No description available.

No description available.

No description available.

by Au Sze Ki. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves xii-xix). / Acknowledgement --- p.i / Table of Content --- p.ii / List of Abbreviation --- p.vi / Abstract --- p.viii / Chapter Chapter One : --- Introduction / Chapter 1.1. --- Osteosarcoma --- p.1 / Chapter 1.1.1. --- Definition --- p.1 / Chapter 1.1.2 --- "Incidence, geographic patterns of distribution and epidemiological consideration" --- p.1 / Chapter 1.1.3 --- "Age, sex and sites" --- p.3 / Chapter 1.1.4 --- Type and grade --- p.5 / Chapter 1.1.4.1. --- Grade --- p.5 / Chapter 1.1.4.2. --- Site --- p.5 / Chapter 1.1.4.3. --- Metastasis --- p.5 / Chapter 1.1.5 --- Histological features --- p.9 / Chapter 1.1.6 --- Clinical features --- p.9 / Chapter 1.1.7 --- Radiological features --- p.11 / Chapter 1.1.8. --- Molecuar genetics --- p.13 / Chapter 1.1.9 --- Treatment --- p.13 / Chapter 1.2 --- Biochemical Markers of Osteosarcoma --- p.14 / Chapter 1.2.1 --- Tumor marker --- p.14 / Chapter 1.2.2 --- Biochemical markers of bone turnover --- p.15 / Chapter 1.2.3 --- Change of biochemical marker in osteosarcoma --- p.17 / Chapter 1.3 --- Alkaline Phosphatase (ALP) --- p.17 / Chapter 1.3.1 --- ALPs Family --- p.20 / Chapter 1.3.2 --- Membrane binding --- p.22 / Chapter 1.3.3 --- Biochemical function and physiological role of ALP --- p.24 / Chapter 1.4 --- Normal values of serum ALP --- p.28 / Chapter 1.5 --- Clinical applications of ALP --- p.28 / Chapter 1.6 --- "Separation, identification and quantification of ALP isozymes" --- p.31 / Chapter 1.6.1 --- Themostability --- p.31 / Chapter 1.6.2 --- Inhibition studies --- p.31 / Chapter 1.6.3 --- Electrophoresis --- p.33 / Chapter 1.6.4 --- Isoelectric focusing --- p.34 / Chapter 1.6.5 --- Affinity precipitation --- p.34 / Chapter 1.6.6 --- Immunological studies --- p.35 / Chapter 1.7 --- Plasma BALP level as biochemical marker of osteosarcoma --- p.35 / Chapter 1.8 --- ALP in malignancies --- p.37 / Aim of study --- p.x / Chapter Chapter Two : --- Methods and Materials / Chapter 2.1 --- Plasma BALP measurement as a biochemical markerin osteosarcoma --- p.40 / Chapter 2.1.1 --- Patient groups --- p.40 / Chapter a) --- Normal subjects --- p.40 / Chapter b) --- Osteosarcoma patients --- p.40 / Chapter 2.1.2 --- Collection and preparation of patient bloods samples of patients --- p.40 / Chapter 2.1.3 --- Plasma total ALP measurement --- p.41 / Chapter a) --- Reagent --- p.41 / Chapter b) --- Procedure --- p.43 / Chapter 2.1.4 --- Plasma BALP measurements --- p.43 / Chapter a) --- Wheat germ lectin precipitation of BALP --- p.44 / Chapter i) --- Reagent / Chapter ii) --- Procedure / Chapter b) --- ABBOTT methods for plasma BALP activity measurement --- p.45 / Chapter c) --- COBAS MIRA methods for BALP measurement --- p.45 / Chapter d) --- ALKPHASE-B method of BALP measurement --- p.46 / Chapter 2.1.5 --- Inter-conversion of plasma BALP activity measurement in different methods --- p.47 / Chapter 2.1.6 --- Statistical analysis --- p.48 / Chapter 2.2 --- Alkaline phosphatase isozymes expression in human osteosarcoma --- p.48 / Chapter 2.2.1 --- In Vitro cultures of human SaOS-2 and U-2 OS osteosarcoma cell line --- p.48 / Chapter a) --- Reagent --- p.49 / Chapter b) --- Procedure --- p.50 / Chapter i) --- Storage of U-2 OS and SaOS-2 / Chapter ii) --- Subculture of confluent monolayer / Chapter 2.2.2 --- Protein assay --- p.51 / Chapter a) --- Standard Assay --- p.51 / Chapter i) --- Reagent / Chapter ii) --- Procedure / Chapter b) --- Mircoassay --- p.51 / Chapter i) --- Reagent / Chapter ii) --- Procedure / Chapter 2.2.3 --- Extraction of ALP from the cultured osteosarcoma cells --- p.52 / Chapter a) --- Reagent --- p.52 / Chapter