Opioid Receptors (ORs) are involved in the pathophysiology of several neuropsychiatric conditions yet remain an untapped therapeutic resource. Although only mu-, delta-, and kappa-OR types have been cloned, additional subtypes result from complexes generated by direct receptor-receptor interactions. Mu- and delta-ORs form a heteromeric receptor complex with unique pharmacological and signalling properties distinct from those of mu- and delta-OR homomers. In these studies, we sought to characterize the ligand binding pocket and agonist-induced internalization profile of the mu-delta heteromer, to investigate mu-delta heteromer-specific signalling in brain, and to interrogate the contribution of this receptor complex to opioid-mediated behavioural effects. In competition radioligand binding studies, delta-agonists displaced high affinity mu-agonist binding from the mu-delta heteromer but not the muOR homomer, suggestive of delta-agonists occupying or allosterically modulating the muOR ligand binding pocket within the heteromer. Delta-agonists induced internalization of the mu-delta heteromer in a dose-dependent, pertussis toxin resistant, and muOR- and deltaOR-dependent manner from the cell surface via the clathrin and dynamin endocytic machinery. Agonist-induced internalization of the mu-delta heteromer persisted following chronic morphine treatment conditions which desensitized the muOR homomer. Using Galpha-specific GTPgammaS binding assays, we demonstrated that mu-delta heteromer signalling previously characterized in cell lines was present in the striatum and hippocampus, and did not desensitize following prolonged morphine treatment conditions which desensitized muOR homomer-mediated signalling. Since delta-agonists which also target the mu-delta heteromer possess antidepressant-like and anxiolytic-like properties, we investigated the role of this receptor complex in mood regulation. We devised a strategy to selectively analyze the effects of the mu-delta heteromer by dissociating it using a specific interfering peptide aimed at a sequence implicated in mu-delta heteromerization. The interfering peptide abolished the unique pharmacological and trafficking properties of delta-agonists at the mu-delta heteromer and dissociated this receptor complex in vitro. Intra-accumbens administration of the interfering peptide disrupted the mu-delta interaction in vivo and allowed for isolation of the mu-delta heteromer contribution to the mood-regulatory effects of a delta-agonist with activity at the heteromer. Activation of the mu-delta heteromer in the nucleus accumbens produced antidepressant-like and anxiolytic-like actions in animal models of depression and anxiety.

Syftet är att studera instrumentallärares syn på hantering av elevers internalisering av musikaliska uttryck. Det sociokulturella perspektivet med fokus på hur redskap påverkar internaliseringen är utgångspunkt för studien. Metoden utgörs av semistrukturerad intervju där fyra formellt och reellt utbildade instrumentallärare som arbetar på musik- eller kulturskola har intervjuats om vilka musikaliska uttryck de har i fokus i undervisningen och deras syn på hur elever lär sig musikaliska uttryck. Resultatet visar att det finns flertalet olika redskap för att nå musikaliska uttryck, bland annat eleven själv, musiken som sådan och den sociala omgivningen. Det viktigaste musikaliska uttrycksmedlet är rytm i förhållande till tempo. Resultatdiskussionen fokuserar på redskapens möjligheter och hinder för att internalisera musikaliska uttryck. Det sociala samspelet på lektionstid och i orkester samt fantasi, kreativitet och bilder framhålls som möjligheter för att internalisera musikaliska uttryck. Notläsning och musikaliska uttryck som sekundärt mål framhålls som hinder för att internalisera musikaliska uttryck. Det finns också varierande metoder att ta sig förbi hindren. / The purpose of the study is to gain an understanding of instrumental teachers’ view on the management of internalization of musical expressions by pupils. The theoretical starting point of the study is the sociocultural perspective, with a focus on how tools influence the internalization. The method used was semi-structured interview. Four formally educated and experienced instrumental teachers who work in music or culture schools were interviewed about which expressions they focused on in teaching, and their view on how pupils internalize musical expressions. The results indicate that there are many tools to learn musical expressions; the pupil, the music itself and the social interaction, among others. The most important expression is rhythm related to tempo. The discussion focus on the possibilities and obstacles of tools to internalize musical expressions. The social interaction during the music lesson and in the orchestra, and fantasy, creativity and images are emphasized as possibilities to internalize musical expressions. Reading music and musical expressions as a secondary goal are emphasized as obstacles which can avert internalization of musical expressions. However, there are also a variety of methods to get pupils past such obstacles.

No description available.

