme et une démarche de validation de ses fonctionnalités à travers l’implémentation de modèles d’automates cellulaires de la littérature. Nous présentons ensuite la construction du modèle de cellule biologique en prenant le temps d’expliquer comment est pris en compte le système biologique, comment nous le modélisons puis comment nous paramétrons le modèle. Nous modélisons les processus internes de la cellule, dont les caractéristiques sont liées à l’information génétique qu’elle porte. Ce modèle de cellule permet de reproduire le comportement d’une cellule isolée, et à partir de là, d’un ensemble de cellules via l'automate. Le modèle est ensuite utilisé pour retrouver les courbes de croissance d'une population de bactéries Escherichia coli. Des valeurs de données de fluxomique ont été exploitées et ont permis la reproduction in silico des expériences in vitro dont elles étaient issues. / This PhD thesis project is part of a research program in the fields of biology, physics and computer science aiming to propose a simulation approach for performing experiments in silico. For this, we propose to develop a software platform dedicated to multi-scale modeling of biological systems that can be combined with particle physics simulation tools. We also propose a general individual-based model of biological cell in which data obtained from in vitro experiments can be used. We present the development of this platform and the validation process of its functionalities through the implementation of cellular automata from the literature. We then present the design of the biological cell model by giving the hypothesis we made, how we model and how we parameterize the model. Starting from a simple biological system, bacteria, observed in liquid culture, our model uses a multi-scale middle-out approach. We focus on the cell and we model internal processes, assuming that all their properties come from genetic information carried out by the cell’s genome. This model allows to consider the cell behavior, and then to obtain the behavior of a cell population. Data from fluxomic experiments have been used in this model to parameterize the biochemical processes. The results we obtain allow us to consider the model as validated as simulation results match the experimental data.
97

Oxydation et dégradation de l'ascorbate chez la tomate et impact sur la croissance et le métabolisme / Oxidation and degradation of ascorbate in tomato and impact on growth and metabolism

Truffault, Vincent 12 November 2015 (has links)
Contrôle de l'oxydation et de la dégradation du pool de vitamine C chez la tomate et impact sur la qualité du fruit et la tolérance au stress. Le métabolisme de l’ascorbate et plus principalement le statut redox du pool d’ascorbate sont impliqués dans la tolérance au stress et dans les processus primaires de croissance et de développement de la plante. La teneur et le statut redox de l’ascorbate chez les plantes sont régulés par (i) ses voies de biosynthèse, (ii) par le cycle ascorbate-glutathion permettant le recyclage des formes semi-oxydées et oxydées de l’ascorbate et (iii) par sa dégradation, l’ensemble de ces processus étant sous le contrôle de l’environnement. Au cours de ce travail de thèse, des méthodes de transgénèse nous ont permis d’identifier, chez différents génotypes de tomate à petit et gros fruits, les bouleversements physiologiques et métaboliques permettant de compenser des modifications de l’activité des enzymes monodéhydroascorbate réductase (impliqué dans le cycle ascorbate-glutathion) et ascorbate oxydase. Nous avons observé d’importantes modifications phénotypiques altérant le rendement en fruits de la plante sous conditions de culture pouvant générer un stress et également en condition normale de culture. Des liens entre l’activité des enzymes précités avec le métabolisme des sucres, la photosynthèse et la conductance stomatique sont révélés. Le déséquilibre entre les activités oxydantes et réductrices de ces enzymes constitue la première étape vers une dégradation de l’ascorbate. Le taux de dégradation se révèle très faible à la lumière, tandis qu’à l’obscurité une forte accumulation des produits de dégradation l’oxalate, le thréonate ainsi que l’oxalyl-thréonate est observé dans les feuilles de tomate. Enfin, l’activité de l’enzyme MDHAR est corrélée au taux de dégradation à l’obscurité. Les travaux de cette thèse mettent en avant l’importance du statut redox du couple ascorbate / monodéhydroascorbate dans les processus de croissance cellulaire et entre dans la régulation du rendement chez la tomate, et influe la dégradation de l’ascorbate. / Ascorbate metabolism and particularly ascorbate redox status are involved in stress tolerance and growth processes of plant cells. The concentration of ascorbate and its redox status are under control of (i) its biosynthetic pathways, (ii) the ascorbate-glutathione cycle allowing recycling of semi-oxidized and oxidized forms of ascorbate and (iii) its degradation rate. These processes are under environmental control. Transgenic lines modified for the activity of monodehydroascorbate reductase (involved in ascorbate-glutathione cycle) and ascorbate oxidase were generated in cherry and large-fruited genotypes of tomato. Physiological and metabolic modifications related to the modification of these enzyme activities were studied. We observed large phenotypic alterations that affected fruit yield under both stress conditions and normal growth conditions. Links between ascorbate recycling and sugar metabolism, photosynthesis and stomatal conductance were also revealed. An imbalance between the oxidizing and reducing activities of these enzymes is the first step leading to ascorbate degradation. We have shown that the degradation rate was very low under light, whereas under darkness the degradation compounds oxalate, threonate and oxalyl-threonate accumulated in tomato leaves. Also, the degradation rate is correlated with MDHAR activity. These results highlight the crucial role of the redox status of the ascorbate / monodehydroascorbate couple in growth processes and yield stability in tomato, and the impact on ascorbate degradation.
98

