by Chan Chuek Kee. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1992. / Includes bibliographical references (leaves 165-201). / Abstract --- p.i / Table of Content --- p.iv / List of Abbreviations --- p.viii / List of Tables --- p.x / List of Figures --- p.xii / Acknowledgement --- p.xiii / Chapter Chapter 1. --- Introduction / Chapter 1.1. --- Discovery and historical events --- p.1 / Chapter 1.2. --- General characteristics of rotavirus --- p.3 / Chapter 1.2.1. --- Virus morphology --- p.3 / Chapter 1.2.2. --- Physicochemical properties --- p.3 / Chapter 1.2.3. --- Virus stability and inactivation --- p.4 / Chapter 1.2.4. --- Genome structure --- p.4 / Chapter 1.2.5. --- Rotavirus proteins --- p.5 / Chapter 1.2.6. --- Morphogenesis --- p.8 / Chapter 1.3. --- Characteristics of rotavirus infection --- p.9 / Chapter 1.3.1. --- Morbidity and mortality --- p.9 / Chapter 1.3.2. --- Clinical features --- p.11 / Chapter 1.3.3. --- Pathogenesis --- p.13 / Chapter 1.3.4. --- Seasonal variation of rotavirus infection --- p.13 / Chapter 1.3.5. --- Nosocomial rotavirus infection --- p.14 / Chapter 1.3.6. --- Route of transmission --- p.14 / Chapter 1.4. --- Classification and epidemiology of rotaviruses --- p.15 / Chapter 1.4.1. --- Rotavirus groups --- p.15 / Chapter 1.4.2. --- Rotavirus subgroups --- p.16 / Chapter 1.4.3. --- Rotavirus serotypes --- p.17 / Chapter 1.4.4. --- Rotavirus electropherotypes --- p.20 / Chapter 1.4.5. --- "Relationship between subgroup, serotype and electropherotype" --- p.23 / Chapter 1.4.6. --- Mechanisms contributing to strain variations --- p.24 / Chapter 1.5. --- Detection of rotavirus --- p.28 / Chapter 1.5.1. --- Electron microscopy (EM) --- p.28 / Chapter 1.5.2. --- Virus isolation --- p.29 / Chapter 1.5.3. --- Serological methods --- p.30 / Chapter 1.5.4. --- RNA analysis --- p.30 / Chapter 1.5.5. --- Nucleic acid hybridization --- p.31 / Chapter 1.6. --- Prevention and control of rotavirus infection --- p.32 / Chapter 1.6.1. --- Host resistance --- p.32 / Chapter 1.6.2. --- Vaccine development --- p.33 / Chapter 1.6.3. --- Passive immunization --- p.35 / Chapter 1.7. --- Objectives of this study --- p.36 / Chapter Chapter 2. --- Materials and methods / Chapter 2.1. --- Materials --- p.38 / Chapter 2.1.1. --- Reagents for tissue culture work --- p.38 / Chapter 2.1.2. --- Reagents for electropherotyping --- p.39 / Chapter 2.1.3. --- Reagents for ELISA --- p.42 / Chapter 2.1.4. --- Reagents for hybridization assay --- p.43 / Chapter 2.2. --- Methods / Chapter 2.2.1. --- Collection of specimens --- p.46 / Chapter 2.2.2. --- Rotavirus strains --- p.46 / Chapter 2.2.3. --- Monoclonal antibodies --- p.48 / Chapter 2.2.4. --- Detection of rotavirus antigen by ELISA --- p.49 / Chapter 2.2.5. --- Rotavirus electropherotyping --- p.50 / Chapter 2.2.6. --- Serotyping of rotavirus by fluorescent foci neutralization --- p.52 / Chapter 2.2.7. --- Serotyping of rotavirus by oligo- nucleotide probes hybridization --- p.55 / Chapter 2.2.8. --- Rotavirus serotyping by ELISA --- p.59 / Chapter 2.2.9. --- Rotavirus subgrouping by ELISA --- p.61 / Chapter Chapter 3. --- Results / Chapter 3.1. --- Age and sex distribution of the study population --- p.63 / Chapter 3.2. --- Detection of rotavirus by ELISA --- p.63 / Chapter 3.3. --- Seasonal distribution of rotavirus infections --- p.67 / Chapter 3.4. --- Genetic diversity of human rotaviruses --- p.74 / Chapter 3.5. --- Subgroup determination by ELISA --- p.99 / Chapter 3.6. --- Rotavirus serotypes by fluorescent foci neutralization (FFN) --- p.102 / Chapter 3.7. --- Rotavirus serotypes by ELISA --- p.107 / Chapter 3.8. --- Rotavirus serotypes as determined by oligonucleotide probes --- p.110 / Chapter 3.8.1. --- Dot hybridization --- p.110 / Chapter 3.8.2. --- Northern blots of electrophoresed RNAs --- p.118 / Chapter 3.9. --- Temporal distribution of rotavirus serotypes --- p.124 / Chapter 3.10. --- "Association between serotype, subgroups and electropherotypes" --- p.128 / Chapter 3.11. --- Unusual rotavirus strains --- p.135 / Chapter 3.12. --- Nosocomial rotavirus infection --- p.135 / Chapter Chapter 4. --- Discussion / Chapter 4.1. --- Seasonal variation of rotavirus infection --- p.141 / Chapter 4.2. --- Molecular epidemiology of rotavirus infection --- p.143 / Chapter 4.3. --- Subgrouping of human rotavirus strains --- p.146 / Chapter 4.4. --- Serotyping rotaviruses by ELISA --- p.147 / Chapter 4.5. --- Serotyping rotaviruses by oligonucleotide probe hybridization --- p.150 / Chapter 4.6 --- Advantage and disadvantage of different methods for serotyping of rotaviruses --- p.152 / Chapter 4.7. --- Distribution of rotavirus serotypes --- p.153 / Chapter 4.8. --- Association between rotavirus serotype and electropherotype --- p.156 / Chapter 4.9. --- Nosocomial rotavirus infection --- p.158 / Chapter 4.10. --- Unusual rotavirus strains --- p.159 / Chapter 4.11. --- Concluding remark --- p.162 / Chapter 4.12. --- Future studies --- p.164 / References --- p.165