Objective: Internalizing the pervasive weight bias commonly directed towards individuals with overweight and obesity, cooccurs with increased psychopathology and impaired quality of life. This study sought to establish population norms and psychometric properties of the most widely used self-report questionnaire, the Weight Bias Internalization Scale (WBIS), in a representative community sample. Design and Methods: In a survey of the German population, N = 1158 individuals with overweight and obesity were assessed with the WBIS and self-report measures for convergent validation. Results: Item analysis revealed favorable item-total correlation of all but one WBIS item. With this item removed, item homogeneity and internal consistency were excellent. The one-factor structure of the WBIS was confirmed using confirmatory factor analysis. Convergent validity was shown through significant associations with measures of depressive and somatoform symptoms. The WBIS contributed to the explanation of variance in depressive and somatoform symptoms over and above body mass index. Higher WBIS scores were found in women than in men, in individuals with obesity than in individuals with overweight, and in those with lower education or income than those with higher education or income. Sex specific norms were provided. Conclusions: The results showed good psychometric properties of the WBIS after removal of one item. Future research is warranted on further indicators of reliability and validity, for example, retest reliability, sensitivity to change, and prognostic validity.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A ocorrência de fenótipos multirresistentes de Corynebacterium pseudodiphtheriticum e sua associação a infecções graves, com elevada mortalidade em pacientes imunocomprometidos, aliados ao escasso conhecimento da virulência e patogenia destas infecções, motivou esta pesquisa, que teve como objetivo investigar mecanismos de virulência e resistência microbiana deste agente entre pacientes de um hospital universitário brasileiro. Um total de 113 amostras de C. pseudodiphtheriticum identificadas por métodos bioquímicos convencionais e sistema API-Coryne isoladas de pacientes de diferentes grupos etários. Os micro-organismos eram, em sua maioria, relacionados a infecções no trato respiratório (27,45%), urinário (29,20%) e sitios intravenosos (18,60%) e cerca de 32,70% das amostras foram provenientes de pacientes com pelo menos uma das condições predisponentes: insuficiência renal; transplante renal, tuberculose em paciente HIV+, câncer, cirrose hepática, hemodiálise e uso de cateter. As amostras testadas revelaram-se multirresistentes sendo a maioria resistente à oxacilina, eritromicina e clindamicina. A adesão das cepas ao poliestireno e ao poliuretano indicou o envolvimento de hidrofobicidade da superfície celular na fase inicial da formação de biofilmes. O crescimento subsequente conduziu à formação de microcolônias, agregados bacterianos densos incorporados na matriz exopolimérica rodeada por espaços vazios, típica de biofilmes maduros. Adicionalmente, a interação do micro-organismo com fibrinogênio e fibronectina humana indica o envolvimento destes componentes séricos na formação de biofilme, sugerindo a participação de diferentes adesinas neste processo e a capacidade deste agente formar biofilme in vivo. A afinidade por esses componentes e a formação de biofilme podem contribuir para o estabelecimento e disseminação da infecção no hospedeiro. Adicionalmente, as cepas de C. pseudodiphtheriticum isoladas de pacientes com infecções localizadas (ATCC10700/Pharyngitis) e sistêmicas (HHC1507/Bacteremia) exibiram um padrão de aderência agregativa-like a células HEp-2, caracterizado por aglomerados de bactérias com aparência de um "empilhado de tijolos". Através do teste FAS e ensaios de interação na presença de inibidores de citoesqueleto, demonstramos o envolvimento da polimerização de actina na internalização das cepas testadas. A internalização bacteriana e rearranjo do citoesqueleto pareceu ser parcialmente desencadeado pela ativação da tirosina-quinase. Finalmente, C. pseudodiphtheriticum foi capaz de sobreviver no ambiente intracelular e embora não tenha demonstrado capacidade de replicar intracelularmente, células HEp-2 foram incapazes de eliminar o patógeno completamente no ambiente extracelular no período de 24 horas. Todas as cepas estudadas foram capazes de induzir apoptose em células epiteliais 24 horas pós-infecção evidenciada pelo aumento significativo no número de células mortas e pela ocorrência de