Dissertations / Theses on the topic 'Hit identification'
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Nordqvist, Anneli. "Hit Identification and Hit Expansion in Antituberculosis Drug Discovery : Design and Synthesis of Glutamine Synthetase and 1-Deoxy-D-Xylulose-5-Phosphate Reductoisomerase Inhibitors." Doctoral thesis, Uppsala universitet, Avdelningen för organisk farmaceutisk kemi, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-155428.
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Since the discovery of Mycobacterium tuberculosis (Mtb) as the bacterial agent causing tuberculosis, the permanent eradication of this disease has proven challenging. Although a number of drugs exist for the treatment of tuberculosis, 1.7 million people still die every year from this infection. The current treatment regimen involves lengthy combination therapy with four different drugs in an effort to combat the development of resistance. However, multidrug-resistant and extensively drug-resistant strains are emerging in all parts of the world. Therefore, new drugs effective in the treatment of tuberculosis are much-needed. The work presented in this thesis was focused on the early stages of drug discovery by applying different hit identification and hit expansion strategies in the exploration of two new potential drug targets, glutamine synthetase (GS) and 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR). A literature survey was first carried out to identify new Mtb GS inhibitors from compounds known to inhibit GS in other species. Three compounds, structurally unrelated to the typical amino acid derivatives of previously known GS inhibitors, were then discovered by virtual screening and found to be Mtb GS inhibitors, exhibiting activities in the millimolar range. Imidazo[1,2-a]pyridine analogues were also investigated as Mtb GS inhibitors. The chemical functionality, size requirements and position of the substituents in the imidazo[1,2-a]pyridine hit were investigated, and a chemical library was designed based on a focused hierarchical design of experiments approach. The X-ray structure of one of the inhibitors in complex with Mtb GS provided additional insight into the structure–activity relationships of this class of compounds. Finally, new α-arylated fosmidomycin analogues were synthesized as inhibitors of Mtb DXR, exhibiting IC50 values down to 0.8 µM. This work shows that a wide variety of aryl groups are tolerated by the enzyme. Cinnamaldehydes are important synthetic intermediates in the synthesis of fosmidomycin analogues. These were prepared by an oxidative Heck reaction from acrolein and various arylboronic acids. Electron-rich, electron-poor, heterocyclic and sterically hindered boronic acids could be employed, furnishing cinnamaldehydes in 43–92% yield.
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Li, Hui. "Algorithms for the selection of optimal spaced seed sets for transposable element identification." Miami University / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=miami1283178156.
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Nobili, Alberto [Verfasser]. "Improving biocatalysts via semi-rational protein design : use of a multiple sequence alignment platform to reduce screening efforts and facilitate hit identification / Alberto Nobili." Greifswald : Universitätsbibliothek Greifswald, 2016. http://d-nb.info/1113294191/34.
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Ehrt, Christiane [Verfasser], Daniel [Akademischer Betreuer] Rauh, and Stefan M. [Gutachter] Kast. "Protein binding site comparison : The impact of binding site similarity on hit identification in early drug discovery / Christiane Ehrt ; Gutachter: Stefan M. Kast ; Betreuer: Daniel Rauh." Dortmund : Universitätsbibliothek Dortmund, 2019. http://d-nb.info/1197903011/34.
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Spadaro, Alessandro [Verfasser], and Rolf W. [Akademischer Betreuer] Hartmann. "Pharmacophore aided hit identification, structural optimization and biological evaluation of benzothiazole derivatives as new potent and selective non-steroidal inhibitors of 17beta-hydroxysteroid dehydrogenase type 1 / Alessandro Spadaro. Betreuer: Rolf W. Hartmann." Saarbrücken : Saarländische Universitäts- und Landesbibliothek, 2012. http://d-nb.info/1052338666/34.
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Chen, Zhiqiang. "Identification of HIF-1 inhibitors." Thesis, University of Manchester, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.503047.
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Seddon, Deborah Ann. "Impossible identifications : Shakespeare and his readers (1830-1930)." Thesis, University of Cambridge, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.616135.
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Benson, Claire Elizabeth. "Genetics of familial hip osteoarthritis :identification of genetic susceptibility factors." Thesis, Queen's University Belfast, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.491996.
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Osteoarthritis is already a major cause of disability worldwide and with an ageing population the associated socio-economic burden is expected to soar. Complex interactions between genetic and environmental factors are responsible for the initiation of osteoarthritis. There is a clear genetic contribution to hip' osteoarthritis but to date no major susceptibility gene has been identified. A large collection of affected sib pairs with hip osteoarthritis had already been recruited from Northern Ireland (416 participants) and Nottingham (115 participants) . Additional unaffected (n=42) family members were recruited. As part of a genome wide screen 160 microsatellite markers from chromosomes 9, 10, 11, 14, 19, 20, 21 and 22 were typed in each family member (n=531)and controls (n=49). Candidate genes were identified in areas suggesting positive linkage. Single nucleotide polymorphism (SNP) typing or sequencing were used to investigate these genes further. The three regions most suggestive of linkage were found in chromosomes 9 (at 74.4mB), 11 (at 11.7mB) and 19 (at 8mB). Ten of the candidate genes identified were typed with SNPs in one member of each family (n=206) and controls (n=49). The ten exons of the other gene, osteoclast stimulating factor 1 (OSTFl), were sequenced in both cases (n=40) and controls (n= 8). The only candidate gene with significantly positive results was the collagen type V, alpha 3 gene (COL5A3) on chromosome 19. One haplotype block was significantly more common in the control population (p=O.Ol). Further analyses of the COL5A3 gene and its neighbouring genes are currently underway. Identification of a major susceptibility gene will improve our understanding of the underlying pathogenesis and potentially identify new targets for drug therapy. The global impact of finding a major susceptibility gene for hip osteoarthritis should encourage us to continue with our search to elucidate the genetics of familial hip osteoarthritis
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Crozier, J. R. "Enjoying Jesus Christ and the riches of his grace do you know how rich you are in Christ? a seven-week adventure /." Online full text .pdf document, available to Fuller patrons only, 2003. http://www.tren.com.
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Ledwaba, Neria Hunadi. "Identification of employees needs to be addressed in the HIV/AIDS programme at Aventis Pharmaceutical Company." Diss., Pretoria : [s.n.], 2003. http://upetd.up.ac.za/thesis/available/etd-11182003-132232.
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Siri, Åhman. "Wait, I'm him now? : Identification and choice in games with more than one protagonist." Thesis, Högskolan i Skövde, Institutionen för informationsteknologi, 2018. http://urn.kb.se/resolve?urn=urn:nbn:se:his:diva-15418.
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This study examines correlations between player choice and identification in a multiple protagonist video game, seeking to determine whether a player’s identification with one or more player characters affects the way they make choices while playing. It discusses various definitions and types of identification as well as ways to create a successful narrative with multiple protagonists. The artefact created for the study is a text-based game with a branching narrative, where the player is required to make choices for three different characters, and a qualitative research method based on interviews with a small group of participants. The results show that players seek to identify with the player character even when there are more than one, and often use this as a basis for the choices they make, either by imagining themselves in the situation of the main character or by imagining that they are the main character. They do not usually base their choices while playing as a character on their identification or lack thereof with another, and regardless of how they made choices, most players made more or less the same ones. However, it did show that lack of identification made making choices more difficult for that character, which lessened their enjoyment of that storyline.Artefakten som användes i arbetet utvecklades i samarbete med Amanda Thim
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Embretsén, Hanna, and Maria Palmberg. "En enfärgad regnbåge : Hur den homosexuella normen formas i HBT-magasinet QX." Thesis, Linnéuniversitetet, Institutionen för samhällsvetenskaper, SV, 2012. http://urn.kb.se/resolve?urn=urn:nbn:se:lnu:diva-24076.
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The HBTQ-society is not a homogeneous group; it contains a range of different sub-groups. QX is Sweden’s biggest magazine aimed at members of the HBTQ-society. The purpose of this study is to analyze if the magazine is more directed towards gay men or women – both by examining how many men versus women appear in QX, but also by looking at the different ways in which the articles are more relevant to either men or women, such as by tone, language or choice of subjects. In this study, we have analyzed twelve issues of QX, from January 2012 to December 2012, in order to get the most current results as possible. We used a quantitative content analysis when examining the division of men and women, and a qualitative analysis to investigate the direction towards each gender in the texts. Identification and power have been significant theories with which to analyse the results. The study clearly shows that QX is more aimed towards homosexual men rather than women, despite its claim to be gender neutral. During 2012, 66 % men and 34 % women appeared in QX. It was also discovered that more articles were aimed at gay men rather than women. The results support the theory that lesbian women could feel excluded from the HBTQ-society by reading QX. Since role models are crucial in the process of identification – and gay women access less of them than gay men – lesbians have a disadvantage in finding representation and identification in QX, which is unequal and therefore a problem.
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Abner, Erik 1986. "Identification of HIV-1 reactivating quinoline compounds as bromodomain inhibitors." Doctoral thesis, Universitat Pompeu Fabra, 2016. http://hdl.handle.net/10803/565528.
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Tras la infección por VIH-1, el establecimiento de un depósito de células T en reposo infectadas latentemente con VIH impide la erradicación del virus en pacientes. Para lograr la erradicación, la terapia retroviral existente debe combinarse con medicamentos que reactiven los virus latentes. Previamente, nuestro grupo describió un nuevo compuesto químico, MMQO (8-metoxi-6-metilquinolin-4-ol) que es capaz de reactivar la transcripción viral a través de un mecanismo desconocido. El objetivo de este proyecto fue identificar los proteínas que interaccionan con MMQO e investigar su papel en la reactivación del VIH-1.
Hemos establecido que MMQO es capaz de inducir la transcripción de minigenomas provirales que carecen de genes para los componentes virales, lo que nos permite plantear la hipótesis de que el compuesto funciona principalmente a través de factores del huésped. La caracterización de los perfiles de transcripción de MMQO mediante microarrays de expresión nos permitió identificar numerosos rasgos provocados por el compuesto. MMQO muestra una robusta naturaleza inmunosupresora que afecta a la proliferación celular debido a la disminución de los niveles proteicos de cMyc y Bcl-2 y la desregulación de genes sensibles a acetilación. Estas características indican que MMQO imita las lisinas acetiladas de histonas y funciona como un inhibidor de bromodominio y dominio extraterminal (BET). Análisis adicionales de la expresión génica y proteómica confirmaron esta hipótesis y demostramos que MMQO desplaza de la cromatina a Brd4, un miembro de la familia BET y antagoniza el papel pro-latente de Brd4 cerca del sitio de inicio de la transcripción de VIH-1. Modelos computacionales de docking también confirmaron la especificidad de MMQO hacia los bromodominios de la familia BET y un ensayo in vitro mediante FRET contra los miembros de la familia, identificó que MMQO tiene una mayor afinidad hacia la proteína Brd9. Por último, hemos establecido que la inhibición de Brd9 tiene un mínimo efecto sobre la expresión proviral, lo que sugiere que la principal función de MMQO sobre VIH-1 se puede atribuir al desplazamiento de Brd4.
Debido a la amplia gama de propiedades de los inhibidores de la familia BET, estas moléculas se están evaluando actualmente en ensayos clínicos contra diversos tipos de cáncer y afecciones inmunitarias. MMQO, con un funcionamiento dual, es un nuevo miembro de esta clase de medicamentos. La estructura minimalista de MMQO puede ser muy prometedora ya que puede ser modificada para optimizar la afinidad hacia Brd9 / 4 y, potencialmente, podría ser de utilidad en la investigación contra una gran variedad de enfermedades, incluyendo el VIH.Upon HIV-1 infection, a reservoir of HIV latently infected resting T cells prevents the eradication of the virus from patients. To achieve eradication, the existing virus suppressing antiretroviral therapy must be combined with drugs that reactivate the dormant viruses. Our group previously described a novel chemical scaffold compound, MMQO (8-methoxy-6-methylquinolin-4-ol), which is capable of reactivating viral transcription through an unknown mechanism. The objective of this project was to identify the molecular binding partners of MMQO and elaborate their role in the reactivation of HIV-1. We established that MMQO is capable of inducing HIV-1 independently of viral proteins by inducing transcription from proviral minigenomes lacking genes for viral components, allowing us to hypothesize that the compound primarily functions through host factors. Characterizing MMQO’s transcriptional profiles with total mRNA expression microarrays, we were able to identify numerous traits provoked by the drug. MMQO displayed a robust immunosuppressive nature, it affected cell proliferation by diminishing cMyc and Bcl-2 protein levels and increased the dysregulation of acetylation sensitive genes. These hallmarks indicated that MMQO mimics acetylated lysines of core histones and functions as a bromodomain and extraterminal domain (BET) protein family inhibitor. Further gene expression and proteomic analysis confirmed this supposition and we demonstrated that MMQO deposes of the BET family member Brd4 from global chromatin and antagonizes the pro-latent role of Brd4 near the transcription start site of HIV-1. Computational docking models also confirmed MMQO’s specificity towards the BET family bromodomains and an in vitro screening against the family members by FRET identified MMQO to have the highest affinity towards the Brd9 protein. Finally, we established that the inhibition of Brd9 had minimal effect on the proviral expression, suggesting that the primary function of MMQO on HIV-1 can be attributed to the displacement of Brd4. Due to the broad range of properties characteristic to BET family inhibitors, these molecules are currently being evaluated in clinical trials against various types of cancers and immune conditions. The dual functioning scaffold compound MMQO is a new member of this class of drugs. The minimalistic structure of MMQO shows promise for it to be further optimized for higher affinities towards Brd9 / 4 and could potentially be of use in research against a variety of diseases, including HIV.
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Grace, Christopher. "Diagnostically significant antigens of Treponema pallidum subsp. pallidum : identification, serological efficacy, and characterisation of the major antigenic determinants." Thesis, Open University, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.311861.
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Marcus, Benjamin J. "Predictors of Engagement in the Community Affected by HIV and AIDS." Scholarship @ Claremont, 2009. http://scholarship.claremont.edu/cgu_etd/6.
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The goal of this study was to explore factors that lead to engagement in the community affected by HIV and AIDS (CAHA). An additional goal of this study was to better understand the relationships between psychological sense of community (PSOC) and social identification (SI), and their connections to two classes of motivations: community concern motivations (CCM) and esteem enhancement motivations (EEM). These constructs predicted two types of engagement in CAHA: AIDS activism and intentions for future participation in community related activity. Analyses were conducted on existing data (Omoto, 2005). The results indicate that PSOC and SI should be considered as independent constructs that uniquely explain community engagement. CCM were found to mediate these relationships, potentially explaining their links.
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Wu, Hao, and 吴昊. "Identification of drug resistant mutations in HIV-1 latently infected patients under successful HAART and in CRF_BC variants selected invitro." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B47149826.
