Submitted by Rosangela Silva (rosangela.silva3@unioeste.br) on 2018-05-25T11:53:11Z No. of bitstreams: 2 Indianara Kawana Bueno.pdf: 2050489 bytes, checksum: 3865a86596759f1ff761267b04d8a615 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-05-25T11:53:11Z (GMT). No. of bitstreams: 2 Indianara Kawana Bueno.pdf: 2050489 bytes, checksum: 3865a86596759f1ff761267b04d8a615 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-03-09 / The research for renewable energy sources became even more essential due the imminent depletion of the fossil fuel sources. In this context Brazil has a prominent position on the world stage, since it has already used ethanol from sugar cane for some decades. The second generation ethanol (2G) is produced from the lignocellulosic biomass of the vegetable, which is composed by cellulose, hemicellulose and lignin. The hydrolysis of these compounds requires a specific and high cost enzymatic cocktail. On this scenario, the Trichoderma reesei fungus gains spotlight, since it is one the microorganisms with the highest potential to produce hydroliytic enzymes. Therefore, the attempt to increase the cellulases production of this fungus is an important for the production of biofuels more attractive to the market. The aim of this work is to confirm the deletion of the sequence which codifies the zinc finger motif of the transcription factor ACE1 for cellulose repression from the T. reesei RUT-C30 strain and to characterize the enzymatic production of these mutant strains named T. reesei RUT-C30Δzface1. The enzymatic quantification was carried using the substrates carboxymethyl cellulose, microcrystalline cellulose and Whatman paper filter. The deletion confirmation occurred by the absence of the amplification gene ace1 on the mutants and the amplification of a 429 pb fragment of the RUT-C30 parental strain when the same primers and PCR conditions where used. These results suggest that the deletion of the zinc finger motif of the from ACE1 transcription factor is a prominent way to achieve an economically viable production of bioethanol. / Com a depleção eminente das fontes de combustíveis fósseis, torna-se cada vez mais imprescindível a busca por fontes renováveis de energia. Neste âmbito, o Brasil tem destaque no cenário mundial, pois já utiliza o etanol a partir da cana-de-açúcar há algumas décadas. O etanol de segunda geração (2G) é produzido a partir da massa lignocelulolítica do vegetal, que é composta de celulose, hemicelulose e lignina. A hidrólise desses compostos necessita de um coquetel enzimático específico e de alto custo. Neste cenário, o fungo Trichoderma reesei ganha destaque, pois é um dos microrganismos com maior potencial para produção de enzimas hidrolíticas. Desta forma, as tentativas de aumentar a produção de celulases desse fungo, torna a produção do bioetanol uma alternativa mais atrativa ao mercado. Este trabalho teve como objetivos confirmar a deleção da sequência que codifica o dedo de zinco do fator de transcrição do repressor de celulase ACE1 da linhagem T. reesei RUT-C30 e caracterizar a produção enzimática dessas linhagens mutantes denominadas T. reesei RUT-C30Δzface1. A confirmação de deleção ocorreu pela ausência de amplificação do gene ace1 nos mutantes e amplificação de um fragmento de 479 pb na linhagem parental RUT-C30, quando utilizados os mesmos primers e condições de reação de PCR. A dosagem enzimática com os substratos carboximetilcelulose (CMC), celulose microcristalina (Avicel®) e papel de filtro Whatman (PF), mostraram que o RUT-C30Δzface1 tem a atividade celulolítica aumentada em até 3,2 vezes em Avicel e 2,1 vezes em CMC e PF em comparação à linhagem parental RUT-C30. Em 24 horas de hidrólise os mutantes apresentaram liberação de açúcar 1,4 vezes maior em relação ao RUT-C30. Estes resultados sugerem que a deleção parcial do fator de transcrição ACE1 é um proeminente caminho para a conquista de uma produção de bioetanol economicamente viável.