Role of Ectodermal-neural cortex 1 protein in human glioma progression, identification of a peptide internalized by human glioblastoma cells and development of an alternative method to generate growth curves of adherent cultures / Papel da proteína Ectodermal-neural cortex 1 (ENC1) na progressão de glioma humano, identificação de um peptídeo internalizado por células de glioblastoma humano e desenvolvimento de um método alternativo para gerar curvas de crescimento celular

Pereira, Túlio Felipe 14 November 2018 (has links)
Gliomas are the most common form of primary intracranial malignancy, among which astrocytomas are the most frequent. Ectodermal-cortex protein 1 (ENC 1), also known as Nuclear Restricted Protein/Brain (NRP/B), was first characterized as a protein which interacts with the cytoskeleton by binding to actin through Kelch-like domains, being related to neural fate specification during development of the nervous system. The first chapter of this thesis confirms ENC1 as a tumor suppression properties by a genomic edition approach, analyses ENC1 expression in a set of patient glioma samples and describes the correlation these data with patients survival and progression-free survival, concluding that ENC1 expression may constitute a biomarker for glioma aggressiveness. The second chapter refers to the identification and in vitro characterization of the LHTNELQ peptide, which was selected by the Phage Display method using human glioblastoma cells. This new peptide is able to be internalized by these cells and features as a new tool for the development of glioma therapeutics. The third chapter report an alternative method to generate growth curves of adherent cell cultures, which is based on the CFSE fluorescence decay over time. It is an alternative method to determine growth curves of cultured cells, with smaller variation among technical replicates than that of counting-based methods. / Gliomas são a forma mais comum de malignidades primárias intracranianas, dentre os quais os astrocitomas são os mais frequentes. A proteína Ectodermal-neural cortex 1 (ENC1), também conhecida como Nuclear Restricted Protein/Brain (NRP/B), foi primeiramente caracterizada como uma proteína que interage com o citoesqueleto por meio de ligação à actina através de domínios Kelch-like, sendo relacionada com diferenciação neuronal durante o desenvolvimento do sistema nervoso. O primeiro capítulo desta tese descreve confirmação da capacidade supressora tumoral de ENC1 por abordagem de edição genômica, analisa a expressão de ENC1 em um conjunto de amostras de pacientes com gliomas e correlaciona esses dados com tempo de sobrevida geral e sobrevida livre de progressão tumoral nos pacientes, concluindo que a expressão de ENC1 pode ser utilizada como um biomarcador da agressividade do glioma. O segundo capítulo apresenta a identificação e caracterização in vitro do peptídeo LHTNELQ, que foi selecionado pela metodologia de Phage display utilizandose de células de glioblastoma humano. Este novo peptídeo é capaz de internalizar-se nestas células e figura como uma nova ferramenta para o desenvolvimento de estratégias terapêuticas para glioblastomas. No terceiro capítulo propõe-se um método alternativo para gerar curvas de crescimento celular de cultura aderente, o qual é baseado no decaimento da fluorescência do reagente CFSE ao longo do tempo. Tratase de um método alternativo para a determinação de curvas de crescimento de culturas aderentes, com menor variação entre as réplicas técnicas do que os métodos baseados em contagem das células.
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Análise da coinfecção entre ureaplasmas e o vírus do Papiloma Humano (HPV) em amostras cervicais e em um modelo de estudo \"in vitro\" de queratinócitos primários humanos (PHK). / Analysis of co-infection among ureaplasmas and the Human Papilloma Vírus (HPV) in cervical samples and in a infection model in vitro in primary human keratinocytes (PHK).

