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Belkai, Sonia. "Recherche d'acteurs lysosomaux impliqués dans la présentation de lipides mycobactériens par CD1b aux lymphocytes T". Electronic Thesis or Diss., Université de Toulouse (2023-....), 2024. http://www.theses.fr/2024TLSES176.
Testo completoAbstract (sommario):
Les lipides peuvent être antigéniques et présentés en surface des cellules présentatrices d'antigènes (CPA). Ces lipides, généralement amphiphiles, sont présentés par les protéines CD1 (CD1a à CD1d), des protéines proches structurellement de la protéine du CMH de classe 1, avec pour principale différence le site de liaison des antigènes. Cette présentation mène à une réponse immunitaire spécifique médiée par des lymphocytes T non conventionnels. Mycobacterium tuberculosis (Mtb), agent causal de la tuberculose, possède dans son enveloppe des lipides antigènes présentés par les protéines CD1, et particulièrement par l'isoforme CD1b. Ces lipides sont chargés sur CD1b au niveau du lysosome des cellules dendritiques (CD), dans leur état natif ou après un apprêtement. Cet apprêtement nécessite des protéines de transfert de lipides (LTP), telles que CD1e et les saposines, mais aussi des enzymes (hydrolases), permettant de digérer les parties polaire ou apolaires des lipides. Peu de LTP et hydrolases lysosomales ont à l'heure actuelle été identifiées, il est donc nécessaire de caractériser d'autres acteurs lysosomaux impliqués dans la présentation des antigènes lipidiques mycobactériens par CD1b. Les objectifs de ces travaux étaient de mettre en place des stratégies pour rechercher et identifier ces acteurs. Ainsi, dans le but d'identifier de nouveaux acteurs lysosomaux impliqués dans le transport ou la maturation des glycolipides de Mtb, deux approches complémentaires ont été mises en place. La première, plus globale, nous a conduit à préparer des fractions enrichies en lysosomes provenant de CPA. Pour valider le protocole d'obtention de ces fractions, elles ont été caractérisées morphologiquement ainsi que par la présence de marqueurs protéiques. Elles ont également servi à réaliser des tests de dégradation de lipides mycobactériens in vitro, confirmant l'action des lipases précédemment caractérisées. Enfin, l'analyse protéomique du contenu de ces fractions a contribué à l'élaboration d'une liste de cinq protéines candidates, possiblement impliquées dans l'apprêtement. Parmi elles, certaines n'ont jamais été décrites dans le contexte de présentation de lipides antigènes, et d'autres sont déjà connues pour contribuer à la présentation de lipides par CD1d. La seconde approche a consisté à définir l'interactome de CD1b dans le lysosome, en réalisant une coimmunoprécipitation de CD1b à partir de fractions lysosomales. L'analyse protéomique des partenaires a permis de proposer quatre autres protéines candidates, toutes non décrites à ce jour dans le cadre de la présentation d'antigènes lipidiques. L'expression de certaines de ces protéines candidates, sera par la suite inactivée pour étudier l'impact de cette inactivation sur la présentation des complexes CD1b-lipides. Parmi ces protéines, la protéine NPC1 (" Niemann-Pick disease C1 "), déjà connue pour être impliquée dans la présentation de lipides par l'isoforme CD1d chez la souris, a retenu notre intérêt. Des essais d'inactivation ont été réalisés grâce à un inhibiteur spécifique, avant d'envisager une inactivation de l'expression par utilisation de siRNA ou CRISPR/Cas9. Dans le but d'apprécier les effets de cette inactivation, il est nécessaire de posséder des outils biologiques capables d'évaluer la participation du candidat dans la présentation d'un lipide particulier. Lors de cette étude, deux outils ont été validés : 1) des anticorps recombinants spécifiques de CD1 présentant le sulfoglycolipide diacylé mycobactérien (Ac2SGL), et 2) différents clones lymphocytaires spécifiques de CD1b en complexe avec divers glycolipides mycobactériensLipids can be antigenic and presented on the surface of antigen-presenting cells (APCs). These lipids, generally amphiphilic, are presented by CD1 proteins (CD1a to CD1d), which are structurally similar to class I MHC proteins, with the main difference being in the antigen-binding site. This presentation leads to a specific immune response mediated by unconventional T lymphocytes. Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, has antigenic lipids in its envelope that are presented by CD1 proteins, particularly by the CD1b isoform. These lipids are loaded onto CD1b in the lysosome of dendritic cells (DCs), either in their native state or after processing. This processing requires lipid transfer proteins (LTPs), such as CD1e and saposins, as well as enzymes (hydrolases) that digest the polar or lipidic parts of the lipids. Few lysosomal LTPs and hydrolases have been identified so far, making it necessary to characterize other lysosomal actors involved in the presentation of mycobacterial lipid antigens by CD1b. The objectives of this work were to develop strategies to search for and identify these actors. To identify new lysosomal actors involved in the transport or maturation of Mtb glycolipids, two complementary approaches were implemented. The first, more global approach, involved preparing lysosome-enriched fractions from APCs. To validate the protocol for obtaining these fractions, they were characterized morphologically and by the presence of protein markers. They were also used to perform in vitro degradation tests of mycobacterial lipids, confirming the action of previously characterized lipases. Finally, proteomic analysis of the contents of these fractions led to the elaboration of a list of five candidate proteins that may be involved in processing. Among them, some have never been described in the context of lipid antigen presentation, while others are already known to contribute to lipid presentation by CD1d. The second approach involved defining the interactome of CD1b in the lysosome by performing co-immunoprecipitation of CD1b from lysosomal fractions. Proteomic analysis of the partners identified four other candidate proteins, none of which have been described to date in the context of lipid antigen presentation. The expression of some of these candidate proteins will subsequently be knocked out to study the impact of this inactivation on the presentation of CD1b-lipid complexes. Among these proteins, NPC1 ("Niemann-Pick disease C1"), already known to be involved in lipid presentation by the CD1d isoform in mice, was considered interesting. Inactivation tests were performed using a specific inhibitor, before considering inactivation of expression using siRNA or CRISPR/Cas9. To evaluate the effects of this inactivation, it is necessary to have biological tools capable of assessing the candidate's role in the presentation of a particular lipid. In this study, two tools were validated: 1) recombinant antibodies specific to CD1 presenting the mycobacterial diacylated sulfoglycolipid (Ac2SGL), and 2) different lymphocyte clones specific to CD1b in complex with various mycobacterial glycolipids
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Haig, Neil Ainslie. "The identification of endogenous lipid antigens associated with CD1 proteins and functional investigation of immune recognition". Thesis, University of Oxford, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.526535.
Testo completo
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Hassan, Namir. "Interactions of the leukocyte cell-surface proteins CD5 and CD6". Thesis, University of Oxford, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.398158.
Testo completo
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Wong, Chung Kai. "The DIX domain protein Ccd1 inhibits JNK activation by axin and dishevelled through distinct mechanisms /". View abstract or full-text, 2004. http://library.ust.hk/cgi/db/thesis.pl?BICH%202004%20WONG.
Testo completoAbstract (sommario):
Thesis (M. Phil.)--Hong Kong University of Science and Technology, 2004.Includes bibliographical references (leaves 60-68). Also available in electronic version. Access restricted to campus users.
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Martins, Soraia Alexandra Araújo. "CD81 interacting proteins". Master's thesis, Universidade de Aveiro, 2016. http://hdl.handle.net/10773/16139.
Testo completoAbstract (sommario):
Mestrado em Biomedicina MolecularFertilization is a multistep and complex process culminating in the merge of gamete membranes, cytoplasmic unity and fusion of genome. CD81 is a tetraspanin protein that participates in sperm-oocyte interaction, being present at the oocyte surface. CD81 has also been implicated in other biological processes, however its specific function and molecular mechanisms of action remain to be elucidated. The interaction between CD81 and its binding partner proteins may underlie the CD81 involvement in a variety of cellular processes and modulate CD81/interactors specific functions. Interestingly, in a Yeast two Hybrid system previously performed in our lab, CD81 has emerged as a putative interactor of the Amyloid Precursor Protein (APP). In the work here described, bioinformatics analyses of CD81 interacting proteins were performed and the retrieved information used to construct a protein-protein interaction network, as well as to perform Gene Ontology enrichment analyses. CD81 expression was further evaluated in CHO, GC-1 and SH-SY5Y cell lines, and in human sperm cells. Additionally, its subcellular localization was analyzed in sperm cells and in the neuronal-like SH-SY5Y cell line. Subsequently, coimmunoprecipitation assays were performed in CHO and SH-SY5Y cells to attempt to prove the physical interaction between CD81 and APP. A functional interaction between these two proteins was accessed thought the analyses of the effects of CD81 overexpression on APP levels. A co-localization analysis of CD81 and some interactors proteins retrieved from the bioinformatics analyses, such as APP, AKT1 and cytoskeleton-related proteins, was also performed in sperm cells and in SH-SY5Y cells. The effects of CD81 in cytoskeleton remodeling was evaluated in SH-SY5Y cells through monitoring the effects of CD81 overexpression in actin and tubulin levels, and analyzing the colocalization between overexpressed CD81 and F-actin. Our results showed that CD81 is expressed in all cell lines tested, and also provided the first evidence of the presence of CD81 in human sperm cells. CD81 immunoreactivity was predominantly detected in the sperm head, including the acrosome membrane, and in the midpiece, where it co-localized with APP, as well as in the post-acrosomal region. Furthermore, CD81 co-localizes with APP in the plasma membrane and in cellular projections in SH-SY5Y cells, where CD81 overexpression has an influence on APP levels, also visible in CHO cells. The analysis of CD81 interacting proteins such as AKT1 and cytoskeletonrelated proteins showed that CD81 is involved in a variety of pathways that may underlie cytoskeleton remodeling events, related to processes such as sperm motility, cell migration and neuritogenesis. These results deepen our understanding on the functions of CD81 and some of its interactors in sperm and neuronal cells.
A fecundação é um processo complexo e faseado que culmina na fusão celular das membranas dos gametas, do citoplasma e do genoma. A CD81 é uma proteína tetraspanina que participa na interacção espermatozóide-oócito, estando presente na superfície do oócito. A CD81 também tem sido associada a outros processos biológicos, contudo a sua função específica e os seus mecanismos de acção não estão elucidados. A ligação entre a CD81 e as suas proteínas interactoras fundamenta o envolvimento da CD81 numa variedade de processos celulares e funções específicas. O desenvolvimento de um sistema de Dois Híbrido em Leveduras, anteriormente realizado no nosso laboratório, mostrou que a CD81 potencialmente interage com a Proteína Percursora de Amilóide (PPA). No presente trabalho, foi realizada a análise bioinformática das proteínas interactoras da CD81. A informação obtida permitiu a construção de uma rede de interações proteína-proteína, bem como a análise de enrequecimento de Ontologia de Genes. Adicionalmente, a expressão da CD81 foi avaliada nas linhas celulares CHO, GC-1 e SH-SY5Y e em espermatozóides humanos. A sua localização subcelular foi também analisada em espermatozóides humanos e na linha de neuroblastoma SH-SY5Y. Foram ainda realizados ensaios de coimunoprecipitacão nas linhas celulares CHO e SH-SY5Y, com a tentativa de provar a intercação física entre a CD81 e a PPA. A interação funcional entre estas duas proteínas foi estudada através da análise do efeito da sobreexpressão da CD81 nos níveis de PPA. Foram também realizados estudos de colocalização entre a CD81 e algumas proteínas interactoras, nos espermatozóides humanos e na linha celular SH-SY5Y. Os interatores analisados, PPA, AKT1 e proteínas relacionadas com o citoesqueleto, foram obtidos da análise bioinformática previamente realizada. O efeito da CD81 na remodelação do citoesqueleto foi avaliado através da monitorização dos efeitos da sobre-expressão da CD81 nos níveis de actina e tubulina, bem como através da análise da colocalização entre a CD81 sobre-expressa e a F-actina. Os nossos resultados mostram que a CD81 está expressa em todas as linhas celulares testadas, providenciando a primeira evidência da presença da CD81 em espermatozóides humanos. A marcação da CD81 foi predominantemente detectada na cabeça do espermatozóde e na peça intermédia, onde colocaliza com a PPA, bem como na região pós-acrossómica. Em adição, a CD81 colocaliza com a PPA na membrana plasmática e nas projecções celulares das células SH-SY5Y, onde a sobre-expressão da CD81 influencia os níveis de PPA, efeito também observado nas células CHO. A análise de proteínas interactoras da CD81, como a AKT1 e proteínas relacionadas com o citoesqueleto, evidenciou que a CD81 está envolvida na remodelação do citoesqueleto, nomeadamente na motilidade dos espermatozóides, na migração celular e na neuritogénese. Estes resultados permitiram aprofundar o conhecimento das funções da CD81 e de alguns dos seus interactores, em espermatozóides e em células neuronais.
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Simões, Inês Tadeu dos Anjos. "Characterization of the in vivo immunomodulatory properties of CD5 and CD6". Doctoral thesis, Universitat de Barcelona, 2017. http://hdl.handle.net/10803/593499.