As mutações no gene PROP1 são a causa genética mais comum da deficiência combinada de hormônios hipofisários. Até o momento, diversas mutações missense e pequenas deleções foram descritas sendo a mutação 301-302 delAG a mais freqüente. Nosso objetivo foi estudar as mutações em DNA de pacientes com hipopituitarismo e padronizar a extração de DNA de células de swab oral, usando um método com NaCl e comparar com um kit comercial (Purigene, Minneapolis, EUA). Amplificamos os 3 exons do gene PROP1 do DNA obtido de células orais e de sangue periférico. Identificamos a mutação 301-302delAG em 6 pacientes, 4 em homozigose (33%) e 2 em heterozigose (16%) e a mutação G51A em heterozigose em um único paciente. Em dois irmãos, filhos de pais consangüíneos, não foi possível amplificar os 3 exons do gene PROP1 enquanto que os os genes LHX3 e LHX4 foram amplificados com sucesso. Para confirmar a hipótese de deleção do PROP1, o Southern blotting foi realizado usando como sonda o produto de PCR do exon 2 do gene PROP1 e um fragmento do gene CYP21A2 como sonda controle. A banda referente ao CYP21A2 estava presente nos pacientes e nos controles enquanto a banda referente ao PROP1 estava ausente nos irmãos e presente na mãe e nos controles. Para definir a extensão da deleção usamos um mapa de STS próximos ao gene e o STS GDB:314805 localizado a 6,0 kb a montante do PROP1 não foi amplificado nos pacientes. Entretanto, o gene Q8N6H0 a 18 kb a juzante e o STS WI-16216 a 59 kb a montante do PROP1 foram amplificados com sucesso nos pacientes e controles indicando que a deleção está localizada dentro de 81 Kb. Para determinar os limites da deleção, várias reações de PCR foram realizadas com primers desenhados progressivamente distantes de gene PROP1, cobrindo toda a região. Isto nos permitiu determinar a região deletada de 9,6 kb a juzante e 11 kb a montante do gene PROP1, com o tamanho máximo deletado de 18,4 kb. Por ambos os métodos de extração obtivemos um DNA de boa qualidade, permitindo o amplificação dos 3 exons do gene PROP1. A extração com NaCl foi mais rápida e mais barata resultando em maior quantidade de DNA quando comparada com o kit comercial. Em conclusão, descrevemos a deleção completa do gene PROP1 em dois irmãos com o fenótipo clássico de hipopituitarismo associado à hipófise hipoplásica ou aumentada e padronizamos a extração de DNA de células de mucosa oral com NaCl, que apresentou custo mais baixo e resultado mais rápido quando comparado a extração por um kit comercial, indicando que o swab oral é uma fonte prática de obtenção de DNA para estudos genéticos. / PROP1 gene mutations are the most common cause of genetic combined pituitary hormone deficiency. To date, several missense mutations and small deletions have been described and the 301-302 del AG is the most frequent. Our objective was to study PROP1 mutations in patients with hypopituitarism and standardize DNA extraction from an oral swab, using the NaCl method, comparing it with a commercial kit (Purigene, Minneapolis, USA). We amplified the 3 exons of PROP1 gene in DNA obtained from oral cells and peripheral blood cells. We identified the delAG301-302 mutation in 6 patients, 4 in homozygous (33%) and 2 in heterozygous (16%) state and G51A mutation in heterozygous state in a single patient. In two siblings, a boy and a girl, born to consanguineous parents we failed to amplify PROP1 gene by PCR whereas LHX3 and LHX4 genes were successfully amplified. To confirm the hypothesis of PROP1 gene deletion, Southern blotting was performed using PROP1 exon 2 gene PCR product as a probe and a fragment of CYP21A2 gene as a control probe. The CYP21A2 band was present in patients and controls whereas PROP1 band was absent in both siblings and present in their mother and in controls. To define the extension of this deletion we used STS mapping approach and no amplification for a STS GDB:314805 6.0 kb downstream of PROP1 was found. However, Q8N6H0 gene located 18 kb upstream and the STS WI-16216 located 59 kb downstream of PROP1 were successfully amplified indicating that the deletion is placed within 81 Kb. To determine the limits of the deletion a number of PCR covering this region were then carried out with primers located progressively distant from PROP1. This allowed us determine the deleted region from 9.6 kb upstream to 11 kb downstream of PROP with a maximum deletion size of 18.4 kb. Both methods yielded good quality DNA, allowing the amplification of 3 exons of PROP1 gene. The NaCl method showed to be faster and less expensive, resulting in a larger amount of DNA when compared with the commercial kit. In conclusion, we describe a complete deletion of PROP1 gene in two siblings with classical hypopituitarism phenotype associated with hypoplastic or enlarged pituitary gland and standardized the DNA extraction of oral cells with NaCl, which presented lower costs and faster results, when compared with the extraction by a commercial kit indicating that oral swabs are a reliable DNA source for genetic studies.