Muitas doenças genéticas permanecem ainda sem sua causa definida. A dificuldade de estudar algumas dessas doenças remontam a problemática da obtenção de material clinico, sugerindo situações extremas para a coleta, dependendo da patologia. Além disso, é necessário que se obtenha material genético em quantidade adequada para os ensaios e que, de preferência, venha de uma fonte celular que não esteja diretamente exposta a mutações. Nesse trabalho estabelecemos o cultivo inédito das células epiteliais de mucosa oral como modelo celular para o estudo de doenças genéticas. As amostras foram coletadas através da raspagem da mucosa oral de voluntários saudáveis. Após estabelecimento do cultivo, as células foram caracterizadas em relação à sua morfologia através de técnica de colorações, ensaios para analisar a viabilidade celular, microscopia eletrônica de transmissão e varredura, analise da expressão de marcadores por imunocitoquímica e por RT-PCR. Os resultados revelaram a expressão de marcadores como citoquetaratinas 4, 13 e 18, conexinas e marcadores de ciclo celular, como PCNA3 e GAPDH. A expressão de marcadores de pluripotência foi negativa, já que essas células são adultas e totalmente diferenciadas. Com a realização deste trabalho, concluímos que essas células são um bom modelo para o estudo de doenças genéticas e futuras terapias gênicas. / Many genetic diseases are still without their cause defined. The difficulty in studying these diseases back to the problem of obtaining clinical material, suggesting extreme situations for the collection, depending on the pathology. Moreover, it is necessary to obtain genetic material in sufficient quantities for testing and preferably come from a cellular source that is not directly exposed to mutations. This work established for the first time, a protocol for culturing of epithelial cells from oral mucosa as a cellular model for studying genetic diseases. The samples were collected by scraping the oral mucosa of healthy volunteers. After the establishment of cultivation, cells were characterized for their morphology by staining technique, tests to analyze cell viability, transmission electron microscopy and scanning, analysis of expression of markers by immunocytochemistry and RT-PCR. The results revealed the expression of markers such as citoquetaratinas 4, 13 and 18, connexins and cell cycle markers, as PCNA3 and GAPDH. The expression of pluripotency markers were negative, as these cells are adult and fully differentiated. With this work, we conclude that these cells are a good model for studying genetic diseases and future gene therapies.