alterações nucleares reveladas através dos métodos de coloração pelo azul Trypan, pelo DAPI e microscopia electrônica de transmissão. Alterações morfológicas incluindo a vacuolização, a fragmentação nuclear e a formação de corpos apoptóticos foram observadas neste período. A citometria de fluxo demonstrou ainda uma diminuição significativa no tamanho das células infectadas e a utilização de dupla marcação (iodeto de propídio / anexina V) permitiu a detecção da ocorrência de necrose e apoptose tardia. Em conclusão, o conhecimento de tais características contribuiu para a compreensão de mecanismos envolvidos no aumento da frequência de infecções graves com elevada mortalidade em pacientes no ambiente hospitalar, por C. pseudodiphtheriticum, um patógeno rotineiramente subestimado em países em desenvolvimento. / The occurrence of multiresistant phenotypes and associated with severe infections, with high mortality in immunocompromised hosts due to Corynebacterium pseudodiphtheriticum, allied to little known about virulence and pathogenesis these infections, led to present investigation. The investigation aims to examine the virulence mechanisms and resistance to antimicrobial agents of C. pseudodiphtheriticum among patients with bacterial infections at a Brazilian teaching hospital. A total of 113 C. pseudodiphtheriticum strains identified by conventional biochemical methods and API-Coryne System were recovered from patients from different age groups. Micro-organisms were mostly related to infections in the respiratory tracts (27.45%), urinary (29.20%) and intravenous sites (18.60%) and approximately 32.70% samples were obtained of patients presenting at least one of the pre-disposing conditions: end-stage renal disease; renal transplant; AIDS and Mycobacterium tuberculosis infection; cancer, hepatic cirrhosis; haemodialysis and catheter use. Antimicrobial susceptibility tests identified multiresistant phenotypes. Most strains were resistant to oxacillin, erythromycin and clindamycin. Adherence to polystyrene and polyurethane indicated the involvement of cell surface hydrophobicity in the initial stage of biofilm formation. Further growth led to the formation of dense bacterial aggregates embedded in the exopolymeric matrix surrounded by voids, typical of mature biofilms. Data also showed C. pseudodiphtheriticum recognizing human fibrinogen (Fbg) and fibronectin (Fn) and involvement of these sera components in biofilm formation in conditioning films. These findings suggest that biofilm formation may be associated with the expression of different adhesins. C. pseudodiphtheriticum may form biofilm in vivo possibly by an adherent biofilm mode of growth in vitro currently demonstrated on hydrophilic and hydrophobic abiotic surfaces. The affinity to Fbg and Fn and the biofilm-forming ability may contribute to the establishment and dissemination of infection caused by C. pseudodiphtheriticum. Additionally, C. pseudodiphtheriticum strains isolated from patients with localized (ATCC10700/Pharyngitis) and systemic (HHC1507/Bacteremia) infections exhibited an aggregative adherence-like pattern to HEp-2 cells characterized by clumps of bacteria with a stacked-brick appearance. The fluorescent actin staining test demonstrated that actin polymerization is involved in the internalization of the C. pseudodiphtheriticum strains. Bacterial internalization and cytoskeletal rearrangement seemed to be partially triggered by the activation of tyrosine kinase activity. Although C. pseudodiphtheriticum strains did not demonstrate an ability to replicate intracellularly, HEp-2 cells were unable to fully clear the pathogen within 24 hours. All samples were able to induce apoptosis in HEp-2 cells 24 h post-infection, evidenced by significant increase in the number of dead cells and nuclear alterations were observed by the Trypan blue assay, DAPI and transmission electron microscopy. Morphological changes in HEp-2 cells observed 24 h post-infection included vacuolization, nuclear fragmentation and the formation of apoptotic bodies. Flow cytometry revealed an significant decrease in cell size of infected HEp-2 cells. Furthermore, a double-staining assay using Propidium Iodide/Annexin V gave information about the numbers of vital vs. early apoptotic cells and late apoptotic or secondary necrotic cells. In conclusion, these characteristics may contribute to understanding of mechanisms involved on increase of severe infection, with high mortality in nosocomial enviroment patients by C. pseudodiphtheriticum, a pathogen usually overlooked in emerging countries.