b) --- Procedure --- p.52 / Chapter 2.2.4 --- "ALP extraction from human liver, placenta and osteosarcoma tissue" --- p.53 / Chapter a) --- Reagent --- p.53 / Chapter b) --- Procedure --- p.54 / Chapter 2.2.5 --- Isoelectric focusing of ALP --- p.55 / Chapter a) --- Preparation of the agarose IEF gel --- p.55 / Chapter b) --- Samples preparation --- p.56 / Chapter c) --- Isoelectric focusing --- p.57 / Chapter d) --- Protein detection --- p.59 / Chapter i) --- Reagent / Chapter ii) --- Procedure / Chapter e) --- Visualization of ALP isozyme --- p.60 / Chapter 2.2.6 --- Biochemical differentiation of ALP expressed in human osteosarcoma --- p.61 / Chapter a) --- Thermodenaturation of ALP --- p.61 / Chapter b) --- Ammino acid inhibition of ALP --- p.61 / Chapter 2.2.7 --- Immunohistostaining of placental ALP in human Osteosarcoma --- p.62 / Chapter a) --- Reagent --- p.62 / Chapter b) --- Preparation of human osteosarcoma cell line --- p.63 / Chapter c) --- Preparation of human osteosarcoma tissue --- p.63 / Chapter d) --- Immunohistostaining --- p.64 / Chapter Chapter Three : --- Results / Chapter 3.1 --- General information of the patients --- p.65 / Chapter 3.1.1 --- Age and sex distribution --- p.65 / Chapter 3.1.2 --- Sites --- p.65 / Chapter 3.1.3 --- Treatment and survival rate --- p.66 / Chapter 3.2 --- Clinical significance of plasma bone-specific alkaline phosphatase (BALP) activity measurementin osteosarcoma patients --- p.71 / Chapter 3.2.1 --- Plasma BALP activity measurement --- p.71 / Chapter 3.2.2 --- Normal reference of plasma BALP determination --- p.72 / Chapter 3.2.3 --- Diagnostic value of plasma BALP measurement in osteosarcoma --- p.75 / Chapter a) --- Plasma BALP level at admission --- p.75 / Chapter b) --- Plasma Total ALP level at admission --- p.78 / Chapter 3.2.4 --- Prognosis value of plasma BALP measurement in osteosarcoma Patients --- p.78 / Chapter a) --- Correlation of plasma BALP-Adm with the local relapse of the disease --- p.78 / Chapter b) --- Correlation of plasma BALP-Adm with survival rate of the patients --- p.90 / Chapter i) --- One year survival Rate / Chapter ii) --- two-year survival Rate / Chapter iii) --- Three-year survival rate / Chapter c) --- Correlation of the plasma BALP-Adm with the tumor volume --- p.90 / Chapter 3.2.5 --- Using plasma BALP measurement for monitoring of the disease --- p.91 / Chapter a) --- Effectiveness of pre-operative chemotherapy --- p.91 / Chapter b) --- Change of plasma BALP level during the treatment --- p.92 / Chapter i) --- Monitoring of pre-operative chemotherapy / Chapter ii) --- Detection of local recurrence and secondary metastasis / Chapter 3.3 --- Alkaline phosphatase isozyme expressionin osteosarcoma --- p.103 / Chapter 3.3.1 --- Isoelectric point (pI) gradient in isoelectric focusing (IEF)gel --- p.10? / Chapter 3.3.2 --- ALP isozyme standard --- p.103 / Chapter 3.3.3 --- Ectopic expression of ALP in human osteosarcoma cell line: U-2 OS and SaOS-2 --- p.104 / Chapter a) --- Isoelectric focusing separation --- p.104 / Chapter b) --- Biochemical differentiation of ALP extracts --- p.110 / Chapter 3.3.4 --- Alkaline phosphatase expression in osteosarcoma patient plasma sample --- p.110 / Chapter 3.3.5 --- Alkaline phosphatase isozyme expression in human osteosarcoma biopsy tissue --- p.111 / Chapter 3.3.6 --- Ectopic expression of placental ALP in human osteosarcoma by immunohistochemistry --- p.111 / Chapter a) --- Ectopic expression of placental ALP in human osteosarcoma cell line U-2 OS --- p.111 / Chapter b) --- Ectopic expression of placental ALP in human osteosarcoma tissue sections --- p.112 / Chapter Chapter Four : --- Discussion --- p.128 / Chapter Chapter Five : --- Conclusion --- p.142 / Bibliography --- p.xii / Appendix --- p.xx