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Ballalai, Rodrigo Clemente. "O jovem no movimento hip hop : espaço potencial de criatividade e identificação? /." Assis : [s.n.], 2009. http://hdl.handle.net/11449/97560.
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Orientador: Olga Ceciliato MattioliBanca: Diana Pancini de Sá Antunes Ribeiro
Banca: Leila Rute Oliveira Gurgel do Amaral
Resumo: A partir de experiência de estágios da graduação em Psicologia, particularmente no trabalho desenvolvido junto ao "Programa de Liberdade Assistida" da FEBEM, unidade Bauru, quando oferecemos um pronto atendimento psicológico a seus participantes, iniciamos o contato com o movimento hip hop. Entrevistamos jovens que se identificam com o movimento hip hop, analisando sua trajetória ao ingressarem nos grupos organizados de hip hop do município de Bauru. Analisamos suas biografias e percepções que estes jovens têm de si e de sua realidade social. O instrumento utilizado foi a entrevista semi dirigida, cuja aplicação ocorreu individualmente O procedimento para a análise das informações se apoiou no método psicanalítico. Analisamos o que leva os jovens a participarem do referido movimento, por meio do conceito psicanalítico de identificação. Verificamos que a significação que os jovens atribuem ao hip hop pode ser entendido como um espaço potencial de criatividade, de acordo com a teoria do psicanalista inglês Donald Winnicott. Encontramos diferentes usos desse movimento cultural, de acordo com a singularidade de cada participante, inclusive manifestado pelos diversos meios de participação artística e ideológica de seus componentes. No campo identificatório consideramos o movimento hip hop como modelo identificatório contemporâneo e significativo para grande parcela da população jovem. Como apresenta Maria Rita Kehl, o tratamento de irmãos ("manos") indica um campo de identificação horizontal com um grupo de pessoas iguais, oriundas de uma mesma realidade dolorosa; distinto de uma identificação vertical, tal como ocorre na grande massa em relação ao líder ou ídolo.
Abstract: During the course of training in psychology, especially in a work with the Program of Freedom Febem Assisted in the unit of Bauru, offering psychological care ready to participants, initiated the contact with the hip hop movement. In this study, we analyzed the motivation of some young people to join this movement, to perform such analysis, we used the psychoanalytic concept of identification. We want to check the meaning given by these young people to the hip hop movement, and if this can be seen as a potential space for creativity. Young people interviewed are identified with the hip hop movement, and why we are looking at their life stories the moment they began to join the hip hop movement organized in the city of Bauru. I reviewed their biographies and their perceptions about themselves and about social reality in which they live. We conducted semi-directed interviews with each couple in particular. We use the psychoanalytic method to analyze the content of the interviews, especially the theory developed by psychoanalyst Donald Winnicot.
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LORAIN, STEPHANIE. "La famille des genes hir : etude chez les eucaryotes et elements d'analyse fonctionnelle par identification de partenaires moleculaires." Paris 6, 1998. http://www.theses.fr/1998PA066556.
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Le gene hira a ete isole pour sa position dans la region du chromosome 22 critique pour le syndrome de digeorge. L'analyse de la sequence proteique de son produit a montre qu'il codait pour un homologue structural des proteines hir1p et hir2p de s. Cerevisiae. Ces proteines repriment la transcription de genes d'histones au cours du cycle cellulaire. Elles agissent ensemble sur la structure de la chromatine, sans doute au sein d'un complexe multiproteique. Mon travail de these s'est articule autour de trois axes principaux : 1 o une tentative de complementation fonctionnelle de mutants hir de levure par la proteine hira humaine. Nos resultats montrent une restauration partielle d'un phenotype sauvage par la proteine humaine. 2 o l'exploration de la famille des genes hir chez les eucaryotes superieurs. En particulier, nous avons isole l'homologue de hira chez la souris, un prerequis a l'etude de son implication dans l'embryogenese ; nous avons egalement elucide la structure genomique du gene hira humain pour permettre la recherche de mutations chez la minorite de patients atteints de syndrome de digeorge presentant un caryotype normal. 3 o la recherche de partenaires de la proteine hira humaine par un test de piege a interactions proteiques. Plusieurs cribles ont abouti a l'isolement de six adnc codant pour des interacteurs de hira. La moitie d'entre eux codait pour des proteines connues : l'histone h2b et le facteur de transcription pax3. Les autres interacteurs ont ete nommes hirip3, hirip4 et hirip5 (pour hira interacting protein). Des tests in vitro ont confirme et precise les interactions detectees dans la levure. Une association de la proteine hira avec une deuxieme histone de la particule cur nucleosomale, l'histone h4, a ete mise en evidence. Par ailleurs, hirip3 interagit avec h2b et h3. Hirip4 appartient a la famille des proteines dnaj qui agissent avec les proteines dnak dont elles stimulent l'activite atpase intrinseque. Une partie des molecules hirip4 etant nucleaires, nous emettons l'hypothese que ces proteines pourraient recruter une activite atpase importante a la fonction d'un complexe nucleaire contenant hira. Ces resultats, ajoutes aux donnees relatives a hir1p et hir2p de levure, indiquent que les proteines hir fonctionnent tres probablement dans toutes les especes au sein de complexes proteiques ayant un effet sur la structure de la chromatine.
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Jacobs, Michael A. "Identification and characterization of a chloroplast-encoded His-Asp signal transduction protein in the toxic stramenopile Heterosigma akashiwo /." Thesis, Connect to this title online; UW restricted, 2000. http://hdl.handle.net/1773/5250.
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Gálvez, Celada Cristina. "Identification of immunovirological factors that determine an extremely low viral reservoir: approaching the cure of HIV-1 infection." Doctoral thesis, Universitat Autònoma de Barcelona, 2021. http://hdl.handle.net/10803/671118.
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Gairebé 40 milions de persones viuen actualment amb el virus de la immunodeficiència humana tipus 1 (VIH-1) al món, i més de 30 milions de morts s’atribueixen al VIH-1 des del començament de l’epidèmia. El desenvolupament de la teràpia antiretroviral combinada (TARc) és un dels principals èxits mèdics del segle XX, ja que suprimeix eficaçment la virèmia plasmàtica, millora la funció del sistema immunològic, redueix la mortalitat i morbiditat, i el risc de transmissió viral. No obstant, la TARc no pot curar la infecció per la presència de cèl·lules infectades de manera latent, el que es coneix com a reservori viral.
El nombre de cèl·lules infectades latentment presents en el cos varia notablement entre diferents individus infectats pel VIH-1. Baixos nivells de reservoris s’han relacionat tradicionalment amb un control del VIH-1 o bon pronòstic de la malaltia, però els factors específics que provoquen aquesta reducció del reservori encara es desconeixen. En aquest context, l’objectiu d’aquesta tesi és caracteritzar els factors clínics, virals i immunogenètics associats a la reducció del reservori del VIH-1 en tres models diferents: [1]controladors naturals de la infecció amb un llarg seguiment (Controladors d’Elit Excepcionals o EEC), [2]individus amb un reservori de VIH-1 espontàniament baix malgrat una progressió normal de la infecció (LoViReT), i [3]persones amb VIH en les que s’observa una inducció de la caiguda del reservori després d’un trasplantament al·logènic de cèl·lules mare hematopoètiques (allo-HSCT).
Els nostres resultats van mostrar que els baixos nivells de reservori observats en els EEC es van deure a la combinació de virus defectius que no evolucionen amb factors genètics protectors de l’hoste juntament amb respostes immunitàries potents específiques del VIH-1. En conjunt, la combinació d’aquests factors sembla important per a controlar la replicació del VIH-1 i aconseguir una cura funcional espontània. D’altra banda, els individus LoViReT semblen tenir un establiment defectiu del reservori de VIH-1 ja que es van observar baixos nivells de reservori en sang (fins i tot abans de l’inici del tractament) i santuaris anatòmics. Això estava recolzat per una distribució esbiaixada de la latència en les diferents subpoblacions de cèl·lules T CD4+ cap a subpoblacions de vida més curta. No obstant això, encara que els individus LoViReT van mostrar un sistema immunològic preservat, no es van observar respostes immunitàries específiques del VIH-1 i, per tant, no s’esperaria que poguessin controlar la replicació del VIH-1 en absència de TARc. Finalment, l’estudi d’individus infectats per VIH-1 que van rebre un trasplantament al·logènic de cèl·lules mare hematopoètiques va mostrar una notable disminució del reservori de VIH-1 en sang perifèrica i compartiments anatòmics, fins i tot en el context de cèl·lules amb un CCR5 salvatge. Això probablement va ser degut a un potent efecte de l’empelt contra VIH-1 causat per factors específics del transplantament com el ràpid quimerisme, cèl·lules procedents d’un donant adult i la malaltia de l’empelt contra l’hoste. Desafortunadament, la falta de respostes immunes potents específiques del VIH-1 sostingudes en el temps resultaria en un rebot viral, com es va observar en un dels individus estudiats trasplantat amb cèl·lules amb un CCR5 salvatge.
En conjunt, demostrem que els factors relacionats amb una funció viral alterada seran importants per la reducció del reservori del VIH-1. Però, per controlar la replicació del VIH en absència de TARc serà necessària la combinació amb respostes immunes potents específiques del VIH-1, com en els EEC. Futurs assajos clínics en persones amb un reservori baix de VIH-1, com els individus LoViReT o que hagin rebut un trasplantament al·logènic de cèl·lules mare, serien de gran interès en estratègies destinades a estimular les respostes immunes específiques del VIH-1 amb l’objectiu final de curar el VIH-1.Casi 40 millones de personas viven actualmente con el virus de la inmunodeficiencia humana tipo 1 (VIH-1) en el mundo, y más de 30 millones de muertes se atribuyen al VIH-1 desde el comienzo de la epidemia. El desarrollo de la terapia antirretroviral combinada (TARc) es uno de los principales éxitos médicos del siglo XX, ya que suprime eficazmente la viremia plasmática, mejora la función del sistema inmunológico, reduce la mortalidad y morbilidad, y el riesgo de transmisión viral. Sin embargo, la TARc no puede curar la infección debido a la presencia de células infectadas de forma latente, lo que se conoce como reservorio viral. El número de células latentemente infectadas presentes en el cuerpo varía notablemente entre diferentes individuos infectados por VIH-1. Bajos niveles de reservorios se han relacionado tradicionalmente con un control del VIH-1 o buen pronóstico de la enfermedad, pero los factores específicos que provocan esta reducción del reservorio aún se desconocen. En este contexto, el objetivo de esta tesis es caracterizar los factores clínicos, virales e inmunogenéticos asociados con la reducción del reservorio del VIH-1 en tres modelos diferentes: [1]controladores naturales de la infección con un largo seguimiento (Controladores de Elite Excepcionales o EEC), [2]individuos con un reservorio de VIH-1 espontáneamente bajo a pesar de una progresión normal de la infección (LoViReT), y [3]personas VIH-1+ en las que se observa una caída del reservorio después de un trasplante alogénico de células madre hematopoyéticas (allo-HSCT). Nuestros resultados mostraron que los bajos niveles de reservorio observados en los EEC se debieron a la combinación de virus defectivos que no evolucionan con factores genéticos protectores del huésped junto con respuestas inmunitarias potentes específicas del VIH-1. La combinación de estos factores parece importante para controlar la replicación del VIH-1 y alcanzar una cura funcional espontánea. Por otro lado, los individuos LoViReT parecen tener un establecimiento defectivo del reservorio de VIH-1 ya que se observaron bajos niveles de reservorio en sangre (incluso antes del inicio del tratamiento) y santuarios anatómicos. Esto estaba acompañado por una distribución sesgada de la latencia en las diferentes subpoblaciones de células T CD4+ hacia subpoblaciones de vida más corta. Sin embargo, aunque los individuos LoViReT mostraron un sistema inmunológico preservado, no se observaron respuestas inmunitarias específicas del VIH-1 y, por lo tanto, no se esperaría que controlaran la replicación del VIH-1 en ausencia de TARc. Finalmente, el estudio de individuos infectados por VIH-1 que recibieron un trasplante alogénico de células madre hematopoyéticas mostró una notable disminución del reservorio de VIH-1 en sangre periférica y compartimentos anatómicos, incluso en el contexto de células con un CCR5 salvaje. Esto probablemente fue debido a un potente efecto de injerto contra VIH-1 causado por factores específicos del trasplante como un rápido quimerismo, células procedentes de un donante adulto y la presencia de enfermedad del injerto contra huésped. Desafortunadamente, la falta de respuestas potentes específicas de VIH-1 sostenidas en el tiempo resultaría en un rebote viral, como se observó en uno de los individuos estudiados que fue trasplantado con células con un CCR5 salvaje. En conjunto, demostramos que los factores relacionados con una función viral alterada son importantes para la reducción del reservorio de VIH-1. Aun así, para controlar la replicación del VIH-1 en ausencia de TARc será necesaria la combinación con respuestas inmunes potentes específicas de VIH-1, como sucede en los EEC. Futuros ensayos clínicos en personas con un reservorio bajo de VIH-1, como individuos LoViReT o que hayan recibido un trasplante alogénico de células madre, serían de gran interés en estrategias destinadas a estimular las respuestas inmunes específicas del VIH-1 con el objetivo de curar
Nearly 40 million individuals are currently living with the human immunodeficiency virus type 1 (HIV-1) worldwide and more than 30 million deaths can be attributed to HIV-1 since the start of the epidemic. The development of combination antiretroviral therapy (cART) was one of the major medical success of the 20th century as it effectively suppresses plasma viremia, improves the immune system function, reduces mortality and morbidity, and the risk of viral transmission. However, cART cannot cure the infection due to the presence of latently infected cells. The number of latently infected cells present in the body, known as the reservoir, varies notably among different HIV-1-infected individuals. Low levels of reservoirs are traditionally related to HIV-1 control or good disease prognosis, but the specific factors provoking a reduction of this reservoir are still unknown. In this context, the aim of this thesis is to characterize the clinical, viral and immunogenetic factors associated with the reduction of the HIV-1 reservoir in three different models: [1] a natural long-term control of the infection (Exceptional Elite controllers or EEC), [2] a natural low HIV-1 reservoir despite regular HIV-1 progression (Low Viral Reservoir Treated individuals or LoViReT), and [3] an induced low reservoir after allogeneic hematopoietic stem cell transplant (allo-HSCT) in persons living with HIV-1. Our results showed that the low-level reservoirs observed in EEC were due to a combination of defective non-evolving virus with protective host genetic factors together with potent HIV-1 specific immune responses. Altogether, the combination of those factors seems important to control HIV-1 replication and reach a spontaneous functional cure. On the other hand, LoViReT individuals seem to have an impaired HIV-1 reservoir establishment since the low levels of reservoir were observed in blood (even before the initiation of treatment) and anatomical sanctuaries. This was supported by a skewed distribution of the provirus among the different CD4+ T cells subpopulations towards short-live subpopulations. However, although LoViReT individuals have a preserved general immune system, they do not have HIV-1 specific immune responses and therefore are not able to control HIV-1 replication in the absence of cART. Finally, the study of HIV-1-infected individuals that received an allo-HSCT showed a remarkably decline of the HIV-1 reservoir in peripheral blood and anatomical compartments, even in the setting of CCR5 wild-type cells. This was most likely due to a potent graft versus HIV-1 effect caused by specific factors of the transplant as a rapid engraftment, an adult donor, or a graft versus host disease. Unfortunately, the lack of a potent HIV-1 specific immune responses sustained in time would result in viral rebound, as observed in one of the studied individuals transplanted with CCR5 wild-type cells. Altogether, we showed that factors related to an impaired viral function will be important to the reduction of HIV-1 reservoir. A combination with a potent HIV-1 specific immune response will be necessary to control the HIV-1 replication in the absence of cART as happened in EEC. Future clinical trials in individuals with a low HIV-1 reservoir, like LoViReT or allo-HSCT individuals, would be of great interest in strategies aimed at boosting HIV-1 immune specific responses with the ultimate goal of curing HIV-1.