Objetivo: Descrever as manifestações clínicas de indivíduos com hipótese diagnostica da Sindrome de deleção 22q11 (SD22q11) confirmados por testes genéticos, na primeira avaliação e durante o acompanhamento dos mesmos em avaliações subsequentes para uma melhor definição do curso natural da doença. Local: Laboratório de Genética e Citogenética Humana do HRAC-USP Bauru/SP. Casuística e metodologia: O presente trabalho é retrospectivo e analisou os dados de prontuários de 72 indivíduos cadastrados no HRAC-USP, os quais receberam hipótese diagnóstica da SD22q11 e foram confirmadas por teste genético (MLPA ou FISH). A avaliação envolveu a analise dos dados relatados por todos os setores do HRAC-USP. Resultados e discussão: Foramanalisados 72prontuários deindivíduos com a SD22q11. Constatamos que a idade media dos indivíduos quando do cadastro no HRAC-USP foi de seis anos. Também constatamos que houve um longo período de tempo entre os retornos ao hospital e que, nesses retornos, nem todas as especialidades foram contempladas. Esses fatos prejudicaram a analise da historia natural da anomalia em questão. Com relação às características fenotipicas, observamos a presença de sinais clínicos típicos, como por exemplo: face alongada, lábios finos, hipoplasia alar, anormalidades menores na orelha, dígitos longos e fendas palpebrais, fissuras labiopalatinas, cardiopatias congenitas, dificuldade de aprendizagem, atraso de linguagem e distúrbios comportamentais. A fissura oral foi à manifestação otorrinolaringológica mais frequente, presente em 75% dos pacientes, onde as fissuras submucosas foram as mais frequentes (43%). As características cognitivas como, atraso de fala (87%), dificuldades de aprendizagem (95%) e distúrbios comportamentais (81%), tiveram um resultado significativo, descritas em quase todos os indivíduos. As cardiopatias congênitas estavam presentes em 47,2% dos prontuários analisados. De um modo geral, comparando a frequência dos sinais clínicos encontrados neste trabalho com dados da literatura, constatamos que as frequências encontram-se dentro do esperado. Conclusão: A maioria dos indivíduos cadastrados no HRAC-USP, pertencentes ao grupo de estudo, apresentou idade superior a 06 anos. Portanto, a observação do curso natural da historia da SD22q11 para avaliar características fenotípicas que surgissem ao longo da evolução clinica do indivíduo e que pudessem ajudar no diagnóstico, ficou prejudicada. Mesmo nos casos onde o indivíduo foi cadastrado no HRAC-USP com idade inferior a dois anos, o diagnóstico foi tardio devido a falta de uma ação multidisciplinar e interdisciplinar no hospital. Mesmo não sendo possível avaliar as características fenotípicas surgidas durante a historia natural da doença, constatamos que as manifestações clínicas relatadas nos prontuários cursam com as características da SD22q11 e em frequências que corroboram com as da literatura / To describe clinical manifestations observed in medical records of individuals registered in the hospital with a diagnostic hypothesis of 22q11.2DS confirmed by genetic tests (MLPA OR FISH), since the first assessment in the HRAC-USP and during the follow up of these individuals in subsequent assessments, in order to achieve a better definition to the natural courses of the disease. Local: Laboratory of Human Genetics and Cytogenetics (HRAC-USP Bauru/SP). Methods: This retrospective study analyzed 72 medical records of individuals registered at the HRAC-USP, who were diagnosed with 22q11DS and who had this diagnosis confirmed by a genetic test (MLPA OR FISH). The assessment concerned the analysis of reported data in all sectors of the HRAC-USP. Results and Discussion: 72 medical records of individuals with 22q11DS were analyzed. It was verified that the average age of individuals when registering at the HRAC-USP was six years old. It was also verified that it took a long period of time for these individuals to return to the hospital and, when they did, not all specialties were contemplated. These facts harmed the analysis of the natural history of the anomaly. About the phenotypic characteristics, some typical clinical signs were observed, such as: long face, thin lips, hypoplasia nasal alar, minor abnormalities in the ear, long digits and narrow palpebral fissures, palatal abnormalities, congenital heart defects, learning disabilities, delay speech and behavioral disorders. An oral cleft was the most frequent otorhinolaryngology manifestation, present in 75% of the patients; among which submucous cleft palate were the most frequent (43%). Cognitive features such as, delay speech (87%), learning disabilities (95%) and behavioral disorders (81%) had a significant result, described in almost all individuals. Congenital heart defects were observed at 4% to 48% of individuals with 22q11.2DS, in this study it was observed in 47.2%. In general, comparing the frequency of some clinical signs observed in this study with the literature data, it was verified that the frequencies were within expectations. Conclusion: Most of the individuals registered at the HRAC belonging to the study group were over 6 years old. Therefore, the observation of natural course of the history of 22q11DS to evaluate the phenotypic characteristics that would arise during the clinical evolution of the individual and that could help in the diagnosis was harmed. Even in cases when the individual was registered at the HRAC-USPunder the age of two, the diagnosis was delayed due to lack of a multidisciplinary and interdisciplinary action in the hospital. Even not being possible to measure the phenotypic characteristics that emerged during the natural history of the disease, it was verified that the clinical manifestations reported in the records occur with the 22q11DS characteristics and in frequencies that corroborate with the literature