Submitted by Emanuel Varela Cardoso (emanuel.varela@ufcg.edu.br) on 2018-08-02T20:14:16Z No. of bitstreams: 1 LUCAS PEREIRA DA SILVA – TESE (PPGCS) 2017.pdf: 2960555 bytes, checksum: d73e936223a1339d1c161747aa50f1d4 (MD5) / Made available in DSpace on 2018-08-02T20:14:16Z (GMT). No. of bitstreams: 1 LUCAS PEREIRA DA SILVA – TESE (PPGCS) 2017.pdf: 2960555 bytes, checksum: d73e936223a1339d1c161747aa50f1d4 (MD5) Previous issue date: 2017-10-31 / O estudo aqui apresentado volta-se para a vida universitária em seu perfil atual, em diálogo com a efetivação das políticas públicas de democratização/interiorização dos espaços formativos nas universidades federais. O objetivo principal da presente pesquisa é analisar as experiências formativas de estudantes universitários das camadas populares, sobretudo, daqueles advindos das escolas públicas do Semiárido paraibano, que entre 2006 a 2015, ingressam, permaneceram no campus de Cuité/UFCG e obtiveram sucesso, ou êxito, na formação, dando continuidade aos estudos numa pós-graduação stricto sensu (mestrado e doutorado), construindo assim, uma “trajetória universitária prolongada”. As especificidades contextuais e constitutivas desse processo em curso cobram estudos que possam subsidiar outras investigações de cunho comparativo. Nesse intuito, assumindo os pressupostos de uma pesquisa qualitativa, que não despreza a importância de elementos quantitativos, buscamos desenvolver um Estudo de Caso. O corpus utilizado para análise das experiências formativas são os relatos autobiográficos enviados por 20 ex-estudantes do campus de Cuité (sujeitos da pesquisa), identificados através de um Questionário Base, aplicado através de uma plataforma online. Os relatos autobiográficos, para além das autobiografias tradicionais, são instrumentos que informam sobre o contexto das práticas universitárias, em observância a construção de um novo perfil do campus brasileiro e de um novo habitus no cenário atual das universidades federais. A base teórica adotada no estudo, propõe uma interface entre Ciências Sociais e Educação. Nesse diálogo, recorre-se à Sociologia da Educação, a partir de Bourdieu e Lahire, referente ao campo universitário, capital cultural, as origens sociais dos estudantes e a construção de um novo habitus nas práticas universitárias e à Teoria Sociohistórica, Vygotsky e Leontiev, com as noções de mediação e atividades mediadoras, buscando compreender a construção das trajetórias de formação dos estudantes do ponto de vista relacional. Com isso, problematiza-se as iniciativas recentes de democratização do acesso ao ensino superior público, considerando três dimensões: a) democratização do ingresso; b) democratização das condições de permanência e c) sucesso ou êxito acadêmico (formação inicial e continuidade dos estudos). A análise evidencia as iniciativas e os indicadores do processo de democratização em curso, mostrando o Estado da Paraíba na dianteira deste processo em relação: 1) às taxas bruta e líquida de matrícula na população de 18 a 24 anos; 2) ao crescimento no número de estudantes matriculados em instituições públicas vindos da escola pública; 3) ao crescimento no número de estudantes que se autodeclaram pardos e pretos na rede federal; 4) ao ingresso na universidade pública por estudantes oriundos de famílias das camadas populares; 5) a importância da formação dos professores e da experiência dos estudantes em atividades de iniciação a pesquisa, extensão, iniciação a docência, monitoria, entre outras, resultando na construção de suas trajetórias. Trata-se de um delineamento pouco explorado no estudo das sociabilidades contemporâneas com foco na formação universitária em sua atual configuração. O estudo questiona ainda, o sentido das práticas na universidade e busca contribuir para a avaliação das políticas de Educação Superior e para o crescimento das Ciências Sociais na abordagem do tema. / The study presented here turns to the university life in its current profile, in dialogue with the implementation of the public policies of democratization / internalization of the educational spaces in the federal universities. The main objective of the current research is to analyze the formative experiences of university students of the popular stratum, especially those coming from public schools in the semi-arid region of Paraíba, that between 2006 and 2015, they entered, stayed in the campus of Cuité / UFCG and obtained success or outcome in the formation, giving continuity to the studies in a stricto sensu postgraduate (masters and doctorate), thus constructing, a “longstading university trajectory”. The contextual and constitutive specificities of this ongoing process require studies that may support other comparative investigations. For this purpose, assuming the assumptions of a qualitative research, which does not disregard the importance of quantitative elements, we seek to develop a Case Study. The corpus used to analyze the formative experiences are the autobiographical reports sent by 20 former students of the Cuité campus (subjects of the research), identified through a Base Questionnaire, applied through an online platform. The autobiographical report, beyond the traditional autobiographies, are instruments that inform the context of the university practices, observing the construction of a new profile of the Brazilian campus and of a new habitus in the current scenery of the federal universities. The theoretical basis adopted in the study proposes an interface between Social Sciences and Education. In this dialogue, the Sociology of Education is used, starting from Bourdieu and Lahire, referring to the university field, cultural capital, the social origins of the students and the construction of a new habitus in university practices and Sociohistorical Theory, Vygotsky and Leontiev, with the notions of mediation and mediating activities, seeking to understand the construction of the trajectories of formation of students from the relational point of view. Thus, the recent initiatives for the democratization of the access to public higher education are problematized, considering three dimensions: a) democratization of the admission; b) democratization of the permanence conditions and c) academic success or outcome (initial formation and continuity of studies). The analysis points the initiatives and indicators of the ongoing democratization process, showing the State of Paraíba at the forefront of this process in relation to: 1) the gross and net enrollment rates in the population aged 18-24 years; 2) the growth in the number of students enrolled in public institutions from the public school; 3) an increase in number of students who declare themselves as mixed race or black in the federal network 4) to the entrance in the public university by students coming from families of the popular layers; 5) the importance of teacher training and the students' experience in initiation, resear ch, extension, teaching, monitoring, among others, resulting in the construction of their trajectories. It is a delimitation little explored in the study of the contemporary sociabilities with focus in the university formation in its current configuration. The study also questions the meaning of practices in the university and seeks to contribute to the evaluation of Higher Education policies and to the growth of Social Sciences on the approach of the theme.