Universitat Autònoma de Barcelona. Programa de Doctorat en Bioquímica, Biologia Molecular i Biomedicina
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Tartour, Kevin. "Identification et caractérisation d’une nouvelle famille de facteur de restriction - les IFITMs." Thesis, Lyon, École normale supérieure, 2014. http://www.theses.fr/2014ENSL0972.
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Le VIH, comme tous les virus, nécessite la coopération de nombreuses protéines cellulaires pour réaliser son cycle viral. Néanmoins, un panel de protéines est spécifiquement dédié à la protection des cellules et ce, de façon autonome. Ces protéines sont nommées facteurs de restriction. Jusqu’à présent, un grand nombre de protéines ayant une action antivirale plus ou moins importante contre le VIH a été identifié. Seules quelques-Unes sont bien étudiées telles que APOBEC3G, Tétherine, TRIM5α ou SamHD1. Toutefois, ces facteurs de restriction ne permettent pas à eux seuls d’expliquer la permissivité de tous les types cellulaires. Notre étude a donc pour objet la découverte de nouvelles protéines agissant contre le VIH. Parmi un grand nombre de candidats, nous avons identifié une famille de protéines agissant contre le VIH : les IFITMs. Il a été précédemment décrit que l’expression des IFITMs protège les cellules de l’infection par le VIH mais aussi par de nombreux autres virus, filovirus, coronavirus, flavivirus ou encore paramyxovirus. Nous décrivons ici un nouveau mécanisme antiviral mis en place par les IFITMs. L’expression des IFITMs dans les cellules productrices de virus affecte la production de particules infectieuses. D’une part, moins de particules sont produites et d’autre part les particules sont moins infectieuses. Nous nous sommes focalisés sur le défaut d’infection et celui-Ci corrèle avec l’incorporation des IFITMs à la membrane des virions. Les virus produits et portant les IFITMs présentent un défaut lors du processus d’entrée dans la cellule cible au cours de l’étape de fusion. Ceci n’est pas dû à un défaut d’incorporation de l’enveloppe virale, mais le mécanisme précis n’est pas encore compris. Il apparaît donc que les IFITMs peuvent jouer un double rôle restrictif, protégeant les cellules où ils sont exprimés et diminuant la production de particules infectieusesHIV, as all viruses, hijacks the cellular machinery to its benefits. Nevertheless, a range of proteins protects cells by an autonomous way, they are called restriction factor and belong to a system called “the intrinsic immunity”. Up to now, a certain number of cellular proteins, most of which belong to this defense system, have been described to be more or less detrimental for HIV replication, as for example, APOBEC3G, Tetherin, TRIM5α and SamHD1. Given that a large number of cellular restriction factors are likely to be discovered, we started to screen to look for new ones. Among a lot of candidates, we have identified a family of proteins interfering with HIV: the IFITMs. Previously, studies described IFITMs as proteins protecting cell against viral entry and fighting a lot of viruses, including HIV. In this thesis, we described a new antiviral mechanism mediated by IFITMs against retroviruses. When IFITMs are expressed in cells producing new virions, there is a high defect of infectious particle release due to a defect of particle release and a defect of the infectivity of particles. We focus on the intrinsic defect of infectivity and we show that particles cannot anymore enter into target cells and this observation is correlated with the incorporation of IFITMs in viral membranes. The defect is located at the fusion step and is not explained by a defect of envelope incorporation.To conclude, IFITMs have a double restrictive effect, protecting cells from incoming viruses and decreasing the production of infectious particles
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Ballalai, Rodrigo Clemente [UNESP]. "O jovem no movimento hip hop: espaço potencial de criatividade e identificação?" Universidade Estadual Paulista (UNESP), 2009. http://hdl.handle.net/11449/97560.
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ballalai_rc_me_assis.pdf: 488510 bytes, checksum: 0f5cd2f861ddf680adab3dd5756129d7 (MD5)A partir de experiência de estágios da graduação em Psicologia, particularmente no trabalho desenvolvido junto ao “Programa de Liberdade Assistida” da FEBEM, unidade Bauru, quando oferecemos um pronto atendimento psicológico a seus participantes, iniciamos o contato com o movimento hip hop. Entrevistamos jovens que se identificam com o movimento hip hop, analisando sua trajetória ao ingressarem nos grupos organizados de hip hop do município de Bauru. Analisamos suas biografias e percepções que estes jovens têm de si e de sua realidade social. O instrumento utilizado foi a entrevista semi dirigida, cuja aplicação ocorreu individualmente O procedimento para a análise das informações se apoiou no método psicanalítico. Analisamos o que leva os jovens a participarem do referido movimento, por meio do conceito psicanalítico de identificação. Verificamos que a significação que os jovens atribuem ao hip hop pode ser entendido como um espaço potencial de criatividade, de acordo com a teoria do psicanalista inglês Donald Winnicott. Encontramos diferentes usos desse movimento cultural, de acordo com a singularidade de cada participante, inclusive manifestado pelos diversos meios de participação artística e ideológica de seus componentes. No campo identificatório consideramos o movimento hip hop como modelo identificatório contemporâneo e significativo para grande parcela da população jovem. Como apresenta Maria Rita Kehl, o tratamento de irmãos (“manos”) indica um campo de identificação horizontal com um grupo de pessoas iguais, oriundas de uma mesma realidade dolorosa; distinto de uma identificação vertical, tal como ocorre na grande massa em relação ao líder ou ídolo.
During the course of training in psychology, especially in a work with the Program of Freedom Febem Assisted in the unit of Bauru, offering psychological care ready to participants, initiated the contact with the hip hop movement. In this study, we analyzed the motivation of some young people to join this movement, to perform such analysis, we used the psychoanalytic concept of identification. We want to check the meaning given by these young people to the hip hop movement, and if this can be seen as a potential space for creativity. Young people interviewed are identified with the hip hop movement, and why we are looking at their life stories the moment they began to join the hip hop movement organized in the city of Bauru. I reviewed their biographies and their perceptions about themselves and about social reality in which they live. We conducted semi-directed interviews with each couple in particular. We use the psychoanalytic method to analyze the content of the interviews, especially the theory developed by psychoanalyst Donald Winnicot.
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Yuan, Tingting, and 袁婷婷. "Identification of intermediate antibodies of broadly neutralizing HIV-1 human monoclonal antibody b12 and characterization of variable loops of HIV-1 envelop glycoprotein." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2013. http://hdl.handle.net/10722/196445.
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Otieno, Fredrick Odhiambo. "Use of modified respondent driven sampling methodology to enhance identification and recruitment of most at risk persons into an HIV prevention trial in Kisumu, Western Kenya." University of the Western Cape, 2016. http://hdl.handle.net/11394/4975.
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Philosophiae Doctor - PhDThis thesis presents research on the use of modified respondent driven sampling (mRDS) methodology to enhance identification and recruitment of key populations (KP) into an HIV prevention trial in Kisumu, western Kenya through a three phase mixed method study. The study was carried out in Kisumu, western Kenya within the Kenya Medical Research Institute (KEMRI) and the US Centers for Disease Control and Prevention (CDC) Research and Public Health Collaboration platform. The three phases included: 1. PHASE I: Identification and determination of categories of KPs and techniques of locating and motivating them to participate in HIV prevention trials.2. PHASE II: Design and Implementation of a mRDS methodology in recruiting Ks into HIV prevention trials.3. PHASE III: Evaluation of the mRDS in recruitment of KPs into an HIV incidence cohort study. Methods Phase I of the study included the conduct of in depth interviews which were used to identify different categories of persons considered to be KPs within Kisumu, identify strategies of locating the KPs and determine motivators and inhibitors of KPs participation in HIV prevention trials. Phase II on the other hand included the administration of a survey that had been refined in Phase I. The survey was used to design a mRDS methodology which was then implemented to recruit KPs into the survey. Phase III evaluated the success of the mRDS in recruiting KPs into an HIV prevention study by assessing the risk profiles for participants screened and enrolled into the Phase III study. Ethical approval for the study was sought from the ethics committee of the Kenya Medical Research Institute, the US CDC and the University of the Western Cape.The study recruited 53 individuals into phase I and was able to 8 identify different categories of people considered to be KPs and the 4 salient strategies that could be used to recruit them into HIV prevention studies. The phase also identified 8 potential motivators and 9 potential inhibitors to participation in HIV prevention research. These categories and salient strategies were used in phase II to develop and pilot a mRDS methodology in recruiting 203 individuals into a survey. The survey was used as a validation tool for the risk levels of persons recruited by the mRDS using the variables of inconsistent condom use and having multiple partners. The validated mRDS was then applied in the recruitment of 1,292 participants in phase III of thestudy. These study participants had characteristics similar to those seen in similar studies and elucidated from phases I and II of the study. HIV seropositivity was used as the variable for validating risk levels of participants in this study and this was found to be higher that that seen in general population and comparable to that seen in other KPs groups in the region. Results: Overall the study was able to identify different categories of people considered to be at high riskof HIV acquisition. The groups identified included people who frequent bars (e.g. bar workers, drunkards, sex workers, businessmen), people who work in transportation (e.g. truck drivers, matatu drivers, motorcycle drivers, taxi drivers, bicycle taxi drivers), fishermen/fishmongers, MSM and hair salon workers. The study also identified using personal contact, link persons, peer mobilisers and leaders as strategies of identifying and locating KPs. The study used the mRDS successfully in recruiting participants with evaluation of inconsistent condom use and multiple sexual partnerships showing the participants to be of high risk behaviour. Of all the females in the study, only 3.3% were pregnant. The prevalence of Chlamydia was 2.9%, gonorrhoea was 5.0%, syphilis was 0.4% and HSV-2 was 46.0%. Those who tested positive for HIV were 26.2% with 42.3% of the HIV positive participants having CD4 counts of between 250 – 500 cells/ml. Recommendations and Conclusion:The mRDS was successful in recruiting KPs in an HIV prevention trial. Majority of the participants reported inconsistent condom use and having multiple sex partners. In addition to MSM, SW and transport industry workers, fisherfolk, discordant couples, widowers, street youth, car washers and police also form part of KPs groups. The HIV prevalence was higher amongst these groups compared to general population with discordant couples having the highest HIV prevalence. The study recommends that mRDS should be used to identify and recruit KPs as it not only allows for faster recruitment of KPs, it also reduces the expense and complexity associated with coupon management in the standard RDS.
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Heyman, Heino Martin. "Identification of anti-HIV compounds in Helichrysum species (Asteraceae) by means of NMR-based metabolomic guided fractionation." Thesis, University of Pretoria, 2013. http://hdl.handle.net/2263/40273.
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The plant kingdom contributes significantly to the natural products that are used for the
treatment of a large number of ailments and disease across the globe. Included in these
species is the Helichrysum genus (Asteraceae), which comprises of more then 600
species across Africa of which 244 species are found in South Africa. Helichrysum
species are used in many cases for the treatment of coughs, colds, fever, infection,
headaches, menstrual pain and are also very popular for wound dressing due to their
potential antibacterial properties. The most common Helichrysum species used in
traditional medicine and for several medicinal purposes are H. cymosum, H. odoratissimum, H. petiolare and H. nudifolium. Previously published research has
shown that several of the Helichrysum species do have antimicrobial activity with the
most relevant to this study being the discovery of antiviral activity of H. aureonitens
against herpes simplex virus type 1 (HSV-1) as well as the reports of anti-HIV (human
immunodeficiency virus) activity of several Helichrysum species. With this knowledge, a
more in-depth study was initiated to identify the possible active constituents in South
African Helichrysum species against HIV. Due to the need to speed up drug discovery
especially against epidemic diseases like HIV, this study investigated a new tool
(nuclear magnetic resonance (NMR) – based metabolomics) to speed up drug
discovery form natural products especially when anti-viral constituents are investigated. odoratissimum, H. petiolare and H. nudifolium. Previously published research has
shown that several of the Helichrysum species do have antimicrobial activity with the
most relevant to this study being the discovery of antiviral activity of H. aureonitens
against herpes simplex virus type 1 (HSV-1) as well as the reports of anti-HIV (human
immunodeficiency virus) activity of several Helichrysum species. With this knowledge, a
more in-depth study was initiated to identify the possible active constituents in South
African Helichrysum species against HIV. Due to the need to speed up drug discovery
especially against epidemic diseases like HIV, this study investigated a new tool
(nuclear magnetic resonance (NMR) – based metabolomics) to speed up drug
discovery form natural products especially when anti-viral constituents are investigated.
In this study very promising anti-HIV results were obtained from several aqueous
extracts (1:1 methanol/water) using a full virus model i.e. Helichrysum populifolium (IC50
12 μg/ml), H. appendiculatum (IC50 17 μg/ml), H. cymosum ssp. clavum (IC50 19 μg/ml),
H. oxyphyllum (IC50 19 μg/ml) and H. cymosum ssp. cymosum (IC50 21 μg/ml). With the
use of NMR-based metabolomics and multivariate data analysis (MVA) the specific
characteristic that differentiated the active extracts from the non-active extracts was
identified by making use of Orthogonal Projections to Latent Structures – Discriminant
Analysis (OPLS-DA). This characteristic was then used as a “blue print” or “fingerprint”
to guide the process of fractionation and purification. H. populifolium showed the highest
anti-HIV activity and thus was selected as the candidate extract for further analysis.