Testo completoAbstract (sommario):
Our goal in this doctoral thesis was to study the immunomodulatory effects of CD5 and CD6, two proteins expressed on the lymphocytes membrane. These two proteins belong to the Scavenger Receptors Cystein- Rich superfamily, characterized by the presence of one or more cysteine rich domains. The interaction between CD6 and ALCAM, its principal ligand, is well established but recently more ligands of CD6 have been described. On the other hand, to date there is no consensus about the CD5 ligand/s, although several candidates have been proposed. Previous work carried out with CD5 and CD6-deficient mice revealed the role as negative modulators of the T/B cell receptor signaling. It was observed how its blockade/absence led to a hyper-activation post T cell activation by increment of cell proliferation and calcium mobilization.
We decided to generate a transgenic mouse that would expresses high levels of the extracellular region of human CD5 constitutively, in order to block the interactions mediated by membrane-bound CD5 with its ligand/s. This “functional knockout" would resemble what would happen in a clinical application, by injecting the soluble recombinant protein to humans. These mice presented indeed soluble human CD5 in serum, which resulted in immunophenotypical changes that were reproduced in wild-type mice after repeated administration of exogenous rshCD5 protein. The transgenic mice did not reveal any gross alterations in major lymphocyte subpopulations. However, a decrease in T and B cell subsets with regulatory function, as well as an increment in NKT cells were observed. They also presented more severe disease outcome to two different induced experimental autoimmune disease models and a better anti- tumor response against murine melanoma cells (B16-F0) and a genetically modified line of thymoma tumor cells (EG7-OVA). We have observed that this effect is due to the increase in number and activity of the effector cells of the immune system as well as the lowering of the regulatory T cells in the tumor draining lymph nodes. On the other hand, wild-type mice challenged with melanoma showed an improved anti-tumor response as well as changes in the tumor draining lymph node after therapeutic administration of the protein. In addition, in this model, a decrease in IL-6 expression levels was observed, and the loss of efficacy obtained with CD5 administration by depleting NK cells.
We also generated another murine model expressing elevated levels of human soluble CD6 in a constitutive way, with the objective of blocking the heterophillic and homophillic interactions of CD6 and its ligands. These mice had a decrease in the spleen and lymph nodes total cell number. Their B cells have reduced proliferative capacity and the regulatory T cell less suppressive activity aside to a reduction of their number. We observed an increment of the anti-tumor response to different tumor cell lines, but their autoimmune response was not exacerbated, as it was expected. These results were reproduced when recombinant soluble human CD6 was injected repeatedly into wild-type mice.
The results obtained demonstrate that counteracting the function of membrane-bound CD5/CD6 with sCD5/CD6 may enhance current immunotherapies for cancer, as well as be used in other possible applications such as in acute or chronic infections (CD5 recognizes fungal and viral components, and CD6 bacterial components) and vaccines.El objetivo de esta tesis doctoral ha sido el estudio de los efectos inmunomoduladores de CD5 y CD6, dos proteínas expresadas en la membrana de los linfocitos. Estas dos proteínas pertenecen a la superfamilia de receptores "Scavenger Receptor Cystein-Rich", caracterizada por la presencia de uno o varios dominios ricos en cisteínas. En cuanto a CD6, están descritos varios ligandos, sin embargo, a día de hoy no existe un consenso acerca del ligando/s de CD5.Trabajos previos llevados a cabo con ratones deficientes para CD5/CD6, pusieron de manifiesto sus papeles como moduladores negativos de la señal del receptor de células T/B. Decidimos generar un ratón que expresara niveles elevados de la región extracelular de CD5 humano constitutivamente, con el objetivo de bloquear las interacciones mediadas por el CD5 de membrana con su ligando/s. Observamos que estos ratones presentaron una respuesta autoinmune exacerbada pero mejor respuesta anti-tumoral frente a células murinas de melanoma y linfoma. Este efecto se debió al aumento de número y actividad de las células efectoras del sistema inmune, así como a la bajada de las células T reguladoras en los ganglios drenantes del tumor. Por otro lado, ratones silvestres a los que se les indujo melanoma mostraron una respuesta anti-tumoral mejorada, así como cambios en el ganglio drenante, tras la administración de la proteína de forma terapéutica. Además, la eficacia obtenida con la administración de CD5 se perdió al deplecionar las células NK. Además, se generó otro modelo murino que expresara niveles elevados de CD6 soluble humano de manera constitutiva, con el mismo objetivo. Estos ratones presentaron una bajada en cuanto al número de células en bazo y nódulos linfáticos debido a la capacidad proliferativa reducida de los linfocitos B. Además de una disminución en número, las células T reguladoras presentaron una menor actividad supresora. Así, observamos un aumento de la respuesta anti-tumoral frente a diferentes líneas tumorales, pero su respuesta autoinmune no se encontraba exacerbada. Estos resultados se reprodujeron cuando se inyectó CD6 soluble a ratones silvestres. Los resultados obtenidos denotan la importancia de CD5 y CD6 en la respuesta inmune, y su posible aplicación en combinación con las actuales terapias inmunomoduladoras.
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Yap, Jessica. "Identification of Plasmodium falciparum protein kinase substrates and interacting proteins". Honors in the Major Thesis, University of Central Florida, 2012. http://digital.library.ucf.edu/cdm/ref/collection/ETH/id/644.
Testo completoAbstract (sommario):
Characterization of PfPKA and PfPK5 substrates, as well as the proteins they interact with, will help us to develop innovative therapies targeting binding sites.; Malaria is a devastating disease that results in almost one million deaths annually. Most of the victims are children under the age of five in Sub-Saharan Africa. Malaria parasite strains throughout developing countries are continually building resistance to available drugs. Current therapies such as mefloquine, chloroquine, as well as artemisinin are becoming less effective, and this underscores the urgency for therapeutics directed against novel drug targets. In order to identify new drug targets, the molecular biology of the malaria parasite Plasmodium needs to be elucidated. Plasmodium exhibits a unique cell cycle in which it undergoes multiple rounds of DNA synthesis and mitosis without cytokinesis. Thus, cell cycle regulatory proteins are likely to be promising pathogen-specific drug targets. It is expected that fluctuating activity of key proteins, such as protein kinases, play an essential role in regulating the noncanonical life cycle of Plasmodium. Consequently, malarial kinases are a prime target for therapy. One way to better understand the role of malarial kinases in Plasmodium cell cycle regulation is to identify putative protein kinase substrates and interacting proteins. Two malarial kinases that have been implicated in regulating malaria parasite cell cycle stages were investigated in this study: P. falciparum CDK-like Protein Kinase 5 (PfPK5) and cAMP-Dependent Protein Kinase A (PfPKA). A transgenic P. falciparum line was created for the expression of epitope-tagged PfPK5 for pull-down analysis. Phospho-substrate antibodies were used to identify physiological substrates of both PfPK5 and PfPKA. Immunoblotting with these antibodies identified several potential substrates. Identities of the PfPKA physiological substrates were determined from the global P. falciparum phosphoproteome dataset that has recently been generated in our laboratory.B.S.
Bachelors
Burnett School of Biomedical Sciences
Molecular and Microbiology
8
Nishimura, Hiroyuki. "Developmentally regulated expression of the PD-1 protein on the surface of double-negative (CD4-CD8-) thymocytes". Kyoto University, 1997. http://hdl.handle.net/2433/202184.
Testo completo
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Carrat, Christophe. "Etude des mécanismes de présentation des lipides mycobactériens par les protéines CD1". Thesis, Toulouse 3, 2019. http://www.theses.fr/2019TOU30293.
Testo completoAbstract (sommario):
Les antigènes lipidiques sont présentés aux lymphocytes T par les protéines CD1 (CD1a à CD1d), exprimées à la surface des cellules présentatrices d'antigènes (CPA), telles que les cellules dendritiques. Dans le cadre de la tuberculose, dont l'agent étiologique est Mycobacterium tuberculosis (Mtb), un certain nombre d'antigènes lipidiques ont été décrits depuis une vingtaine d'années et sont, pour la plupart, présentés par l'isoforme CD1b. Plus récemment, il a été montré que ces antigènes peuvent être apprêtés par des enzymes lysosomales de la CPA, à l'instar de ce qui est connu pour la présentation de peptides par le complexe majeur d'histocompatibilité. L'apprêtement des antigènes lipidiques fait intervenir des protéines de transfert de lipides, telle que la protéine CD1e, une cinquième isoforme soluble nécessaire à la présentation des phosphatidyl-myo-inositol mannosides (PIM). Cependant, les mécanismes de l'apprêtement des antigènes lipidiques ainsi que les épitopes réellement présentés par CD1b à la surface des CPA sont à l'heure actuelle encore mal connus. Afin d'identifier les épitopes lipidiques présentés par CD1b lors d'une infection par Mtb, nous avons développé une stratégie innovante permettant d'isoler les complexes CD1b: lipides présents en surface des CPA, en s'affranchissant de l'utilisation de détergents, proscrits ici. Cette stratégie repose sur la construction d'une CPA comportant une forme clivable de la protéine CD1b, exprimant CD1e ou non. Les conditions de clivage et de purification de ces complexes ont été mises au point au cours de ce travail. Les lipides sont ensuite extraits et analysés par HPLC couplée à la spectrométrie de masse. Cette approche a permis l'analyse des lipides endogènes présents dans CD1b. Elle a ensuite été validée par des expériences de stimulation des CPA par des lipides mycobactériens purifiés. Afin d'améliorer la détection des épitopes par spectrométrie de masse, des optimisations des conditions de stimulation des CPA sont actuellement en cours. Le second axe développé au cours de ma thèse a porté sur l'étude des enzymes impliquées dans l'apprêtement des antigènes lipidiques. Les glycolipides mycobactériens peuvent être le substrat d'enzymes lysosomales. Les PIM en sont l'exemple le mieux caractérisé, car des enzymes intervenant dans l'hydrolyse de la partie saccharidique et de la partie hydrophobe du lipide ont été identifiées. Pour déterminer de nouvelles activités enzymatiques impliquées dans l'apprêtement des glycolipides de Mtb, nous avons cherché à générer une fraction lysosomale issue de CPA pour réaliser des tests de dégradation in vitro. [...]Lipid antigens are presented to T cells by CD1 proteins (CD1a to CD1d), expressed at the surface of antigen presenting cells (APC), such as dendritic cells. Mycobacterium tuberculosis (Mtb) is the etiological agent of tuberculosis (TB). Mtb lipid antigens have been described over the last twenty years and are, most of them being presented by the CD1b isoform. Recent studies in humans have shown that specific T cell responses to Mtb lipid antigens play a role in the immune response to infection, and contribute to the creation of a reservoir of memory cells. More recently, it has been shown that these antigens can be processed by lysosomal enzymes in APC, similarly to that described for the presentation of peptides by the major histocompatibility complex. Lipid antigens processing involves lipid transfer proteins, such as CD1e, a fifth soluble isoform of CD1 necessary for the presentation of phosphatidyl-myo-inositol mannosides (PIM). However, the mechanisms of lipid antigen processing as well as the repertoire of epitopes presented by CD1b at the surface of APC are still poorly understood. In order to identify the lipid epitopes presented by CD1b during Mtb infection, we developed an innovative strategy to isolate the CD1b:lipid complexes at the surface of APC, by avoiding the use of detergents. This strategy is based on the construction of APC cell lines. Different APC were constructed, that may or may not express CD1e. The conditions of CD1b cleavage and purification, as well as the extraction of the lipids and their analysis by mass spectrometry were set up and optimized. The lipids are then extracted and analyzed by HPLC coupled to mass spectrometry. In a first step, this approach was used to study the endogenous lipids presented by CD1b. It was then validated by APC stimulation experiments with purified mycobacterial lipids. Detecting Mtb epitopes during infection requires a high sensitivity in mass spectrometry. Therefore, optimizations of the APC stimulation efficiency are in progress. The second axis developed during my PhD focused on the study of enzymes involved in lipid antigen processing. Mycobacterial glycolipids may be the substrates of lysosomal enzymes. Among them, the PIM family is the best characterized example, with enzymes involved in the hydrolysis of both the saccharide and the lipid part. To characterize new enzymatic activities involved in processing of Mtb glycolipids, we sought to generate a lysosomal fraction from APC to perform in vitro digestion tests. Digestion tests for tetra-acylated PIM2 yielded di and tri-acylated PIM2 generated by lipase activities.[...]
10
Gohring, John Thomas Fan Xudong. "Detection of CD4 and CD8 t-lymphocytes and HER2 breast cancer biomarker using the opto-fluidic ring resonator biosensor". Diss., Columbia, Mo. : University of Missouri--Columbia, 2009. http://hdl.handle.net/10355/6661.
Testo completoAbstract (sommario):
Title from PDF of title page (University of Missouri--Columbia, viewed on March 10, 2010). The entire thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file; a non-technical public abstract appears in the public.pdf file. Thesis advisor: Dr. Xudong Fan. Includes bibliographical references.
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Prince, Amanda L. "The Role of Inducible T Cell Kinase (Itk) in the Development of Innate T Cells and in the Formation of Protective Memory Responses: A Dissertation". eScholarship@UMMS, 2013. https://escholarship.umassmed.edu/gsbs_diss/660.