Introdução: A Holoprosencefalia (HPE) é uma malformação craniofacial decorrente de falhas no processo de formação do sistema nervoso durante o desenvolvimento embrionário. Sua etiologia é heterogênea e complexa, envolvendo fatores ambientais e genéticos. Estudos recentes sugerem que alterações subteloméricas possam ser um dos fatores etiológicos para o aparecimento da HPE. Objetivos: Investigar possíveis alterações (microdeleções/duplicações) nas regiões subteloméricas em indivíduos com diagnóstico de HPE. Metodologia: Avaliação genética de 25 amostras de DNA de indivíduos matriculados no HRAC-USP, com diagnóstico de HPE através da técnica de MLPA (Multiplex ligation-dependent probe amplification), segundo protocolo proposto por Schouten et al. (2002). Estes indivíduos já foram previamente triados para mutações/deleções nos genes candidatos à HPE (SHH, ZIC2, TGIF, GLI2 e PTCH1) através do sequenciamento direto de Sanger e da técnica de MLPA, sem resultado positivo para qualquer alteração. Resultados: Dentre os 25 indivíduos analisados, o fenótipo predominante foi a microforma da doença. Os principais achados clínicos da amostra estudada foram: hipotelorismo, microcefalia e fissura de lábio e palato (100%), nariz achatado (84%), presença de um incisivo central único (24%) e ponte nasal baixa (64%). Quatro pacientes apresentaram comprometimento no SNC (16%). Nenhum indivíduo apresentou quaisquer alterações, como microdeleções e/ou duplicações nos genes contidos nas regiões subteloméricas, através da análise pela técnica de MLPA. Dessa forma, estes permanecem sem diagnóstico genético definido. Discussão: A não detecção de alterações subteloméricas sugere que o fenótipo predominante na amostra, microforma, possa não apresentar alterações subteloméricas relacionadas ao aparecimento da doença. Entretanto, deve-se salientar que o universo amostral é relativamente pequeno, de forma que este possa não ser um exemplo fiel dos casos de microforma da HPE. Reforça-se, ainda, a grande variedade de fatores envolvidos no surgimento desta patologia, bem como o envolvimento de outros genes ainda não estabelecidos, além das causas ambientais, ainda não completamente elucidadas. Conclusões: Não foram encontradas alterações subteloméricas nos pacientes com diagnóstico de HPE estudados. A técnica de MLPA consiste em uma metodologia rápida, sensível, eficaz e de baixo custo, quando comparada a outras técnicas, sendo indicada para o uso em laboratórios de diagnóstico genético, uma vez que diversos estudos já mostraram que consiste em um método confiável de diagnóstico. Devido ao tamanho relativamente pequeno da amostra utilizada neste estudo, e os dados ainda inconsistentes da literatura atual, estudos adicionais são necessários para que seja possível a realização de um diagnóstico diferencial, explicando o amplo espectro fenotípico observado nesta doença, conforme sugerido pela hipótese dos múltiplos fatores. / Introduction: Holoprosencephaly (HPE), a craniofacial malformation, results from flaws in formation process of the nervous system during embryonic development. The etiology is heterogeneous and complex, involving environmental and genetic factors. Recent studies suggest that subtelomeric aberrations could be an etiological factor to the onset of HPE. Objectives: Investigate possible changes (microdeletions/duplications) in subtelomeric regions in individuals diagnosed with HPE. Methodology: Genetic evaluation of 25 DNA samples from individuals enrolled at HRAC-USP, diagnosed with HPE (performed by Syndromology Division HRAC/USP) by MLPA technique, as proposed by Schouten et al (2002). Patients were previously screened for mutations/deletions in HPE candidate genes (SHH ZIC2, TGIF, GLI2 and PTCH1) by direct Sanger sequencing and MLPA technique, without any match. Analyses were performed at the Laboratory of Molecular Genetics, Hospital for Rehabilitation of Craniofacial Anomalies, HRAC-USP. Results: Among the 25 individuals analyzed, the predominant phenotype was HPE microform. The main clinical findings of the study sample were: hypotelorism, microcephaly, and cleft lip/palate (100%); flat nose (84%); presence of a single central incisor (24%) and low nasal bridge (64%). Four patients had CNS commitment (16%). No subtelomeric mutations were found in our sample, such as microdeletions/duplications of genes analyzed by MLPA technique. Thus, they remain without defined genetic diagnosis. Discussion: Subtelomeric changes were not found, suggesting that the predominant sample phenotype, microform HPE, could not be related with subtelomeric changes associated to the disease outbreak. However, it should be noted that the sample universe is relatively small, so this may not be a true example of HPE microform cases. It should also be reinforced the wide variety of factors involved in the onset of this pathology, as well as the involvement of other genes not yet established and environmental causes, not completely understood. Conclusions: No subtelomeric mutations were found in the HPE individuals studied. MLPA technique consists in a rapid, sensitive and cost effective methodology, when compared to other techniques being suitable for use in genetic diagnostic laboratories, since several studies have shown that consists of a reliable method of diagnosis. Due to the relatively small sample size used in this study, and even inconsistent data in literature, further studies are needed to make it possible to perform a differential diagnosis, explaining the wide phenotypic spectrum observed in this disease, as suggested by the multiple hit hypothesis.