After a very quick and simple chromatographic fractionation process, seven fractions
were compared against the activity profile by making use of their NMR profiles, which
then visually indicated which of the fractions had the highest similarity. Fraction 6 had
the most similar “fingerprint”. The compounds of this active fraction were then identified
with the use of liquid chromatography – ion trap – time of flight (LC-IT-TOF) for quick
identification. The analysis revealed the presence of five chlorogenic type compounds,
3,4-dicaffeoyl quinic acid (DCQA), 3,5-DCQA, 4,5-DCQA, 1,3,5- tricaffeoyl quinic acid
(TCQA) and 5-malonyl-1,3,4-TCQA of which several are well known to have anti-HIV activity ranging from 0.85μM to 12μM. We were thus able to show with this study the
possibility of using NMR-based metabolomics guided fractionation to guide the process
of fractionation and identification from an active characteristic profile to the active
constituents within the active H. populifolium extract.Thesis (PhD)--University of Pretoria, 2013.
gm2014
Plant Science
unrestricted
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Jegede, Oyebisi. "Identification and Characterization of Novel Antiretroviral Compounds: from Small Molecule Library Screening to Rationally Designed Compounds." [Kent, Ohio] : Kent State University, 2007. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=kent1185563176.
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Thesis (Ph.D.)--Kent State University, 2007.Title from PDF t.p. (viewed Mar. 11, 2009). Advisor: Miguel Quiñones-Mateu. Keywords: HIV/AIDS, drug discovery, small molecule library screening, characterization of new antiretroviral drugs, highly active antiretroviral therapy. Includes bibliographical references (p. 180-200).
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Fleta, Soriano Eric 1983. "Broad-spectrum host-acting antivirals: identification and characterization of anti-HIV drugs." Doctoral thesis, Universitat Pompeu Fabra, 2015. http://hdl.handle.net/10803/402212.
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Hundreds of host factors related to viral infections like HIV, hepatitis C virus, dengue virus or West Nile virus have been identified. As many of these host factors are shared by different viruses, chemical blockade of key virus-associated cellular components may effectively act as broad-spectrum antiviral treatment. Broad-spectrum host-acting antivirals (HAAs) may reduce treatment complexity and costs, increase adherence to the therapy and may pose a higher barrier to develop resistance. In this thesis a high-throughput anti-HIV assay was used to screen for virus inhibitory effects of a library of secondary metabolites derived from myxobacteria. Compounds with high anti-HIV activity and low toxicity were classified as hits and two of them (ratjadone A and soraphen A) were selected for further analysis. The mechanism of HIV inhibition of both compounds is described here. The results presented in this thesis show that broad-spectrum HAAs are a feasible option for antiviral treatment and that the compounds identified can be further studied for hit-to-lead compound development.Cientos de factores del huésped relacionados con infecciones virales por VIH, hepatitis C, dengue o virus del Nilo occidental han sido identificados. Como muchos de esos factores del huésped son compartidos por diferentes virus, el bloqueo químico de un componente celular clave asociado al virus podría actuar de forma efectiva como un tratamiento antiviral de amplio espectro. Antivirales de amplio espectro contra factores del huésped podrían reducir la complejidad y el coste del tratamiento, incrementar el cumplimiento de la terapia y pueden suponer una barrera mayor al desarrollo de resistencia. En esta tesis un cribado de alta capacidad anti-VIH fue aplicado a una librería de metabolitos secundarios de myxobacteria. Compuestos con alta actividad anti-VIH y baja toxicidad fueron clasificados como hits y dos de ellos (ratjadone A y soraphen A) fueron seleccionados para posteriores estudios. El mecanismo de inhibition de VIH de ambos compuestos es descrito aquí. Los resultados presentados en esta tesis muestran que usar antivirales de amplio espectro contra factores del huésped es un opción viable para tratamientos antivirales y que los compuestos identificados pueden ser estudiados para el desarrollo de fármacos.
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Dadgar, Majid. "Pattern Language: Identification of design opportunities for the child with Autism Spectrum Disorder (ASD) to develop his/her social skills." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1313619497.
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Álvarez, Marimón Mª Elena. "Structural studies of Heteromeric Amino acid Transporters (HATs): Validation of the first 3D structural model of a HAT (human 4F2hc/LAT2) and identification of new HAT targets for 3D‐crystallization." Doctoral thesis, Universitat de Barcelona, 2014. http://hdl.handle.net/10803/284082.
Full textAbstract:
Heteromeric amino acid transporters (HATs) mediate the transport of amino acids through the plasma membrane. They are composed of two subunits (a heavy and a light one) linked by a conserved disulfide bridge. Genetic defects in the genes coding these HATs may affect its functionality or expression, leading to inherited aminoacidurias. Thus, solving the structure of the Eukaryotic HATs has become of great importance. However structural information about interactions between the heavy and light subunits of HATs is scarce.
In this work, human 4F2hc/L-type amino acid transporter 2 (LAT2) first low resolution 3D model obtained by single particle negative-staining transmission electron microscopy (TEM) was validated. In order to assess the interaction between both subunits of the heterodimer, crosslinking experiments between cysteine residues in both moieties was tried. Namely, two chemical spacers of different length (10.5 and 14.3 Å) were tested and crosslinking was observed for those mutants with pairing positions between 8 and 17.5 Å. Indeed, specific residues that crosslinked 4F2hc and LAT2 nearly completely (>80%). As a result of the positive results (as compared to the appropriate controls) the idea that 4F2hc-ED almost completely covers the extracellular surface of the transporter subunit LAT2 is reasonable. Moreover, further varied evidences (TEM, SPA and docking experiments) were in line with the obtained results, revealing that the extracellular domain of 4F2hc interacts with LAT2, almost completely covering the extracellular face of the transporter. The interaction of 4F2hc with LAT2 gives insights into the structural bases for light subunit recognition and the stabilizing role of the ancillary protein in HATs. In addition, it has been demonstrated that the ectodomain of 4F2hc suffices the stabilization of the light subunit.
The second goal of the thesis was to find a suitable HAT candidate to perform crystallization trials and posterior structure elucidation, since human 4F2h/LAT2 was not stable enough for this aim. Until now, only the human 4F2hc ectodomain atomic structure has been solved (Fort et al., 2007), and some low sequence amino acid identity prokaryotic homologues of LATs.
In order to identify putative good eukaryotic light subunits for 3D crystallization the adopted GFP‐based Saccharomyces cerevisiae protocol for our transporters resulted successful since it allowed to find three putative good candidate eukaryotic light subunits for 3D crystallization studies. GFP technology allowed quick expression screening, membrane protein-detergent solubilization screening and finally another screening step including assessment of the stability by ultracentrifugation dispersity sedimentation.
Once the candidates selected in the best conditions, further purification was required (size exclusion chromatography) before attempting crystallization. In this sense, further efforts were delivered in order to try to enhance the stability (and minimize aggregation). Thus, addition of lipids in the solubilization step and during protein purification was used to mimic the protein membrane environment and reduce the aggregation. Really interesting is the stabilizing effect that cholesterol has on almost all light subunits tested. This is in concordance with the fact that 4F2hc has been found located in lipid rafts where membrane are rich in cholesterol. Removal of reactive cysteine and generation of truncated versions of the protein in the C and N terminal were introduced to increase the crystallization probability. This work ended with the finding of 3 good candidates for crystallization screenings, and the best candidate was optimized in terms of stability and protein flexibility for crystallization studies. Just preliminary crystallization trials were done.Los transportadores heteroméricos de aminoácidos (HAT) median el transporte de aminoácidos a través de la membrana plasmática. Representan el único ejemplo de transportadores de solutos formado por dos subunidades distintas unidas por un puente disulfuro. Debido a su gran relevancia en fisiología (asociados a aminoacidurias, infección por virus, cáncer,…) el estudio de su estructura-función resulta clave. Debido a su naturaleza, son proteínas difíciles de cristalizar, de las que sólo se conoce la estructura atómica del ectodominio de 4F2hc humano. En este escenario la tesis se centró en la validación del primer modelo 3D a baja resolución de un HAT humano (4F2hc/LAT2), mediante experimentos de crosslinking entre subunidades, e identificación de nuevos candidatos para cristalización 3D. Para ello se seleccionaron 24 subunidades ligeras de distintas especies eucariotas y se testaron en un proceso de selección para determinar el/los mejores candidatos. El primer objetivo de la tesis concluyó con la determinación de residuos concretos en 4F2hc y LAT2 cercanos a una distancia de 3-14 Å mediante la utilización de crosslinkings de cisteínas. Finalmente, tres líneas distintas: el modelo 3D obtenido por microscopía electrónica de transmisión y tinción negativa de partículas individuales (en colaboración con Dr. Fotiadis), los experimentos de crosslinking, y el docking generado en colaboración con Dr. Fdz-Recio, demostraron que 4F2hc-ED cubre, casi completamente, la superficie extracelular de LAT2. Además, se demostró que el ectodominio de 4F2hc es suficiente para estabilizar LAT2. Como resultados del segundo objetivo, tres subunidades ligeras fueron seleccionadas, tras adaptar el protocolo desarrollado por Drew et al.,2008, como mejores candidatas para estudios de cristalización 3D. Posteriormente, distintas estrategias se siguieron para mejorar la estabilidad de la mejor candidata: eliminación de la cisteína reactiva, adición de lípidos a la muestra, cambio de sistema de expresión para aumentar su expresión a células de insecto Sf9. Además, se generaron mutantes delecionados en N y C terminal para reducir su flexibilidad y aumentar la probabilidad de cristalización. Se concluyó en encontrar un buen candidato para estudios de cristalización.
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Janvier-Chemy, Blandine. "Identification d'épitopes B des protéines de core des virus HIV-1 et HIV-2 : application au développement de tests pronostiques." Compiègne, 1990. http://www.theses.fr/1990COMPD326.
Full textAbstract:
L'épitope mapping de 9 ACM dirigés contre la protéine majeure de core du virus HIV- 1, réalisé selon la technique développée par H. M. Geysen et al. A permis l'identification de 4 épitopes distincts de la p24, couvrant Les acides aminés 179-188, 209_217, 260-267 et 281-286 du précurseur gag p55. Les épitopes AAEWDRVHP (209-217) et EIYKRWII (260-267) se sont révélés immuogéniques lors de l'infection naturelle par HIV-1. La détection et le tirage des anticorps anti-AAEWDRVHP lors de l'infection par les virus HIV de types 1 et 2, a été rendue possible grâce à l'utilisation de l'ACM 23A5G4 ~< f reconnaissant la séquence AAEWDRVHP conjugué à la peroxydase dans 2 tests immunoenzymatiques par inhibition séquentielle, SIEIA-l et SIEIA-2. L'étude de sérums de patients se trouvant à différents stades de l'infection par HIV nous a permis de montrer que la séquence 209-217, ou le domaine antigénique couvrant cette séquence, était un épitope majeur des protéines p24 et p26 des virus HIV-1et HIV-2, reconnu à un stade précoce de l'infection virale. L'épitope mapping de 2 ACM humains anti-p24 a permis de localiser 2 épitopes supplémentaires, résidus 144-152 et 292-302 du précurseur gag p55. Un épitope de la protéine de core p17 a également été identifié en position 64-73 du précurseur gag, Les résultats d'épitope mapping des ACM murins et humains ont montré une bonne corrélation avec les programmes de prédictions d'épitopes globalisés ainsi qu'avec les modèles de structure tridimensionnelle des protéines p17 et p24. Deux grandes boucles exposées à la surface de la protéine p24 (190-228 et 257-295) semblent particulièrement immunogènes au cours de l'infection naturelle par HIV-1.
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Gramatica, Andrea. "HIV-1: identification of a pathway for virus release and development of a new nanotechnological strategy to counteract the infection." Doctoral thesis, Humboldt-Universität zu Berlin, Lebenswissenschaftliche Fakultät, 2014. http://dx.doi.org/10.18452/17013.
Full textAbstract:
Das Humane Immundefizienz-Virus 1 (HIV-1) rekrutiert Wirtszellproteine und -Signalwege für seinen eigenen Lebenszyklus und beeinträchtigt viele Funktionen von Immunzellen, darunter die Phagozytose von Pathogenen durch Makrophagen. Diese Schwächung der Immunabwehr verursacht das acquired immunodeficiency syndrome (AIDS).Der Zusammenbau von HIV-1 geschieht sowohl an der Plasmamembran als auch in endosomalen Kompartimenten und wird durch das Struktur-Polyprotein Gag kontrolliert, das auch für die Freisetzung virusähnlicher Partikel (VLPs) aus der Wirtszelle sorgt. Vorangegangene Studie haben gezeigt, dass durch hohe induzierte Calciumkonzentrationen im Zytoplasma die Anzahl an VLPs in endolysosomalen Kompartimenten steigt und die VLP-freisetzung drastisch zunimmt. Der verantwortliche Mechanismus ist jedoch bisher unbekannt. Die vorliegende Arbeit zeigt zunächst, dass Calciumausschüttung aus Lysosomen die Fusion von Endosomen und Lysosomen - und damit die Exozytose der in Lysosomen enthaltenen VLPs - auslöst. Dieser Prozess wird durch Synaptotagmin VII reguliert und verhindert den Abbau eines Teils der in späten Endosomen und Lysosomen eingschlossenen VLPs. Der zweite Teil dieser Arbeit beschreibt die Entwicklung eines nanobiotechnologischen Systems zur Eliminierung von Env/Gag-VLPs (HIV-VLPs) mit Hilfe von Makrophagen. Dieses basiert auf Immunoliposomen, die HIV-VLPs über anti-Env-Antikörper binden und von Makrophagen dank membranständigem Phosphatidylserin (PS), einem apoptotischen Signal, phagozytiert werden. Die Lipososmen imitieren apoptotische Zellen und induzieren ihre Internalisierung und die lysosomale Aufnahme von HIV-VLPs. Das System nutzt einen effizienten Internalisierungsweg, der während der HIV-1-Infektion nicht beeinträchtigt ist. Diese Ergebnisse bieten neue Einblicke in die intrazellulären Prozesse der HIV-1 Freisetzung und präsentieren PS-Immunoliposomen als neuen potentiellen nanomedizinischen Ansatz zur Virusbeseitigung und HIV-Antigenpräsentation.Human immunodeficiency virus 1 (HIV-1) hijacks proteins and signaling pathways of the host cell for its own life cycle, thereby impairing many functions of immune cells, including pathogen-phagocytosis by macrophages. The overall weakening of immune functions eventually results in the development of acquired immunodeficiency syndrome (AIDS). HIV-1 assembly takes place at the plasma membrane as well as in endosomal compartments and is governed by the structural polyprotein Gag, which is also sufficient for the release of virus-like particles (VLPs) from the host cell. It was shown that an induced high cytoplasmic calcium concentration increases the amount of VLPs in endo-lysosomal compartments and results in a dramatic enhancement of VLP release [1,2]. However, the mechanism by which calcium can promote the release of VLPs remains to be determined. The first part of this work shows that release of calcium from lysosomes causes fusion between endosomes and lysosomes as well as exocytosis of lysosomeentrapped VLPs. This mechanism is regulated by Synaptotagmin VII and prevents degradation of a part of the late endosome- and lysosome-entrapped VLPs. The second part focuses on the development of a nanobiotechnological system for the clearance of Env/Gag-VLPs (HIV-VLPs) by macrophages. This system is based on immunoliposomes that (1) bind HIV-VLPs via anti-Env antibodies and (2) are phagocytosed by macrophages due to the presence of phosphatidylserine (PS), an apoptotic signal. Essentially, the PS-liposomes mimic apoptotic cells thereby inducing internalization and lysosomal delivery of bound HIV-VLPs. These immunoliposomes exploit an efficient internalization pathway not impaired upon HIV-1 infection. The results of this thesis provide new insights into the intracellular pathways controlling HIV-1 release and demonstrate that PS-immunoliposomes can represent a novel nanomedical approach for viral clearance and HIV antigen presentation.