Testo completoAbstract (sommario):
T cell development in the thymus produces multiple lineages of cells, including conventional naïve CD4+ and CD8+ T cells, regulatory T cells, and innate T cells. Innate T cells encompass γδ T cells, invariant natural killer (iNKT) cells, mucosal-associated invariant T (MAIT) cells, and H2-M3-restricted cells (Berg, 2007). Although they are a minor subset of all thymocytes, innate T cells develop in the thymus and share characteristics of the innate and adaptive immune systems (Berg, 2007). These lymphocytes undergo antigen receptor rearrangement and are able to exert their effector function immediately upon ex vivo stimulation (Berg, 2007). However, in several strains of mice harboring mutations in T cell signaling proteins or transcriptional regulators, conventional CD8+ T cells develop as innate cells that share characteristics with memory T cells (Atherly et al., 2006b; Broussard et al., 2006; Fukuyama et al., 2009; Gordon et al., 2011; Verykokakis et al., 2010b; Weinreich et al., 2010). One of these signaling proteins, inducible T cell kinase (Itk) is a nonreceptor protein tyrosine kinase that signals downstream of the T cell receptor (TCR) (Berg et al., 2005). Upon TCR activation, Itk is activated and recruited to the TCR signaling complex, where Itk interacts with Src homology 2 (SH2) domain-containing leukocyte phosphoprotein of 76 kDa (SLP-76), linker for activation of T cells (LAT), and phospholipase C γ1 (PLCγ1) (Berg et al., 2005). Thus, in Itk-deficient mice, TCR signaling is disrupted, which results in mature CD4- CD8+ (CD8SP) thymocytes that are CD44high, CD62Lhigh, CD122+, and CXCR3+ and that express high levels of the transcription factor, Eomesodermin (Eomes) (Atherly et al., 2006b; Broussard et al., 2006; Weinreich et al., 2010). Recently, it was determined that the development of these innate CD8SP thymocytes in itk-/- mice is dependent on IL-4 produced in the thymic environment by a poorly characterized subset of CD3+ thymocytes expressing the transcriptional regulator, promyelocytic leukemia zinc finger (PLZF) (Gordon et al., 2011; Verykokakis et al., 2010b; Weinreich et al., 2010). Here we show that a sizeable proportion of mature CD4+ CD8- (CD4SP) thymocytes in itk-/- mice also develop as Eomesodermin+ innate T cells. These Eomes+ innate CD4+ T cells are CD44high, CD62Lhigh, CD122+, and CXCR3+ (Atherly et al., 2006b; Broussard et al., 2006; Dubois et al., 2006; Weinreich et al., 2010). Surprisingly, neither CD4SP nor CD8SP innate thymocytes in itk-/- mice are dependent on γδ T cells for their development as was previously hypothesized (Alonzo and Sant'Angelo, 2011). Instead, both subsets of innate itk-/- T cells require the presence of a novel PLZF-expressing, SAP-dependent thymocyte population that is essential for the conversion of conventional CD4+ and CD8+ T cells into Eomesodermin-expressing innate T cells with a memory phenotype. This novel subset of PLZF-expressing SAP-dependent innate T cells preferentially home to the spleen and mesenteric lymph nodes and have a restricted TCR repertoire. Thus, we have christened this subset as CD4+ PLZF + MAIT-like cells. We have characterized multiple subsets of innate T cells that expand in the absence of Itk. Therefore, we were interested in how innate T cells respond to infection. Although Itk KO mice have defects in cytolytic function and cytokine production during an acute infection, these mice are able to clear viral infections (Atherly et al., 2006a; Bachmann et al., 1997). Hence, we hypothesized that Itk-deficient memory CD8+ T cells would be able to provide protection upon a challenge infection. Conversely, we found this not to be true although Itk-deficient memory CD8+ T cells were present in similar frequencies and cell numbers as WT memory CD8+ T cells at 42 days post-infection. Furthermore, Itk-deficient memory CD8+ T cells were able to produce IFNγ and exert cytolytic function upon stimulation. Although the function of Itk-deficient memory CD8+ T cells appeared to be intact, we found that these cells were unable to expand in response to a challenge infection. Remarkably, conventional memory CD8+ T cells lacking Itk were able to expand and form protective memory responses upon challenge. Thus, the inability of Eomes+ innate CD8+ T cells to form protective memory responses does not appear to be intrinsic to cells deficient in Itk. This thesis is divided into six major chapters. The first chapter will provide an introduction to T cell development and the role of Itk in T cell development. Additionally, it will introduce a variety of innate T cell subsets that will be discussed throughout this thesis and will provide an overview of CD4+ and CD8 + T cell differentiation during infection. This section will explain the role of Itk in CD4+ helper T cell differentiation and describe how Itk-deficient CD8+ T cells respond to acute infection. The introduction will also discuss the generation of conventional memory CD8+ T cells. The second chapter will provide the details of the experimental procedures used in this thesis. The third chapter will describe the characterization and development of Eomes+ innate CD4+ T cells that develop in the absence of Itk. Additionally, this chapter will address the subset of PLZF+ innate T cells that induce the expression of Eomes in innate T cells. The fourth chapter will further characterize and explore the development of itk-/- CD4+ PLZF+ MAIT-like T cells. The fifth chapter will examine the role of Eomes + innate CD8+ T cells in protective memory responses. Chapters three through five will display work that is in preparation to be submitted to a peer-reviewed journal. The sixth chapter will discuss the results of this thesis and their implications.
12
Prince, Amanda L. "The Role of Inducible T Cell Kinase (Itk) in the Development of Innate T Cells and in the Formation of Protective Memory Responses: A Dissertation". eScholarship@UMMS, 2002. http://escholarship.umassmed.edu/gsbs_diss/660.
Testo completoAbstract (sommario):
T cell development in the thymus produces multiple lineages of cells, including conventional naïve CD4+ and CD8+ T cells, regulatory T cells, and innate T cells. Innate T cells encompass γδ T cells, invariant natural killer (iNKT) cells, mucosal-associated invariant T (MAIT) cells, and H2-M3-restricted cells (Berg, 2007). Although they are a minor subset of all thymocytes, innate T cells develop in the thymus and share characteristics of the innate and adaptive immune systems (Berg, 2007). These lymphocytes undergo antigen receptor rearrangement and are able to exert their effector function immediately upon ex vivo stimulation (Berg, 2007). However, in several strains of mice harboring mutations in T cell signaling proteins or transcriptional regulators, conventional CD8+ T cells develop as innate cells that share characteristics with memory T cells (Atherly et al., 2006b; Broussard et al., 2006; Fukuyama et al., 2009; Gordon et al., 2011; Verykokakis et al., 2010b; Weinreich et al., 2010). One of these signaling proteins, inducible T cell kinase (Itk) is a nonreceptor protein tyrosine kinase that signals downstream of the T cell receptor (TCR) (Berg et al., 2005). Upon TCR activation, Itk is activated and recruited to the TCR signaling complex, where Itk interacts with Src homology 2 (SH2) domain-containing leukocyte phosphoprotein of 76 kDa (SLP-76), linker for activation of T cells (LAT), and phospholipase C γ1 (PLCγ1) (Berg et al., 2005). Thus, in Itk-deficient mice, TCR signaling is disrupted, which results in mature CD4- CD8+ (CD8SP) thymocytes that are CD44high, CD62Lhigh, CD122+, and CXCR3+ and that express high levels of the transcription factor, Eomesodermin (Eomes) (Atherly et al., 2006b; Broussard et al., 2006; Weinreich et al., 2010). Recently, it was determined that the development of these innate CD8SP thymocytes in itk-/- mice is dependent on IL-4 produced in the thymic environment by a poorly characterized subset of CD3+ thymocytes expressing the transcriptional regulator, promyelocytic leukemia zinc finger (PLZF) (Gordon et al., 2011; Verykokakis et al., 2010b; Weinreich et al., 2010). Here we show that a sizeable proportion of mature CD4+ CD8- (CD4SP) thymocytes in itk-/- mice also develop as Eomesodermin+ innate T cells. These Eomes+ innate CD4+ T cells are CD44high, CD62Lhigh, CD122+, and CXCR3+ (Atherly et al., 2006b; Broussard et al., 2006; Dubois et al., 2006; Weinreich et al., 2010). Surprisingly, neither CD4SP nor CD8SP innate thymocytes in itk-/- mice are dependent on γδ T cells for their development as was previously hypothesized (Alonzo and Sant'Angelo, 2011). Instead, both subsets of innate itk-/- T cells require the presence of a novel PLZF-expressing, SAP-dependent thymocyte population that is essential for the conversion of conventional CD4+ and CD8+ T cells into Eomesodermin-expressing innate T cells with a memory phenotype. This novel subset of PLZF-expressing SAP-dependent innate T cells preferentially home to the spleen and mesenteric lymph nodes and have a restricted TCR repertoire. Thus, we have christened this subset as CD4+ PLZF + MAIT-like cells. We have characterized multiple subsets of innate T cells that expand in the absence of Itk. Therefore, we were interested in how innate T cells respond to infection. Although Itk KO mice have defects in cytolytic function and cytokine production during an acute infection, these mice are able to clear viral infections (Atherly et al., 2006a; Bachmann et al., 1997). Hence, we hypothesized that Itk-deficient memory CD8+ T cells would be able to provide protection upon a challenge infection. Conversely, we found this not to be true although Itk-deficient memory CD8+ T cells were present in similar frequencies and cell numbers as WT memory CD8+ T cells at 42 days post-infection. Furthermore, Itk-deficient memory CD8+ T cells were able to produce IFNγ and exert cytolytic function upon stimulation. Although the function of Itk-deficient memory CD8+ T cells appeared to be intact, we found that these cells were unable to expand in response to a challenge infection. Remarkably, conventional memory CD8+ T cells lacking Itk were able to expand and form protective memory responses upon challenge. Thus, the inability of Eomes+ innate CD8+ T cells to form protective memory responses does not appear to be intrinsic to cells deficient in Itk. This thesis is divided into six major chapters. The first chapter will provide an introduction to T cell development and the role of Itk in T cell development. Additionally, it will introduce a variety of innate T cell subsets that will be discussed throughout this thesis and will provide an overview of CD4+ and CD8 + T cell differentiation during infection. This section will explain the role of Itk in CD4+ helper T cell differentiation and describe how Itk-deficient CD8+ T cells respond to acute infection. The introduction will also discuss the generation of conventional memory CD8+ T cells. The second chapter will provide the details of the experimental procedures used in this thesis. The third chapter will describe the characterization and development of Eomes+ innate CD4+ T cells that develop in the absence of Itk. Additionally, this chapter will address the subset of PLZF+ innate T cells that induce the expression of Eomes in innate T cells. The fourth chapter will further characterize and explore the development of itk-/- CD4+ PLZF+ MAIT-like T cells. The fifth chapter will examine the role of Eomes + innate CD8+ T cells in protective memory responses. Chapters three through five will display work that is in preparation to be submitted to a peer-reviewed journal. The sixth chapter will discuss the results of this thesis and their implications.
13
Clark, Krista Lea. "Characterization of CD81 associated proteins on T cells /". Search for this dissertation online, 2003. http://wwwlib.umi.com/cr/ksu/main.
Testo completo
14
Sjögren, Tove. "Structural Plasticity and Function in Cytochrome cd1 Nitrite Reductase". Doctoral thesis, Uppsala universitet, Institutionen för naturvetenskaplig biokemi, 2001. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-1394.
Testo completoAbstract (sommario):
Cytochrome cd1 nitrite reductase is a bifunctional enzyme, which catalyses the one-electron reduction of nitrite to nitric oxide, and the four-electron reduction of oxygen to water. The latter is a cytochrome oxidase reaction. Both reactions occur on the d1 haem iron of the enzyme. Time resolved crystallographic studies presented here show that the mechanisms of nitrite and oxygen reduction share common elements. This is of interest from an evolutionary point of view since aerobic respiratory enzymes are thought to have evolved from denitrifying enzymes. Despite of similarities, the results also imply different requirements for the timing of electron transfer to the active site in these reactions. Quantum chemical calculations suggest that nitric oxide, the product of nitrite reduction, is not spontaneously released from the haem iron while this is not the case with water. Reduction of the haem while nitric oxide is still bound to it would result in a tight dead-end complex. A mechanism must therefore exist for the selective control of electron transfer during the reaction. Structural studies with a product analogue (carbon monoxide) combined with flash photolysis of the complex in solution revealed an unexpected proton uptake by the active site as the neutral CO molecule left the enzyme. This led to the suggestion that the increased positive potential of the active site triggers preferential electron transfer when the active site is empty. Crystallisation and structure determination of the reduced enzyme revealed extremely large domain rearrangements. These results offer insights into the role of tethered electron shuttle proteins in complex redox systems, and suggests a mechanism for conformational gating in catalysis.
15
Ni, Tao. "Structural and functional study of MACPF/CDC superfamily proteins". Thesis, University of Oxford, 2016. https://ora.ox.ac.uk/objects/uuid:50793dea-6aa6-4922-b7d8-43c50079639b.