Objetivos: estudar indivíduos com (G1) e sem (G2) sinais da Síndrome Velocardiofacial (SVCF) para verificar diferenças entre eles quanto à extensão e espessura velar, profundidade nasofaríngea, razão entre profundidade nasofaríngea e extensão velar (PNF/EV), tamanho da falha velofaríngea, ângulo velar, movimento do véu palatino e das paredes laterais e posterior da faringe e à presença da tonsila faríngea; diferenças para as medidas de extensão e espessura velar, profundidade nasofaríngea e razão PNF/EV dos grupos estudados com os valores de normalidade propostos por Subtelny (1957); e correlação entre o tamanho da falha velofaríngea e a razão PNF/EV. Material e Método: estudo prospectivo com 60 indivíduos de ambos os sexos sem fissura palatina evidente e com disfunção velofaríngea (DVF), não operados, sendo 30 com sinais clínicos da SVCF (G1) (idade de 5,4 a 51 anos, com média de 15,7±9,5 anos) e 30 sem os sinais da SVCF (G2) (idade de 4,5 a 33 anos, com média de 15±7,6 anos). O exame videofluoroscópico foi realizado nas projeções lateral e frontal para análise da extensão e espessura velar, profundidade nasofaríngea, falha velofaríngea e ângulo velar, movimento do véu palatino, paredes laterais e posterior da faringe e presença da tonsila faríngea. Resultados: quanto às medidas de extensão e espessura velar, profundidade nasofaríngea e razão PNF/EV, não houve diferença entre os grupos quando se comparou os casos maiores de 18 anos, bem como os menores de 18 anos pareados por idade; mas quando a idade não foi pareada nos casos menores de 18 anos, a espessura velar foi menor (p=0,019) e a razão PNF/EV foi maior (p=0,048) no G1 e, ao se analisar independente da faixa etária, a razão PNF/EV foi maior no G1 (p=0,024). Em relação às medidas de normalidade, a extensão velar foi menor no G1 (p=0,007), a espessura velar foi menor no G1 (p=0,000) e G2 no (p=0,000), a profundidade nasofaríngea e a razão PNF/EV foram maiores no G1 (p=0,000) e no G2 (p=0,000), entretanto ao se considerar a variação de 2 desvios-padrão em relação aos valores de normalidade, não houve diferença entre os grupos para todas as medidas. Também não houve diferença entre os grupos quanto ao ângulo de elevação velar (p=0,232) e presença (p=0,698) e tamanho da falha velofaríngea (p=0,293), movimento velar (p=0,085) e das paredes laterais (p=0,763) e posterior (p=0,237) da faringe, além do tamanho da tonsila faríngea (p=0,307). Não houve correlação entre a falha velofaríngea e a razão PNF/EV no G1 (p=0,153) e no G2 (p=0,598). Conclusões: a razão PNF/EV foi maior nos indivíduos com DVF e sinais da SVCF comparados aos indivíduos com DVF sem os sinais da síndrome, sugerindo ser este um indicador velofaríngeo para a SVCF, enquanto os aspectos funcionais da velofaringe não diferiram entre os indivíduos com e sem os sinais da SVCF. Não houve correlação entre o tamanho da falha no fechamento velofaríngeo e a razão PNF/EV nos grupos com e sem sinais da SVCF. / Objective: to study individuals with (G1) and without (G2) signs of velocardiofacial syndrome (VCFS), so as to verify differences in terms of length and thickness of the soft palate, nasopharyngeal depth, ratio of nasopharyngeal depth to velar length (PD/VL), velopharyngeal gap size, velar angle, soft palate movement, lateral and posterior pharyngeal walls movement and the presence of adenoidal tissue; differences for the measurements of velar length and thickness, nasopharyngeal depth and PD/VL ratio for the groups studied with the normality values proposed by Subtelny (1957); and correlation between the size of velopharyngeal gap and the PD/VL ratio. Methods: a prospective study with 60 subjects from both genders, with no evident cleft palate, with velopharyngeal dysfunction (VPD), non operated, being 30 with clinical signs of VCFS (age range 5.4 to 51 yrs, mean 15.7±9.5 yrs), and 30 with no signs of VCFS (age range 4.5 to 33 yrs, mean 15±7.6 yrs). The videofluoroscopy was