Submitted by Marcia Bacha (marcia.bacha@fgv.br) on 2012-01-17T18:45:23Z No. of bitstreams: 1 1425453.pdf: 5060804 bytes, checksum: 3fca53c0ba321ccb58589df2289e9fd3 (MD5) / Approved for entry into archive by Marcia Bacha (marcia.bacha@fgv.br) on 2012-01-17T18:45:37Z (GMT) No. of bitstreams: 1 1425453.pdf: 5060804 bytes, checksum: 3fca53c0ba321ccb58589df2289e9fd3 (MD5) / Approved for entry into archive by Marcia Bacha (marcia.bacha@fgv.br) on 2012-01-17T18:45:44Z (GMT) No. of bitstreams: 1 1425453.pdf: 5060804 bytes, checksum: 3fca53c0ba321ccb58589df2289e9fd3 (MD5) / Made available in DSpace on 2012-01-17T18:45:51Z (GMT). No. of bitstreams: 1 1425453.pdf: 5060804 bytes, checksum: 3fca53c0ba321ccb58589df2289e9fd3 (MD5) Previous issue date: 2010 / Thís paper aims to offer an insight into the viability of the metropolitanízation of the Pernambuco's Central Wasteland Regíon. Therefore, ít is presented an initial analysis on the ways of survival of small production centers in the constant and metamorphic process 01' globalization. lt is also shown some processes of regionalization, passíng by its concept to newer forros of this phenomenon, reaching to metropolitanization as an alternative to urban centers to reach a new social, political, economic and cosmopolitan leveI, which best suits its realíty. It is presented here, the current statc of Pernambuco's Central Wasteland Regíon in several respects, especiaIIy on the local economy, which is the drivíng [orce in the region, and the main argument o[ those who defend its metropolítanization. Thereforc, it is necessary to consíder also the recent process of ínternalization of dcvclopment in Brazi!, and in paliicular in the State ofPernambuco. It also made a historical, socio-cconomic and politica! analysis of the main cities in the region, which are Santa Cruz Capibaribc, Toritama and Caruaru, focusing on thc latter, whom by vocation, would be the host cily of thc metropolitan arca of alleged Pernambuco's Central Wasteland Region. Next, it examines the interaction of the cities involved, and the data are compared with the three major metropolitan areas in the interior of the Country, and is drawn a parallel between the eurrent cluster development and the likely benefits of so-called metropolitan area, and finally, analyzed the perception of politicians, on the subject, to present the vision of thís work on the feasibility of metropolitanizatíon in the region studied, and the alternative proposaIs. / Este trabalho tem por finalidade apresentar uma visão sobre a viabilidade da metropolização do Agreste Central de Pernambuco. Para tanto, trata-se aqui de apresentar uma análise inicial sobre o atual estágio da globalização os seus efeitos e formas de sobrevivência dos pequenos centros produtivos neste constante e metamórfico processo. Trata ainda dos processos de regionalização, passando-se pela sua conceituação até as formas mais recentes desse fenômeno, alcançando-se a metropolização como alternativa de centros urbanos para atingir um novo patamar social, político, econômico e cosmopolita, que melhor se adéqua à sua realidade. É apresentada neste trabalho, a situação atual do Agreste Central de Pernambuco, sob diversos aspectos, especialmente quanto à economia local, que é a mola propulsora da região, e o principal argumento daqueles que defendem a sua metropolização. Para tanto, faz se necessário analisar, também, o recente processo de interiorização do desenvolvimento por que passa o Brasil, e, em especial, o Estado de Pernambuco. É feita, ainda, uma avaliação histórica, sócio-econômica e política das principais cidades desta região, que são Santa Cruz do Capibaribe, Toritama e Caruaru, com foco nesta última, que, por vocação, seria a cidade sede da pretensa Região Metropolitana do Agreste Central de Pernambuco. Em seguida, é feita uma análise de interação das cidades envolvidas, e os dados são comparados com as três principais regiões metropolitanas do interior do País. É traçado um paralelo entre o atual pólo de desenvolvimento e os prováveis benefícios da pretensa região metropolitana, c, por fim, analisada a percepção dos Agentes Políticos, sobre o tema, para se apresentar a visão deste trabalho sobre a viabilidade da metropolização da região estudada, c as propostas alternativas.