Testo completoAbstract (sommario):
The thesis mainly focuses on structural study of proteins in membrane attack complex- perforin/cholesterol-dependent cytolysins (MACPF/CDC) superfamily, in particular, Astrotactins from human and perforin-like proteins (PLPs) from Toxoplasma and Plasmodium. Both of these subfamily proteins are implicated in human diseases and structurally uncharacterised before. Astrotactins (ASTNs) have been shown to play a crucial role in enabling neural migration along glial fibres. While ASTN1 directly forms contacts between the neuron and the fibre, ASTN2 is responsible for extracting ASTN1 from contacts at the lagging edge of the cell and recycling them to the leading edge. ASTN2 is associated with endosomes, with the majority of its structure (at the C-terminus) projecting into their lumen, anchored by a pair of transmembrane â-helices. Here we present the structure of this "endodomain" region of ASTN2, and find it to consist of a unique combination of polypeptide folds comprising a membrane attack com- plex/perforin (MACPF) domain, an EGF-like domain, a previously-unobserved form of fibronectin type III (Fn(III)) domain and an annexin-like domain. Taken together, the structural characterisation provides a framework for better understanding the mechanism of the ASTNs and related proteins in neural and other forms of vertebrate development. Perforin-like proteins from Toxoplasma and Plasmodium (TgPLP1 and PPLPs) are critical for normal life cycle progression of these parasites, and knockout out of any of them results in significant defects in their life cycle, entrapping the parasites within the host cells and thus limiting their ability to egress. Here we present the crystal structures of TgPLP1 MACPF domain and C-terminal domain at 2.0 Å and 1.1 Å, respectively. We also presents the MACPF domain assembled in helical and hexameric ring form, which indicates the possibility of pore-formation by 6 subunits. This is the first structure of perforin-like protein from Apicomplexan parasites and provides a structural basis to elucidate the function of PLPs in toxoplasmosis and malaria pathogenesis. The final section is a continuation of a previous study in our lab on pleckstrin homology (PH) domain of kindlins. We determined the crystal structure of kindlin-3 PH domain and characterised its lipid and membrane binding properties. Using nanodiscs incorporated with different lipids as model membrane system to study the interaction between inositol phosphate lipids and kindlin-3 PH domain, together with molecular dynamic simulation studies (in collaboration with Mark Sansom's group, University of Oxford), we propose that a subset of PH domains is able to bind to multiple inositol phosphates simultaneously and so via an avidity effect have its interaction with target membranes strengthened.
16
Jérôme, Kerzerho. "ÉVALUATION DES PROTEINES MIDKINE ET SURVIVINE SUREXPRIMEES DANS LES CELLULES TUMORALES COMME CIBLES DE L'IMMUNITE CELLULAIRE ANTI-TUMORALE". Phd thesis, Université Paris Sud - Paris XI, 2009. http://tel.archives-ouvertes.fr/tel-00383751.
Testo completoAbstract (sommario):
Peu d'antigènes tumoraux présentent à la fois une expression dans un grand nombre de cellules tumorales et exercent un rôle vital pour leur développement. Mon travail de thèse a consisté à étudier les réponses en lymphocytes T dirigées contre deux de ces protéines : la Survivine et la Midkine. Nous avons démontré que ces deux protéines induisent spécifiquement des lymphocytes T CD4+ et CD8+ capables de reconnaître des cellules tumorales. Nous avons également identifié les séquences immunogéniques (épitopes T) dans ces antigènes. Deux épitopes CD4 et CD8 de la Midkine sont localisés dans le peptide signal. Notre étude montre que la Midkine constitue une nouvelle cible pour la vaccination contre de nombreux cancers et propose des séquences peptidiques originales comme candidats vaccins.
17
Kieselbach, Brit. "Molekulare Effekte der Immunmodulation mit einem anti-CD4-Antikörper". Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2004. http://dx.doi.org/10.18452/15074.
Testo completoAbstract (sommario):
Das grundlegende Problem in der Transplantationsimmunologie ist es, die Langzeitakzeptanz eines fremden (allogen) Organs zu erreichen, ohne die sonstige Immunkompetenz des Empfängers zu beeinträchtigen. Die Induktion einer solchen spenderspezifischen Toleranz würde eine Alternative zum Langzeiteinsatz von Immunsuppressiva darstellen. Deswegen versucht man, während der Transplantation die Aktivierung der für die Abstoßung entscheidenden T-Helferzellen zu unterdrücken, bis eine Akzeptanz des Spenderorgans etabliert ist. Wichtig für eine Aktivierung der T-Zellen ist das für alle T-Helferzellen typische Zelloberflächenmolekül CD4. Antikörper gegen CD4 können in Tiermodellen eine Transplantattoleranz induzieren. Ein besonderes Interesse gilt der Charakterisierung der genauen Mechanismen dieser induzierten Transplantatakzeptanz, da diese noch wenig verstanden sind. Der von uns verwendete nicht-depletierende Maus-anti-Ratten-CD4mAk (RIB5/2) besitzt im allogenen Nierentransplantationsmodell der Ratte eine hohe toleranzinduzierende Wirkung und erzielt eine permanente Transplantatakzeptanz bei >80% der Empfängertiere. In dieser Arbeit wurde versucht, die Effekte dieses monoklonalen Antikörpers auf die T-Zellaktivierung näher zu untersuchen. Ausdruck der blockierten T-Zellaktivierung ist eine verminderte T-Zell-Proliferation und die Reduzierung der Synthese von TH-1-Effektorzytokinen, welche eine zelluläre Immunantwort fördern. Zu diesen für die Abstoßung gefährlichen Th-1-Effektorzytokinen gehören Interleukin 2 (=IL-2, Hauptwachstumsfaktor aktivierter T-Zellen) und Interferon gamma (=IFNgamma, wichtiger Aktivator von APC''s). Während die IL-2 Produktion vollständig verhindert wird, ist die Alloantigen-induzierte IFNgamma mRNA Expression nicht reduziert. Allerdings kommt es unter dem Einfluss des Antikörpers nicht zur IFNgamma Proteinsekretion. Wird jedoch das fehlende IL-2 ersetzt, kann sowohl die defekte Proliferation als auch die posttranskriptionelle Blockade der IFNgamma Produktion wieder aufgehoben werden. Das spiegelt sich auch in vivo wieder, da rekombinantes IL-2 auch hier den Toleranzstatus brechen kann. In dieser Arbeit konnte ein Kandidat dieser IFNgamma Translationskontrolle ermittelt werden. Zusätzlich wurde das Kochaperon p23, Teil eines Hsp90-Komplexes, in unsere Untersuchungen miteinbezogen, da es als ein differentiell reguliertes Gen in allogenen und anti-CD4mAk-behandelten T-Zellen identifiziert wurde. Hsp90 stabilisiert z.B. Kinasen, die wichtige Mediatoren der Signaltransduktion sind. p23 könnte aufgrund seiner Funktion als Kofaktor von Hsp90 an der Regulierung dieser Kinasen beteiligt sein, ist jedoch bisher kaum im Zusammenhang mit T-Zellaktivierung analysiert worden. Meine Untersuchungen ergaben, dass die p23 Expression ebenfalls durch den anti-CD4mAk reduziert wird. Da die Proliferation/p23 und die IFNgamma Synthese IL-2-abhängig reguliert werden, wurden IL-2-induzierte Signalwege auf ihre Relevanz für Proliferation und IFNgamma Regulation hin untersucht. Die Aufklärung der molekularen Mechanismen der anti-CD4mAk-Behandlung auf die T-Zellaktivierung soll mit dazu beitragen, Grundlagen für ein besseres Verständnis des Abstoßungsprozesses und damit Transplantatfunktions-Monitoring zu schaffen.The major problem in transplantation immunology is the development of long-term donor-specific nonresponsiveness without reduction of the normal recipient immunocompetence. A tolerant state would obviate the need for continuing immunosuppressive therapy. One level of immunosuppression for inducing graft acceptance involves antibodies specific for T-cells of the recipient leading to donorspecific tolerance (e.g. by using of anti-CD4 monoclonal antibodies = aCD4mAb). CD4+ T cells play a predominant role in the cascade of immune processes following transplantation of foreign tissues. The anti-rat CD4 mAb RIB5/2 is very potent in inducing allo-specific tolerance to renal and heart allografts in rat recipients. Here I investigated the molecular mechanisms underlying anti-CD4 antibody mediated inhibition of allo-specific T cell activation and how this is antagonised by exogenous IL-2. IL-2 acts as growth factor for antigen-activated T cells by inducing the expression of cell cycle proteins and also enhances the expression of cytokines, e.g. IFNgamma in T cells. IFNgamma profoundly affects a variety of immune responses, including activation of antigen presenting cells. Anti-CD4 treatment, in vivo and in vitro, completely abrogated IL-2 production by alloreactive T cells. In contrast, anti-CD4 treated allo-activated T cells showed similar IFNgamma mRNA expression as untreated allo-activated T cells, but did not secrete any protein. Thus, the anti-CD4 antibody cannot prevent IFNgamma mRNA expression but is interfering with posttranscriptional mechanisms controlling IFNgamma production during allo-activation of T cells. The investigations revealed a candidate of these IFNgamma translation control. Additionally I investigated the heat shock protein 90 (Hsp90)-associated cochaperone p23. p23 upregulation during T cell activation is also abrogated by anti-CD4 treatment. Hsp90 chaperoning is critical for proper folding, stabilization and trafficking of a number of cellular signaling proteins as e.g. kinases. Hsp90-kinase complexes play an important role in T-cell signal transduction and little is known about the importance or even regulation of Hsp90-cochaperones like p23 during T-cell activation. I analysed the regulation of p23 and downstream effects on different kinases involved in T-cell signaling. These findings are supposed to contribute to a better understanding of the mechanisms underlying tolerance induction.
18
Lucas, Julie Ann. "The Role of Itk in T Cell Development: A Dissertation". eScholarship@UMMS, 2005. https://escholarship.umassmed.edu/gsbs_diss/91.
Testo completoAbstract (sommario):
Itk is a member of the Tec family of non-receptor tyrosine kinases. It is expressed in T cells, NK cells, and mast cells. The purpose of this study was to determine the role of Itk in T cell development. Previous work from our lab and others has demonstrated that Itk is involved in signaling downstream of the T cell receptor and initial analysis of Itk-deficient mice revealed that these mice had some defects in T cell development. There are two stages of T cell development, the pre-T cell stage and the CD4+ CD8+ double positive stage, at which signals downstream of the T cell receptor are important. At the CD4+ CD8+ double positive stage, these signals direct two concurrent, but distinct processes known as repertoire selection and CD4/CD8 lineage commitment/differentiation. I show that there are only slight defects in development at the pre-T cell stage, presumably due to reduced TCR signaling. However these results clearly demonstrate that Itk is not essential at this stage of development. In contrast, repertoire selection, in particular positive selection, is significantly affected by the absence of Itk. Similarly, I show that Itk plays a role in lineage differentiation, although commitment to the appropriate lineage occurs normally in the absence of Itk.
19
Atherly, Luana O. "The Role of ITK and RLK in CD8+ T Cell Development and Function: a Dissertation". eScholarship@UMMS, 2004. https://escholarship.umassmed.edu/gsbs_diss/120.
Testo completoAbstract (sommario):
Itk and Rlk are members of the Tec kinase family of non-receptor protein tyrosine kinases that are preferentially expressed in T cells. Numerous previous studies have demonstrated that these proteins play an important a role in the regulation of signalling processes downstream of TCR activation in CD4+ T cells, particularly in the phosphorylation of PLCγl. In addition, Itk and Rlk have both been shown to be important for CD4+ T cell development, differentiation, function and homeostasis following TCR activation. In the absence of Itk and Rlk, CD8+ SP thymocytes and T cells develop a memory/previously activated phenotypic profile, however, very little is known about the influence of Itk and Rlk on CD8+ T cell development and function. This study illustrates a previously unappreciated role for Itk and Rlk in the regulation of cytokine signals during CD8+ SP thymocyte maturation, and in the development of the memory CD44hi profile of Itk -/- and Itk -/- Rlk -/- CD8+ SP thymocytes and CD8+ T cells. This study also provides the first detailed study of the role of loss of Itk and particularly both Itk and Rlk in CD8+ signalling and function and shows that these Tec kinase family members play an important role in the maintenance of CD8+ T cell fitness and function, particularly in the ability of CD8+ T cells to accumulate in response to infection. Collectively, my studies demonstrate a critical role for Itk and Rlk in the generation of optimal CD8+ T cell responses. They also raise the novel observation that these proteins may be involved on the regulation of cytokine signals in T cells.
20
Hintze, Thorsten. "Einfluss des clostridialen C3 Toxins auf die Dendritenmorphologie und Spinebildung von CA1 Pyramidenzellen in Hippocampus-Schnittkulturen der Maus - eine quantitative lichtmikroskopische Untersuchung". Doctoral thesis, Universitätsbibliothek Leipzig, 2010. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-62305.