performed in lateral and frontal views, for the analysis of velar length and thickness, nasopharyngeal depth, velopharyngeal gap, velar angle, soft palate movement, lateral and posterior pharyngeal walls movement and the presence of adenoidal tissue. Results: there was no difference in the measurements of velar length and thickness, nasopharyngeal depth and PD/VL ratio, between the groups, when cases over 18 yrs, as well as those under 18, paired by age, were compared; however, when age was not paired in the cases under 18 yrs, the velar thickness was smaller (p=0.019) and the PD/VL ratio was greater (p=0.048) in G1 and, by analyzing regardless the age range, the PD/VL ratio was greater in G1 (p=0.024). In relation to normality measurements, the velar length was smaller in G1 (p=0.007), the velar thickness was smaller in G1 (p=0.000) and G2 (p=0.000), the nasopharyngeal depth and the PD/VL ratio were greater in G1 (p=0.000) and G2 (p=0.000), nevertheless, when considering the variation of 2 standard deviations in relation to the normality values, there was no difference between the groups for all measurements. No difference was seen between the groups, as to the velar angle (p=0.232) and presence (p=0.698) and size of velopharyngeal gap (p=0.293), velar movement (p=0.085) and lateral (p=0.763) and posterior (p=0.237) pharyngeal walls movement, besides the size of adenoidal tissue (p=0.307). No correlation was seen between the velopharyngeal gap and the PD/VL ratio in G1 (p=0.153) and G2 (p=0.598). Conclusions: the PD/VL ratio was greater in individuals with VPD and signs of VCFS, as compared to individuals with VPD with no signs of the syndrome, suggesting that this is a velopharyngeal indicator for VCFS, whereas velopharyngeal functional aspects did not differ between individuals with and without signs of VCFS. There was no correlation between the size of the velopharyngeal gap and the PD/VL ratio, in the groups with and without signs of VCFS.

Orientadores: Andréa Trevas Maciel Guerra, Maricilda Palandi de Mello / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-23T00:38:47Z (GMT). No. of bitstreams: 1 Santos_AnaPaulados_M.pdf: 2717833 bytes, checksum: 8a3a2ff5cccd42ca60c0bc51ba3b0067 (MD5) Previous issue date: 2013 / Resumo: As disgenesias gonadais parcial (DGP) e mista (DGM) caracterizam-se por ambiguidade genital e presença de gônada disgenética associada a testículo disgenético ou dois testículos disgenéticos. Na DGP o cariótipo é 46,XY; na DGM, há mosaico 45,X/46,XY ou suas variantes (mais de duas linhagens e (ou) anomalias estruturais do cromossomo Y). Esses mosaicos podem determinar, ainda, fenótipo feminino com síndrome de Turner (ST), distúrbio da diferenciação do sexo ovotesticular (DDS OT) e esterilidade em homens com genitais normais. Independentemente do fenótipo gonadal e genital, esses indivíduos apresentam outros sinais clínicos decorrentes da linhagem 45,X, como baixa estatura, dismorfismos, anomalias cardíacas e renais e diversas afecções adquiridas. Nos últimos anos surgiram evidências de ligação entre microdeleções do Y e o mosaicismo com linhagem 45,X. Há, ainda, indicações de que a instabilidade cromossômica trazida por essas deleções possa ser mais pronunciada nas gônadas. O objetivo deste trabalho foi investigar a presença de microdeleções do Y em indivíduos com DGP e naqueles com mosaico 45,X/46,XY ou suas variantes e diferentes fenótipos. A casuística constou de 15 indivíduos com DGP e 15 com mosaicismo, dos quais a maioria apresentava DGM (11 casos). Foram analisados 38 sequence tagged sites (STS) cobrindo a região específica masculina (MSY, male specific region) em Yp, centrômero e Yq por meio da técnica de reação em cadeia da polimerase (PCR) multiplex e individual. Todos os STS investigados nos indivíduos com DGP tiveram amplificação positiva, porém havia STS de Yq ausentes em seis indivíduos com mosaicismo e DGM, dos quais dois sem alterações estruturais