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A ocorrência de fenótipos multirresistentes de Corynebacterium pseudodiphtheriticum e sua associação a infecções graves, com elevada mortalidade em pacientes imunocomprometidos, aliados ao escasso conhecimento da virulência e patogenia destas infecções, motivou esta pesquisa, que teve como objetivo investigar mecanismos de virulência e resistência microbiana deste agente entre pacientes de um hospital universitário brasileiro. Um total de 113 amostras de C. pseudodiphtheriticum identificadas por métodos bioquímicos convencionais e sistema API-Coryne isoladas de pacientes de diferentes grupos etários. Os micro-organismos eram, em sua maioria, relacionados a infecções no trato respiratório (27,45%), urinário (29,20%) e sitios intravenosos (18,60%) e cerca de 32,70% das amostras foram provenientes de pacientes com pelo menos uma das condições predisponentes: insuficiência renal; transplante renal, tuberculose em paciente HIV+, câncer, cirrose hepática, hemodiálise e uso de cateter. As amostras testadas revelaram-se multirresistentes sendo a maioria resistente à oxacilina, eritromicina e clindamicina. A adesão das cepas ao poliestireno e ao poliuretano indicou o envolvimento de hidrofobicidade da superfície celular na fase inicial da formação de biofilmes. O crescimento subsequente conduziu à formação de microcolônias, agregados bacterianos densos incorporados na matriz exopolimérica rodeada por espaços vazios, típica de biofilmes maduros. Adicionalmente, a interação do micro-organismo com fibrinogênio e fibronectina humana indica o envolvimento destes componentes séricos na formação de biofilme, sugerindo a participação de diferentes adesinas neste processo e a capacidade deste agente formar biofilme in vivo. A afinidade por esses componentes e a formação de biofilme podem contribuir para o estabelecimento e disseminação da infecção no hospedeiro. Adicionalmente, as cepas de C. pseudodiphtheriticum isoladas de pacientes com infecções localizadas (ATCC10700/Pharyngitis) e sistêmicas (HHC1507/Bacteremia) exibiram um padrão de aderência agregativa-like a células HEp-2, caracterizado por aglomerados de bactérias com aparência de um "empilhado de tijolos". Através do teste FAS e ensaios de interação na presença de inibidores de citoesqueleto, demonstramos o envolvimento da polimerização de actina na internalização das cepas testadas. A internalização bacteriana e rearranjo do citoesqueleto pareceu ser parcialmente desencadeado pela ativação da tirosina-quinase. Finalmente, C. pseudodiphtheriticum foi capaz de sobreviver no ambiente intracelular e embora não tenha demonstrado capacidade de replicar intracelularmente, células HEp-2 foram incapazes de eliminar o patógeno completamente no ambiente extracelular no período de 24 horas. Todas as cepas estudadas foram capazes de induzir apoptose em células epiteliais 24 horas pós-infecção evidenciada pelo aumento significativo no número de células mortas e pela ocorrência de alterações nucleares reveladas através dos métodos de coloração pelo azul Trypan, pelo DAPI e microscopia electrônica de transmissão. Alterações morfológicas incluindo a vacuolização, a fragmentação nuclear e a formação de corpos apoptóticos foram observadas neste período. A citometria de fluxo demonstrou ainda uma diminuição significativa no tamanho das células infectadas e a utilização de dupla marcação (iodeto de propídio / anexina V) permitiu a detecção da ocorrência de necrose e apoptose tardia. Em conclusão, o conhecimento de tais características contribuiu para a compreensão de mecanismos envolvidos no aumento da frequência de infecções graves com elevada mortalidade em pacientes no ambiente hospitalar, por C. pseudodiphtheriticum, um patógeno rotineiramente subestimado em países em desenvolvimento. / The occurrence of multiresistant phenotypes and associated with severe infections, with high mortality in immunocompromised hosts due to Corynebacterium pseudodiphtheriticum, allied to little known about virulence and pathogenesis these infections, led to present investigation. The investigation aims to examine the virulence mechanisms and resistance to antimicrobial agents of C. pseudodiphtheriticum among patients with bacterial infections at a Brazilian teaching hospital. A total of 113 C. pseudodiphtheriticum strains identified by conventional biochemical methods and API-Coryne System were recovered from patients from different age groups. Micro-organisms were mostly related to infections in the respiratory tracts (27.45%), urinary (29.20%) and intravenous sites (18.60%) and approximately 32.70% samples were obtained of patients presenting at least one of the pre-disposing conditions: end-stage renal disease; renal transplant; AIDS and Mycobacterium tuberculosis infection; cancer, hepatic cirrhosis; haemodialysis and catheter use. Antimicrobial susceptibility tests identified multiresistant phenotypes. Most strains were resistant to oxacillin, erythromycin and clindamycin. Adherence to polystyrene and polyurethane indicated the involvement of cell surface hydrophobicity in the initial stage of biofilm formation. Further growth led to the formation of dense bacterial aggregates embedded in the exopolymeric matrix surrounded by voids, typical of mature biofilms. Data also showed C. pseudodiphtheriticum recognizing human fibrinogen (Fbg) and fibronectin (Fn) and involvement of these sera components in biofilm formation in conditioning films. These findings suggest that biofilm formation may be associated with the expression of different adhesins. C. pseudodiphtheriticum may form biofilm in vivo possibly by an adherent biofilm mode of growth in vitro currently demonstrated on hydrophilic and hydrophobic abiotic surfaces. The affinity to Fbg