Resumo: Vassoura-de-bruxa e podridão de Moniliophthora, causadas pelos fungos Crinipellis perniciosa e Moniliophthora roreri, respectivamente, são as doenças de maior impacto econômico da cultura do cacau e estão presentes na maioria dos países produtores do Continente Americano. Evidências biológicas e moleculares comprovam que estes fitopatógenos estão intimamente relacionados. O uso de resistência genética através de dones resistentes de cacaueiro, é a medida mais eficiente no controle destas doenças. O conhecimento sobre as populações destes fungos é importante na geração de informações para o programa de melhoramento genético do cacau visando resistência. Marcadores moleculares RAPO e SSR foram usados para analisar a estrutura genética de populações destes fitopatógenos. No geral, as populações do Brasil, Equador, Peru e Trinidad agruparam-se de acordo com o país de origem, apresentando maior variabilidade dentro e não entre países, com presença de subpopulações. A população do Brasil apresentou maior diversidade genotípica em comparação com as demais. A transferibilidade de pares de primers SSR de C. perniciosa para M. roreri foi satisfatório. Populações de M. roreri do Equador e Peru apresentaram alta diferenciação genética interpopulacional, sendo que a do Peru apresentou maior variabilidade. / Abstract: Witches' broom and fresty pod hot, caused by Crinipellis perniciosa and Moniliophthora roreri, are the most important disease of cacao in the American Continent. Biological and molecular data have shown that these pathogen are closely related. Resistance is the most efficient method to control these diseases. Therefore, information about the population structure of these cacao pathogen are important to support the breeding programo Molecular markers such as RAPD and SSR were used to analyzed the genetic structure of C. perniciosa and M. roreri frem the American Continent. Populations of C. perniciosa clustered according to their country of origin, with more variability within than between countries, revealing the presence of subpopulations. C. perniciosa Brazilian populations presented higher genotypic diversity than C. perniciosa from other countries. The transferability of C. perniciosa-SSR to M. roreri was positive. On the contrary, high interpopulation variability was observed between Ecuador and Peru, being M. roreri from Peru much more diverse than Ecuador. / Orientador: Carlos Ruggiero / Coorientador: Karina Peres Gramacho / Banca: João Carlos de Oliveira / Banca: Antonio de Goes / Banca: Maria Lúcia Carneiro Vieira / Banca: João Alexio Scarpare Filho / Doutor