Testo completoAbstract (sommario):
Lokale Pyramidenzellen sind die Hauptneurone des Hippocampus und können durch ihre Position und die Morphologie ihrer Dendriten als CA1 und CA3 Pyramidenzellen identifiziert werden. Die Dendriten der exzitatorischen Pyramidenzellen sind mit postsynaptischen Vorwölbungen, den so genannten Spines, bedeckt, welche in einem spezifischen Verteilungsmuster angeordnet sind. Neurotoxine wie das C3 Toxin von Clostridium botulinum sind funktionelle Substanzen, die die neuronale Morphologie verändern und die neuronale Funktion beeinflussen können. In dieser Studie wurden die morphologischen Veränderungen von intrazellulär mit Biocytin gefüllten CA1 Pyramidenzellen qualitativ und quantitativ analysiert. Die hippocampalen Schnittkulturen, in denen sich bekanntermaßen Pyramidenzellen ähnlich entwickeln wie in vivo, wurden dazu herangezogen, die Effekte der C3bot Toxin-Applikation auf die Verzweigung der Dendriten sowie Anzahl und Dichte der dendritischen Spines zu untersuchen. Drei Gruppen von Zellen wurden verglichen: Erstens Neurone, die in serumhaltigem Medium inkubiert worden waren, zweitens Nervenzellen, die in einem Medium ohne Serum inkubiert worden waren und drittens Zellen, die unter Serumentzug dem C3bot Toxin ausgesetzt worden waren. Die Inkubation dauerte 14 Tage, während die Dauer der Toxinexposition zwischen vier und sechs Stunden betrug. Mit Hilfe eines Computers wurden zweidimensionale Nachbildungen der biocytin-markierten CA1 Pyramidenzellen erstellt, und die Gesamtlänge der Dendriten, die Anzahl der dendritischen Verzweigungspunkte und die Gesamtzahl und Dichte der dendritischen Spines gemessen und statistisch ausgewertet. Signifikante Unterschiede wurden zwischen der mit C3 Toxin behandelten Gruppe und der serumhaltig inkubierten Kontrollgruppe beobachtet. Diese signifikanten morphologischen Veränderungen traten selektiv an den Apikaldendriten der toxinbehandelten CA1 Pyramidenzellen auf. Aus der Behandlung resultierte eine Reduktion der Anzahl apikaler Verzweigungspunkte, der Anzahl der apikalen Spines, der Gesamtzahl (basal und apikal addiert) der Spines sowie der Gesamtspinedichte. Im Gegensatz dazu ergaben sich keine signifikanten Unterschiede zwischen der toxinbehandelten Gruppe und der ohne Serum inkubierten Kontrollgruppe, obwohl der Serumentzug im Vergleich zur serumhaltig inkubierten Kontrollgruppe die Entwicklung der Zellen beeinflusste. Auf Grundlage der beobachteten Veränderungen können wir schließen, dass die Behandlung mit C3 bot einen starken Einfluss selektiv auf die Morphologie der Apikaldendriten ausübt. Der Mechanismus, der dieser selektiven Empfindlichkeit der Apikaldendriten gegenüber dem C3 bot Toxin zugrunde liegt, wird Gegenstand weiterer Untersuchungen sein.
21
Barbu, Corina. "Der Einfluss des Tau-Proteins auf die Morphologie von Nervenzellen". Doctoral thesis, Universitätsbibliothek Leipzig, 2012. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-100213.
Testo completoAbstract (sommario):
Tau ist ein Mikrotubuli-assoziiertes Protein, das die Polymerisation von Tubulin fördert und die Mikrotubuli stabilisiert. Folglich wird angenommen, dass Tau essentiell für die neuronale Morphogenese ist, vor allem für die Axonenausbildung und -aufrechterhaltung. Mittels tangentieller Nissl-gefärbter Schnitte von Mäusegehirnen konnte in der vorliegenden Arbeit gezeigt werden, dass Tau-knockout Mäuse die regelhafte thalamokortikale Barthaar-projektion („Barrel“ Konfiguration) entwickeln. Der Einfluss von Tau auf die Entstehung von Dendriten wurde anhand von Golgi-gefärbten Präparaten untersucht. Die Sholl-Analyse der gefärbten CA1-Pyramidenzellen zeigte, dass die Komplexität apikaler Dendriten durch das Fehlen von Tau reduziert wurde, während die Basaldendriten unbeeinflusst blieben. Das Tau-Protein scheint demzufolge unwesentlich für die Entstehung von axonalen Verbindungen im embryonalen Gehirn zu sein, ist aber beteiligt an der Steuerung des dendritischen Verzweigungsmusters. Ferner wurde beobachtet, dass sowohl die adulte Neurogenese, als auch die Mikrotubuli-Stabilität in den Apikal- und Basaldendriten und die Synapsen von dem Fehlen des Tau-Proteins unbeeinflusst blieben. In primären Zellkulturen aus dem Kleinhirn von Tau-knockout und Tau-wildtyp Mäusen konnten zwischen den zwei Genotypen keine signifikanten Unterschiede in der Länge oder im Verzweigungsmuster der Dendriten und der Axone von Körnerzellen beobachtet werden. Die Untersuchung der Effekte einzelner Tau-Isoformen auf die Morphologie von N2A-Zellen zeigte, dass es Unterschiede sowohl zwischen Tau-defizienten und Tau-positiven Zellen, als auch zwischen Zellen mit den verschiedenen Tau-Isoformen gibt. Das Tau-Protein übt demnach in vivo einen wichtigen Einfluss auf die Morphologie der Nervenzellen und besonders der Dendriten aus, welcher in vitro weiter analysiert wurde.
22
Xu, Qifang. "Statistical Analysis of Biological Interactions from Homologous Proteins". Diss., Temple University Libraries, 2008. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/25686.
Testo completoAbstract (sommario):
Computer and Information SciencePh.D.
Information fusion aims to develop intelligent approaches of integrating information from complementary sources, such that a more comprehensive basis is obtained for data analysis and knowledge discovery. Our Protein Biological Unit (ProtBuD) database is the first database that integrated the biological unit information from the Protein Data Bank (PDB), Protein Quaternary Server (PQS) and Protein Interfaces, Surfaces and Assemblies (PISA) server, and compared the three biological units side-by-side. The statistical analyses show that the inconsistency within these databases and between them is significant. In order to improve the inconsistency, we studied interfaces across different PDB entries in a protein family using an assumption that interfaces shared by different crystal forms are likely to be biologically relevant. A novel computational method is proposed to achieve this goal. First, redundant data were removed by clustering similar crystal structures, and a representative entry was used for each cluster. Then a modified k-d tree algorithm was applied to facilitate the computation of identifying interfaces from crystals. The interface similarity functions were derived from Gaussian distributions fit to the data. Hierarchical clustering was used to cluster interfaces to define the likely biological interfaces by the number of crystal forms in a cluster. Benchmark data sets were used to determine whether the existence or lack of existence of interfaces across multiple crystal forms can be used to predict whether a protein is an oligomer or not. The probability that a common interface is biological is given. An interface shared in two different crystal forms by divergent proteins is very likely to be biologically important. The interface data not only provide new interaction templates for computational modeling, but also provide more accurate data for training sets and testing sets in data-mining research to predict protein-protein interactions. In summary, we developed a framework which is based on databases where different biological unit information is integrated and new interface data are stored. In order for users from the biology community to use the data, a stand-alone software program, a web site with a user-friendly graphical interface, and a web service are provided.
Temple University--Theses
23
Ren, Lei. "Lipopolysaccharide-binding protein and CD14 in human gingiva". Click to view the E-thesis via HKUTO, 2005. http://sunzi.lib.hku.hk/hkuto/record/B31374281.
Testo completo
24
Sarrab, Ramadan. "Role of CD2 associated protein in podocyte differentiation". Thesis, University of Bristol, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.550317.
Testo completoAbstract (sommario):
The glomerular ultrafiltration barrier contains highly terminally differentiated podocytes with major processes and foot processes interlinked by ultrathin slit diaphragms. A number of molecules that are associated with nephrosis and podocyte damage have been described and these discoveries have given insight into the mechanisms that lead to podocyte injury. One of these molecules is CD2-associated protein (CD2AP), which is a crucial protein for slit- diaphragm assembly and function. In spite of the fact that CD2AP knockout causes nephrotic syndrome in mice and the heterozygous +/- mouse is prone to proteinuria, little is known about the relevance of this molecule in human renal pathology. In this thesis I studied the effect of a disease causing CD2AP mutation on the human podocyte phenotype. I identified the dramatic effects of the CD2AP mutation on the morphology of the cells and on the expression of mesenchymal, epithelial and other markers. I found that in contrast to wild type podocytes, CD2AP mutant podocytes acquired the characteristics of dedifferentiated cells. I compared the phenotypic characteristics of the CD2AP mutant podocytes with podocytes carrying mutations in the transcription factor WTI which is essential for normal nephrogenesis. Surprisingly, this study detected a similar de-differentiated phenotype in the CD2AP mutant podocytes to that seen in podocytes carrying a mutation in the WTl gene. In view of these phenotypic similarities I studied whether there is a functional link between CD2AP and WTI which confer these similarities in cellular phenotype. CD2AP is also known to bind to WTIP (WTl interacting protein), a molecule that binds WTI and can shuttle to the nucleus during podocyte injury. In this study, I found that WTIP might provide the functional link between CD2AP and WTl. In addition, in this study I have established the first conditionally immortalized human glomerular mesangial cell line with unique migratory properties, which will be an important adjunct in studies of representative glomerular cells, as well as in eo-culture studies. Overall this thesis demonstrates that there are clear implications for a novel role for CD2AP in the development and progression of glomerular disease. This thesis also discusses a novel conditionally immortalized human mesangial cell line that represents a new tool for the study of human mesangial cell biology in vitro.
25
Ren, Lei, e Ph D. 任蕾. "Lipopolysaccharide-binding protein and CD14 in human gingiva". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2005. http://hub.hku.hk/bib/B31374281.
Testo completo
26
Trautmann, Susanne. "Functions of the Cdc14-Family Phosphatase Clp1p in the Cell Cycle Regulation of Schizosaccharomyces pombe: A Dissertation". eScholarship@UMMS, 2005. https://escholarship.umassmed.edu/gsbs_diss/10.
Testo completoAbstract (sommario):
In order to generate healthy daughter cells, nuclear division and cytokinesis need to be coordinated. Premature division of the cytoplasm in the absence of chromosome segregation or nuclear proliferation without cytokinesis might lead to aneuploidy and cancer.
The cyclin dependent kinases, CDKs, are a main regulator of the cell cycle. Timely increase and decrease in their activity is required for cell cycle progression. To enter mitosis, mitotic CDK activity needs to rise. CDK activity stays elevated until chromosome segregation is completed and exit from mitosis requires decrease in CDK activity.
Observations in several experimental systems suggest that coordination of cytokinesis with the nuclear cycle is regulated through CDK activity. Prolonged high CDK activity, as it occurs when chromosome segregation is delayed, was found to oppose cytokinesis. Prevention of cytokinesis through high CDK activity may therefore provide a mechanism to prevent precocious cell division in the absence of chromosome segregation. To prevent polyploidy when cell division is delayed, progression through the next nuclear cycle should be inhibited until cytokinesis is completed, presumably by the inhibition of CDK activity.
In the fission yeast Schizosaccharomyces pombe, a signaling cascade called Septation Initiation Network (SIN) is required for the coordination of cytokinesis with the nuclear cycle. The SIN is essential for cytokinesis, triggering the execution of cell division through constriction of the actomyosin ring. The activation of the SIN signaling cascade, and thus cytokinesis, is opposed by high CDK activity, preventing precocious cytokinesis.
S. pombe delay entry into the next nuclear division in response to delayed cytokinesis due to defects in the contractile ring until cytokinesis is completed thereby preventing the accumulation of multinucleate, non viable cells. This safeguard against multinucleate cells is termed the cytokinesis checkpoint. The cytokinesis checkpoint keeps CDK activity low, preventing nuclear cycle progression. The SIN is required for the cytokinesis checkpoint and therefore is a key coordinator between nuclear cycle and cytokinesis. How the SIN functions in the cytokinesis checkpoint was not known.
Cdc14-family phosphatases are highly conserved from yeast to humans, but were only characterized in Saccharomyces cerevisiae at the time this thesis was initiated. Cdc14 had been identified as the effector of a signaling cascade homologous to the SIN, called the mitotic exit network (MEN), which is required for exit from mitosis. This thesis describes the identification of the S. pombe Cdc14-like phosphatase Clp1p as a component of the cytokinesis checkpoint. Clp1p opposes CDK activity, and Clp1p and the SIN activate each other in a positive feedback loop. This maintains an active cytokinesis checkpoint and delays mitotic entry. We further found that Clp1p regulates chromosome segregation.
Concluding, this thesis describes discoveries adding to the characterization of the cytokinesis checkpoint and the function of Clp1p. While others found that Cdc14-family phosphatases, including Clp1p, have similar catalytic functions, we show that their biological function may be quite different between organisms, possibly due to different biological challenges.
27
Giavina-Bianchi, Mara Huffenbaecher. "Análise da expressão da proteína NY-ESO-1 no melanoma cutâneo". Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/5/5133/tde-20062016-111434/.