There is a large variety of diseases caused by single-gene mutations. Although most of these conditions are rare, together they impose a significant burden to the population. This thesis describes clinical and genetic studies of two single-gene diseases: 1) Adult-onset autosomal dominant leukodystrophy with autonomic symptoms (ADLD) caused by LMNB1 gene duplications, and characterized by autonomic, pyramidal and cerebellar symptoms. Spinal cords of patients with ADLD were studied by MRI and found to be thin, with high signal intensity in white matter. Histopathology showed loss of myelinated fibres with some reactive gliosis. DNA samples from four different families with ADLD were obtained, and the LMNB1 gene was screened for duplications. Single nucleotide polymorphism array revealed LMNB1 duplications in all ADLD families. LMNB1 mRNA and protein levels were assessed in white blood cells using quantitative polymerase chain reaction and Western blot, and increased levels of LMNB1 mRNA and lamin B1 protein could be demonstrated. We concluded that spinal cord atrophy in patients with ADLD is a valuable differential diagnostic sign, and that increased levels of LMNB1 can be detected in peripheral blood. 2) Rippling muscle disease (RMD) is caused by CAV3 gene mutations. Clinical features are percussion-induced muscle mounding, –rapid contractions and undulating muscle contractions (rippling). The CAV3 gene was sequenced in 38 members of a family with RMD. Twenty-two individuals had clinical features of RMD. No muscle weakness was seen. All patients with signs of RMD carried the p.A46T CAV3 mutation, showing that the p.A46T mutation was benign and that the diagnosis can be made clinically. In vitro contracture test results from 10 of the subjects were collected, but no association between pathological test results and RMD was found.

Darbo tikslas ir uždaviniai: Surinkti ir išanalizuoti informaciją apie esančius Lietuvos kačių registrų duomenų bazes; Ištirti kačių veislių populiarumą Lietuvoje; Ištirti kačių skaičiaus kitimo tendencijas populiariausiose veislėse 2009 – 2013 m.; Ištirti kokios dažniausiai pasitaiko genetinės ligos veisiant kates ir kokiose veislėse. Tyrimo metodika: mokslinės literatūros analizė, statistinių duomenų rinkimas ir apdorojimas. Rezultatai ir išvados: Nustatyta, kad per 2009 – 2013 metus, buvo užregistruoti 34618 gyvūnai. Lietuvoje buvo užregistruotos 7297 katės iš kurių 4198 priklausė dešimčiai populiariausių kačių veislių. RegiVet duomenų bazėje per penkis metus buvo užregistruotos 3409 katės. Vilniaus miesto gyvūnų duomenų bazėje užregistruotos 711 katės. 15 kačių veislynuose, per penkis metus buvo išvestos 372 katės. Tyrimo metu buvo atrinktos populiariausios kačių veislės ir sudarytas šių veislių dešimtukas. 2009 – 2013 metais dažniausiai aptinkama kačių veislė buvo Persų katės. Taip pat vienos iš populiariausių kačių buvo Britų trumpaplaukės, Siamo, Kornvalio reksų ir Egzotų katės. Per tiriamuosius metus mažiau populiarios katės buvo Meino meškėnai, Rusų mėlynosios, Abisinijos, Bengalijos, bei Turkų angoros veislių katės. Buvo nustatytos veislės, kurioms prognozuojama skaičiaus didėjimo tendencija, tai Persų, Britų trumpaplaukėms ir Siamo veislių katėms. Taip pat nustatytos veislės kurioms prognozuojama nežymi skaičiaus didėjimo tendencija, tai... [toliau žr. visą tekstą] / Research methodology: Analysis of scientific literature, scientific articles, and statistical data. Different articles and research material offering information on the issue under consideration were used as the basis for the research. Results and conclusions: It was found that during the 2009 - 2013 year has registered 34618 animals. Lithuania was recorded in 7297 from a 4198 cat belonged to the ten most popular cat breeds. RegiVet database within five years was recorded in 3409, cat. Vilnius city animal database recorded 711 cats. 15 cattery over five years has been derived 372 cats. The study has been selected for the most popular breed of cat and consists of a dozen varieties. 2009 - 2013, the most common breed of cat has been found in Persian cats. It is also one of the most popular British Shorthair cat was, Siamese, Cornish rex and Exotic cats. Over the years, research has been less popular cat Maine Coon, Russian Blue , Abyssinian, Bengal, and the Turkish Angora cat breed. Varieties have been identified which predicted increase in the number of trendsetting is Persian, British shorthair and Siamese cats. It has also established a variety of which predicted a slight increase in the number of trendsetting, the Cornish Rex, Maine Coon, Exotic, and the Bengal cat breed. And set out a variety of monitored depopulation trendsetting, this Russian Blue, Abyssinian, Turkish Angora cat breed. The study showed the likelihood of genetic diseases in the most popular cat breeds... [to full text]

Diss. (sammanfattning)--Linköping : Linköpings universitet, 2006. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Washington, 1998. / Vita. Includes bibliographical references (p. [119]-127).

Akademisk avhandling--Uppsala. / Extra t.p., with thesis statement, inserted. Bibliography: p. 55-60.