Submitted by Marlene Santos (marlene.bc.ufg@gmail.com) on 2014-10-03T21:12:43Z No. of bitstreams: 2 Dissertação - Adriana Helena de Matos Abe - 2011.pdf: 852814 bytes, checksum: 8db7f03ca663f93c40dd3a61fd3c27fe (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Jaqueline Silva (jtas29@gmail.com) on 2014-10-03T21:37:01Z (GMT) No. of bitstreams: 2 Dissertação - Adriana Helena de Matos Abe - 2011.pdf: 852814 bytes, checksum: 8db7f03ca663f93c40dd3a61fd3c27fe (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2014-10-03T21:37:01Z (GMT). No. of bitstreams: 2 Dissertação - Adriana Helena de Matos Abe - 2011.pdf: 852814 bytes, checksum: 8db7f03ca663f93c40dd3a61fd3c27fe (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2011-04-29 / Introduction: Dengue is the most important emerging and reemerging disease in morbidity and mortality today. Among those affected. 2.5% die, and is increasing the number of deaths in children annually. In this group, in particular, early diagnosis is more difficult. Objectives: To identify clinical and laboratory markers for diagnosis and severity of dengue infection cases reported in the period 2001 to 2009 in children under 15 years old living in the city of Goiania, laboratory confirmed, describe sociodemographic data, tourniquet test and admissions performed. Methodology: Database Information System for disease surveillance of the City Health Department to Goiania was used to do a retrospective, descriptive, exploratory quantitative analysis, using the test of statistical significance t-student test, with significance level 5% and the final statistical analysis in SPSS 17.0. Results: In the period 2001 to 2009 were reported in the city of Goiania, 124,794 cases of dengue, of these 113,744 (91%), referring to residents of the capital. Among those under 15 years reported in this period, 22,278 were residents of Goiania. The tourniquet test was performed in 7428 (33.34%) of these children, 1,415 (6.36%) reported not having performed this test. 13,435 (60.30%), had this spot in blank and 1,222 (16.45%) were described as positive. Were documented 1043 hospitalizations of children in this period. Laboratory confirmation occurred in 10,756 cases. As a final classification was found 77% of Classic Dengue, Dengue with 7.4% of complications, 0.4% of Dengue Hemorrhagic Fever and 0.14% deaths, 15% were not classified. Among the laboratory markers found were registered 1,209 cases with IgM positive cases em108 DEN 1, DEN 2 in 14 cases and DEN 3 in 67 cases, Histopathology was cited positive in 4 cases, the hematocrit ranged from 21% to 81%, among the sociodemographic markers the race / ethnicity most marked in this population was white, children over 11 years were most affected, students complete basic education. The more clinical markers reported were fever, headache. myalgia, prostration; among hemorrhagic manifestations: petechiae and epistaxis; as signs of vascular leakage, ascites, pleural and pericardial effusion; warning sign most frequently reported were abdominal pain; the complications of gravity as myocarditis, shock and neurological manifestations have been reported among others. DEN 2 was the responsible for the highest number of severe symptoms. Conclusions: Were identified clinical markers for diagnosis and severity for Dengue in the period studied. The population of children showed severe signs of vascular leakage and neurological impairment. Despite the incompleteness of some data, it did not limit the study, which serves as the basis for a greater knowledge about dengue in children in this region. / Introdução: A dengue é a doença emergente e reemergente mais importante em morbidade e mortalidade na atualidade. Dentre os acometidos 2,5% morrem, e é crescente o número de óbitos em crianças anualmente. Neste grupo, em particular, o diagnóstico precoce é mais difícil. Objetivos: Identificar os marcadores clínicos e laboratoriais de diagnóstico e gravidade da infecção pelo vírus dengue nos casos notificados no período de 2001 a 2009 em menores de 15 anos residentes no município de Goiânia, confirmados laboratorialmente, descrever dados sócio demográficos, prova do laço e internações realizadas. Metodologia: Foi utilizado o Banco de Dados do Sistema de Informação de Agravos de Notificação da Secretaria Municipal de Saúde de Goiânia para um estudo retrospectivo, descritivo, exploratório de análise quantitativa, utilizando o teste de significância estatística t-student, com nível de significância de 5% e análise estatística final no programa spss 17.0. Resultados: No período de 2001 a 2009 foram notificados no município de Goiânia, 124.794 casos de Dengue, destes 113.744 (91%), referentes a moradores da capital. Dentre os menores de 15 anos notificados neste período, 22.278 eram residentes da Goiânia. A prova do laço foi realizada em 7.428 (33,34%) destas crianças, 1.415 (6,36%) informaram não ter realizado este exame. 13.435 (60,30%), tinham este campo em branco e 1.222 (16,45%) foram descritas como positiva. Foram documentadas 1.043 internações de crianças neste período. A confirmação laboratorial ocorreu em 10.756 casos. Como classificação final encontrou-se 77% de Dengue Clássica, 7,4% de Dengue com Complicações, 0,4% de Febre Hemorrágica da Dengue e 0,14% de óbitos, 15% não foram classificados. Dentre os marcadores laboratoriais encontrados foram registrados 1.209 casos com 1001 positiva, DEN 1 em108 casos, DEN 2 em 14 casos e DEN 3 em 67 casos, Histopatologia citada positiva em 4 casos; o hematócdto variou entre 21% a 81%, dentre os marcadores sócio demográficos a raça/etnia mais assinalada nesta população foi a branca, crianças maiores de 11 anos foram mais atingidas e alunos do ensino fundamental completo. Os marcadores clínicos mais reportados foram febre, cefaléia, mialgia, prostração; dentre as manifestações hemorrágicas: petéquias e epistaxe; como sinais de extravasamento vascular, ascite, derrame pleural e pericárdico; sinal de alerta relatado com maior frequência foi dor abdominal; as complicações de gravidade como miocardite, choque e manifestações neurológicas foram reportadas dentre outras. DEN 2 foi responsável pelo maior número de sintomas graves. Conclusões: Foram identificados marcadores clínicos de diagnóstico e gravidade para Dengue no período estudado. A população infantil apresentou sinais graves de extravasamento vascular e comprometimento neurológico. Apesar da incompletude de alguns dados, a mesma não limitou o estudo, que serve como base para um maior conhecimento sobre a Dengue em crianças nesta região.