Testo completoAbstract (sommario):
INTRODUÇÃO: o câncer é a doença que mais mata pessoas com idade abaixo de 85 anos e é um problema de saúde pública. Os tumores podem expressar em determinada fase de seu desenvolvimento proteínas anômalas que podem ser alvo de métodos diagnósticos e de intervenções terapêuticas. A expressão de NY-ESO-1 é detectada em 20 a 40% dos melanomas. Há evidências que esta expressão é mais freqüente em tumores de estágios mais avançados e está associada a um pior prognóstico. OBJETIVOS: determinar a frequência de expressão da proteína NY-ESO-1 no melanoma cutâneo e tentar correlacioná-la com o índice de Breslow, aspectos histopatológicos do melanoma, incluindo o infiltrado linfocítico tumoral, e a morbi-mortalidade dos pacientes. MÉTODOS: o presente estudo é longitudinal de coorte retrospectiva e foi realizado de agosto de 2009 a outubro de 2015. Foram selecionados 89 melanomas de 87 pacientes do Ambulatório de Tumores do Departamento de Dermatologia da FMUSP, divididos em 3 grupos, sendo: grupo 1: 34 melanomas com índice de Breslow <= 1,0 mm; grupo 2: 29 melanomas com índice de Breslow entre 1,1 - 4,0 mm e grupo 3: 26 melanomas com índice de Breslow >= 4,0 mm. As lâminas dos exames anátomo-patológicos destes pacientes foram revisadas quanto ao diagnóstico de melanoma, seu índice de Breslow e a presença de infiltrado linfocítico tumoral. A seguir, realizou-se exame de imunohistoquímica para a determinação da presença do antígeno NY-ESO-1 em todos os 89 tumores coletados e em mais 20 nevos (11 displásicos e 9 intradérmicos) escolhidos ao acaso. Através da revisão dos dados do prontuário, foram obtidos os dados clínicos de: idade, sexo, raça, fototipo da pele, local de aparecimento do melanoma, status do linfonodo sentinela quando realizado, desenvolvimento de metástases e sobrevida dos pacientes. Os dados anátomo-patológicos do tumor analisados foram: tipo histológico, presença de ulceração, e tipo de infiltrado linfocítico tumoral. Nos melanomas que apresentavam infiltrado linfocítico tumoral, foram realizados testes imunohistoquímicos para pesquisa de células CD3+, CD8+, FoxP3+ e CD8+FoxP3+ (duplamente positivas). RESULTADOS: O antígeno NY-ESO-1 esteve presente em 19% dos melanomas cutâneos primários e não foi detectado em nenhum dos 20 nevos pesquisados. A expressão do antígeno NY-ESO-1 esteve estatisticamente relacionada a tumores com espessuras maiores. Apresentou também uma associação inversa com o tipo extensivo superficial em relação aos outros tipos histológicos. O infiltrado linfocítico tumoral dos melanomas NY-ESO-1 positivos continha menor número de células CD3+, que se encontravam isoladas ou arranjadas em pequenos grupos de até 5 células, o que contrastava significantemente com os tumores NY-ESO-1 negativos, com maior densidade de células CD3+, dispostas em grandes grupos, com 6 ou mais células. A expressão da proteína NY-ESO-1 não esteve associada à idade, ao sexo, ao fototipo, ao sítio primário do tumor, à presença de ulceração, ao status do linfonodo sentinela, ao desenvolvimento de metástases ou à sobrevida. CONCLUSÕES: Há expressão de NY-ESO-1 em uma porcentagem considerável dos melanomas, principalmente nos mais espessos. O menor número de células CD3+ no infiltrado linfocítico tumoral, acrescido ao fato destas células estarem isoladas ou em pequenos grupos, sugere que embora imunogênico, a expressão do antígeno NY-ESO-1 não resulta num estímulo eficaz do sistema imune no combate ao tumor. O desenvolvimento de uma vacina para estes pacientes poderá, no futuro, aumentar as possibilidades terapêuticas do melanomaINTRODUCTION: cancer is the disease that leads to the greatest number of deaths in people over 85 years old and it has become a major public health problem. Tumors may express aberrantly proteins during certain phases of their development, which can be target for diagnostic or treatment purposes. NY-ESO-1 is detected in 20 to 40% of melanomas. There is evidence that it is more frequent in advanced stages and that is associated with a worse prognosis. OBJECTIVES: to determine the frequency of NY-ESO-1 protein expression in cutaneous melanoma and to try to correlate it to Breslow index, melanoma histopathological aspects, including the tumor infiltrating lymphocytes, and patients morbi-mortality. METHODS: the present study is longitudinal of retrospective cohort. The research was carried on from August 2009 to October 2015. Eighty nine melanomas were selected from 87 patients in Oncology Outpatient Clinic, Dermatology Division, University of São Paulo and divided in 3 groups, such as: group 1: 34 melanomas with Breslow index <= 1,0 mm; group 2: 29 melanomas with Breslow index between 1,1 - 4,0 mm e group 3: 26 melanomas with Breslow index >= 4,0 mm. All specimens were reviewed for diagnostic, Breslow index and tumor infiltrating lymphocytes. After that, immunohistoquimical test for the presence of NY-ESO-1 antigen was performed in all 89 melanomas collected and in 20 nevi (11 dysplastic nevi and 9 dermal nevi) that were randomly chosen. By reviewing clinical charts, the following data was obtained: age, sex, skin phototype, site of the tumor, lymph node sentinel status, development of metastases and survival of the patients. The histological data analyzed was: histological melanoma type, presence of ulceration, grade of tumor infiltrating lymphocytes. In those melanomas that had tumor infiltrating lymphocytes, we performed immunohistoquimical tests for the presence of CD3+, CD8+, FoxP3+ and CD8+FoxP3+ (double positive) cells. RESULTS: antigen NY-ESO-1 was present in 19% of primary cutaneous melanomas and none of the 20 nevi. The expression of antigen NY-ESO-1 was statistically related to thicker melanomas. It presented also an inverse association with superficial spreading melanoma type compared to other subtypes. Tumor infiltrating lymphocytes of NY-ESO-1 positive melanomas had fewer CD3+ cells, that were isolated or arranged in small groups up to 5 cells, which was significantly different from tumors NY-ESO-1 negatives, with higher density of CD3+ cells, displayed in large groups of 6 or more cells. The expression of NY-ESO-1 protein was not associated to age, sex, phototype, site, ulceration, lymph node sentinel status, development of metastases and survival. CONCLUSIONS: A considerable amount of melanomas express NY-ESO-1, mainly thicker tumors. The fewer number of CD3+ cells in the tumor infiltrating lymphocytes, added to the fact of those cells being isolated or in small groups suggest that, although immunogenic, the expression of NY-ESO-1 antigen does not result in a efficient stimulus of the immune system to fight the tumor. The development of a vaccine to those patients may, in the future, enhance the roll of therapeutic possibilities for melanoma
28
Head, Jared G. "Structure and function of the #beta#-sheet proteins, titin and CD2". Thesis, University of Bristol, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.265387.
Testo completo
29
Garcia, Diaz Yoel R. "Synthesis of Novel Glycolipid Agonists of the Protein CD1d". Thesis, University of Birmingham, 2010. http://etheses.bham.ac.uk//id/eprint/544/.
Testo completoAbstract (sommario):
Invariant NKT (iNKT) cells are a subset of T lymphocytes that express an invariant \(\alpha\) \(\beta\) T cell receptor (TCR) as well as an NK1.1 marker. They play an important role in autoimmune diseases, such as type I diabetes and lupus. In contrast to conventional CD4+ and CD8+ T lymphocytes that recognise foreign peptides bound to the major histocompatibility complex (MHC) class I or MHC class II, iNKT cells recognise a range of foreign lipids and glycolipids bound to CD1d proteins. \(\alpha\) \(\beta\)-Galactosyl Ceramide (\(\alpha\) \(\beta\)GalCer), originally isolated from a marine sponge, is a powerful agonist of CD1d capable of triggering an immune response that results in the proliferation of a range of regulatory cytokines, including IFN-\(_y\) (Th1), as well as IL-4 (Th2). This mixed cytokine response (i.e. Th1 and Th2), combined with the “unresponsive state” of iNKT cells after activation with \(\alpha\)GalCer, limits the therapeutic potential of this agonist. To address some of these issues, we have also synthesised an \(\alpha\)GalCer analogue, namely Threitol Ceramide (ThrCer), that exhibits attenuated activity relative to \(\alpha\)GalCer. ThrCer is a truncated analogue of \(\alpha\)GalCer that conserves the stereochemistry of the hydroxyl functions present in \(\alpha\)GalCer and that exhibit a much stronger ether bond under acidic hydrolysis linking the sugar moiety with ceramide than the glycosidic bond in \(\alpha\)GalCer. We have labelled ThrCer with a biotin residue and with a 14C radiolabel, and our collaborators have used these derivatives to show that ThrCer behaves similarly to \(\alpha\)GalCer in endogenous lipid trafficking and its tissue distribution in vivo. We have also made advances towards the stereoselective synthesis of recently discovered natural agonists of iNKT cells from pathogenic origin, namely \(\alpha\)-galactosyl diacylglycerol (\(\alpha\)GalDAG).
30
Thorpe, Helena M. "Site-specific recombination in Streptomyces temperate phage #pi#C31". Thesis, University of Nottingham, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.285769.
Testo completo
31
Triantafilou, Kathy. "Homotypic and heterotypic interactions of HLA-DR, CD74 and CD14 molecules : biochemical and fluorescence imaging analysis". Thesis, University of Essex, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.285855.
Testo completo
32
Sinotte, Christopher Matthew. "Construction, expression, and purification of soluble CD16 in bacteria". Thesis, Available online, Georgia Institute of Technology, 2006, 2006. http://etd.gatech.edu/theses/available/etd-05142006-222347/.
Testo completoAbstract (sommario):
Thesis (M.S.)--Bioengineering, Georgia Institute of Technology, 2007.Zhu, Cheng, Committee Chair ; Selvaraj, Periasamy, Committee Member ; Orville, Allen, Committee Member ; Butera, Robert, Committee Member.
33
Deftos, Michael Laing. "Notch signaling in T cell development /". Thesis, Connect to this title online; UW restricted, 2001. http://hdl.handle.net/1773/8364.
Testo completo
34
Van, Dyken Steven John. "CD8⁺ T lymphocyte apoptosis is regulated by protein O-glycosylation". Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2006. http://wwwlib.umi.com/cr/ucsd/fullcit?p3226770.
Testo completoAbstract (sommario):
Thesis (Ph. D.)--University of California, San Diego, 2006.Title from first page of PDF file (viewed October 11, 2006). Available via ProQuest Digital Dissertations. Vita. Includes bibliographical references (p. 83-92).
35
Wainberg, Zev Aryeh. "Stress protein modulation in HIV-1 infected CD4-expressing cells". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq29807.pdf.
Testo completo
36
Wainberg, Zev. "Stress protein modulation in HIV-1 infected CD4-expressing cells". Thesis, McGill University, 1996. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=27432.
Testo completoAbstract (sommario):
Heat shock or stress proteins (HSPs), are a large family of phylogenetically conserved molecules that can be constitutively expressed at high levels under normal physiological conditions. These proteins are selectively synthesized following metabolic and environmental insult. This study was designed to determine whether cognate and/or inducible HSP production are altered in CD4-expressing lymphocytic cell lines concomitant with acute and chronic human immunodeficiency virus type 1 (HIV-l) infection. Our findings indicate that in CEM.NKR cells, HSP27 production is selectively altered at early stages of acute HIV-infection (6-24h post-infection), subsequent to virus internalization but prior to synthesis of viral progeny. Levels of HSP27 induction were viral dose-dependent and were not accompanied by any alterations in cellular proliferation. In contrast, acute HIV-infection was not associated with significant quantitative changes in the constitutive expression of HSP60, HSP70, or HSP90. Nevertheless, a transient, marked induction of select HSP70 subspecies was evident at early stages of infection, finally disappearing by 48-72h. Acute-infection of Jurkat cells resulted in similar patterns of de novo induction of HSP27 and HSP70 isoforms. Uninfected and chronically-infected CEM extracts showed little detectable constitutive HSP27. However, synthesis of select HSP27 and HSP70 homologues in chronically-infected cells was observed following exposure to a mild heat shock and doses of TNF$ alpha$. Similar HSP70 homologues arose in chronically-infected cells treated with heat-shock and TNF$ alpha$. These findings indicate that HSP pathways are uniquely modulated in CD4+ cells as a consequence of acute and chronic HIV-1 infection.
37
Garland, Russell John. "The ex-vivo expansion of human CD8'+ cytotoxic T lymphocytes to herpes simplex virus". Thesis, University of Bristol, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.324367.
Testo completo
38
Bayard, Florence. "Etude des réponses cellulaires T spécifiques de la protéine hepatitis B spliced-generated protein (HBSP) du virus de l'hépatite B (VHB) et caractérisation de nouveaux épitopes du VHB". Paris 7, 2009. http://www.theses.fr/2009PA077089.