Amorim, Aline Teixeira 30 April 2015 (has links)
O desenvolvimento do câncer cervical depende da exposição ao HPV, fator necessário, mas não suficiente. Outras bactérias, tais como ureaplasmas, têm sido associadas como cofatores. O objetivo deste estudo foi avaliar a presença de ureaplasmas em mulheres com lesão cervical, e observar alterações em PHK causadas pela infecção por ureaplasmas. 140 swabs vaginais foram coletados. O material foi submetido a PCR para a detecção de HPV, Mollicutes, U. urealyticum, U. parvum e seus sorotipos, e outras bactérias de importância ginecológica; e qPCR para U. urealyticum e U. parvum. Também foi realizada a infecção de ureaplasmas em PHK transformados com HPV. As células foram contadas e realizou-se a dosagem das citocinas IL1-&beta;, IL-6 e TNF-&alpha;. HPV, Mollicutes, U. parvum, sorotipos 1 e 6 de U. parvum, T. vaginalis e G. vaginalis, além de alguns fatores socioeconômicos, foram associados com lesão cervical. Verificou-se maior carga de U. parvum entre mulheres com lesão. Houve diminuição do número de células e maior liberação de IL-6 e TNF-&alpha; nos grupos infectados. Com os resultados obtidos neste estudo, foi possível verificar uma associação entre os ureaplasmas e HPV no início das lesões cervicais, contudo mais estudos precisam ser realizados para aprimorar essa hipótese. / The development of cervical cancer depends on the exposure to HPV, necessary factor, but not enough. Other bacteria, such as ureaplasmas, have been associated as cofactors. The aim of this study was to evaluate the presence of ureaplasmas in women with cervical injury, and observe changes in PHK infected by ureaplasmas. 140 vaginal swabs were collected. The material was subjected to PCR for detection of HPV, Mollicutes, Ureaplasma urealyticum, U. parvum (and serotypes) and other bacteria gynecological importance; qPCR for U. urealyticum and U. parvum was made. PHK transformed by HPV was infected by ureaplasma. Cells were counted and it was done titration of IL1-&beta;, IL-6 and TNF-&alpha;. HPV, Mollicutes, U. parvum, serotypes 1 and 6 U. parvum, T. vaginalis and G. vaginalis, and some socioeconomic factors were associated with cervical injury. Besides this, it was detected higher load U. parvum among women with injury. There was decrease in cell number and increased release of IL-6 and TNF-&alpha; in infected groups. With the results of this study, we found an association among HPV and ureaplasmas at the beginning of cervical lesions, but more studies are needed to enhance this hypothesis.
100

Análise da coinfecção entre ureaplasmas e o vírus do Papiloma Humano (HPV) em amostras cervicais e em um modelo de estudo \"in vitro\" de queratinócitos primários humanos (PHK). / Analysis of co-infection among ureaplasmas and the Human Papilloma Vírus (HPV) in cervical samples and in a infection model in vitro in primary human keratinocytes (PHK).

Aline Teixeira Amorim 30 April 2015 (has links)
O desenvolvimento do câncer cervical depende da exposição ao HPV, fator necessário, mas não suficiente. Outras bactérias, tais como ureaplasmas, têm sido associadas como cofatores. O objetivo deste estudo foi avaliar a presença de ureaplasmas em mulheres com lesão cervical, e observar alterações em PHK causadas pela infecção por ureaplasmas. 140 swabs vaginais foram coletados. O material foi submetido a PCR para a detecção de HPV, Mollicutes, U. urealyticum, U. parvum e seus sorotipos, e outras bactérias de importância ginecológica; e qPCR para U. urealyticum e U. parvum. Também foi realizada a infecção de ureaplasmas em PHK transformados com HPV. As células foram contadas e realizou-se a dosagem das citocinas IL1-&beta;, IL-6 e TNF-&alpha;. HPV, Mollicutes, U. parvum, sorotipos 1 e 6 de U. parvum, T. vaginalis e G. vaginalis, além de alguns fatores socioeconômicos, foram associados com lesão cervical. Verificou-se maior carga de U. parvum entre mulheres com lesão. Houve diminuição do número de células e maior liberação de IL-6 e TNF-&alpha; nos grupos infectados. Com os resultados obtidos neste estudo, foi possível verificar uma associação entre os ureaplasmas e HPV no início das lesões cervicais, contudo mais estudos precisam ser realizados para aprimorar essa hipótese. / The development of cervical cancer depends on the exposure to HPV, necessary factor, but not enough. Other bacteria, such as ureaplasmas, have been associated as cofactors. The aim of this study was to evaluate the presence of ureaplasmas in women with cervical injury, and observe changes in PHK infected by ureaplasmas. 140 vaginal swabs were collected. The material was subjected to PCR for detection of HPV, Mollicutes, Ureaplasma urealyticum, U. parvum (and serotypes) and other bacteria gynecological importance; qPCR for U. urealyticum and U. parvum was made. PHK transformed by HPV was infected by ureaplasma. Cells were counted and it was done titration of IL1-&beta;, IL-6 and TNF-&alpha;. HPV, Mollicutes, U. parvum, serotypes 1 and 6 U. parvum, T. vaginalis and G. vaginalis, and some socioeconomic factors were associated with cervical injury. Besides this, it was detected higher load U. parvum among women with injury. There was decrease in cell number and increased release of IL-6 and TNF-&alpha; in infected groups. With the results of this study, we found an association among HPV and ureaplasmas at the beginning of cervical lesions, but more studies are needed to enhance this hypothesis.

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