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Long, Kimberly Renee. "Identification and Characterization of Agv1, a Pre-Metazoan Arf GAP: A Dissertation." eScholarship@UMMS, 2007. https://escholarship.umassmed.edu/gsbs_diss/339.
Full textAbstract:
Human immunodeficiency virus type 1 (HIV-1) is a member of the lentivirus subfamily of retroviruses. HIV-1 expresses multiple genes from a single provirus by alternative splicing. Early in viral expression, fully spliced 2-kb viral RNA is exported from the nucleus and encodes the viral regulatory protein, Rev, which is essential for nuclear transport of partially spliced and unspliced genomic-length RNA. Rev binds to an RNA structural element called the Rev response element (RRE) and mediates nuclear export through the leucine-rich nuclear export signal (NES) pathway. The human Rev Interacting Protein (hRIP) interacts specifically with the Rev NES. Rev NES mutants that are unable to export Rev-dependent RNAs are also unable to bind to hRIP. The hRIP cDNA encodes a 562 amino acid protein containing an N-terminal zinc finger with homology to Arf GAP domains, a central serine and threonine rich region, and C-terminal phenylalanine-glycine (FG) repeats characteristic of nucleoporins.
To identify an hRIP ortholog in a genetically tractable organism, we performed database searches using the N-terminal zinc finger of hRIP. Using this approach, we identified a novel gene in Schizosaccharomyces pombe. Alignment of the entire reading frame of the putative ortholog with hRIP indicates similarity with the serine/threonine rich region and with the FG repeats, suggesting that S. pombecould be a good model system to study the cellular function of hRIP.
We find that the S. pombe ORF is an essential gene, which encodes a 483 amino acid protein that is also able to interact with the NES of HIV-1 Rev. Based on being an essential gene, and the presence of a putative Arf GAP domain, the ORF was named an Arf GAP essential for viability, agv1+. We show that Agv1 is not directly involved in the nuclear export of poly(A+) RNA or 5S rRNA, nuclear export of leucine-rich NES-containing proteins, or nuclear import of nuclear localization signal (NLS)-containing proteins. However, Agv1 does appear to play a role in the cytoplasmic localization of 5S rRNA.
We demonstrate that loss of Agv1 alters the localization of endoplasmic reticulum (ER) membrane and Golgi membrane resident proteins, accumulates intracellular membrane, and blocks processing of carboxypeptidase Y. Furthermore, the S. cerevisiae ADP-ribosylation factor (Arf) GTPase activating protein (GAP) Glo3, but not a catalytically inactive Glo3 mutant [R59K], is able to partially compensate for the loss of Agv1 function in temperature sensitive strains, indicating that Agv1 is an S. pombe Arf GAP with some functional features similar to S. cerevisiae Glo3.
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Vazquez, Ana Carolina. "Identification and characterization of compounds with antiviral activity against influenza viruses." [Kent, Ohio] : Kent State University, 2008. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=kent1227644336.
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Thesis (Ph.D.)--Kent State University, 2008.Title from PDF t.p. (viewed Dec. 14, 2009) Advisor: Miguel E. Quinones-Mateu. Keywords: biomedical research, cellular biology, molecular biology, virology. Includes bibliographical references (p. 201-228)
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Shehadeh, Nancy. "Ethnic Identification: Its Impact on HIV/AIDS Risk among Migrant and Seasonal Workers in South Florida." FIU Digital Commons, 2012. http://digitalcommons.fiu.edu/etd/722.
Full textAbstract:
Migrant workers are one of the most rapidly growing populations in the United States (U.S.) and have been significantly affected by HIV/AIDS. More than 9 million people in the U.S., primarily concentrated in Texas, Florida, Washington, California, Oregon, and North Carolina, are migrant farm workers. High prevalence rates are also suspected among migrant worker communities where risky health behaviors appear to be common. Constant mobility, isolation, limited education, substandard housing, and poverty are some of the factors that migrant workers experience and in many cases increases their HIV risk. Recent studies have suggested that ethnic identity or the level of attachment with one’s ethnic group may influence engagement in HIV risk behaviors, a fact that may be important in the development of interventions among ethnic minorities. This study assesses the relationship between ethnic identity and HIV risk behaviors in two different samples; one assesses this relationship at baseline with a total of 431 African American migrant and seasonal workers in Immokalee, Florida. The second analyzes changes in ethnic identity and HIV behaviors in a sample of 270 Hispanic and African American migrant and seasonal workers in Immokalee, Florida. Data from baseline and 6-month follow-up were used in the analyses presented. The results suggest that individuals with higher levels of ethnic identity report lower levels of engagement in some, but not all, of the risky behaviors examined. These findings point to a potentially protective role for ethnic identity among this sample.
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Birckhead, Traci M. "For the Sake of His House| The Role of Leadership Development of Lay Leaders of an African American Protestant Church in Enhancing Organizational Identification." Thesis, The George Washington University, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10931942.
Full textAbstract:
This research study examined the organizational identification process of lay church leaders. The organizational identification process is important to all organizations, as it increases an individual’s connection to the organization and thus has the potential to increase attendance, commitment, group behaviors, and the enactment of organizational policies. Lay leaders are key to the success of churches, as the weight of leadership lies within these structured leadership positions of small, established ministries.
One research question guided this study: What is the role of leadership development of lay leaders in enhancing their organizational identification? The research site for this single descriptive case study was First Baptist Church of Highland Park, a large, African American Protestant church located near Washington, DC. Data were collected through interviews with lay leaders of three ministries of this church—deacons, ministers, and trustees—as well as observation and document analysis. Lay leaders were selected as the participants for this study since most roles and positions of leadership within the church involve this volunteer, lay leader base, and these individuals are trained for the ministry through an extensive leadership development process developed and provided by the church.
This study found that (1) the early life of lay leaders made a difference in how the leadership development program affected their organizational identification; (2) the associated ministry roles and responsibilities, as refined in the leadership development process, enhanced their process of organizational identification; (3) the self-esteem of the lay leaders developed through their leader training process and was an important factor in further enhancing their process of organizational identification; and (4) the church’s identity and specific aspects of its mission and values were central in the organizational identification process.
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Gentili, Matteo. "Identification of viral and host mechanisms that determine innate immune activation by the DNA sensor cGAS." Thesis, Sorbonne Paris Cité, 2017. http://www.theses.fr/2017USPCB023.
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Les acides nucléiques sont des activateurs puissants des réponses immunitaires innées. Pour lutter contre les infections, les organismes multicellulaires ont développé de multiples façons de détecter les agents pathogènes. L'une d'entre elles est la reconnaissance de l'ADN dans le cytoplasme. La présence d'ADN dans le cytoplasme est généralement liée à des infections par des bactéries ou des virus comme le VIH. La GMP-AMP synthase cyclique (cGAS) est un capteur cytosolique essentiel de l'ADN chez les mammifères. Lors de la reconnaissance de l'ADN, cGAS synthétise le petit second messager cyclique GMP-AMP (cGAMP), qui active les voies de signalisation de l’immunité innée. De plus, cGAS joue un rôle central en réponse aux microbes et au développement de maladies auto-immunes comme les interféronopathies. Cette thèse examine le rôle de cGAS en réponse aux virus et à l’ADN du soi. En explorant la réponse médiée par cGAS en présence du VIH, nous avons constaté que dans les cellules produisant du virus, le cGAMP synthétisé par cGAS est contenu dans des particules virales et des vésicules extracellulaires. Les particules virales délivrent efficacement cGAMP aux cellules cibles et activent une réponse antivirale puissante. Le transfert de cGAMP nécessite une activité catalytique de cGAS et une fusion des particules virales avec les cellules cibles. Nous avons également montré que dans le contexte d'une infection, les virus à ADN tels que MVA et mCMV peuvent conditionner cGAMP. Par conséquent, le transfert de cGAMP médié par une infection virale est un mécanisme de défense généralisé du système immunitaire inné. Nous avons également étudié l'activation du cGAS par l'ADN en se concentrant sur l'ADN de la chromatine nucléaire. Dans les cellules eucaryotes, l'ADN est compartimenté dans deux organelles: le noyau et les mitochondries. La compartimentation nucléaire de l'ADN peut être transitoirement perdue lors de la rupture de l'enveloppe nucléaire pendant la division cellulaire et lors des ruptures d'enveloppes nucléaires dans les cellules dendritiques migrantes (DC). Nous avons constaté que la rupture de l'enveloppe nucléaire ou la perte de l'intégrité de l'enveloppe nucléaire entraînent un recrutement de cGAS sur l'ADN nucléaire. Nous avons également montré que les DC présentent un pool nucléaire de cGAS à l'état basal. La rupture de l'enveloppe nucléaire pendant le cycle cellulaire conduit au recrutement de cGAS à l'ADN nucléaire. En jouant sur la localisation cGAS, nous avons montré que le cGAS est peu actif lorsqu'il se trouve dans le noyau et la transfection d'ADN exogène permet son activité enzymatique complète. L'expression du cGAS localisé dans le noyau des DC a conduit à la maturation des DC et à la production d'interféron de type I (IFN). La région N-terminale de cGAS régule la distribution nucléaire / cytoplasmique de cGAS dans les cellules en interphases et nous avons établi que les lamines A / C sont requises pour l'accès de cGAS à l'ADN nucléaire dans les DC migrantes. De façon inattendue, nous montrons que cGAS n'est pas activé lors de la rupture de l'enveloppe nucléaire. Enfin, nous identifions l'hétérochromatine pericentromérique comme étant l'ADN de liaison préférentielle pour le cGAS nucléaire exprimé dans les DC. Au total, notre étude montre que le noyau agit comme un site intracellulaire immunitaire privilégié pour l'activation de cGAS. Collectivement, nous avons montré une pertinence de l'activation de cGAS lors d'une infection virale et découvert un mécanisme « cheval de Troie » de l'incorporation de cGAMP dans la progénie virale. Nous avons également décrit des mécanismes, encore à définir, qui limitent les réponses à l'ADN de la chromatine dans le noyau. À mesure que les preuves augmentent sur la prise en charge de cGAS en réponse à des agents pathogènes, dans des maladies auto-immunes, en réponse aux dommages causés par l'ADN et dans le développement de la sénescence cellulaire, (...)Nucleic acids are potent activators of innate immune responses. To control infections, multicellular organisms have developed multiple ways to detect pathogens. One of these is the recognition of DNA in the cytoplasm. Presence of DNA in the cytoplasm is usually linked to infections by bacteria or viruses, such as HIV. Cyclic GMP-AMP synthase (cGAS) is an essential cytosolic sensor for DNA in mammals. Upon DNA recognition, cGAS synthetizes the small second messenger cyclic GMP-AMP (cGAMP), which activates innate immune signaling pathways. cGAS plays a pivotal role in response to microbes and in the development of autoimmune diseases such as interferonopathies. This thesis investigate the role of cGAS in response to viruses and to self DNA. By exploring the cGAS-mediated response to HIV, we found that in cells producing virus, cGAS-synthesized cGAMP is packaged in viral particles and extracellular vesicles. Viral particles efficiently deliver cGAMP to target cells and activate a potent antiviral response. The transfer of cGAMP requires cGAS catalytic activity and fusion of the viral particles with the target cells. We also showed that in the context of an infection, DNA viruses such as MVA and mCMV can package cGAMP. Therefore viral mediated cGAMP transfer is a generalized defense mechanism of the innate immune system. We further investigated cGAS activation by DNA focusing our attention on nuclear chromatin DNA. In eukaryotic cells, DNA is compartmentalized in two organelles: the nucleus and the mitochondrion. Nuclear compartmentalization of DNA can be transiently lost upon nuclear envelope breakdown during cell division and upon nuclear envelope ruptures in migrating dendritic cells (DCs). We found that nuclear envelope breakdown or loss of nuclear envelope integrity leads to cGAS recruitment on the nuclear DNA. We also showed that DCs present with a nuclear pool of cGAS at steady state. Nuclear envelope breakdown during cell cycle leads to cGAS recruitment to the nuclear DNA. By manipulating cGAS localization, we showed that cGAS is poorly active when in the nucleus, and that providing exogenous DNA unmasked its full enzymatic activity. Expression of nuclear localized-cGAS in DCs leads to DCs maturation and Type I interferon (IFN) production. The N-terminal region of cGAS regulates cGAS nuclear/cytoplasmic distribution in interphase cells and we established Lamin A/C as required for cGAS accessibility to nuclear DNA in migrating DCs. Unexpectedly, we show that cGAS is not activated upon nuclear envelope rupture. Finally, we identify the pericentromeric heterochromatin as the preferential binding DNA for expressed nuclear cGAS in DCs. Altogether, our study shows the nucleus to act as an intracellular immune-privileged site for the activation of cGAS. Collectively, we have shown relevance of cGAS activation upon viral infection and uncovered a Trojan horse mechanism of cGAMP incorporation in the viral progeny. We have also described yet to be defined regulatory mechanisms that limit responses towards chromatin DNA in the nucleus. As evidence grows for cGAS involvement in response to pathogens, in autoimmune diseases, in response to DNA damage and in the development of cell senescence, fully understanding the mechanisms of distinction between self and pathogen related DNA by the sensor will be important to develop effective means to regulate the pathway
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Bonaventure, Boris. "Cribles CRISPR/Cas9 à l'échelle du génome : identification de l'hélicase à ARN DDX42 comme nouvel inhibiteur viral." Thesis, Montpellier, 2020. http://www.theses.fr/2020MONTT035.