Testo completoAbstract (sommario):
L'infection chronique par le VHB reste un problème de santé publique majeur, touchant 400 millions de personnes dans le monde. La présence d'anticorps spécifiques de la protéine « Hepatitis-B spliced-generated protein » (HBSP) chez 30% des porteurs chroniques du VHB a été associée à la sévérité de la fibrose hépatique, suggérant un rôle possible de la réponse immunitaire spécifique de HBSP dans la fibrogenèse. Nous avons identifié plusieurs épitopes dérivés de HBSP capables d'activer une réponse T CD8 chez des souris transgéniques pour les molécules HLA-A2 et HLA-B7 immunisées à l'aide de vecteurs codant pour HBSP. Nous avons ensuite montré que ces épitopes étaient également apprêtés et reconnus chez les porteurs chroniques du VHB. Nous recherchons actuellement les liens éventuels entre la réponse T spécifique de HBSP et la sévérité de la fibrose hépatique et/ou la réplication virale. Dans une deuxième partie, nous avons étudié la fonction auxiliaire de deux épitopes du VHB présentés par HLA-DR1. Le transport et l'apprêtement efficaces de ces épitopes grâce à la chaîne invariante de nos vecteurs a permis d'aider au développement d'une réponse T CD8 polyfonctionneîle spécifique de l'enveloppe virale. Les épitopes décrits ici sont de puissants activateurs des réponses T CD8 et/ou CD4. Leur utilisation au sein de vaccins polyépitopiques pourrait améliorer les réponses cellulaires activées par ces vaccins. De plus, l'identification de réponses T cellulaires spécifiques de HBSP corrélant avec l'évolution de la maladie hépatique, pourrait avoir un impact clinique notamment dans le développement d'un outil de diagnostique de la progression de la fibroseChronic HBV infection remains a Worldwide health problem, as 400 million people are chronic HBV-carriers. Detection of antibodies against the recently identified "Hepatitis B spliced-generated protein" (HBSP) in 30% of HBV chronic carriers has been related to the severity of fîbrosis. This suggests a possible role of HBSP-specific T cell immune response in fibrogenesis. We first studied the HBSP-specific T cell immune response in HLA-A2 and HLArB7:transgenic mice immunized with HBSP-encoding vectors. Several epitopes activated CD8 T cell responses in immunized mice. Then, we showed that these epitopes were efficiently processed and recognized by T cells from HBV chronic carriers. We are currently investigating the possible link of these immune responses with severe fibrosis and/or viral replication by exploring the HBSP-specific T cell immune response/in patients with different clinical settings. In a second part, we studied the helper potential of two HLA-DRl-restricted epitopes derived from HBV. Invariant chain of our vectors allowed efficient transport and processing of HLA-DR1 epitopes. These epitopes efficiently helped to develop a polyfunctional CD8 T cell response specific for HBV envelope. Ail the epitopes described here are strong activators of CD8 and/or CD4 T cell immune responses. They could be included in polyepitopic DNA vaccine constructs to increase the cellular responses primed by such vaccines. Moreover, if HBSP-specific T cell responses are correlated with outcome of disease, it could have a great clinical impact, for example to develop a diagnostic tool for fibrosis progression
39
Steinhoff, Ulrich Johannes. "Von Toleranz zur Autoimmunität". Doctoral thesis, Humboldt-Universität zu Berlin, Medizinische Fakultät - Universitätsklinikum Charité, 2002. http://dx.doi.org/10.18452/13835.
Testo completoAbstract (sommario):
Immunologische Toleranz ist eine elementare Eigenschaft des Immunsystems, die primär durch die klonale Deletion autoreaktiver T-Zellen im Thymus gewährleistet wird. Neben diesem als zentrale Toleranz bezeichneten Mechanismus, verfügt ein Organismus gleichzeitig über periphere Toleranzmechanismen wie Ignoranz, Anergie und regulatorische T-Zellen. Trotz dieser Kontrollmechanismen können in bestimmten Situationen autoreaktive CD4+ und CD8+ T-Zellen aktiviert werden und meistens zu örtlich und zeitlich begrenzten Autoimmunreaktionen führen. Ursache hierfür kann die hormonelle Regulation oder das gewebespezifische Vorkommen eines Selbsttantigens sein. Am Beispiel von HSP60-kreuzreaktiven CD8+ T-Zellen konnte gezeigt werden, dass der Transfer dieser T-Zellen in Tiere zu einer Entzündung des Dünndarms aber nicht des Dickdarms führt, obwohl das Selbstantigen im letzteren wesentlich stärker exprimiert wird. Die Gewebespezifität der Autoimmunpathologie konnte durch die in den Organen unterschiedliche, proteasomale Antigenprozessierung, erklärt werden. Proteinbiochemische und immunologische Analysen ergaben, dass sich die 20S Proteasomen verschiedener Organe strukturell und funktionell deutlich unterscheiden und somit jedes Gewebe ein individuelles Repertoire von MHC-Klasse I restringierten Peptiden präsentiert. Damit wurde ein weiterer Mechanismus entdeckt, durch den Reaktivität von protektiven und pathologischen CD8+ T-Zellen kontrolliert wird.Immunological tolerance which is primarily mediated by the clonal deletion of autoreactive T cells in the thymus is a key feature of the immune system. Besides this central tolerance, several mechanisms act also in the periphery including ignorance, anergy and regulatory T cells. Despite all these checkpoints, autoreactive CD4+ and CD8+ T cells may still be activated causing local and time restricted autoimmune-reactions. This may refer primarily to self-antigens which are hormonally regulated or tissue-specifically expressed. Adoptive transfer of crossreactive, hsp60-specific CD8+ T cells into mice induced an local inflammation of the small intestine but not the colon despite elevated expression of hsp60 in the latter organ. The pathology could be explained by the finding that the proteasomal antigen processing varies between different organs. Biochemical and immunological analyses revealed that 20S proteasomes of different organs vary in their structural and functional properties indicating that every tissue displays an individual and distinct repertoire of MHC class I peptides. This represents a new mechanism by which the activity of protective and pathological CD8+ T cell responses may be controlled.
40
Fontenot, Jason David. "The role of Foxp3 in CD4⁺ T cell development and function /". Thesis, Connect to this title online; UW restricted, 2003. http://hdl.handle.net/1773/8336.
Testo completo
41
Mayack, Shane Renee. "The role of Janus Kinase 3 in CD4+ T Cell Homeostasis and Function: A Dissertation". eScholarship@UMMS, 2004. https://escholarship.umassmed.edu/gsbs_diss/94.
Testo completoAbstract (sommario):
This dissertation addresses the role for Janus Kinase 3 (Jak3) in CD4+ T cell homeostasis and function. Jak3 is a protein tyrosine kinase whose activity is essential for signals mediated by the γc dependent cytokines IL-2, -4, -7, -9, -15, and -21. Previous data have demonstrated that peripheral CD4+ T cells from Jak3-deficient mice have a memory phenotype and are functionally impaired in both proliferative and IL-2 responses in vitro. Interestingly, Jak3/γc activity has been previously shown to play a role in the prevention of T cell anergy.
These studies were initiated to more precisely define the role for Jak3/γc cytokines in the prevention of T cell anergy and the maintenance of functional CD4+ T cell responses. We began to address this question by assessing global gene expression changes between wild type and Jak3-/- CD4+ T cells. These data indicate that Jak3-/- CD4+ T cells have an increase in gene expression levels of inhibitory surface receptors as well as immunosuppressive cytokines.
Further analyses confirmed that Jak3-deficient T cells express high levels of PD-1, secrete a Trl-type cytokine profile following direct ex vivo activation, and suppress the proliferation of wild type T cells in vitro. These characteristics indicate that CD4+ Jak3-/- T cells share properties with regulatory T cell subsets that have an important role in peripheral tolerance and the prevention of autoimmunity.
We next addressed whether these regulatory characteristics were T cell intrinsic or rather the result of expanding in a Jak3-deficient microenvironment characterized by a number of immune abnormalities and a disrupted splenic architecture. Jak3-/- CD4+ T cells proliferate in vivoin a lymphopenic environment and selectively acquire regulatory T cell characteristics in the absence of any additional activation signals. While the precise mechanism by which Jak3-deficient T cells acquire these characteristics remains unclear, our data indicate that one important component is a T cell-intrinsic requirement for Jak3 signaling.
These findings indicate several interesting aspects of T cell biology. First, these studies, demonstrate that the homeostatic proliferation of CD4+ T cells is not dependent on signaling via γc-dependent cytokine receptors. And, second, that the weak activation signals normally associated with homeostatic expansion are sufficient to drive Jak3-/- T cells into a non-conventional differentiation program. Previous data indicate that, for wild type T cells, signaling through both the TCR as well as γc-dependent cytokine receptors promote the homeostatic proliferation of T cells in lymphopenic hosts. Since Jak3-/- T cells are unable to receive these cytokine signals, their proliferation is likely to be wholly dependent on TCR signaling. As a consequence of this TCR signaling, Jak3-/- T cells proliferate, but in addition, are induced to up regulate PD-1 and to selectively activate the IL-10 locus while shutting off the production of IL-2. Since this fate does not occur for wild type T cells in a comparable environment, it is likely that the unique differentiation pathway taken by Jak3-/- T cells reflects the effects of TCR signaling in the absence of γc-dependent cytokine signaling.
Interestingly, wild type T cells undergoing homeostatic expansion in lymphopenic hosts show many common patterns of gene expression to freshly-purified unmanipulated Jak3-/- T cells. For instance, micro array analysis of gene expression in wild type CD4+ T cells after lymphopenia induced homeostatic expansion show a similar pattern of upregulation in surface markers (PD-1 and LAG-3), and cytokine signaling molecules (IL-10 and IFN-γ cytokine, receptors, and inducible gene targets) to that of Jak3-/- CD4+ T cells immediately ex vivo. These data suggest that the process of homeostatic proliferation normally induces immune attenuation and peripheral tolerance mechanisms, but that full differentiation into a regulatory T cell phenotype is prevented by γc-dependent cytokine signals.
Taken together these data suggest that Jak3 plays an important role in tempering typical immune attenuation mechanisms employed to maintain T cell homeostasis and peripheral tolerance.
42
Miller, Andrew Todd. "The Function of the Tyrosine Kinase, Itk, in CD4+ T Cell Differentiation and Death: a Dissertation". eScholarship@UMMS, 2003. https://escholarship.umassmed.edu/gsbs_diss/58.
Testo completoAbstract (sommario):
The Tec family tyrosine kinase, Itk, plays an important role in signal transduction following T cell receptor engagement. Several prior studies have established the importance of Itk in immune system processes, such as T cell development and T cell activation. Additional biochemical studies have found that Itk specifically functions within a multi-molecular signalosome complex, which ultimately functions to provide a platform by which Itk can phosphorylate and activate PLC-γ1, a crucial step in T cell activation. To further study how Itk regulates distinct immune outcomes via T cell effector processes within the peripheral immune system, and to further understand how Itk functions in T cells in response to a physiological ligand-receptor interaction, I crossed Itk-deficient mice to mice transgenic for a TCR specific for a moth cytochrome C peptide. My studies have established a unique role for Itk in several important aspects of T cell function. Following T cell activation, I identified an imperative role for Itk in activation-induced cell death via FasL, a mechanism of immune homeostasis. Furthermore, I found Itk plays a unique role in the process of T cell differentiation, where Itk positively regulates the induction of cytokine genes, such as IL-4, while negatively regulating the induction of T-bet, a transcription factor important for Th1 differentiation. Lastly, following T cell differentiation, I found that Itk mRNA and protein are up-regulated during Th2 differentiation, while Rlk, a related Tec kinase, disappears rapidly from Th2 cells, indicating a critical role for Itk in Th2 cell function. Collectively, my thesis work has more clearly defined an important function for Itk not only in TCR signaling, but also in immune processes such as T cell differentiation and activation-induced cell death that are required for proper immune function.
43
VALERE, THOMAS. "Transgenese somatique chez la souris par la proteine humaine cd4 soluble". Paris 7, 1995. http://www.theses.fr/1995PA077313.
Testo completoAbstract (sommario):
La therapie genique emploie couramment des transgenes a produit stable in vivo. Les transgenes non selectionnables, ou dont le produit a une pharmacocinetique defavorable ne font pas l'objet de tentatives a long terme. Nous avons mene une telle tentative, avec comme marqueur la molecule scd4, de demi-vie de quelques heures. Des fibroblastes primaires murins explantes, puis infectes in vitro par un vecteur retroviral de scd4, ont ete emballes dans un gel de collagene contenant des fibres synthetiques. La structure obtenue, un organoide secreteur de scd4, a ete reimplantee dans la cavite peritoneale. Les serums des souris implantees ont montre durant deux mois une concentration de scd4 significative et reproductible, avec un pic a un mois post-implantation. C'est a notre connaissance la premiere transgenese somatique exprimant durablement un produit a courte demi-vie. Independamment, nous avons obtenu des resultats interessants sur la propagation d'un vecteur retroviral par un virus replicatif. Le vecteur retroviral de scd4 utilise est de type auto-inactivant. Ces vecteurs s'integrent sous une forme privee des deux amplificateurs viraux, diminuant theoriquement les risques d'empaquetage et de propagation de l'arn du vecteur, par d'eventuels retrovirus surinfectants (dans une optique de securite). Nos test in vitro ont montre que le vecteur etait empaquete, propage, et fonctionnel (pour l'expression du marqueur scd4) par toute une palette de virus assistants replicatifs, contrairement a ce qui est generalement attendu. La securite pretee aux vecteurs retroviraux auto-inactivants est donc mise en cause in vitro dans notre systeme experimental. Cependant, les experiences in vivo, qui consistaient a employer le virus assistant le plus efficace (f-mulv) pour propager le vecteur chez des souris, n'ont pas abouti a la secretion de scd4 dans les tissus cibles. Ce resultat confirme la securite des vecteurs auto-inactivants in vivo, malgre les fuites observees in vitro.