Abstract available in the Journal of Strength and Conditioning Research.

Both small molecules and antibodies are powerful tools for research in biological mechanisms and therapeutics. The discovery of such molecules involves two opposite starting points: one being specific targets and the other being phenotypic screens. The first part of this thesis focuses on drug development starting with a specific target. The second part of this thesis focuses on identification of ferroptosis biomarkers by phenotypic screen. The specific target highlighted in the first part of this thesis is KRAS (Kirsten rat sarcoma viral oncogene homolog), the most commonly mutated
 oncogene in human pancreatic cancers, colorectal cancers, and lung cancers. The high prevalence of KRAS
mutations and its prominent role in many cancers make it a
potentially attractive drug target; however, it has been difficult
 to design small molecule inhibitors of mutant K-Ras proteins. Here, we identified a putative small molecule binding site on
K-RasG12D, which we have termed the P110 site (due to its adjacency to proline 110), using computational analyses of the protein structure. We then confirmed that one compound, named K-Ras Allosteric Ligand KAL-21404358, might bind to the P110 site of K-RasG12D using a combination of computational
 and biochemical approaches. The phenotypic screen used in the second part of this thesis focus on the process of ferroptosis, a form of regulated cell death process driven by the iron-dependent accumulation of polyunsaturated-fatty-acid-containing phospholipids (PUFA-PLs). Currently, there is no way to selectively stain ferroptotic cells in tissue sections to characterize relevant models and diseases. To circumvent this problem, we immunized mice with membranes from diffuse large B Cell lymphoma (DLBCL) cells treated with piperazine erastin (PE), and screened the generated monoclonal antibodies. The results suggested that for the first time we could detect cells undergoing ferroptosis in human tissue sections. In summary, these two projects illustrate how molecular screening and design starting from either a specific target or a phenotype screen aid in drug and biomarker development.

No description available.

Recent theoretical developments have led to a renewed interest in the potential role of chromosomal rearrangements in speciation. Australian morabine grasshoppers (genus Vandiemenella, viatica species group) provide an excellent study system to test this potential role, because they show extensive chromosomal variation: 12 chromosomal races/species with parapatric distributions. The research in this thesis involves the application of molecular genetic analyses to examine patterns of gene introgression among chromosomal races of Vandiemenella at three different spatial scales: local-scale hybrid zone analysis, island-scale phylogeography, and continental-scale phylogeography. The aims of these multi-scale analyses are to investigate whether chromosomal races represent genetically distinct taxa with limited gene flow, and to infer the historical biogeography of Vandiemenella and evolutionary origins of their parapatric distributions. Karyotype and 11 nuclear markers revealed a remarkably narrow hybrid zone with substantial linkage disequilibrium and strong deficits of heterozygotes between the chromosome races P24(XY) and viatica17 on Kangaroo Island, suggesting that the zone is maintained by a balance between dispersal and selection against hybrids (tension zone). Selection that maintains the stable hybrid zone is unlikely to be operating only on loci linked to rearranged chromosomes. Island-scale and continental-scale phylogeography using multiple nuclear markers indicated that Vandiemenella chromosome races/species generally represent genetically distinct taxa with reduced gene flow between them. In contrast, analyses of a mitochondrial gene showed the presence of distinctive and geographically localised phylogroups that do not correspond with the distribution of the Vandiemenella taxa. These discordant population genetic patterns are likely to result from introgressive hybridization between the taxa and range expansions and contractions. Overall, our molecular analyses favour the allopatric mode of diversification for the evolution of Vandiemenella and do not support the stasipatric speciation model of White (1978). Patterns of genetic differentiation between the chromosomal races analysed at three different spatial scales show dynamic responses of the grasshoppers to past climatic fluctuations, leading to opportunities for long-term isolation and allopatric fixation of new chromosome variants and molecular mutations at many loci. Further analyses are necessary to assess potential roles of chromosomal rearrangements in facilitating diversification in Vandiemenella by reducing recombination within the rearranged chromosome segments.