de Y evidentes ao cariótipo. Essas deleções se localizavam em regiões contendo genes relacionados à espermatogênese (AZFb e AZFc - azoospermia factor). A ausência de deleções nos indivíduos com DGP não confirma a hipótese de que a instabilidade desse cromossomo nas gônadas seja uma das causas dessa afecção. Por outro lado, as deleções encontradas no segundo grupo indicam, em alguns casos, associação entre alterações estruturais do Y detectáveis somente a nível molecular e o surgimento de mosaicismo. Caso sejam criados no sexo masculino e busquem procedimentos de fertilização in vitro, há risco de que esses indivíduos transmitam cromossomos Y instáveis na divisão celular / Abstract: Partial and mixed gonadal dysgenesis (PGD and MGD) are characterized by genital ambiguity and the finding of either a streak gonad and a dysgenetic testis or two dysgenetic testes. In PGD there is a 46,XY karyotype, whereas in MGD there is a 45,X/46,XY mosaic or its variants (more than two lineages and/or structural abnormalities of the Y chromosome). These mosaics are also compatible with a female phenotype and Turner syndrome, ovotesticular disorder of sex development, and infertility in men with normal external genitalia. Regardless of the gonadal and genital phenotypes, these individuals present other clinical features associated with the 45,X cell line, including short stature, dysmorphisms, cardiovascular and renal anomalies and various acquired diseases. During the last few years, evidences of a link between Y microdeletions and 45,X mosaicism have been reported. There are also indications that the instability caused by such deletions might be more significant in germ cells. The aim of this work was to investigate the presence of Y chromosome microdeletions in individuals with PGD and in those with 45,X/46,XY mosaicism or its variants and variable phenotypes. Our sample comprised 15 individuals with PGD and 15 with mosaicism, most of them with a MGD phenotype (n=11). Thirty-eight sequence tagged sites (STS) spanning the male specific region (MSY) on the Y chromosome (Yp, centromere and Yq) where analyzed by multiplex PCR and some individual reactions. All STS showed positive amplifications in the PGD group. Conversely, in the group with mosaicism, six individuals with MGD had been identified with Yq microdeletions, two of them did not have structural abnormalities of the Y chromosome recognized by routine cytogenetic analysis. The deleted STSs were located within AZFb and AZFc (Azoospermia Factor) regions, which harbor several genes responsible for spermatogenesis. Absence of deletions in individuals with PGD does not confirm the hypothesis that instability of the Y chromosome in the gonads could be one of the causes of such condition. However, deletions identified in the second group indicate that mosaicism may be associated with Y chromosome abnormalities detectable only at the molecular level. If patients with mosaicism and Y microdeletions reared as males decide to undergo in vitro fertilization, Y chromosomes which tend to be unstable during cell division may be transmitted to offspring / Mestrado / Ciencias Biomedicas / Mestra em Ciências Médicas

We propose a new, continuous model of the fractionation process (duplicate gene deletion after polyploidization) on the real line. The aim is to infer how much DNA is deleted at a time, based on segment lengths for alternating deleted (invisible) and undeleted (visible) regions. After deriving a number of analytical results for "one-sided" fractionation, we undertake a series of simulations that help us identify the distribution of segment lengths as a gamma with shape and rate parameters evolving over time. This leads to an inference procedure based on observed length distributions for visible and invisible segments. We suggest extensions of this mathematical and simulation work to biologically realistic discrete models, including two-sided fractionation.

This is essentially a phonological analysis of the Dholuo loanwords derived from English and Swahili. This study examines loanword adaptation at three levels: phonemic, phonotactic and prosodic. The study analyses the strategies that the language has used in adapting the foreign phonemes to the native phoneme system. It also examines the way foreign consonant and vowel clusters are adapted to the Dholuo system and how the stress systems in the source languages are adapted to the Dholuo tonal pattern. The Dholuo principles of syllabification are also examined. On adaptation of incoming sounds into the language, the study determined that Dholuo replaces such foreign segments with native sounds which are acoustically and auditorily closest to the foreign sounds. Some foreign sounds, however, are adopted into the sound system of the language, either to fill some phonological gaps in the language or for non-linguistic factors, like the prestige value. The study found that the native speaker-hearer has knowledge of the possible phonetic sequences in his language and performs the simplest possible adaptation in the loanword to make it correspond to these well-formed sequences. This extends to the insertion or deletion of foreign segments to make a loanword conform to the syllable structure constraints of the native system. The study reveals that Dholuo employs several strategies to nativize unnatural, non-canonic syllable structures: epenthetic vowel insertion, extrasyllabic consonant or vowel deletion, devocalization of unnatural vowel sequences, addition of a final vowel, and in some cases, consonant clusters may be tolerated. At the suprasegmental level, the study reveals that stress in the source languages is generally rendered as high tone in the language, while the stressed vowel in the loanword generally determines the ATR harmony in the loanword. The study revealed that if the first syllable in the loanword is stressed, then the loanword will be rendered with +ATR in Dholuo, while an unstressed first syllable will lead to a loanword with –ATR harmony. The study concludes that the means employed by a given language for the adaptation of unnatural, non-canonic syllable shapes are, in a general sense, peculiar to that language, and have nothing to do with the internally-motivated morpheme structure or phonological rules of the target language. The study also concludes that foreign phonemes are directly mapped onto corresponding native phonetic forms, and there is very scarce evidence in the data to support the theory that loanwords are nativized at the abstract phonological level of the target language. / Thesis (DLitt (African Languages))--University of Pretoria, 2005. / African Languages / unrestricted

The study is to establish whether there is any impact on the social media company that is implementing the deceptive design on the account deletion process. By conducting the qualitative research with Instagram users and analyzing users’ feelings, opinions, and potential actions, the study finds out using the deceptive design on account deletion process on social media has a weak influence on the social media company.

Branding dates back to centuries ago when traders were trying to distinguish their products from others in order to promise a higher quality to their consumers. Today, brands are considered as intangible resources that can have a significant contribution to the firm performance. Based on the Resource-Based Theory (RBT), valuable, rare, inimitable, and non-substitutable brands are strategic resources that create superior value and play a key role in achieving a sustainable competitive advantage over rivals. In the process of developing and maintaining strong brands, brand managers constantly need to make multiple decisions. Whether to add, delete or retail brands are among the routine decisions that brand managers face in managing their brand portfolios. Brand managers need to regularly assess their brand portfolios in order to make sure they are not selling redundant brands. Through brand portfolio assessment, brand managers can recognize weak brands and delete the unprofitable brands from the portfolio in order to free up resources and reinvest them in their stronger and more successful brands to gain competitive advantage in the market. This admonition is in line with the RBT of competitive advantage. This dissertation builds upon and extends previous literature on RBT in the context of brand deletion to achieve three main objectives. The first objective is to find the answer to why companies decide to delete brands from their portfolios. Thus, the focus of the first objective is to identify the organizational (i.e., firm, managerial, and brand) factors that drive the brand deletion strategy in a company. The second goal is to find the answer to the when question through identifying the environmental (i.e., market) factors associated with brand deletion decision making in a company. Finally, the third objective is to go deeper and investigate the different types of brand deletion strategy (i.e., merge, sell, milk, and kill). In other words, the third objective seeks to find the answer to the how question. Deleting brands from the portfolio of a company, being the most sensitive issue in strategic brand portfolio management, is yet understudied in the brand portfolio management literature. This study adds to the literature of strategic brand portfolio management by a) applying the Resource-based Theory (RBT) in the context of brand deletion decision making and b) empirically testing the relationships among the drivers of brand deletion strategies. The findings of this dissertation provide a better understanding on how each of these factors are associated with the brand deletion decision making process in companies. The current dissertation provides practitioners with several managerial insights as well. First, the study identifies and empirically tests several organizational-level factors that drive brand deletion decisions in companies. This will help brand managers be familiar with factors that they need to consider when evaluating their poor-performing brands. Breaking these factors into internal (brand and firm) and external (market) drivers provides practitioners with a better understanding of the brand deletion decision making process. In addition, the findings of this study help managers realize their own role (in terms of their attitude toward deletion and their commitment to the brand) in the brand deletion process. Finally, the identification and discussion of the four types of brand deletion strategy help companies have a clearer picture of how they can remove brands from their portfolios.