and Fn and the biofilm-forming ability may contribute to the establishment and dissemination of infection caused by C. pseudodiphtheriticum. Additionally, C. pseudodiphtheriticum strains isolated from patients with localized (ATCC10700/Pharyngitis) and systemic (HHC1507/Bacteremia) infections exhibited an aggregative adherence-like pattern to HEp-2 cells characterized by clumps of bacteria with a stacked-brick appearance. The fluorescent actin staining test demonstrated that actin polymerization is involved in the internalization of the C. pseudodiphtheriticum strains. Bacterial internalization and cytoskeletal rearrangement seemed to be partially triggered by the activation of tyrosine kinase activity. Although C. pseudodiphtheriticum strains did not demonstrate an ability to replicate intracellularly, HEp-2 cells were unable to fully clear the pathogen within 24 hours. All samples were able to induce apoptosis in HEp-2 cells 24 h post-infection, evidenced by significant increase in the number of dead cells and nuclear alterations were observed by the Trypan blue assay, DAPI and transmission electron microscopy. Morphological changes in HEp-2 cells observed 24 h post-infection included vacuolization, nuclear fragmentation and the formation of apoptotic bodies. Flow cytometry revealed an significant decrease in cell size of infected HEp-2 cells. Furthermore, a double-staining assay using Propidium Iodide/Annexin V gave information about the numbers of vital vs. early apoptotic cells and late apoptotic or secondary necrotic cells. In conclusion, these characteristics may contribute to understanding of mechanisms involved on increase of severe infection, with high mortality in nosocomial enviroment patients by C. pseudodiphtheriticum, a pathogen usually overlooked in emerging countries.

L’objectif de cette thèse est de développer des nanoparticules mésoporeuses de silice (MSNs) capables de libérer un anticancéreux, le cisplatine, dans le milieu intracellulaire de tumeurs solides. Une étude paramétrique montre que les propriétés morphologiques, structurales et texturales des MSNs de type MCM-41, synthétisées par voie sol-gel à partir d’un mélange d’un précurseur de silice (TEOS) et de tensioactif (CTAB), dépendent du pH, de la vitesse d’agitation, de la température et du mode d’extraction du CTAB. La synthèse doit avoir lieu sous atmosphère contrôlée pour éviter la présence en solution d’ions carbonates et d’éthanol à l’origine de ponts entre les particules qui altèrent la stabilité colloïdale des MSNs.Les MSNs ont été fontionnalisées pour contrôler l’affinité chimique du cisplatine pour la surface des MSNs et pour améliorer la stabilité colloïdale des MSNs en milieu physiologique. Des organosilanes (APTES, TESP, MPTES, CEST) et des polymères neutres ou ionisés (PEG, PEI) ont été ajoutés par co-condensation ou par greffage post synthèse tout en préservant les propriétés structurales et texturales des MSNs. Les quantités de cisplatine encapsulées par imprégnation sont plus élevées que par adsorption et dépendent de la fonctionnalisation des MSNs. Les MSNs greffées en surface par la PEI permettent une libération progressive de l’anticancéreux ainsi qu’une cytotoxicité comparable à celle du cisplatine administré seul.Des tests in vitro montrent que les MSNs en l’absence de cisplatine ne présentent pas d’effet cytotoxique jusqu’à une concentration de 200 µg/mL. Une étude par microscopie confocale montre une internalisation des MSNs à partir de 2h de mise en contact avec les cellules. / The aim of this PhD thesis is to elaborate mesoporous silica nanoparticles (MSNs) able to sustain the release of cisplatin into the intracellular compartments of solid tumors. The parametric study shows that morphological, structural and textural properties of MSNs-MCM-41, synthesized by sol-gel reaction by using TEOS as a silica source and CTAB as a structure-directing agent, depend on pH, stirring speed, temperature and extraction process of CTAB. The synthesis atmosphere has to be controlled in order to avoid the presence of ethanol and carbonate species which are responsible of necks between particles generating unstable suspensions of MSNs.MSNs were functionalized in order to control the interactions between cisplatin and MSNs or to improve their colloidal stability in physiological media.Various silanated organic molecules (APTES, TESP, MPTES, CEST) and neutral or ionised polymers were grafted into the MSNs using a co-condensation approach or a postsynthetic functionalization method while keeping structural and textural properties of MSNs unchanged.The amount of cisplatin encapsulated by using impregnation are higher than those obtained by using adsorption and depend on MSNs fonctionalization. MSNs grafted by PEI allow a sustained drug release and cytotoxic effects similar to cisplatin. In vitro assays show that MSNs devoid of cisplatin are not cytotoxic up to 200 µg/mL. Confocal microscopy study reveals that MSNs internalization is efficient after about 2h of contact with cells.

Submitted by Kátia Marques Da Silva (azeretolum@outlook.com) on 2018-04-23T23:48:25Z No. of bitstreams: 1 Tese - Katia Marques.pdf: 4561081 bytes, checksum: df3b1f8ff238748716a20d44b5315c27 (MD5) / Approved for entry into archive by Elza Mitiko Sato null (elzasato@ibilce.unesp.br) on 2018-04-24T15:54:07Z (GMT) No. of bitstreams: 1 silva_km_dr_sjrp.pdf: 4561081 bytes, checksum: df3b1f8ff238748716a20d44b5315c27 (MD5) / Made available in DSpace on 2018-04-24T15:54:07Z (GMT). No. of bitstreams: 1 silva_km_dr_sjrp.pdf: 4561081 bytes, checksum: df3b1f8ff238748716a20d44b5315c27 (MD5) Previous issue date: 2018-03-09 / Esta pesquisa, um estudo de caso etnográfico, teve por objetivo investigar, segundo os princípios da perspectiva sociocultural de formação de professores de línguas, apoiada na psicologia sócio-histórica de Vygotsky (1978, 1988, 1987, 1986, 1981, 1992, 1990) e em Johnson e Golombek (2016, 2013, 2011, 2011a, 2011b, 2002, 2003), Johnson (2015, 2009) e Golombek (2011, 2015), a relação da ensinagem de uma professora formadora de Metodologia de Ensino de Língua Estrangeira - Inglês com a ensinagem de um aluno-professor de um Curso de Letras nas aulas de Metodologia e no Estágio Supervisionado. Para atingir esse objetivo principal, foram investigadas as possíveis ocorrências de (co)construção de conceitos nas atividades teórico-práticas desenvolvidas. Os dados foram gerados a partir de diários, desenho pictórico, gravações em áudio, entrevistas, questionários e documentos (planos de aula) produzidos pelo participante da pesquisa. A análise dos dados, com base em conceitos advindos da perspectiva sociocultural, permitiu identificar e selecionar, no curso desses diferentes momentos de observações teórico-práticas, a construção, a coconstrução e a reconstrução de conceitos cotidianos e científicos nas ações do aluno-professor de língua inglesa em formação e a relação destes com a ensinagem da professora formadora. Estes conceitos, chamados de acadêmicos e cotidianos (JOHNSON; GOLOMBEK, 2016), são necessários para o desenvolvimento dialógico do professor em formação inicial, por serem parte de um processo único de construção do conhecimento (VYGOTSKY, 1978). / This study research, an ethnographic case study, aims at investigating the relation of the teaching of a teacher educator of the subject course Metodologia de Ensino de Língua Estrangeira - Inglês to the teaching of a student-teacher in the classes of methodology and practicum of a Curso de Letras of a federal university of the interior of the state of Minas Gerais, based on the principles of a sociocultural perspective of second language teacher education, supported by Vygotsky (1978, 1988, 1987, 1986, 1981, 1992, 1990), and by Johnson and Golombek (2016, 2013, 2011, 2011a, 2011b, 2002, 2003), Johnson (2015, 2009) and Golombek (2011, 2015). Therefore, theoretical-practical activities developed during the second semester of 2014 and the first semester of 2015 were observed and monitored. The data were collected from diaries, drawings, audio recordings, interviews, questionnaires, and also from documents (lesson plans) produced by the participant of this research. The analysis of the data, oriented by the concepts of a sociocultural perspective, contributed to the identification and selection in different moments of theoretical-practical observations, the construction, coconstruction, and reconstruction of academic and everyday concepts in the action of the student-teacher in preservice education in relation to the teaching of the teacher educator. These concepts, named academic and everyday concepts (JOHNSON; GOLOMBEK, 2016), are necessary to the student-teacher’s dialogical development, as they are part of a unique process of construction of knowledge (VYGOTSKY, 1978).