Full textAbstract:
L’utilisation de cribles à l’échelle du génome basé sur la technologie CRISPR/Cas9 est une approche puissante, permettant l’identification de nouveaux facteurs impliqués dans des phénotypes d’intérêt. Afin d’identifier de nouveaux inhibiteurs du VIH-1, nous avons développé un crible CRISPR/cas9 à l’échelle du génome en utilisant l’IFN de type I pour créer un environnement cellulaire hostile à la réplication virale. Ce crible nous a permis d’identifier DDX42, membre de la famille des hélicases à motif DEAD, comme inhibiteur intrinsèque du VIH-1. Les cellules invalidées pour DDX42 deviennent plus susceptibles à l’infection par le VIH-1, indépendamment du système IFN. Cette activité antivirale de la protéine DDX42 endogène a été observée en lignées modèles, mais aussi en lymphocytes T CD4 primaires et en macrophages dérivés de monocytes, cellules cibles du VIH-1. Par ailleurs, l’expression ectopique de DDX42 inhibe efficacement l’infection par le VIH-1. L’effet positif de la déplétion de DDX42 sur l’infection du VIH-1 est directement corrélé à une augmentation de l’accumulation d’ADN viral, suggérant que DDX42 affecte la transcription inverse ou la stabilité du génome. De façon intéressante, des essais de ligation de proximité ont montré que DDX42 était dans l’environnement proche de la Capside du VIH-1 en macrophages primaires infectés. L’activité antivirale de DDX42 affecte également d’autres lentivirus que le VIH-1, ainsi que le rétrovirus MLV et les rétrotransposons de type LINE-1. Enfin, nous démontrons que DDX42 est capable d’inhiber l’infection par d’autres virus tels que les alphavirus, les flavivirus et les coronavirusGenome-wide CRISPR/Cas9 knock-out genetic screens are powerful approaches to unravel new regulators of various cellular processes, including viral replication cycles. With the aim of identifying new cellular inhibitors of HIV-1, we have developed a whole-genome CRISPR/Cas9 knock-out screen using the ability of type I interferon to potently inhibit HIV-1 infection in order to create a cellular environment hostile to HIV-1 replication. This screen led to the identification of the DEAD-box RNA helicase, DDX42, as an intrinsic inhibitor of HIV-1. Cells depleted for DDX42 became more susceptible to HIV-1 infection both in the presence and in the absence of interferon. The ability of endogenous DDX42 to potently inhibit HIV-1 infection was observed in cell lines and in physiological targets of HIV-1, primary CD4+ T cells and monocyte-derived macrophages. Similarly, DDX42 overexpression potently inhibited HIV-1 infection. The positive impact of DDX42 depletion on HIV-1 infection was directly correlated to a substantial increase in viral DNA accumulation, pinpointing an inhibitory role of DDX42 on reverse transcription. Within cells, DDX42 was found primarily in the nucleus but was also present in the cytoplasm. Interestingly, proximity ligation assays showed that DDX42 was in the close vicinity of HIV-1 Capsid during infection of primary monocyte-derived macrophages. Investigation on DDX42 specificity showed that different lentiviruses, the retrovirus MLV and the retrotransposition of LINE-1 were inhibited by DDX42. Finally, we reveal that DDX42 was able to decrease infection with a variety of viruses including alphaviruses, flaviviruses and coronaviruses
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Younan, Patrick. "Identification and Characterization of SNAPIN as a Novel Antagonist of HIV-1 Egress: A Dissertation." eScholarship@UMMS, 2010. https://escholarship.umassmed.edu/gsbs_diss/460.
Full textAbstract:
Vpu has been shown to possess two distinct roles in the pathogenesis of HIV. First, Vpu has been shown to down-regulate the expression of CD4 molecules at the plasma membrane of infected cells by targeting CD4 molecules for degradation in the endoplasmic reticulum. Second, Vpu promotes viral egress in specific cell lines termed non-permissive cells by mechanism that remain relatively unclear.
Therefore, experiments were conducted in order to identify cellular factors involved in the Vpu-dependent phenotype. Using full-length Vpu as bait in yeast two-hybrid experiments, several candidate cellular factors were identified. One protein, SNAPIN, was identified as a cellular factor putatively involved in the Vpu-dependent phenotype. Further experiments determined that not only do SNAPIN and Vpu interact, but that Vpu also leads to the degradation of SNAPIN by both proteasomal and lysosomal degradation pathways.
Over-expression of SNAPIN in cell lines that do not normally require Vpu expression for viral production resulted in a Vpu-dependent phenotype. While over-expression of SNAPIN in otherwise permissive cell lines significantly reduced Vpu-deficient virus production, wild type levels remained relatively constant. Importantly, no defective viral structural protein production was observed; however, intracellular p24/p55 did not accumulate suggesting that in SNAPIN expressing cells, Gag is also targeted for degradation.
In addition, the reduction of SNAPIN expression in non-permissive cell lines significantly increased viral titers in supernatants. Of particular interest, even in cells expressing Bst-2 (a previously identified cellular factor involved in the Vpu-phenotype), siRNA mediated knockdown of SNAPIN led to increased viral titers. In addition, the co-transfection of siRNAs targeting both SNAPIN and Bst-2 resulted in an additive effect, in which Vpu-deficient viral titers were nearly equivalent to wild-type titers. Surprisingly, siRNA-mediated knockdown of SNAPIN in Jurkat cells was sufficient to overcome any restriction in viral egress imposed by the deletion of Vpu. Conversely, siRNA targeting Bst-2 had little or no effect on viral titers in Jurkat cells regardless of whether it was transfected alone or in combination with siRNAs targeting SNAPIN.
These experiments provide evidence of an alternate cellular restriction mechanism involved in viral egress that is countered by the HIV-1 accessory protein, Vpu. In addition, this research may provide further insight into the complex cellular networks involved in the trafficking of Gag through cellular endosomal pathways.
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Eloraby, Ghada. "Identification of protein targets of nevirapine reactive metabolites using click chemistry and mass spectrometry-based differential proteomics." Mémoire, Université de Sherbrooke, 2016. http://hdl.handle.net/11143/8995.
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Abstract : Adverse drug reactions (ADRs) are undesirable effects caused after administration of a single dose or prolonged administration of drug or result from the combination of two or more drugs. Idiosyncratic drug reaction (IDR) is an adverse reaction that does not occur in most patients treated with a drug and does not involve the therapeutic effect of the drug. IDRs are unpredictable and often life-threatening. Idiosyncratic reaction is dependent on drug chemical characteristics or individual immunological response. IDRs are a major problem for drug development because they are usually not detected during clinical trials.
In this study we focused on IDRs of Nevirapine (NVP), which is a non-nucleoside reverse transcriptase inhibitor used for the treatment of Human Immunodeficiency Virus (HIV) infections. The use of NVP is limited by a relatively high incidence of skin rash. NVP also causes a rash in female Brown Norway (BN) rats, which we use as animal model for this study. Our hypothesis is that idiosyncratic skin reactions associated with NVP treatment are due to post-translational modifications of proteins (e.g., glutathionylation) detectable by MS. The main objective of this study was to identify the proteins that are targeted by a reactive metabolite of Nevirapine in the skin.
The specific objectives derived from the general objective were as follow:
1) To implement the click chemistry approach to detect proteins modified by a reactive NVP-Alkyne (NVP-ALK) metabolite. The purpose of using NVP-ALK was to couple it with Biotin using cycloaddition Click Chemistry reaction.
2) To detect protein modification using Western blotting and Mass Spectrometry techniques, which is important to understand the mechanism of NVP induced toxicity.
3) To identify the proteins using MASCOT search engine for protein identification, by comparing obtained spectrum from Mass Spectrometry with theoretical spectrum to find a matching peptide sequence.
4) To test if the drug or drug metabolites can cause harmful effects, as the induction of oxidative stress in cells (via protein glutathionylation). Oxidative stress causes cell damage that mediates signals, which likely induces the immune response.
The results showed that Nevirapine is metabolized to a reactive metabolite, which causes protein modification. The extracted protein from the treated BN rats matched 10% of keratin, which implies that keratin was the protein targeted by the NVP-ALK.Résumé : Les effets indésirables (EI) sont les effets indésirables causés après l'administration d'une dose unique ou une administration prolongée du médicament ou le résultat de la combinaison de deux médicaments ou plus. La Réaction idiosyncratique (IDR) est une réaction indésirable qui ne se produit pas dans la plupart des patients traités avec un médicament et qui ne comporte pas l'effet thérapeutique du médicament. IDR sont imprévisibles et peuvent mettre la vie du malade en danger. Cette réaction dépend des caractéristiques chimiques du médicaments et/ou de la réponse immunitaire individuelle du patient. IDR est un problème majeur pour le développement de médicaments car ils ne sont généralement pas détectés au cours des essais cliniques. Dans cette étude, nous nous sommes concentrés sur la Réaction idiosyncratique de névirapine (NVP) qui est un inhibiteur de transcriptase inverse non nucléosidique utilisé pour le traitement du virus d'immunodéficience humaine (VIH). L'utilisation de NVP est limitée par une incidence relativement élevée d'éruption cutanée. NVP provoque également une éruption cutanée chez les rats femelles de souche Brown Norway. Notre étude vise à mieux comprendre les IDRs induites par l'administration de NVP chez l'animal. La présente étude vise à vérifier l'hypothèse que les problèmes cutanés associés à la prise de NVP soient attribuables à la modification post-traductionnelle de protéines détactable par spectrométrie de masse. Les principaux objectifs de ce projet étaient : 1) Déterminer si la Nevirapine alcynes (NVP-ALK), un analogue de la NVP peut développer la même éruption cutanée que la NVP. La NVP-ALK a été couplé avec de la biotine en utilisant la réaction chimique (click chemistry). 2) Détecter les modifications post-traductionelles des proteines par Western blot et des techniques de spectrométrie de masse, pour comprendre le mécanisme de la toxicité induite par la NVP. 3) Identifier les protéines modifiées en utilisant le moteur de recherche MASCOT pour l'identification des protéines, en comparant le les spectres de masse obtenus avec les spectres théoriques pour trouver une séquence correspondante de peptide. 4) Tester si la NVP et ses métabolites peuvent provoquer des effets nocifs, comme l'induction d'un stress oxydatif dans les cellules (par la mesure de la glutathionylation des protéines). Les résultats ont montré que la névirapine est métabolisé en métabolite réactif ce qui provoque une modification de la kératine. Ainsi nos résultats suggèrent que la kératine est la cible des métabolites de la NVP-ALK.
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Paoletti, Audrey. "Étude des étapes précoces de l’infection par le VIH-1 : identification d’un nouveau point de contrôle immunitaire immunitaire impliquant le récepteur P2Y2 et la protéine NLRP3." Thesis, Université Paris-Saclay (ComUE), 2015. http://www.theses.fr/2015SACLS240.
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Plus de 34 millions de personnes dans le monde vivent avec le virus de l’immunodéficience humaine de type 1 (VIH-1). Cette pandémie est partiellement contrôlée par l’utilisation de combinaison d’agents antirétroviraux spécifiques. Cependant l’émergence de nouvelles souches virales multi-résistantes nécessite le développement de nouvelles stratégies antirétrovirales. Notre laboratoire porte une attention particulière à la compréhension des évènements cellulaires et viraux impliqués dans les étapes précoces du cycle réplicatif du VIH-1. Récemment, nous avons révélé l’existence d’une nouvelle voie de signalisation cellulaire, impliquant un canal membranaire (la Pannexine-1), un signal de danger (l’ATP extracellulaire) et un récepteur purinergique (P2Y2) participant à l’entrée du VIH-1 dans ses cellules cibles. Etant donnée que ces trois évènements cellulaires sont également des acteurs de la réponse immunologique, nous avons décidé de poursuivre l’étude du rôle des protéines de la réponse immunologique innée dans les étapes précoces d’infection par le VIH-1.Au cours de mes travaux de thèse, nous avons révélé une interaction entre le récepteur purinergique P2Y2 et la protéine de l’inflammasome NLRP3. Dans un premier temps, nous avons démontré que la migration cellulaire dépendante du récepteur purinergique P2Y2 est réprimée pendant l’activation de l’inflammasome NLRP3. A l’inverse, nous avons également observé que la polarisation des macrophages, la sécrétion de l’interleukine-1β et la pyroptose déclenchées par l’activation de NLRP3 sont sous le contrôle de l’autophagie induite par l’activation du récepteur purinergique P2Y2. Ces résultats suggèrent que l’interaction entre NLRP3 et P2Y2 constitue un nouveau point de contrôle immunologique qui régule les fonctions des macrophages. A la suite de ces travaux, nous avons analysé le rôle de ce point de contrôle immunologique lors de l’infection par le VIH-1 et avons démontré que l’activation de l’inflammasome NLRP3 empêche l’activation de la voie de signalisation purinergique qui implique l’ATP, la pannexine-1 et le récepteur P2Y2, et qui permet l’entrée du VIH-1 dans ses cellules cibles. Nos travaux de recherche mettent ainsi en lumière la capacité de l’inflammasome NLRP3 à représenter un nouveau facteur de restriction inductible du VIH-1. L’ensemble de mes travaux recherche souligne l’existence au niveau des macrophages d’un nouveau point de contrôle du système immunitaire impliquant la protéine NLRP3 et le récepteur P2Y2 et qui peut être moduler afin de développer de nouvelles approches thérapeutiques permettant de lutter contre l’émergence de virus résistants aux traitements rétroviraux classiquesIn 3 decades infection with the virus of the human immunodeficiency of type 1 (HIV-1) caused over than 34 million deaths and the surge of new multiresistant virus strains require the development of novel antiretroviral strategies.Our laboratories revealed a new signaling pathway involving in the early step of HIV-1 infection, involving a hemichannel (Pannexin-1), a common danger signal (extracellular ATP) and a purinergic receptor (P2Y2). These three cellular events are also players in the immune response; we decided to continue the study of proteins involved in the innate immune response during the early stages of infection by HIV -1.Here we demonstrated during this work a new interaction between the purinergic receptor P2Y2 and protein of the inflammasome NLRP3. We demonstrate that P2Y2-stimulated migration of macrophages is inhibited by NLRP3 inflammasome activation. Conversely, NLRP3-dependent macrophage polarization, interleukin-1 β secretion and pyroptosis are under the control of P2Y2-induced autophagy.Finally, the results suggest that the interaction between NLRP3 and P2Y2 is a new immunological checkpoint that regulates macrophage functions. Following this work, we analyzed the role of this immunological control during infection by HIV -1 and have demonstrated that activation of the inflammasome NLRP3 prevents the activation of the purinergic signaling channel involving ATP, pannexin -1 and the P2Y2 receptor, and which allows the entry of HIV -1 in its target cells. Our research and bring to light the capacity of the NLRP3 inflammasome to represent a new inducible restriction factor of HIV-1.All of this research work highlights the existence in macrophages of a new immune system checkpoint involving NLRP3 protein and P2Y2 receptor and can be modulated in order to develop new therapeutic approaches to fight against the emergence of viruses resistant to conventional retroviral treatments
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Dupont, Maeva. "Identification of novel factors involved in the exacerbation of HIV-1 infection and spread among macrophages in the tuberculosis context." Thesis, Toulouse 3, 2019. http://www.theses.fr/2019TOU30211.
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Mycobacterium tuberculosis (Mtb), la bactérie responsable de la tuberculose (TB), et le virus de l'immunodéficience humaine (VIH-1), l'agent du syndrome de l'immunodéficience acquise (SIDA), accélèrent leurs progressions mutuelles chez les patients co-infectés. Alors que de nombreuses données cliniques rapportent une augmentation de la charge virale dans les sites anatomiques co-infectés, les mécanismes qui en sont responsables restent insuffisamment décrits. Mtb cible principalement les macrophages. Nous émettons l'hypothèse que l'infection des macrophages par Mtb créé un microenvironnement propice à la réplication du VIH-1 au niveau des sites co-infectés. Pour le montrer, j'ai utilisé un modèle in vitro précédemment établi par mes équipes (le cmMTB - pour " conditioned media of Mtb-infected macrophages "). Celui-ci permet de mimer un environnement tuberculeux, par la différenciation et l'activation des macrophages vers un profil " M(cmMTB) ", largement retrouvé dans les poumons lors d'une tuberculose active. En rejoignant le laboratoire, j'ai participé à l'étude des mécanismes responsables de l'augmentation de la réplication virale dans le contexte de co-infection, en utilisant ce modèle. Nous avons trouvé que les M(cmMTB) forment de nombreux nanotubes (ponts intercellulaires), leur permettant de transférer plus de virus d'un macrophage à l'autre, et conduit à une forte augmentation de la production virale. L'objectif principal de ma thèse a donc été d'identifier, dans un contexte tuberculeux, de nouveaux facteurs impliqués dans l'augmentation de la réplication du VIH-1 dans les macrophages. Pour cela, une analyse transcriptomique des M(cmMTB) a été réalisée, révélant deux facteurs essentiels : le récepteur Siglec-1 et les interférons de type I (IFN-I) via STAT1. Dans un premier temps, j'ai étudié le rôle de Siglec-1 dans la synergie entre Mtb et le VIH-1 dans les macrophages. D'abord, j'ai montré que son expression de surface était augmentée par le cmMTB, de façon dépendante des IFN-I. Ensuite, j'ai établi que l'abondance des macrophages alvéolaires exprimant Siglec-1 chez les primates non-humains co-infectés avec Mtb et le virus de l'immunodéficience simienne corrélait avec la sévérité de la pathologie, et était associée à la signalisation des IFN-I, via l'activation de STAT1. De plus, j'ai identifié une nouvelle localisation de Siglec-1 le long d'un sous-type de nanotubes.[...]Mycobacterium tuberculosis (Mtb), the bacteria causing tuberculosis (TB), and the human immunodeficiency virus type 1 (HIV-1), the etiological agent of acquired immunodeficiency syndrome (AIDS), act in synergy to exacerbate the progression of each other in co-infected patients. While clinical evidence reveals a frequent increase of the viral load at co-infected anatomical sites, the mechanisms explaining how Mtb favours HIV-1 progression remain insufficiently understood. Macrophages are the main target for Mtb. Their infection by the bacilli likely shapes the microenvironment that favours HIV-1 infection and replication at sites of co-infection. To address this issue, I took advantage of an in vitro model mimicking the TB-associated microenvironment (cmMTB, "conditioned media of Mtb-infected macrophages") previously established in the laboratory; a model that renders macrophages susceptible to intracellular pathogens like Mtb. Upon joining the team, I participated in the study on how Mtb exacerbates HIV-1 replication in macrophages, using this model. We found that cmMTB-treated macrophages (M(cmMTB)) have an enhanced ability to form intercellular membrane bridges called tunneling nanotubes (TNT), which increase the capacity of the virus to transfer from one macrophage to another, leading to the exacerbation of HIV-1 production and spread. The principal objective of my PhD thesis was to identify novel factors that are involved in the exacerbation of HIV-1 replication in macrophages in the context of tuberculosis. To this end, a transcriptomic analysis of M(cmMTB) was conducted, and revealed two key factors: the Siglec-1 receptor and type I interferon (IFN-I)/STAT1 signaling. The first part of my PhD thesis dealt with the characterization of Siglec-1 as a novel factor involved in the synergy between Mtb and HIV-1 in macrophages. First, I demonstrated that its increased expression in M(cmMTB) was dependent on IFN-I. Second, in Mtb and simian immunodeficiency virus co-infected non-human primates, I established a positive correlation between the abundance of Siglec-1+ alveolar macrophages and the pathology, associated with the activation of the IFN-I/STAT-1 pathway. [...]
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Ahuka, Mundeke Steve. "Identification et caractérisation moléculaires des rétrovirus simiens et évaluation du risque de transmission à l'homme en Afrique Centrale." Thesis, Montpellier 1, 2011. http://www.theses.fr/2011MON1T002.
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De nombreux primates d'Afrique sont infectés par les SIV et SLTV en particulier par ceux reconnus comme les ancêtres du VIH et HTLV à l'origine de graves épidémies chez l'homme. Des humains en Afrique continuent d'être exposés à ces virus lors des activités liées à la chasse. Ainsi le risque de transmissions inter-espèces des rétrovirus des primates aux humains persiste toujours dans cette région. Nous avons montré que le mangabey agile est infecté par un SIVagi phylogénétiquement très proche du SIVrcm infectant le mangabey à collier blanc au Cameroun. Nous avons montré aussi que les SIVdeb infectant les cercopithèques de Brazza se regroupent phylogénétiquement selon leurs régions géographiques d'origine non seulement à travers l'Afrique Equatoriale mais aussi à l'intérieur même du Cameroun. Nous avons adapté et validé un outil sérologique (Luminex) qui permet de tester près de 34 antigènes SIV/HIV simultanément. Cet outil et d'autres nous ont permis ensuite de documenter une prévalence globale élevée d'infection SIV et STLV chez les singes chassés pour la viande de brousse en RDC, particulièrement chez les espèces les plus consommées. De nouvelles lignées de SIV et STLV ont aussi été décrites. Par ailleurs, nous avons montré pour la première fois que les fécès peuvent être utilisés pour la détection des STLV chez les bonobos qui sont naturellemnt infectés par les STLV-2 et 3. En revanche, aucune évidence d'infection SIV chez les bonobos n'a été observée. Les travaux de cette thèse contribuent à l'amélioration des connaissances sur les infections rétrovirales chez les primates non humains au Cameroun et en RDC, complètent les informations disponibles sur les réservoirs du VIH-1 et HTLV et enfin fournissent des éléments d'appréciation du risque de transmission de ces virus à l'homme en RDC et au CamerounSIVs and SLTVs infecting apes and monkeys in Africa are the progenitors of HIV and HTLV. Numerous African non-human primates are infected with SIV and STLV and humans continue to be exposed to these viruses by hunting and handling of primate bushmeat. Therefore the risk of cross-species transmissions from primates to humans is still persistent. We showed that SIVagi infecting captive agile mangabey is most closely related to SIVrcm from a wild-caught red capped mangabey from Cameroon. We observed also phylogeographic clustering among SIVdeb strains from Cameroon, DRC and Uganda, but also among distinct areas in Cameroon. We adapted and evaluated a novel high troughput immune assay that included 34 different HIV and SIV antigens in a single well. Using this tool and others, we found a high SIV and STLV prevalence especially among the most hunted monkeys in DRC. We identified also new SIV and STLV lineages. On the other hand, we did not find any evidence of SIV infection in bonobos. However, we showed, for the first time, that fecal samples could be used to detect STLV infection in bonobos that are naturally infected with STLV-2 and 3. The results obtained during this thesis contribute to the improvement of our knowledge on retroviral infections in nonhuman primates from Central Africa, complete information on HIV and HTLV reservoirs and provide background information on human transmission risk of these infections in central Africa especially in DRC
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Ligot, Damien. "Trans-musicalité « taike » : Distinction d’une nouvelle « taiwanité » au sein d'un underground local (1990-2010)." Thesis, Lyon 3, 2012. http://www.theses.fr/2012LYO30022/document.
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Dans les années 1990 et 2000, la scène musicale underground taiwanaise a vu naître successivement le Taik – ou Rock Taike 台客搖滾 taike yaogun – puis le Taiwan Traditional Rap, ou 台灣味唸歌 Taiwan wei niange. Ces courants « trans-musicaux » se rejoignent sur de nombreux points tels que la revendication de « traditions » populaires locales, accompagnée cependant d'une forte complaisance au métissage culturel, et par-dessus tout d'un besoin de se définir – de s'identifier – en dehors des clichés aliénants du bon-goût, dictés de manière hégémonique par la culture « dominante » centrée depuis la fin des années 1940 sur le modèle de la République de Chine. Appuyée par un travail de terrain réalisé entre 2005 et 2010 selon les principes de l'observation participante, et pensée à la lumière des cultural studies et d'ouvrages d'auteurs tels que Dick Hebdige et Stuart Hall, cette thèse propose une approche « sensible » d'une sous-culture particulière, tempérée d'autre part par une critique des théories développées par Pierre Bourdieu dans La Distinction, Critique sociale du jugement. Elle tente ainsi – au travers du prisme trans-musical – de définir la place occupée par la sous-culture locale « Taike » dans l'histoire globale d'autres sous-cultures comparables, et s'impose alors en contexte tel un trait d'union entre Taiwan et le reste du mondeIn the 1990's and 2000's, the underground music scene in Taiwan successively gave birth to the Taik – also called Taike Rock 台客搖滾 taike yaogun – then to Taiwan Traditional Rap 台灣味唸歌 Taiwan wei niange. These "trans-musical" currents are similar in many issues such as the claim to local folk traditions, accompanied however by accomodating a strong cultural mix and by a need to define – to identify – themselves outside the alienating good taste, so dictated since the late 1940's by the hegemonic "dominant" culture focused along the lines of the Republic of China. This thesis proposes a ''sensitive'' approach to a particular subculture, furthermore tempered by a critique of theories developed by Pierre Bourdieu in Distinction: A Social Critique of the Judgment of Taste. It has been backed up by field studies conducted between 2005 and 2010 according to the principles of the participant-observer and trends of thought enlightened by cultural studies and works of authors such as Dick Hebdige and Stuart Hall. It attempts also – through the prism of trans-musicality – to define the place of the local "Taike" subculture in the global history of other comparable subcultures, as an essentiel link between Taiwan and the rest of the world
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Morphet, Marilynn Norma. "Method for identification of effective first-line treatment for HAART naïve HIV/AIDS patients." Thesis, Queensland University of Technology, 2002.
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Picard, Léa. "Identification of Novel Viral Interacting Proteins through the Detection of Genetic INNovations (DGINN) combined with functional assays." Thesis, Lyon, 2020. http://www.theses.fr/2020LYSEN031.
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L'identification de protéines cellulaires qui interfèrent avec la réplication virale est un enjeu majeur en virologie. Parmi ces protéines, la découverte de celles engagés dans des interactions hôte-virus à long terme et en co-évolution est d'intérêt particulier. Chez l'hôte, la présence de pressions sélectives de ce type se traduit par l'apparition de différentes innovations génétiques, telles que la sélection positive sur sites spécifiques, la variation du nombre de copies du gène, la recombinaison, etc. Selon l'hypothèse que des innovations génétiques dans le cadre de l'immunité innée apparaîtraient plus particulièrement dans les protéines interagissant avec les protéines virales, nous avons développé un pipeline permettant de récupérer les séquences orthologues, de les aligner et de reconstruire leur phylogénie, suivi par la détection d'innovations génétiques. Cette procédure simplifiée permet pour la première fois de combiner la détection de gènes paralogues, de points de recombinaison, et de signatures de sélection positive avec plusieurs méthodes fréquemment utilisées. Nous avons validé ce pipeline évolutionnaire et prédictif sur des gènes dont les profils de sélections étaient déjà connus. Ensuite, nous avons analysés deux jeux de données de gènes candidats. Le premier comportait 56 gènes dont la diminution d'expression impactait la réponse interféron à l'infection virale. Le second était composé de 60 gènes surexprimés dans des macrophages résistants à l'infection par le VIH. Nous avons identifié plusieurs gènes présentant d'importantes marques de conflit génétique, et qui encodent donc potentiellement de nouvelles protéines interagissantes avec les protéines virals. Deux de ces candidats sont en cours de caractérisation fonctionnelle pour leur rôle dans le cycle réplicatif du VIH, et d'autres sont en attente d'être étudiés plus avant. Au final, nous avons développé un pipeline complet et extrêmement flexible, disponible au public, capable d'analyser de grands jeux de données et permettant aux chercheurs et chercheuses de classer leurs gènes candidats afin de hiérarchiser leurs expérimentations fonctionnellesThe identification of cellular proteins that interfere with virus replication is a key challenge in virology. Amongst them, finding those engaged in long-term virus-host interaction and co-evolution is of particular interest. In the host, such selective pressures induce diverse genetic innovations, such as site-specific positive selection, gene copy number variation, recombination, etc. Under the hypothesis that genetic innovations in innate immunity may particularly occur in viral interacting proteins, we developed a pipeline for retrieving orthologous sequences, aligning them and reconstructing their phylogeny, followed by the detection of genetic innovations. This streamlined procedure uniquely allows for the detection of paralogous genes, recombination breakpoints, and signatures of positive selection with several widely-used methods. We validated this evolutionary and predictive pipeline on genes with known selection profiles. Furthermore, we screened two datasets of candidate genes. The first one was composed of 56 genes which knock-downs impact the interferon response to viral infection. The second one was composed of 60 genes upregulated in macrophages resistant to HIV infection. We found numerous genes presenting important marks of genetic conflict, thus potentially encoding for novel Viral Interacting Proteins. Two of these candidates are undergoing functional characterization for their role in the HIV replicative cycle, and others are pending further investigation. Overall, we designed a complete and highly-flexible pipeline, available to the public, that can screen large datasets and allow researchers to rank candidate genes in order to prioritize their wet-lab experiments
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Truter, Jan Hendrik Lodewyk. "'n Teologies-empiriese ondersoek na die rol wat gemeentebou in die beplanning en oprigting van kerkkomplekse binne die Nederduitsch Hervormde Kerk van Afrika die afgelope tien jaar gespeel het." Diss., Pretoria : [s.n.], 2005. http://upetd.up.ac.za/thesis/available/etd-05232006-162503/.
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Gray, Laura. "Stéréotypes liés à l’activité physique et la fatigue chez les personnes vivant avec le VIH : une identification des mécanismes psychologiques et physiologiques impliqués." Thesis, Université Côte d'Azur (ComUE), 2018. http://www.theses.fr/2018AZUR4217/document.