Muitas doenças genéticas permanecem ainda sem sua causa definida. A dificuldade de estudar algumas dessas doenças remontam a problemática da obtenção de material clinico, sugerindo situações extremas para a coleta, dependendo da patologia. Além disso, é necessário que se obtenha material genético em quantidade adequada para os ensaios e que, de preferência, venha de uma fonte celular que não esteja diretamente exposta a mutações. Nesse trabalho estabelecemos o cultivo inédito das células epiteliais de mucosa oral como modelo celular para o estudo de doenças genéticas. As amostras foram coletadas através da raspagem da mucosa oral de voluntários saudáveis. Após estabelecimento do cultivo, as células foram caracterizadas em relação à sua morfologia através de técnica de colorações, ensaios para analisar a viabilidade celular, microscopia eletrônica de transmissão e varredura, analise da expressão de marcadores por imunocitoquímica e por RT-PCR. Os resultados revelaram a expressão de marcadores como citoquetaratinas 4, 13 e 18, conexinas e marcadores de ciclo celular, como PCNA3 e GAPDH. A expressão de marcadores de pluripotência foi negativa, já que essas células são adultas e totalmente diferenciadas. Com a realização deste trabalho, concluímos que essas células são um bom modelo para o estudo de doenças genéticas e futuras terapias gênicas. / Many genetic diseases are still without their cause defined. The difficulty in studying these diseases back to the problem of obtaining clinical material, suggesting extreme situations for the collection, depending on the pathology. Moreover, it is necessary to obtain genetic material in sufficient quantities for testing and preferably come from a cellular source that is not directly exposed to mutations. This work established for the first time, a protocol for culturing of epithelial cells from oral mucosa as a cellular model for studying genetic diseases. The samples were collected by scraping the oral mucosa of healthy volunteers. After the establishment of cultivation, cells were characterized for their morphology by staining technique, tests to analyze cell viability, transmission electron microscopy and scanning, analysis of expression of markers by immunocytochemistry and RT-PCR. The results revealed the expression of markers such as citoquetaratinas 4, 13 and 18, connexins and cell cycle markers, as PCNA3 and GAPDH. The expression of pluripotency markers were negative, as these cells are adult and fully differentiated. With this work, we conclude that these cells are a good model for studying genetic diseases and future gene therapies.

Familial hypercholesterolemia (FH) is an autosomal dominant disorder in which the primary defect is a mutation in the LDL receptor. Heterozygous FH is among the most common inborn errors of metabolism and remains as the best example of an inherited defect causing premature coronary artery disease (CAD). This thesis describes the physical and biochemical characteristics of heterozygous FH in a large cohort consisting of 208 women and 156 men. The influence of both genetic and environmental factors on the clinical expression of FH were investigated to better understand the phenotypic variation within FH and thus improve the prediction and treatment of CAD in affected individuals. The general incidence of CAD in this population was lower compared to previous reports but the differences between the sexes were expected. It was shown that men had a much higher frequency of CAD (31%) compared to women (13%) despite having lower concentrations of total and LDL cholesterol. In addition, the average age of onset of coronary symptoms was delayed in females, 55 years compared to 48 years for males. A greater risk of developing CAD for men was associated with low levels of HDL cholesterol and a history of smoking. In women, however, CAD was associated with elevated triglyceride levels and the presence of hypertension. In order to efficiently assess the influence of the co-inheritance of the apolipoprotein E polymorphism in this large FH population, a novel apo E phenotyping procedure was developed. Phenotypes were determined directly from plasma which was neuraminidase treated, delipidated and focused in polyacrylamide minigels. The accuracy of this method was confirmed by making a comparison to the established procedure of phenotyping by isoelectric focusing of delipidated VLDL. The low cost, speed and simplicity of the minigel methodology provided ideal conditions to phenotype a large patient population. The frequencies of the ɛ2, ɛ3 and ɛ4 alleles of apolipoprotein E in 125 unrelated FH subjects did not differ significantly from the normal population. In addition, there was no apparent relationship between apo E4 and the concentration of any of the parameters in the plasma lipid profile. However, the presence of the E2 isoform was associated with significantly elevated triglycerides in both sexes. From this study, it is evident that the mutant FH gene exerts its effect within a system of interacting environmental and polygenic factors that are known to modify atherosclerotic risk. It has been established that the dissimilarity in the frequency of CAD between men and women is related to differences between the impact of known risk factors and the incidence of CAD. Therefore, the importance of the influence of these risk factors and the differences between men and women should be emphasized when treating and predicting the development of CAD in patients with FH. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19625号 / 医博第4132号 / 新制||医||1015(附属図書館) / 32661 / 京都大学大学院医学研究科医学専攻 / (主査)教授 高田 穣, 教授 松田 文彦, 教授 小泉 昭夫 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM