La thèse présentée décrit la préparation et l’étude d’auto-assemblages élaborés à partir de conjugués amphiphiles Tat-b-poly(triméthylène carbonate) (Tat-b-PTMC) doués de propriétés d’internalisation cellulaire conférées par le segment peptidique Tat. L’objectif principal de ces travaux était d’établir et de comprendre les liens entre la structure macromoléculaire, les caractéristiques colloïdales et l’activité biologique de ces systèmes. Les efforts de précision moléculaire et de caractérisation fournispour la synthèse des chimères Tat-b-PTMC a permis de corréler finement leurs structures chimiquesaux paramètres physico-chimiques des nanoparticules obtenues. Grâce à une approche expérimentale transverse combinant des études de biologie cellulaire et de biophysique, le mécanisme d’interaction in vitro de ces nanoparticules avec les cellules HeLa a pu être en partie élucidé. Enfin, un camouflage électrostatique pH-sensible a été mis au point pour tenter de moduler leur activité et d’augmenter leur sélectivité vis-à-vis de l’environnement tumoral. / This thesis work deals with preparation and study of cell-penetrating self-assemblies from amphiphilicpolymer-b-peptide Tat-b-poly(trimethylene carbonate) conjugates. Tat-b-PTMC chimeras withtunable hydrophilic fractions were synthesized, thoroughly characterized and self-assembled inaqueous buffer into size-tunable, highly monodisperse core-shell nanoparticles, presenting a full Tatcorona. Their physico-chemical profiles were assessed by complementary imaging (AFM, TEM) andscattering techniques (multiangle DLS, SANS) and correlated with their molecular architectures.Their transduction ability in vitro on HeLa cells and interaction mode with phospholipid membraneswere studied with a view to correlate their physico-chemical profiles with their biological properties.This interdisciplinary approach partially shed light on the interactions at play in the cellular uptakeprocess. With the ultimate goal of improving pharmacological characteristics, we finally endeavoredto develop an ON/OFF PEGylation strategy to harness the cell penetrating power of thosebiomacromolecular self-assembled systems.

No description available.

No description available.

No description available.

Le cancer gynécologique pelvien est une pathologie qui se loge dans les organes génitaux internes de la femme : vagin, utérus, col de l'utérus et ovaires. Ces organes malades sont traités chez les patientes de l'étude par hystérectomie plus ou moins élargie (ablation des organes génitaux), chimiothérapie, radiothérapie et curiethérapie. C'est donc une castration que ces femmes subissent dans la réalité d'un corps qui souffre. Cancer et traitements sont autant d'effractions corporelles qui réitèrent une représentation psychique de la pénétration comme de l’accueil passif de l'autre en soi. Cette figure de l'étranger à l’intérieur du moi convoque des fantasmes, affects et représentations pénétrantes qui endommagent l'image du corps, génère des images de passivité et ravive des expériences de passivation. De plus, cette effraction s'opère spécifiquement sur le lieu intime du sexe, là où s'incarne le devenir femme puis le devenir mère, convoquant le féminin de la femme dans sa pluralité, mais aussi dans ses rapports à la féminité. Après avoir dégagé, dans un référentiel littéraire très large, les rares travaux qui mettent en perspective l'effraction d'un cancer gynécologique et ses traitements avec la sexualité des femmes, nous avons situé notre propre recherche dans une approche de psychopathologie psychanalytique et dans ses rapports avec la psychosomatique. S'ensuit un repérage de la théorie psychanalytique conséquent des deux grands champs sollicités par le sujet de l'étude : la sexualité féminine et le corps dans la maladie. La méthodologie originale mise en place s'inscrit dans une démarche exploratoire étayée de la passation de deux tests projectifs : le Rorschach et le TAT, ainsi que d'entretiens cliniques de recherche. Il s'agit d'une méthodologie longitudinale sur trois temps choisis du parcours thérapeutique de ces femmes. Les résultats, traités dans un premier temps dans la dimension singulière de l'étude de cas, sont regroupé ensuite en fonction des hypothèses pour faire apparaître des aménagements communs chez ces patientes. L'analogie entre la situation de l'infans et celle de ces femmes sous contraintes de passivité accompagne la description des trois temps de l'étude. Celle-ci répond à l'hypothèse d'une mobilisation de modalités psychiques spécifiques du traitement de l'effraction corporelle qui convoquent, malmènent, blessent et suppriment les organes génitaux internes. L'étude révèle l'actualisation d'angoisses primitives spécifiquement féminines, telle que l'angoisse de pénétration, traitées par des modalités limites, narcissiques et anales autour d'un fantasme de corps troué et d'une intériorité dangereuse. En définitive, l'étude explore l'ensemble du jeu pulsionnel pour en décrypter les mouvements d’intrication et de désintrication sous-tendant le désinvestissement érotique du génital, convoquant le féminin dans ses potentialités passives et accompagnant l'infléchissement de la relation d'objet sous l'influence de la pulsion de mort. / Pelvic gynaecological cancer is a disease that fits into internal reproductive organs of women : vagina, uterus, cervix and ovaries. These cancers are treated in patients of the study by hysterectomy, extended or not (removal of genitals), chemotherapy, radiotherapy and brachytherapy. It is a castration that these women experience in the reality of a body that suffers. Cancer and treatments are corporal effractions that reiterate a psychic representation of the penetration as the passive reception of the other in oneself. This figure of the foreigner inside the ego summons fantasies, affects and penetrating representations that damage body image, generates passivity images and rekindles passivation experiences. Moreover, this effraction takes place specifically on the intimate sex place, where the growing into a woman and then into a mother incarnates itself, summoning the feminine of the woman in his plurality but also in his connections to feminity. After clearing, in a very broad literary repository, the few studies that put into perspective the effraction of the gynaecological cancer and its treatments with feminine sexuality, we set our own research in the approach of psychoanalytic psychopathology in its relations with psychosomatics. A consistent identification of psychoanalytic theory of the two major fields requested by the subject of the study ensues : feminine sexuality and body in the disease. The original methodology implemented fits into an exploratory process supported by the administration of two projective tests : Rorschach and TAT, along with clinical research interviews. This is a longitudinal methodology at three selected steps of the therapeutic course of these women. The results, treated initially in the singular dimension of the case study are then grouped according to the hypotheses to show the shared adjustments in these patients. The analogy between the situation of the infans and the women under passivity constraints accompanies the description of the three steps of the study. It responds to the hypothesis of a mobilization of psychic modalities specific to corporal effraction treating process which summon, mistreat, wound and remove internal genitalia. The study reveals specifically feminine primitive anxiety such as penetration anxiety, treated by limit modalities, narcissistic and anal around a holed body fantasy and a dangerous interiority. Finally, the study explores the entire set of drives in order to figure out fusion and defusion movements subtending the erotic decathexis of the genital, summoning the feminine in its passive potentialities and accompanying the reorientation of the object-relation under the death drive influence.

No description available.

These studies were aimed at investigating the potential application of nanoparticles engineered from oil-in-water microemulsion precursors for enhancing immune responses to HIV-1 Tat and Gag p24 proteins. Both of the HIV-1 proteins have been reported to be critical in the virus life cycle and are being evaluated in clinical trials as vaccine candidates. Anionic nanoparticles were prepared using emulsifying wax as the oil phase and Brij 78 and sodium dodecyl sulfate as the surfactants. The resulting nanoparticles were coated with Tat and were demonstrated to produce superior immune responses after administration to BALB/c mice compared to Tat adjuvanted with Alum. Similarly, cationic nanoparticles were prepared using emulsifying wax and Brij 78 and cetyl trimethyl ammonium bromide as the surfactants. The cationic nanoparticles were investigated for delivery of immunostimulatory adjuvants, namely three Toll-like receptor ligands, for obtaining synergistic enhancements in immune responses to a model antigen, Ovalbumin (OVA). In vitro and in vivo studies were carried out to elucidate possible mechanisms by which nanoparticles may result in enhancements in immune responses. In vitro studies were carried out to evaluate the uptake of nanoparticles into dendritic cells and to assess the release of pro-inflammatory cytokines from dendritic cells in the presence of nanoparicles. In vivo studies were carried out using a MHC class I restricted transgenic mouse model to investigate the potential for nanoparticles coated with OVA to enhance presentation of the protein to CD8+ T cells compared to OVA alone. Finally, the preparation of nanoparticles with a low amount of surface chelated nickel for high affinity binding to histidine-tagged (his-tag) proteins was investigated. It was hypothesized that this strengthened interaction of his-tag protein to the nickel chelated nanoparticles (Ni-NPs) would result in a greater uptake of antigen in vivo; therefore, enhanced immune responses compared to protein bound to anionic nanoparticles. In vivo evaluation of his-tag HIV-1 Gag p24 bound to Ni-NPs resulted in enhanced immune responses compared to protein either adjuvanted with Alum or coated on the surface of nanoparticles.

More than 1.2 million people are estimated to be currently living with the human immunodeficiency virus (HIV) in the United States of America. The gastrointestinal (GI) tract is both a major target and an important component of HIV pathogenesis. The GI processes that are dysregulated during HIV infection are controlled by the enteric nervous system (ENS). Indeed, both clinical and experimental studies have implicated the ENS in HIV and simian immunodeficiency virus (SIV) pathogenesis. In addition to direct viral effects, the HIV virus also indirectly affects the GI tract via cellular and/or viral toxins released by infected cells. Trans-activator of transcription (Tat) is a viral toxin that plays an important role in replication of the HIV virus. While, the HIV virus does not directly infect neurons, Tat has been shown to modulate neuronal function. HIV infection in the gut is accompanied by: translocation of bacteria and bacterial products from the gut lumen to peripheral blood, immune activation and inflammation. Lipopolysaccharide (LPS) is a major bacterial product that is used to determine the rate of bacterial translocation and to drive inflammation. Despite reports of enteric ganglionitis in SIV infected monkeys and autonomic denervation in the jejunum of HIV patients, little is known of the mechanism underlying enteric neuropathogenesis in HIV and the role of the ENS in HIV pathogenesis. In the present study, we assessed the effects of Tat on enteric neuronal excitability and how Tat and LPS interact in the ENS to bring about inflammation and GI motility problems observed in HIV patients. We show that Tat significantly increased enteric neuronal excitability by modulating sodium channels expressed on enteric neurons. Tat sensitized ENS cells to LPS-mediated increase in pro-inflammatory cytokines via a TLR4-mediated pathway involving MyD88. Mice expressing the tat transgene (Tat+) had faster GI transit rates and significantly higher frequencies of diameter changes in the proximal ileum than controls (Tat-). Tat+ mice were also more sensitive to LPS-mediated decreases in colonic transit rate. This study highlights the role of viral and bacterial proteins in HIV pathogenesis in the gastrointestinal tract and also demonstrates a critical role of the ENS in HIV pathogenesis.

Orientador: Deilson Elgui de Oliveira / Banca: Raphael Bessa Parmigiani / Banca: Enrique Mario Boccardo Pierutivo / Banca: Carlos magno Castelo Branco Fortaleza / Banca: Claudia Aparecida Rainho / Resumo: Introdução: Pacientes portadores do vírus da imunodeficiência humana (HIV) apresentam risco aumentado de desenvolverem neoplasias malignas ligadas ao herpesvírus associado ao sarcoma de Kaposi (KSHV), entre elas o sarcoma de Kaposi (SK). Digno de nota, o sarcoma de Kaposi associado a aids (SK-AIDS) apresenta-se mais agressivo que as outras formas de SK, sugerindo a existência de interação sinérgica entre os produtos do KSHV e do HIV-1. O ciclo biológico do KSHV é dividido em fase latente e lítica. As proteínas vFLIP e LANA expressas na fase latente e as proteínas líticas Rta, vGPCR e K1, apresentam propriedades oncogênicas. Graças a habilidade da proteína tat do HIV-1 em estimular a angiogênese e outros fenômenos inflamatórios, é plausível que esta proteína modifique significativamente a expressão gênica do KSHV, proporcionando alterações fenotípicas que contribuem para o desenvolvimento do SK-AIDS. Para melhor entendimento dos efeitos da proteína tat do HIV-1 em células infectadas pelo KSHV, por meio da análise das reações em cadeia da PCR quantitativa acoplada a transcrição reversa (Quantitative real time reverse transcriptase polymerase chain reactions - qRT-PCR), o presente trabalho descreveu as alterações na expressão dos genes codificadores de vFLIP, LANA, Rta, vGPCR e K1 em células endoteliais de veia umbilical humana imortalizada pela telomerase e infectada pelo KSHV a longo prazo (TIVE-LTCs) expostas à proteína tat recombinante do HIV-1 por 12, 24, 48 e 72h. Resultados: Em termos descritivos, a expressões diferencial dos genes latentes na vigência de exposição à proteína tat recombinante do HIV-1 em relação à proteína tat denaturada, aumentou após 12 e 24h para LANA e após 24 e 72h para vFLIP. Para os genes líticos a expressão diferencial aumentou consistentemente até atingir 48h e diminuiu nas próximas 12h para ORF-50. Enquanto que para... / Abstract: Not available / Doutor

HIV is capable of causing a set of neurological diseases collectively termed the HIV Associated Neurocognitive Disorders (HAND). Worsening pathology is observed in HIV+ individuals who use opioid drugs. Memory problems are often observed in HAND, implicating HIV pathology in the hippocampus, and are also known to be exacerbated by morphine use. HIV-1 Tat was demonstrated to reduce spatial memory performance in multiple tasks, and individual subsets of CA1 interneurons were found to be selectively vulnerable to the effects of Tat, notably nNOS+/NPY- interneurons of the pyramidal layer and stratum radiatum, PV+ neurons of the pyramidal layer, and SST+ neurons of stratum oriens. Each of these interneuron subsets are hypothesized to form part of a microcircuit involved in memory formation. Electrophysiological assessment of hippocampal pyramidal neurons with Tat and morphine together revealed that Tat caused a reduction in firing frequency, however, chronic morphine exposure did not have any effect. When morphine was removed after chronic exposure, non-interacting effects of Tat and morphine withholding on firing frequency were observed, suggesting that a homeostatic rebalancing of CA1 excitation/inhibition balance takes place in response to chronic morphine exposure independently of any Tat effects. Additionally, differential morphological effects of Tat and morphine were observed in each of the three major dendritic compartments, with SR being less affected, suggesting complex circuit responses to these insults reflecting local change and potentially changes in inputs from other brain regions. Behaviorally, Tat and morphine interactions occur in spatial memory, with morphine potentially obviating Tat effects.