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L'activité physique (AP) aurait de nombreux effets bénéfiques chez les personnes vivant avec le VIH (PVVIH), notamment sur la fatigue, l'un des principaux symptômes et effets secondaires des traitements dans cette population. Néanmoins, les PVVIH restent insuffisamment actives au regard des recommandations. En s’appuyant sur des théories contemporaines des stéréotypes (e.g., Stereotype Embodiment Theory, Levy, 2009; Stereotype Threat Theory, Steele & Aronson, 1995) et sur une approche intégrative de la fatigue (Kluger et al., 2013), ce travail doctoral a postulé que l’endossement ou l’activation de stéréotypes liés à l’exercice influenceraient le niveau d’AP des PVVIH ainsi que leur performance lors de tâches fatigantes. La première partie de cette thèse a identifié le contenu des stéréotypes liés à l’AP et au VIH à travers une approche qualitative (Etude 1) et a permis le développement et la validation d’une échelle spécifique (Etude 2). La deuxième partie avait pour objectif d’identifier les mécanismes en jeu dans la relation entre stéréotypes liés à l’exercice, fatigue et AP. Les résultats montrent que les stéréotypes liés à l’exercice pourraient influencer l’AP par le biais du mécanisme d’ego depletion, indexé par la fatigue perçue et, être modérés par l’auto-efficacité (Etude 3). La fatigue perçue et la fatigabilité dépendraient également du niveau d’AP des PVVIH, les PVVIH les plus actives étant caractérisées par une fatigue perçue et une fatigabilité moindres (Etude 4). Enfin, basée sur la menace du stéréotype, la dernière partie de ce travail doctoral a indiqué que la menace du nonexerciser stereotype pouvait affecter la fatigabilité des PVVIH moins actives (Etude 5), mais pas celle d’adultes sains (Etude 6). De plus, l’auto-efficacité a modéré l’effet de menace du nonexerciser stereotype chez les PVVIH moins actives (Etude 5). Ces résultats enrichissent la littérature relative aux stéréotypes dans le domaine de l’AP tout en soulevant de nouvelles questions importantes pour le développement des modèles actuels de la fatigue dans des pathologies chroniquesPhysical activity (PA) has been reported to have many beneficial effects among people living with HIV (PLHIV), especially on fatigue, which is one of the main prevalent symptom and possible side effects of treatments in this population. Nevertheless, PLHIV remain insufficiently active according to PA recommendations. Based on contemporary theories of stereotypes applied to health behaviours (e.g., Stereotype Embodiment Theory, Levy, 2009; Stereotype Threat Theory, Steele & Aronson, 1995) and an integrative approach of fatigue (Kluger et al., 2013), this doctoral work postulated that exercise stereotypes endorsement or activation would play a role in the PA level of PLHIV and in their performance in fatiguing tasks. The first part of this dissertation identified the content of stereotypes related to PA and HIV through a qualitative approach (Study 1) and conducted the development and validation of a specific scale (Study 2). The second part aimed at identifying the mechanisms at play in the relationship between exercise stereotypes, fatigue and PA. Results showed that exercise stereotypes might influence PA through ego depletion mechanisms, as indexed by perceived fatigue and, be tempered by exercise self-efficacy (Study 3). Perceived fatigue and performance fatigability might also depend on the PA level of PLHIV, the more physically active PLHIV being characterised by lower perceived fatigue and performance fatigability (Study 4). Finally, based on stereotype threat theory, the last part of this doctoral work indicated that the nonexerciser stereotype threat could affect performance fatigability in less active PLHIV (Study 5) but not in healthy adults (Study 6). Furthermore, self-efficacy tempered the effect of the nonexerciser stereotype in less active PLHIV (Study 5). These results enrich the literature on stereotypes in the PA domain while raising interesting questions for contemporary models of fatigue in chronic diseases
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Davis, Robert M. "Asymmetrical Information Market Failure Triggered by the Chicago School's Profit Motive Paradigm: A Case Study of Virginia's Public Higher Education Market and Media Identification of Public Value Failure." Thesis, Virginia Tech, 2013. http://hdl.handle.net/10919/23281.
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This paper presents a case study examination of the Commonwealth of Virginia\'s public higher education market and the use of asymmetrical information flows between providers and consumers by college and university institutions to intentionally create market failures to maximize brand building through increased revenue collections via profit maximization behaviors. Existing economic research in the financial services market hold that asymmetry of information generates inefficient allocation of goods and subsequent identification of market failure conditions. Market failures can lead to tipping points which may result in public values failures as threats to human subsistence (i.e. food, clothing, education) and imperfect public information. Market failures resulting in public values failures warrant government intervention to correct market inefficiency and ensure pareto efficiency in the allocation of goods. Mandatory non-educational fees increase the cost to attend a post-secondary institution which subsequently aid in increased student debt and reduced access and affordability for low income classification groups thus exacerbating societal cleavages identified as public values failures. This research identifies the application of economic and public administration theory to construct a policy recommendation to mitigate asymmetrical information and improve pareto efficiency involving transactions in the public higher education market.Master of Public Administration
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Zheng, Yue. "Identification d'une plante médicinale africaine par le DNA barcoding et étude de composés à activité anti-HIV de cette plante." Thesis, Strasbourg, 2015. http://www.theses.fr/2015STRAJ095/document.
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Mon travail de thèse porte sur l'identification d'un arbre médicinal africain par le DNA barcoding et l'étude de composés à activité anti-VIH de cet arbre. Une première analyse de la séquence du marqueur ITS2 déterminée à partir d'ADN extrait de copeaux de bois a suggéré que la plante pourrait appartenir au genre Cassia ou au genre apparenté Senna. En analysant la séquence de ce marqueur ITS2 et aussi celle du trnHpsbA spacer d'une cinquantaine d'espèces des genres Cassia et Senna j'ai pu identifier la plante comme étant Cassia abbreviata. L'alignement de ces séquences m'a permis d'identifier, pour les deux marqueurs,des structures particulières spécifiques aux espèces du genre Cassia, permettant donc de les différencier des espèces du genre Senna. J'ai utilisé ces alignements pour effectuer une étude phylogenetique qui illustre que,pour les deux marqueurs, les Cassia forment en effet un clade bien séparé du clade des Senna qui peut être divisé en plusieurs sous-clades. Dans un deuxième temps j'ai étudié les effets anti-VIH de l'extrait alcoolique ainsi que de 57 composés purifiés obtenus au laboratoire. L'extrait brut ainsi qu'un des composés purifiés, le piceatannol, ont montré un grand spectre d’activités antivirales pour le VIH et le virus de l’herpès. Ils inhibent,à un stade précoce, l'infection par le VIH de lignées cellulaires de référence et d'isolats cliniques, ceci indépendamment de l'utilisation du co-récepteur (IC50: 10.47-40.77 μg/ml, CC50>1000 μg/ml; IC50: 8.04-47.46 μM, CC50>300 μM, respectivement). Ni l'un ni l'autre n'a d'effet sur CD4 et CCR5/CXCR4. L'extrait brut,mais pas le piceatannol, bloque l'interaction CD4-gp120, suggérant que l'extrait brut cible gp120 alors que le piceatannol agit comme un inhibiteur non-spécifique d'attachement du virus. Aussi, dans un modèle in vitro de tract génital femelle, le piceatannol inhibe l'infection de cellules cibles TZM-Bl par le VIH et n'active pas les cellules PBMCs, suggérant qu'il pourrait être un bon candidat comme microbicide. D'autres composés à activité anti-VIH dans l'extrait comportent l'acide oléanolique, l'acide palmitique, la taxifoline, ainsi que troiscomposés dont la structure est en train d'être élucidéeMy thesis project deals with the identification, by DNA barcoding, of an African medicinal plant and the study of anti-HIV compounds from this plant. A first analysis of the ITS2 marker sequence determined from DNA extracted from the wood suggested that the plant could belong to the Cassia or the related Senna genus. A further analysis of ITS2 as well as of trnH-psbA spacer sequences from about 50 species of the two genera allowed me to identify the plant as Cassia abbreviata. The sequence alignments, which reveal unique features present in the Cassia but not the Senna sequences, were used to construct phylogenetic trees showing the clear separation of the species of the Cassia and the Senna genus. The two markers therefore allow a quick discrimination between the species of the Cassia and the Senna genus and appear to be excellent barcode markers for identification of these species. Following the identification of the plant I have tested the crude ethanol extract as well as 57 purified compounds from the plant for an anti-HIV activity. The extract, as well as one of the compounds, namely piceatannol, showed a broad spectrum of antiviral activities for HIV and HSV. They inhibited HIV-1 infection at the early stage against various reference strains and resistant clinical isolates independent of the co-receptor usage (IC50: 10.47-40.77 μg/ml, CC50>1000 μg/ml; IC50: 8.04-47.46 μM,CC50>300 μM, respectively). Neither the crude extract nor piceatannol had an effect on CD4 and CCR5/CXCR4. The crude extract blocked CD4-gp120 interaction while piceatannol did not, indicating that CE may target gp120 and piceatannol may act as a non-specific viral attachment inhibitor. Moreover, piceatannol inhibited HIV infection of TZM-Bl target cells in an in vitro female genital tract model and did not activate PBMCs, suggesting that it may represent a good candidate as microbicide. Other anti-HIV compounds found in Cassia abbreviata include oleanolic acid, palmitic acid, taxifolin and three other compounds the structure of which is presently being elucidated
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Fouche, Jean-Paul. "A diffusion tensor imaging study in HIV patients with and without apathy." Thesis, Stellenbosch : University of Stellenbosch, 2010. http://hdl.handle.net/10019.1/5146.
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Thesis (MScMedSc (Biomedical Sciences. Medical Physiology))--University of Stellenbosch, 2010.ENGLISH ABSTRACT: HIV/AIDS is a global epidemic that accounts for a large percentage of the mortality in South Africa every year. Since the implementation of anti-retroviral treatment, HIV positive individuals have been living longer, and the cognitive impairment associated with the disease is becoming increasingly apparent. During the initial systemic infection of HIV, the virus migrates through the blood-brain barrier and inflicts axonal injury by causing upregulation of cytokines and neurotoxic proteins. HIV-associated dementia is a neuropsychological classification of cognitive impairment in HIV and a variety of symptoms have been classified as a part of the dementia complex. One of these is apathy, which is thought to be a precursor for dementia in HIV patients. Three groups of individuals have been recruited and scanned using magnetic resonance imaging (MRI) to examine changes in the brain. These are an HIV non-apathetic cohort, an HIV apathetic cohort and a healthy control cohort. Diffusion tensor imaging (DTI) is an MRI technique used to quantitatively assess white matter (WM) integrity using metrics such as fractional anisotropy (FA). Voxel-based analysis, tract-based spatial statistics (TBSS) and tractography are three established DTI analysis methods that have been applied in numerous studies. However, there are certain methodological strengths and limitations associated with each technique and therefore all three of these techniques were used to compare WM differences across groups. The frontal-subcortical pathways are known to be abnormal in apathy, and this has been demonstrated in a number of imaging studies. Most of these studies have examined apathy in the context of neurodegenerative disorders such as Alzheimer’s disease and Parkinson’s. However, to our knowledge this is the first DTI study in HIV apathetic patients. With the tractography method, the anterior thalamic radiation and the corpus callosum were reconstructed for each individual to determine whether there were any global changes in these tracts. No significant changes were found. However, a variety of regions in the WM were significantly abnormal in the HIV cohorts when comparing the data at a voxel-based level and using TBSS. This included areas such as the genu and splenium of the corpus callosum, the internal capsule and corona radiata. Changes in frontal WM for the HIV apathy group are an indication of dysfunction in the frontal-striatal circuits, and previous literature has implicated these circuits in the neuropathology of apathy in a variety of central nervous system (CNS) disorders.
AFRIKAANSE OPSOMMING: MIV/VIGS is `n wêreldwye epidemie wat verantwoordelik is vir `n hoë sterftesyfer in Suid- Afrika elke jaar. Sedert die inleiding van anti-retrovirale behandeling, het die MIV-positiewe populasie se lewensduur verleng. Tesame met langer lewensduur, het die kognitiewe verswakking wat geassosieer word met die siekte ook meer prominent na vore gekom. Gedurende die beginstadium van sistemiese infeksie in MIV is daar `n migrasie van die virus deur die bloed-breinskans. MIV kan indirek verantwoordelik wees vir aksonale beskadiging deur verhoging van neurotoksiese proteine en sitokinien te induseer. MIV-geassosieerde demensie is `n neurosielkundige klassifikasie van kognitiewe verswakking in MIV en verskeie simptome is al geïdentifiseer as deel van die demensie kompleks. Een van die simptome is apatie en daar word gespekuleer dat dit `n voorloper is vir demensie in MIV pasiënte. Drie groepe individue was gewerf vir die studie en geskandeer deur magnetiese resonansie beeldvorming (MRB) om sodoende veranderinge in die brein te ondersoek. Die groepe was onderskeidelik `n HIV nie-apatiese kohort, `n HIV apatiese kohort en `n gesonde kontrole kohort. Diffusie tensor beelding (DTB) is `n MRB tegniek wat toegepas word om witstof integriteit te meet deur gebruik te maak van maatstawwe soos fraksionele anisotropie (FA). “Voxel-based analysis”, “tract-based spatial statistics (TBSS)” en “tractography” is drie gevestigde DTB analitiese metodes wat al in talle studies toegepas was. Daar is egter sekere metodologiese voordele en beperkings verbonde aan elke tegniek en daarom is al drie tegnieke gebruik om witstof verskille tussen groepe te vergelyk. Die frontale-subkortikale roetes in die brein is bekend vir abnormaliteite in apatie en dit was ook al gedemonstreer in verskeie studies. Die meeste van die studies het apatie ondersoek in die konteks van neurodegeneratiewe siektes soos Alzheimer se siekte en Parkinson se siekte. Maar sover ons weet is hierdie die eerste DTB studie in MIV pasiënte met apatie. Met die “tractography” metode was die anterior thalamic radiation en corpus callosum herbou vir elke individu. Dit was om te bepaal of daar enige globale veranderinge is in hierdie gebiede, maar geen beduidende veranderinge is gevind nie.`n Verskeidenheid van gebiede in die witstof was beduidend abnormaal in die MIV kohorte wanneer die data vergelyk was met “TBSS” en “voxel-based analysis.” Dit het gebiede ingesluit soos die genu en splenium van die corpus callosum, die internal capsule en die corona radiata. Veranderinge in die frontale witstof vir die MIVapatie groep is `n aanduiding van disfunksie in die frontale-striatale bane. Vorige literatuur impliseer dat hierdie bane betrokke is in die neuro-patologie van apatie in verskeie sentrale senuweestelsel (SS) steurings.