44
Small, Lawrence Edward. "PAR Proteins Regulate CDC-42-Dependent Myosin Dynamics During C. elegans Zygote Polarization". The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1461086954.
Testo completo
45
PORCU, ALESSANDRA. "Characterization of Rimonabant effects on G protein activity". Doctoral thesis, Università degli Studi di Cagliari, 2016. http://hdl.handle.net/11584/266891.
Testo completoAbstract (sommario):
G protein-coupled receptor (GPCRs) is the largest class of cell-surface receptors, and represents today the target of 40% of the drugs in the pharmaceutical market. In the absence of agonists, many GPCRs have found to exhibit spontaneous activity, which can be blocked by ligands that are referred to as inverse agonists (Milligan, 2003). Cannabinoid CB1 receptor is one of the most abundant GPCR in the central nervous system, and is coupled to Gi/o proteins to inhibit adenylyl cyclase, activate mitogen-activated protein kinase (MAPK), inhibit voltage gated Ca2+ channels and activate inwardly rectifying K+ channels (Howlett et al., 2000; Pertwee, 2010). The first selective and potent CB1 antagonist Rimonabant (also known SR141716A, SR) at high micromolar concentrations behaves as an inverse agonist, i.e. decreases [35S]GTPγS binding in rodent and human cerebral cortex and in Chinese hamster ovary (CHO) cells transfected with CB1 receptors (Rinaldi-Carmona, 1994). However, in vitro and in vivo studies performed using CB1 knockout (KO) and CHO cells not expressing CB1 receptors suggest that inverse agonist activity of SR is CB1 receptor independent (Pertwee, 2005). Several hypotheses have been postulated to explain the inverse agonism of SR, including its action on different receptors (i.e GPCR mainly coupled to Gi/o proteins) and/or its negative modulation of the constitutive activity of CB1 receptors.
Alternatively, SR might explicate these “inverse agonist effect” in a manner receptor-independent acting directly on G protein level. The present study aimed to determine whether the CB1 receptor-independent effects of SR are mediated via GPCRs, in particular GABAB and dopamine D2 receptors, that share the same Gαi/o signaling pathways, or if SR acts directly on G protein subunits. For this purpose we investigated the molecular mechanisms of SR on G protein activity in native and recombinant systems by using different experimental approaches (i.e., GTPγS binding, bioluminescence Resonance Energy Transfer (BRET), electrophysiological recordings). In particular, we first evaluated the effects of SR on basal and agonist-stimulated [35S]GTPγS binding
in systems containing CB1, GABAB and D2 receptor populations (i.e., rat membrane homogenates and CHO stable transfected with GABAB or D2 receptors), and in systems lacking CB1 and GABAB receptors (i.e., CB1- and GABAB-KO mice).
Then, using BRET approach we monitored dissociation between Gαo and Gβγ subunits and their conformational rearrangements before and after GABAB receptors activation. In addition, using the same assay we studied the molecular interaction between D2 receptor and Gαi1 protein subunits (Gαi1-60, Gαi1-91 and Gαi1-121). Next, we evaluated the effects of SR on adenylate cyclase activity, using BRET with the CAMYEL sensor, a recent technique developed to detect the level of cAMP in living cells. Specifically, the inhibitory effect of SR on Gi and Gs protein pathways measuring BRET signal in cells transfected with CAMYEL and GABAB, D2 or D1 receptors was investigated. Finally, whole cell voltage clamp recordings from midbrain dopamine neurons in acute rat brain slices ex vivo were performed to evaluate the effects of SR on baclofen and quinpirole-induced outward K+ current both in wild-type (WT) and CB1-KO mice. In addition, in order to demonstrate that SR induced the inhibition of GIRK channel activity by acting directly on G protein, we use a GPCR-free experimental setup (i.e. whole cell patch clamp experiments were performed in CHO cells transfected with GIRK1/2). The main finding of this study is that SR, at micromolar concentrations, prevented GPCR-G protein signaling through a direct interaction with the G proteins mainly with the subunits αi/o.
46
Santos, Andressa Cristina Antunes. "Efeito da desnutrição proteica sobre aspectos da mobilização, migração e sinalização celular. Papel da glutamina na modulação desses processos". Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/9/9136/tde-28042016-104055/.
Testo completoAbstract (sommario):
A desnutrição é uma condição nutricional que pode afetar muitos aspectos da resposta imunológica, como alterações na migração celular, na fagocitose, na resposta bactericida, mudanças na produção de radicais livres e espécies de nitrogênio e na produção de citocinas pró-inflamatórias. Logo, indivíduos desnutridos apresentam maior susceptibilidade a infecções. Visto que a glutamina é um aminoácido de extrema importância para a funcionalidade de diversas células do sistema imune e que as mesmas apresentam aumento da utilização desse aminoácido durante processos infecciosos, investigou-se, neste trabalho, quais os efeitos da glutamina sobre alguns aspectos da mobilização, migração e sinalização celular em um modelo experimental de desnutrição proteica. Para tanto, utilizou-se camundongos da linhagem BALB/c machos, os quais foram divididos em dois grupos, Controle e Desnutrido, que passaram a receber dietas isocalóricas contendo 12% (normoproteica) e 2% de caseína (hipoproteica), respectivamente, durante 5 semanas. Para as avaliações in vivo, animais de ambos os grupos receberam por via endovenosa 100µL de solução contendo 1,25µg de LPS e após 1 hora 0,75mg/Kg de L-glutamina (GLUT). Após o período de desnutrição ou de indução ao processo inflamatório, os animais foram eutanasiados e as amostras biológicas coletas. Foram avaliados nos animais estimulados in vivo hemograma, mielograma, as citocinas IL-10 e TNF-α circulantes e a expressão de CD11b/CD18 nos granulócitos do sangue periférico. Foi avaliado, in vitro, a capacidade migratória, a expressão de CD11b/CD18 de polimorfonucleados da medula óssea e do sangue periférico, bem como a síntese de citocinas IL-1α, IL-6, IL-10, IL-12 e TNF-α e a expressão de NF-κB e IκBα em células cultivadas em meio com 0; 0,6; 2 e 10 mM de GLUT. Os animais desnutridos apresentaram anemia, leucopenia, hipoplasia medular e diminuição na concentração sérica de proteínas, albumina e pré-albumina. A GLUT, in vitro, apresentou capacidade de reduzir a produção de IL-1α e IL-6, bem como a ativação da via do NF-κB. No modelo in vivo a GLUT, em animais estimulados com LPS, alterou a cinética de migração neutrofílica e reduziu a expressão de CD18, bem como diminuiu os níveis de TNFα circulantes.Malnutrition is a nutritional condition that can affect many aspects of immune responses, affecting cell migration, phagocytosis, bactericidal response and changing free radicals production as nitrogen species and production of proinflammatory cytokines. Therefore, malnourished individuals are more susceptible to infections. Once glutamine is an amino acid of extreme importance to the functionality of various immune cells and those cells exhibit increased use of this amino acid during infectious processes. In this work was investigated, the effects of glutamine in some aspects of mobilization, cell migration and signaling in an experimental model of protein malnutrition. For this purpose, we used BALB/c mice, which received isocaloric diets, normoproteic or hypoproteic, containing respectively, 12% (Control group) and 2% (Malnourished group) of protein for a period of 5 weeks. The animals in both groups, for in vivo evaluations, received intravenous 100 µl of a solution containing 1.25µg of LPS and after 1 hour 0.75mg/kg of L-glutamine (GLUT). After the malnutrition period or the inflammatory process induction, the animals were euthanized and biological samples were collected. Were evaluated blood count, bone marrow, the cytokines IL-10 and TNF-α circulating and expression of CD11b/CD18 in granulocytes from peripheral blood of animals stimulated in vivo. In vitro were evaluated the migratory capacity, the expression of CD11b/CD18 polymorphonuclear bone marrow and peripheral blood, as well as the cytokines synthesis IL-1α, IL-6, IL-10, IL-12 and TNF-α and the expression of NF-κB and IκBα in cultured cells in media with 0; 0.6; 2 and 10 mM GLUT. Malnourished animals presented anemia, leukopenia, marrow hypoplasia and lower serum proteins, albumin and prealbumin. The GLUT in vitro has the capacity to reduce IL-1α and IL-6 as well as the activation of the NF-κB. In in vivo model, the GLUT altered neutrophil migration kinetics and reduced the expression of CD18, as well as decreased levels of circulating TNF-α in animals stimulated with LPS.
47
Beaumier, Coreen Michele. "Cross-Reactive Memory CD4+ and CD8+ T Cells Alter the Immune Response to Heterologous Secondary Dengue Virus Infections in Mice: A Dissertation". eScholarship@UMMS, 2008. https://escholarship.umassmed.edu/gsbs_diss/350.
Testo completoAbstract (sommario):
Dengue virus (DENV) infects 50-100 million people worldwide every year and is the causative agent of dengue fever (DF) and the more severe dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). There are four genetically and immunologically distinct DENV serotypes (DENV-1, DENV-2, DENV-3, and DENV-4). Evidence suggests that an increased risk for DHF/DSS during secondary infection with a heterologous DENV serotype is due to an immunopathological response mediated by serotype-cross-reactive memory T cells from the primary infection. Furthermore, epidemiological studies have shown that the sequence of infection with different DENV serotypes affects disease severity. Though much has been learned from human studies, there exist uncontrollable variables that are intrinsic in this system such as genetic factors and unknown infection histories. These factors can skew experimental results, making interpretations difficult. Therefore, a murine model to study the immunologic aspects of sequential dengue infections would be an asset to the field of dengue research.
To examine the effect of sequential infection with different DENV serotypes on the CD8+ T cell response, we immunized Balb/c mice with a primary DENV infection on day 0 and subsequently challenged with a heterologous secondary DENV infection on day 28. We tested all possible sequences of infection with the four serotypes. We analyzed the T cell response to two previously defined epitopes on the DENV E (Ld-restricted) and NS3 (Kd-restricted) proteins. Using ELISPOT and intracellular cytokine staining, we measured the frequency of T cells secreting IFNγ and TNFα in response to stimulation with these epitopes during three phases: acute primary, acute secondary, and the memory phase after primary infection. We found that the T cell response in heterologous secondary infections was higher in magnitude than the response in acute primary infection or during the memory phase. We also found that the hierarchy of epitope specific responses, as measured by IFNγ secretion, was influenced by the sequence of infections. The adoptive transfer of immune serum or immune splenocytes suggested that memory T cells from the primary infection responded to antigens from the secondary infection. In vitroexperiments with T cell lines generated from mice with primary and secondary DENV infections suggested the preferential expansion of crossreactive memory T cells.
In testing all of the different possible sequences of infection, we observed that two different sequences of infection (e.g., DENV-2 followed by DENV-1 versus DENV-2 followed by DENV-3) resulted in differential CD8+ T cell responses to the NS3 peptide even though both secondary infection serotypes contain the identical peptide sequence. To investigate this phenomenon, we examined the role of CD4+ T cell help on the memory CD8+ T cell response. We found that CD4+ T cell cytokine responses differ depending on the sequence of infection. In addition, it was also shown that crossreactivities of the CD4+ T cell response are also sequence-dependent. Moreover, denguespecific memory CD4+ T cells can augment the secondary CD8+ T cell response. Taken together, we demonstrated that this serotype sequence-dependent phenomenon is the result of differential help provided by cross-reactive memory CD4+T cells.
The findings in this novel mouse model support the hypothesis that both CD4+ and CD8+ serotype-cross-reactive memory T cells from a primary dengue virus infection alter the immune response during a heterologous secondary dengue virus infection. These data further elucidate potential mechanisms whereby the specific sequence of infection with different dengue virus serotypes influences disease outcomes in humans.
48
Raman, Malavika. "Identification of intracellular signaling pathways regulated by the TAO family of mammalian STE20p kinases". Access to abstract only; dissertation is embargoed until after 5/15/2007, 2006. http://www4.utsouthwestern.edu/library/ETD/etdDetails.cfm?etdID=163.
Testo completo
49
Murray, Alison Jane. "Mutational studies on the dimerization and structure of the metastable protein CD2 domain 1". Thesis, University of Bristol, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.262836.
Testo completo
50
Sleater, Michelle Leigh. "Cellular and molecular effector mechanisms of islet allograft rejection /". Connect to full text via ProQuest. IP filtered, 2006.
Cerca il testo completoAbstract (sommario):
Thesis (Ph.D. in Immunology) -- University of Colorado at Denver and Health Sciences Center, 2006.Typescript. Includes bibliographical references (leaves 151-168). Free to UCDHSC affiliates. Online version available via ProQuest Digital Dissertations;